DOCUMENT RESUME L - ERIC · POP DE. O. Jun 80, NOTE 250p.; Contains ... U.S DEPARTMENT OF EDUCATION NATIONAL INSTITUTE OF EDVCATION ... Laboratory: Proportional Counting. of Mixed

DOCUMENT RESUME L

ED 209 119 SE 035 928A .

TITLE' . Microscopic Analysis, of Activated Sludge. Training- Manual. IP

'INSTITUTION A Office of Viatr--4graik Operations (EPA)r, Cincin

.L'Ohio. FatipAil Training and Operational Technology ,

'. '''- Center. :',

3, FEPORT NO EPA-430/1-80-007 ,,

*,

POP DE Jun 80ONOTE 250p.; Contains,occasional light and broken type.

-Pages'1-198 ',Field Key to Some Genera of Algae', and. ',Ciliated isiotoZoa removed due.to copyright %

'restrictions.H AVAILABLE FROM EPA Indtructional'Reiources Center, 1200 Chambers

, Rd., 3rd,ylbor, Columbus, OH 432124$1.00 plus $0.:03. .

per Otge). .....

tr

.

EDRS PRICE..

MF01/PC10 Plus Postage. * .

DESCRIPTORS .Biological Stiences; Data Analysis; *LaboratoryEquipment; *Laboratory Procedures; AIMicrobiology; .

. - *Microscopes:, Postsecondary Education; *Water ,, 'Pollution; :water lesouices

'IDENTIFIERS Activated Sludge; Analytical Methods; *Waste WaterTreatment

a7tBSTRACT ---/-:*

4,

AThis trIiniA4 manual pwents material on the use Of

a compound microscope to analyze microscope communities, present inwastewater tteatient procefses, for operational control. Coursetopics includei sampling techniques, sample ha moiling, lairdratoryanalysis, iderftificatiot, of organisms, data interpretation,. and use'4 heecomOcund microscope: This manual contains 26 chaptersincluding reading material,- 'laboratory activities, 'and selectedreferences. Prior expgriende ip-microscopy is not necessary. (Co)

..

'

O

**********i***************************************4*************i.****** Reproductions suppliedrbEDRS atethe best that can be made,. *.--* fromIthe'original document.-**********,**************ts***********************Ig*******************

t °

United States National TrainingEnvironmental Protection and OperationalAgen Cy Technology Center

Cincinnati OH 45268

EPA-430/1-80-007June 1980 ,

Water

aEPA MicroscopicAnalysis of

or-4 Activated SludgerJ

Training ManualrA

S

(7-

A

U.S DEPARTMENT OF EDUCATIONNATIONAL INSTITUTE OF EDVCATION

EDUCATIONAL RESOURCES INFORMATIONCENTER (ERIC,

This document has been reproduced asreceived from the person or organizationonginatino a

XMinor changes have been made to improve

reproduction guat

4.Points of view of 01)16'10ns stated in this docu

mem do not necessarily represent official N1E

position or policy,

_IL J1_ __NJLLI __ __

4

li

N

4

S

.

%

EPA-430/1-80-007June-198Q

Microicopic Anasis of Activated Sludge

This course is for anyone who needs the skills touse a compound' microscope toanalyze microscopiccommunities, present in wastewater treatmentprocesses, for operational control. Prior experiencein microscopy 'is not necessary:.

Àfter successfully.completing the course; the studentwill be able to relate microscopic communities presentin the wastewater treatment process to operational .controls. The student will also be capable of instructingtreatril4nt plant personnel in the more proficient use of

. the compound mitroscope and re1ating communitiespresent to operational control.

4 The training, includes classroom instruction andlaboratory practice.

4'

U. S. ENVIRONMENTAL PROTECTION' GENCYOffice of, Water Program Operations

National Training and Operational Technology Center

f

A

1.

.,

4. r

DISCLAIMER

Reference to commercial products, trade names, or

manufacturers` is for_purposes of example and illustration.

Such reference's do not constitute ehdorgement by the

Officeof Water Program Operations, U. S. Environmental4201). .

Protection Agency.

z

0

,

CONTENTS

4

rf Title or Description -Outline Number

The Aquatic Environrdent

Classification of Communities, Ecosystems, .and Trophic LevelsLimnology and Ecology'of Plankton

Biology', of Zooplankton

Optics and the Microscope

Strubture and Function. of Cells

Bacteria and Protozoa as Tokicological Indicatorsiiamentous Bacteria

Fungi and the "Sewage Fungus" Community

Protozoa, Neniatodes, and Rotifers'Activated sludge Protozoa

Free-Living Amoebae and Nematodes

Animal Plankton

Preparation and Enumeration of Plankton in the LaboratoryAttached Growths

Effect of WaVewater Treatment Plant Effluent on Small StreamsEcology iol WaLste Stabilization Processes

Ecology Primer 18

The Laivs of Ecology

Application Orlifological Data

Signilicance of 9Limiting Factors" to POPulation Variation

Algae and Cultural Eutrophication

Calibration and Use of Plankton Counting Equipment

-

1

2

3

4

5

6

7

0

11 A

12

13

14sg

15

16

17

20

21

22

23

24. . t

25

26.

Laboratory: Proportional Counting. of Mixed Liquor .

Key to Selected' Groups of Freshwater Ant ....

,A Key for the Initial Separation o of Sortie C on Plankton Organismsi . .-

Field Key to Some Genera of Algae

Ciliated, Protozoa

r 5 /

1

Z 'AQUATICAQUATIC ENVIRONMENT

Part 1: e Nature and Behavior of Water

I INTRODUCTION

The earth is physic y divisible into thelithospherekor land ma ses, and thehydrosphereiwhich includes the oceans,lakes, stre(ms, and subterranean waters;and the atmosphere.

to Upon the hydrospere are based a numberof sciences which represent differentapproaches. Hydrology is the generalscience of water itself with its variousspecial fields suclkas hydrography, -hydri.tros, etc. These in turn mergeinto physical chemistry and chemistry.

B Limnology and Oceanography combineaspects of all of thegei and deal not oaly,with the physical liquid water and itsvarious naturally occurring solutions and

. forms, but also with living Organisms'and infinite interactions that occurbet hem and their environment.

C Water quality management, includingpollution control, thus looks to allbranches of aquatic science in effortsto coordinate and improve man'srelationship with his aquatic environment.

II SOME FACTS ABOUT WATER

AN.Water is the only abundant liquid on otaplanet( It has many properties mostunustial for liquids, upon which dependmost df the familiar aspebts of the worldabout us as we know it. ZSee Table 1)

TABLE 1

UNIQUE PROPERTIES OF WATER

.

Pro2erty_Highest heat capacity (specific heat) of anysulici or liquid (except NH3)

SignificanceStabilizes temperatures of oilanismstand

',geographical regions

Highest latent heat of fusion (except NH /3

Highest heat.of evaporation of any substance

The only substance that has its maximumdensity as a liquid (4°C)

-

1,

21g23. 80

t,

Thermostatic effect at freezing point

Important in heat and water transfer ofatmosphere

Fresh and brackish waters have maximumdensity abode freezing point. This isimpo ;tant in- vertical circulation patternin lager.

Highest surface tension of any liquid

Dissolves more substances in greaterquantity than any othei. liquid

Controls surface and drop phenomena.iniportant in cellular physiology .

Makes complex biological system possible.Important for transportation of materials

'in solution.

Pure water has the bfgheat di- electric,natant Of any liquid .

Leads to high dissociation of inorganicsubstances in solution .

Very little electrolytic dissaciat

Relatively transparent

Neutral, yet contains,both ft+ and OH ions

AbsOrbs much,energy In infra red and ultraviolet ranges, bta little in visible range.Bence ."color less" .

`,1

4

a

I

The Aquatic Envitomnent

i 4 a

B ysical-Factors of Significance

1 Water substance

Water is not, simply "H20" but inreality is a mixture of some 33different substances involving threeisotAeS each of hydrogen and oxygen(ordinary hydrogen 111,, deuteriumand tritium H3; ordinary oxygen al°,oxygen 17; and oxygen 18) plus 15known ty }es of ions. The moleculesof a water mass tend to associatethemselves as polymers 'rather thanto remain as discrete units.(See Figure 1) .

6

SUBSTANCE OF PURE WATER

Figure I

I

2 Density

a TeMperature and density: Ice.Water is the only known substancein which the solid.state will floaton the liquid state. (See Table 2)

(

TABLE 2

EFFECTS OF TEMPERATURE ON DENSITY- OF PURE WATER AND ICE*

Temperature (°C)

Water

Density

-10 .99815 .9397- 8 99869 .9360,-6 9991'2 .9020

.99945 .92772 .99970 .92,29

'0 .99987 - --)9168'2 .99997

4 1.000006 .99997

.9998816 .99973206: .9982340 .9922560 .9832480_ .97183

100 '" .195838

* *

Tabular values for density, etc.; representestimates brvarious workers rather thanabsolute values, due to the variability ofwater.

Regular Ice is knoivn as "ice I. Four ormore other "forms" of ice are known.tq

_exist (ice II, ice III, etc. ), having densities °

at 1 atm pressure ranging'from 1.1595 -

-,to 1.67. These are of.extremely restrictedoccurrence and. may be ignored in mostroutine operations. -

This epsures°That ice usuallyforms on top of a body of waterand tends'to insulate the renfain-ing water mass from fuither_lossof heat. Did ice sink, therecould be little or no carryover ofaquatic life from.season to seasonin-the higher latitudes. Frazil orneedle ice forms colloid fly at afew Thousandths of a- degree

-below CI° C. It is adhesiye aridmay build up on submerged objectsas "anchor ice's', but it is still

.typical-ice (ice I):

Seasonal increase in solarradiation annually warmssurface waters in summerwhile other factors result in .

winter cooling. The densitydifferences resulting establishtwo classic layer's: the epilimnion.or surface layer,' and thehypolimnion or lower layer, andin between.is the thermoclineor shear-plane.

2) While for certain theoreticalpurposes d'"thermocline" isdefined as a zone in which thetemperature changes onedegree centigrade for eachmeter of depth, in practice,any transitional layer betweentwo relatively stable massesof water of different temper-atures may be regarded as athermOcline.

3) Obviously the greater thetemperature differencesbetween epilimnion andhypolimnion and the sharperthe gradient in the thermOcline,the more stable will thesituation be.

4) From information given above,it should be evident that Whilethe temperature of thehypolimnion rarely dropsmuch below 4° C, theepilimnion may range from0° C upw' ard.

r 5) When epilimnion and hypolimnionachieve the same temperature,stratification no longer exists.

)3The entire body of water ehayeshydrologically asaunit, andtends to assume uniform chemicaland physical characteristics.Even a light breeze may then,cause the entire body of waterto circulate, Such events. are calledoverturns, and usually result in .water quality changes Of consider-able physical, chernicar, and

t.biological significance.

6)

The Aquatic Enviionmentt

Mineral-rich water from thehypolimnion, for example,,,,is mi.Red with oxygenatedwater from the epilimnion..This usually triggers asudden growth or.ubloom"of. plankton organisms.

war

When stratification is present,however, each layer belgiaves,relativelytindependently, andsignificant quality 'differences.may develop.

'7) Thermal stratification asdescribed above has noreference to the size of thewater mass; it is found Inoceans and puddles.

b The relative denSities of thevarious isotopes of waterinfluence its molecular com-position. For example, thelighter 016 tends to go offfirst in the process of evaporation,leading to the relative enrichmentof air by, 016 acid the enrichmettof water by 017 and 018. Thiscan lead to a measurably' higher018 content in warmer climates.Also, the temperature of waterin past geologic ages can beclosely estimated from the ratio.,"of 018 in the. carbonate of molluscshells.

c Dissolved and /or suspended solidsmay,,also affect the density ofnatural water masses (see Table ,3)

TAI3LE 3EFFECTS OF DISSOLVED SOLIDS

ON D ENSITY

Dissolved Solids(Grams,per liter)

Density(at 4°C)

0 1.00000'

1 1.00085

2 1.00169

3 1.00251

10 . 1.0081835 (mean for sea water) . 1.02'822

vo.

1

'I

The Aquatic Environment4

I

d Types-of density stratification ';-____

1) Dersity differences producestratification which may be

,_,permanent, transient, orseasonal.

, 2) 'Permanent stratificationexists'for example where

`there is a heavy mass Of'brine in the deeper areas ofa basin which does not respondto seasonal'or other changingconditions. t'

t ,

3) Transient stratification mayoccur with the recurrentinflux of tidal Water in anestuary,for example, or theoccasional influx of coldmuddy water into a deep lakeor reservoir.

.

4) Seasonal stratification istypically t rynal in nature,

f and thyolv the-annualestablisAm t of the epilimnion,hypolimnion, and thertnoclineas described above.

5) Density stratification is not'limited to two-layered systems;three, four, or ven morelayers may be encountered inlarger bodies of water.

e A "pltinge line' (sometimes called"thezmal line) niay develop at "'the mouth of a stream. Heavierwater flowing into>,a lake orreservoir plunges below the 'lighter Water mass ofthe epiliminiumto flow along at a lower level. Sucha line is usually'marked by anaccumulation of floating debris._

f Stratification may be modifiedor ,entirely -suppressed in somecases *hen deemed expedient, bymean- s of a simple. air lift.'

,3 The viscosity of Water is greater at

lower temperOures (see Table 4).

This is important not only in situationsinvolving the control of flowing wateras in a sand filter, but also since, -overcoining resistance to flow gen-erates heat, it is significant in theheating of water by internal frictionfrom wave and.current action.Living organisms more easily supportthemselves in the more viscous(and also denser) cold waters of thearctic than in the less vis cous warmwaters of the tropic. (See Table 4).

TABLE 4

VISCOSITY OF WAT R (In millipoises at 1 attn)

Temp. 0 C

pissted solids in g/L-

0 1 5 10 30

-10 26.0 1

..- 5. I21.4 7,--- ---- - - --

0 '17, 94 18,1 18.24 18.7

5 15/19 15.3 15.5 16.0

10 13 ,10 13.2 13.4 1.3.8:

30 8,00 8,1 8,2 8.6.100

4 Surface-tension has biological as,wellas physical-significance. Organismswhose body gurfaces cannot be wet bywater can either ride on the surfacefilm or in some instances may be"trapped" on the surface film andbeunable to re-enter the water.,

4. 5 :Heat or energy

Incident solar radiation is;the'primeSource of energy for virtually allorganic and most inorganic processeson earth. For the earth as a wholeth'e total amount (of energy) received

_annually must exactly balance thatlost by reflection and radiation intospace if climatic and related con-ditions are to remain relativelyconstant over geologic time.

9

a' For a given body of water,immediate sources of energyinclyle in addition to solarirradiation terrestrial, heat,transformation of kinetic energy(wave and current action) to heat,'chemical and biochemicalreactions, convection from theatmosphePe, and condensation of

ater vapor.

b the progortion'of light reflecteddepends on the angle of incidence,the temperature, color, and otherqualities of the water; and thepresence or absence of films,. ('

of lighter liquids such as oil.In general, "as the ddpth increasesarithmetically, the light fends todecrease geometrically. Blues,greens, and yellows tend topenetrate most deeply while ultraviolet, violets, and orange-redsare most quickly absorbed. On

,the order of 90% of the totalillUmination which penetrates thesurface film is absorbed in thefirst 10 meters of even the clearestwater, thus tending to warm theupper layer,s.

6 Water movements

a Waves or rhythmic movement

$

4) The best known are travelingwaves caused 'by wind. Theseeffective only against objects nearthe surface. They have little-effect on the movement of large

-masses of water.

are

2) Seiches t

Standingwaves seiches occurin lairs, estuaries, and other,e - ed bodies of water, but are

ge enough to beobserved. An "internal wave orseicli" is an oscillation in a,subrhersed mass of water suchasaa hypblimmon, accompaniedby compenSating oscillation in theoverlying Water so, thatdo,

0. V.

1 4 10.

SThe Aquatic Environment %

significant change in surface.level is detected. Shifts insubmerged water masses ofthis type can have severe effectson the biota and also on humanwater uses where withdrawalsare.confined toa given depth,Descriptions and analyses ofmany other types arid sub-typesof waves and wane -like movementsmay be found in the literature.

b Tides

1) Tides are the longest wavesknown, and are responsible forthe orrce or twice a day rythmicrise'and fall of the',Ocean levelon most shores, around the world.

2) Wylie part, and parcel of thesame phenomenon, it is'oftenConvenient to refer to the riseand fall of the water level as .

"tide, " and to the resultingA currents as "tidal currents. ".1

3) Tides are basically caused by theattraction of the sun and moon onwater masses, large and small;however, it is abnly in the oceansand peksibly certain of the largerlakes that time. tidal action hasbeen,demonstrated. The patternsortidal action are enormouslycomplicated by local, topography,

.interaction with seiches, ancrotherfa,ctors. The literature on tidesis very large.

'a

c Currents (except tidal currents) -

are steady arythrni-c water movelneAswhich have had maRir study only inoceanography although they are,most often observed, in rivers andstream's. 'They are "primarilyconcerned with the tranilocation ofwater masses. They May bp generated,internally by virtue of density changes,or externally by wind or terrestrialtopography,' Turbulence phenomenaor eddy currents are largely respbn-Sible. for, late rals.Mixing in a curtrent .

These are of far more importancein the econry of a body of water thanmere laminar flow.

.0

0 .

1-5 ,

S.

ti

The Aquatic Environment

;- 1-6

d °Coriolis force it a result of inter-action between the rotation of theearth, and the movement of massesor bodies on the earth. The netresult is a slight tendency for movingobjects to veer to the right.in thenorthern hemisphere, and to the\left in the southern hemisphere.While the result in fresh waters isusually negligible, it may be con-siderable in marine waters.. Forexample, other factors permitting,ifiebre is a tendenby in estuaries forfresh waters-to move toward theocean faster along the right bank,while salt tidal waterstend tointrude farther inland,along the.left bank. Effects are even' moredramaticin the open oceans.

e Langmuire spirals (or Langinuirecirculation) are a relativelymassive cylindrical motion imparte dto surface waters under_ the influenceof wind.' Thq axes of the cylindersare parallel to the .direction of the.wind, and their depth and velocity ere

depend on the depth of the water,the velocity and duration of thewind, and other factors. The netresult is that adjacent cylinderstend to rotate in oppOsite directionslike meshing cog wheels. Thus,the water between two given spinalsmaybe meeting and sinking, whilethat between spirjls on either sidewill be meeting and rising. Waterover the sinking, while that betweenspirals on either side will be meet-ing and rising:, Water over.thesinking areas tends to accumulateflotsam and jetsam on the surface'in long conspicuous lines. '

°a This phenomenon is of consider-able importance to those samplingforplankton (or even chemicals)near the surface when the. windis blowing. Grab samples fromeither dance might obviouslydiffer considerably, artkeif' a plankton tow is contemplated

t it should be made,,across thewind in order that thesnetmay pass Through a successionof both dances.

b y b-

-

WATER

SURFACE

WATER WATER

RiSitIG SINK1N,G

Figure 2. 'Langmuitilpiralsh.. Blue dna, water r.dance., water sinking, floating o

-swimming.,ohiects concentrated.I

..\\1

S

b Langmuire spirals. are notsually established until the

wind has either beenk blowingfor an ,extended period, or

, else,is blowing rather hard.Their presence can be detectedby the lines of foam andother floating material whichcoincide with the directionof the wind.

6 The pH of pure water hasheen deter-,mined between 5.7 and 7.01-by various.workers. The latter value is most

,,widely accepted at the present time.Natural waters of coarse vary widelyaccording to circumstances.

C The elements of hydrology mentionedabove represent a selecton of some ofthe more conspicuous physical factorsinvolved in working with water quality:Other }hems no spebificallif Mentionedinclude:. molecular structure of waters,interaction of water and radiation,internal pressur,e, acoustical charac-teristics, presVure-volume-temperature'

. relationships, refractivity, luminescence,color, dielectrical characteristics and

:phenomena, solubility, action and inter.-actions .of gases; liquids and 'solids,

. water vapor, phenomena of hydrostaticsand liy.drodynamics in general.

REFERENCES

The Aquatic Environment,

Buswell, A. M. and Rodebush, W: H.Water. Sci, Am. April 1956.

2 Dorsey, N. ,Ernest. Ribperties ofOrdinary Water Substance.Reinhold Publ. Corpew. York.pp. 1-673. 1940.

3 Fowle, Frederick E. SmithsonianPhyMcal XabIes. -SmithsonianMiscellaneous Collection, 71(1),7th revised ed., 1929.

4 Hutcheson, George:E. A Treatise onLimnology. John Wiley Company.

/1 .

1957.

°

4

,

.

/7.

.4

(

et

Vo

Part 2: The Aquatic Environment as an Ecosystem')

I INTRODUCTION

Part-1 introduced the lithosphere and thehydrosphere. Part 2 will deal with certaingeneral aspects of th.e biosphere, or thesphere of life on this earth, which photp_--graphs from space have showri is a finiteglobe in infinite spaces

This is the habitat of man and the otherorganisms. His relationships with tieaquatic biosphere are our common concern.

,II THE BIOLOGICA I., NATURE OF THE-WQRLD WE LIVE IN

4 A We can only imagine what'this worldmusthave been like before there was life:

B The" ircSrld as we know it is largely shapedby the forces of life.

1 Primitive forms of life created organicmatte?.-and estab/i'shed soil.

D Plants cover the lands and "enormouslyinfluehte the forces of erosion.

3 The nature and rate of 'erosion affect'the redistribution of materialE .

(and mass) on the Surface of -theearth (topographic changes).

4 Organismstie up vast quantities ofcertain chemicals, such as carbon'and oXygen. Ss,

.

5 Respiration of plants and animalsreleathes carbon dioxide to theatmosphere in influential quantities...

6 CO2

affects the heat transmisthori ofthe atmosphere.

C ,Organisms respond to and in turn affect- their environment.. Man is one a the

-'rhosi. influential.

I

III ECOLOGY IS THE STUDY OF THEINTERRELATIONSHIPS BETWEENORGANISMS, AND BETWEEN ORGA-NISMS AND THEIR ENVIRONMENT.

A The ecosystem is the basic functional'unit of ecology. Any area of nature thatinclud s ing organisms and nonlivingsubstances nteracting to produce an*exchange o material8 between the livihgand nonliv g pd.rtse,constitutes anecosystem. (Odurp, 1959) -

1 From a structural standpoint, it isconvenient to recognize fourcpnstituents as composing anecosystem (Figure.1).

a Abiotic NUTRIENT MINERALSwhich ape the physical stuff of,which living protoilasdi will besynthe.sized.

b utotrophic (self-nourishing) orPRODUCER organisms. Theseare largely the -green plants .-(holophytes), .but other minorgroups must also be included

..(See Figure 2). They assimilatethe nutrient, rpinerals,..by the useof conSiderable energy, and combinetheir into living organic, substance.'Heterotrophic (Ole r-nourishing)coNsulAgas (holozoic), are chieflythe animals. They ingest (or eat)and digest organic matter, releasingconsiderablk energy in the process.

C

. d Heterotrophic REDUCERS are chieflybUcteria and fungi that returncomplex organic compounds back tothe originaribiotic mineral condition,thereby reletising the remainingchemical energy.

2 From a functional standpoint, an ,I ecosystem 'ha.8 two parts (Figure 2)

1 -9


PRODUCERS

NUTRIENT. MINERALS

FIGURE 1

a The -autotrophic or producter,organisms, which utilize lightenergy or the oxidation of in-organic bompounds, as theirsole energy source.

b The heterotropic or consumerand reducer organisms which

eutiliies organic compounds forits energy and carbon requirements.

3 Unless the autotrophic and hetero-trophic phases of the cycle approximatea dynamic equilibrium, the ecosystem'and the environment will change. .

B Each dethesegtoups includes simple,single-celled representativei, persisting

4arr at lower levels on the evolutionary stemsof the higher organisms. (Figure 2)

1 These groups span the gaps between thehigher kingdoms with a multitude oftransitional forms: They are collectivelycalled.the PROTISTA and MONERA.

REDUCERS

2 These two groups can be `defined onthe basis of relative complexity ofstructure.

a The bacteria andblue-gieenlacking a nuclear membrane are .

the Monerit.

b The single-celled algae andprotozoa are Protista.

C Distrilpited throughout these groups willbe found most of the traditionalof classic biology.

IV

A

FUNCTIONING OF THE ECOSYSTEM

A food chain is the transfer of food energyfrom plants through a aeries of ,organismsWith repeated bating and being eaten.Food chains are rrot isolated sequences butaxe interconnected.

14

1

1

1

1

1

r "


RELATIONSHIPS BETWEEN FREE LIVING AQUATIC ORGANISMS

Energy'Flows from, eft to Right, Gal Evolutionary Sequence is Upward

PRODUC S I , CONSUMERS..OrganicNiaerial PrOdtced, Arganic Material Ingested orUsuallyN PhotosTifFirt Consumed.

Digested Ipternally

REDUCERSOrganic Material Reducedby Extracellular Digestionand Intracellular Metabolibth,to Mineral Condition

ENERGY STORED ENERGY RELEASED ENERGY RELEASED

Flowering Plants andGymnosperps

Club Mosses, Ferns4

Liverworts, Mosses

fylulticellular GreenAlgae

Red Algae

Brown Algae

Arachnids

Insects

Crustaceans

Segmented Worms

Molluscs

Bryoz oa

Rotifers

Roundworms

Flatworms

Sponges

Mammals

Birds

Reptiles

Amphibians

Fishes

PrimitiveC'bordates

Echinoderms

Coelenterates

Basidiomycetes

Fungi Imperfecti.

Ascomycetes

Higher Phycomycetes

DEVELOPMENT OF NIULT1C'ELLULAR OR COENOCYTIC STRUCTURE.

Unicellular Green Algae

Diatoms

Pigmented Flagellates

A nioet;oic.1

PROTISTAP r o 10 v 0 a

Flagellated, Suctoria -(nnii-pigmented)'

Lower

Phycoznycetes

(Chytridiales, et. al. )'

DEVELOPMENT OF A NUCLEAR MEMBRANE

Blue Green Algae

Phototropic Bacteria

Chenwtropie Bacteria

131. ECO. pl. 2a. 1.69

MONERA

1-4

I

Saprophyt lcBacterial

e s -

I

FIGURE 2 .

Actinomycetes

"IpimenaMes

p

'1*

- .

/The Aquatic EmAroliment.

`.?:?311,

B A food web is the interlOcking pattern offood chains in an ecosystein. (Figures 3, 4)In complex natural communities, organismswhose food is obtained by the same numberof steps are said to belong to the sametrophic (feeding) level.

C Trophic Levels

1 First - Green plants (producers)(Figure 5) fix biochemical energy andwlthesine basic organic subotances.This is primary production .

2 Second - Plant eating animals (herbivores)depend on the producer organisms Mtfood.

3 Third - Primary cazpivores, aninialswhich feed on herbivores.

4 Fourth --Secondary carnivores feed onprimary carnivores.

5 Last Ultimate carnivores are the lastor. ultimate level of consumers.

D _Total Assimilation

The amount of energy which flows througha trophic level is distributed between theproduction of biothass (living substance),and the demands of respiration (internalenergy use by living organisms) in a ratioof approximately 1:10.

E Trophic Structure of the Ecosystem

The interaction of the food chainphenomena (with energy loss at eachtransfer) results in various communitieshaving definite trophic structure or energylevels. Trophic structure- may bemeasured and described either in termsof the standing crop per unit area or interms of energy fixed per unit area perunit time at successive trophic leWis. ,Trophic structure and function can beshown graphically by means of ecologicalpyraMids (Figure 5).

.6

Figure 3. Diagram cf the pond ecosystem. Basic units are asifollows: 1, abfotic substancesbasic inorganic and

organic compounds; IIA, producers--rooted vegetation; IIB, producersphytoplankton; III-1A. primary consumers(berbivores)bottom forms; III.1B, primary con (herbivores)zooplankton; 111.2, secondary consumers (car- .`

&Imes); 1114, tartluy consumers (secondary carnivores); IV, decomposersbacteria and fungi of decay.

*t:

1

1

1

.1

1

e


-**. /Nutrient Esup,*

Phytoplankton

4

\ 1

Bacterialaction ti1/4

x.....-..,_. -.....e,A3''',;!

/Death and decay\ /

Mollusks

topr ts,1

eVe,.,,lte

771.r./1%..4 Ajo,

WItta 4,01. 1.....

17410/111114.1;11101 LI,t1

so.

Figure 4. A MARINE ECOSYSTEM (After Clark, 1954 and Patten, 1966)

17

.3

13

;04

3

k.

The Aquatic EfiVironment

Decomposers(a)

Carnivores (Secondary)

Carmvores (Primary"fierbworekProducers

(b)

'Figtire 5. HYPOTHETICAL PYRAMIDS of(a)'Numbers of individuals, (b) Bioniass, andlc) Energy (Shading Indidatet Energy:Loss).

V BIOTIC COMMUNITIES

.pA Plankton are the macroscopic andmicroscopic animals, Plants, ba teria,etc., floating free in the open wat r.Many clog filters, cause tastes, odors,.and other troubles in watel-supplies.Eggs and larvae of larger forms areoften presetit.

1 Phytoplankton are ,T4..eseare the dominarit producers of thewaters, fresh and salt, "the grass ,of the seas ".'

2; Zooplankton are animal-like.Includes many different animal types,range in size from minute protozoato gigantic marine jellyfishes. ;

tst Periphyton (orAufwuchs) - The communitiesof microscopic organisnis associated.withsubmerged surfaces of any type or depth.

,1.+4,14-

Includes bacteria, algae, .protozoa andother micnoscopic animals, and often theyoung or embryonic stages of algae' andother organisms that normally grow. upto become a part of the benthos (see below).

any planktonic types will also adhereto surfaces as periphyton, and somety.pical periphyton may break off andbe collected as plankters:

C' Benthos are the plants and animals livingon, in, or closely associated with thebottom. They include plants and ,,

-invertebrates.

D Nekton are the community of strongaggressive swimmers of the open waters,often called pellagic. Certain fishes,whales, and invertebrates such asshrimps and squids are included here.

E The marsh community is based on larger"higher" plants,` floating and ernergent.Both marine and freshwater marshes areareas of enormous biological production.

. Collectively known at "wetlands", theygap between the waters and the

dry tan s.

VI PRODUCTIVITY

A The biological resultant of all physicaland chemical factors in the quantity ofHire that may actually be present. Theabilityto produce this "biomass" isoften referred to as the "productivity"of a body of water. This is neither goodnor bad per se. &water cflow'pro-duCtivity is a "poor" water biologically,and also a relatively "pure" or "clean"water; hence desirable as a water supplyor a bathing beach. ,A productive wateron the other hand may be a nuisance toman or highly desirable. it is a nuisanceif foul odors and/or weed-chockedwaterways result, it is 'desirable ifbumper crops of bass, catfish, oroysters are prodliced. Open oceans havea low level of productivity in general.

.

S

VII PERSISTENT CHEMICALS' IN THEENVIRONMENt

*,Increasingly complex manufacturing processes,coupled with rising industrialization, createhealth hazards for humans and aquatic life.

Compounds besides being toxic (acutely orchrOnic) may produce 'mutagenic effects

' including Cancer, tumors, and teratogenicity(embryo defects). Fortunately there are,tests,such as the Aniis test, to screen chemicalcompounds for these effects.

A Metals - current levers of cadmium, leadand other substances constitute a mount-ing concern. Mercury pollution, as atMinimata., Japan has been fully documented.

Pesticides

1 A pesticide and its metabolites may ,move through an ecosystem in manyways. Hard (pestidides which arepersistent, having a long half-life inthe environment includes the organo--chlotines,' ex., DDT) pesticidesingested or otherwise borne by thetarget species will stay in the'nvironment, possibly to be recycledor concentrated further through thehattiral action of food chains if thespecies is eaten., Most of the volumeOf pesticides do not reach their targetat all. ,

2 Biological magnification

Initially, low levels of persistent '\:pesticides in air, soil, and water' may

.. be concentrated at every step up thefood chain. Minute aquatic organismsand scavengers, which screen water andottom mud having pegticide levels of a

few parts peribillion, can accumulatelevels measured in parts per milliona.thousandfold increase. Thesedimentsincluding fecal deposits are continuouslyrecycled by the bottom animals.

A,' I


a Oysters,Vor instance, will'con-centrafe DDT 70,000 times higher

\ in their tissues than it's concentrationin surrounding water. They can.also partially cleanse themselvesin water free-of DDT.

b Fish feeding on lower organismsbuild up concentrations in theirvisceral fat which may reach severalthousand parts per million and levelsin their edible flesh of hundreds ofparts per million.

o Larger animals, such-as fish-eatinggulls and other birds, can-furtherconcentrate the chemicals. .A surveyon organochlorine residtes in aquaticbirds in the Canadian prairie provincesshowed that California and ring-billed.gulls were among the most contaminated.Since gulls breed in colonies, .breedingpopulation changescan be detected andrelated to levels of chemical con-tamination. Ecological research oncolonial birds to monitor the effectsof chemical pollution on the environ-

. inent is useful.

C "Polychlorinated biphenyls" (PCB's).PCB's were.used in plasticizers, asphalt,ink, paper, and a host of other products.Action was Taken to curtail their releaseto the environment. sincedtheir effectsare similarto hard pesticides. Howeverthis doesn't solve thofroblems of con-taininated sedinients and ecosystems andfinal fate of the PCB's still circulating.

D There are numerous,other compoundswhich are toxic and-accumulated in theecosystem.

'1-15

CIA r le-

The AqUa-tic Environment

FERprcEs

1 Clarke;' G. L. ents of Ecology.',john Wiley & Sons, New York. 1954.

2 Cooke, W. B. ''rickling Filter Ecology.Ecology 40(2):273-291. 1959.

3 Hanson, E.D. Animal Diversity.Prentice-Hall, Inc New Jersey. 1964.

4 Hedgpeth, J.W. Aspects of the EstuarineEcosystem. Amer. Fish. Soc., Spec.Publ. No. 3. 1966.

5 Odum, E.P. Fundamentals of Ecology.W. B. Saunders Company,Philadelphia and Londont 1959.

6 Patten, B. C. Systems Ecology.j_kige,Stance. 16(9). 1966.

7 Whittaker, R.H. New Concepts ofKingdoms. Science 163:150-160. 1969.

a 4.

e. part 3. The Freshwater Environment

I INTRODUCTION

The freshwater environment as consideredherein refers to those inland waters notdetectably diluted by ocean waters, althoughthe lower portions of rivers are subject tocertain 'tidal flow effects..

Certain atypical inland waters such as salineor alkaline lakes,, springs, etc., are not-treated, as the main objective here is tyPical....inland water.

All waters have certain basic biological cyclesand types of interactions most of Which havealready beed presented, hence this outlinTwill concentrate on aspects essentially,peculiar to fresh inland waters.

II PRESENT WATER QUALITY AS A ,FUNCTION OF THE EVOLUTION OFFRESH.WATERS

A The history of ,body of water determinesits present condition. Nattficarwaterd haveevolved in the course of geologic timeinto what :we know today.

B Streams,

In the course of their evolution, setealiffsin general pass through four stages of

. development which may be called: birth,'youth, maturity, and old age.

' These terms or conditions may beemployed or considered in two contexts: I/temporal, or spatial. In terms of geologic 4time, a given point in a stream may passthrough each of the stages described belowor: at any given time, these various stagesof development can be loosely identifiedin successive reaches of a stream travelingfrom its he dwaters tb base level in oceanor major la e.

,

-

1 Establishment eir birth. *ismight be a :'dry runt' or headwater'.stream-bed,. before it had erodeddown to the level of ground watery

During periods of run-off after arain or snow -melt, -buch a gulley

-would have a flow of Water whichmight range from torrential 'to a

mere trickle. Erosion mayproceedrapidly as there is no permanentaquatic Llora or 'fauna to stabilizestreambed materials. On the otherhand, terrestrial grass or forestgrowth may retard 'erosion. When

"Ate run-off has passed, however,- the "streambed" is dry.

2 Youthful streams. When thestreambed is- eroded be/ow theground water level, spring orseepage water enters, and thestream becomes permanent. Anaquatic cflora and fauna developsand water flows the year round,.Yout hful streams typically have arelatively steep gradient, rocky.beds,with rapids, falls°, and small pools.

14,

3 Mature streams. Maiure stred.mshave wide valleys, a developedflood plain, are 'deeper, moreturbid, and usually have warmer 'water, sand, rilud, silt, O claybottoin materfalb Ng& shift with . 4increase' in flow,- In Jheir morefavorable reaches,- streams in thisoondition are at a peak of biologicalproductivity. Gradients' are moderate,_riffles, or rapids are often separatedby long pools. -

4 age, ',streams have approachedgeologic Vase level, usually theocean. During -flood siOgge they scourtheir beds and deposit. materials onthe flood Plain which may be verybread and flat. During normal flowthe channel is refilled and:Manyshifting bars are developed. *eandersand ox-bow lakes are often formed.

1-17

The A uatic Environment

4=4f,(Under the influ ce of man this

Pattern may be b oken up, ortemporarily inte upted. Thus anessentially "youthful" stream might...take on some of the *characteristicsof_ a "mature" stream following soilerosion, organic enrichment, andincreased surface runoff. Correctionof these-conditions might likewise befollowed by at least a partial reversionto the "original" Condition).

C Lakes and Resertroirs

GeologiCal factors.iirhioh significantlyaffedt the flatting of either a stream or-lake include toe. following:

1 The geographical locatiOn of the. ' drainage basin or watershe .

became a lake. Or; heglacier may,-actually scoop out a."'hole.. Landslidesmay dam valleys, extinct volcanoes;maycollapse, etc., etc.-

2 Maturing or natural eutrophication oflakets.

IPa If not Airoor present shoal areas

are developed through erosionand deposition of the shore materialby wave action and undertow.

currents produce bars across baysand thud cut off irregular areas.

Silt brought in by tributary streamssettles out in. the quiet lake water

ed to-eurfacet,apd floating free plankton. Deadorganic matter begins to accumulate.on,the bottom.

b.

c

2 The size and shape of the drainagebasin.

45 The general topography, i.e.,'mountainous or plains.

4 The character of the bedrocks andsoils.

5 The character, amount, annualdistribution, and rate' of precipitation,.

6, the natval vegetative cover of theland_ is, of course, responsive to andresponsible for many of the above 'factors and is also severely subjectto.the whims of civilization. This .

is one of the major factors determiningrun-off versud soil absorption, etc.

D Lakes have a developmental history whichsomewhat parallels that'of streams. Thisprocess is often referred to as naturaleutrophication:'

1 The methods of formation vary greatlyfbid all influence the character and ''7subsequent.chibitory of the lake.

In glaciated areas, :for example, ahuge block of ice may have been coveredwith till. The glacier !retreated, theice melted, 'and the resulting hole

!!,.

1-18

e Rooted aquatic plants grow onshoals and bars, and in doing somit off bays and.coqribute to thefilling of the lake.

Dissolyed carbonates and othermaterial's are precipitateld in thedeeper popions of the lake in partthfough the action o plants.'

When filling is well.a anced,,mats of sphagnum mods may extendoutward from the shore. These .

mats are followed by sedies andgrasses which finally con ter_ thelake marsh.

3 _Extinction of lakes. After lakes reachmatukty4, their progress tolyerdfillirig up is accelerated. They become

f

4

. g

extinct through:`41

a- the. downcutting of thec outlet.a

b Fkllfng with detritus erode fromthe shores or brought in btributary streams.

c Filling by the a ccumu,lation'Ok the .

remains of vegetable materialsgrering.in the lake itself.(Often two or three proceSdes mayact concurrently)

.1

C

HP PRODUCTIVITY IN FRESH WATERS

A Fresh waters'in general and undernatural conditions by definition have alesser supply of dissolved substancesthan marine waters, and thus a lesserbasic potential forthe growth.of aqUaticorganiSims By tile same token, theST.maybe said to be more sensitive to tileaddition of extraneous materials(pollutants, nutrients, etc. ), Thefollowing notes are directed towardnatural geological and other environ-mental faqtorS as they ffect theproductivityòf fres% $ters.

B Factors Affecting Stream Productivity(See Table 1)

TABLE' I°

EFFECT OF SUBSTRATE ON STRAM.PRODUCTIVITY*

(The productivity of sand bottoms istaken as 1)

Bottom Material #RelativeProductivity

SandMarlFine GravelGravel and silt ,Coarse gravel 'Moss on fine gravel

. .

Fiasidens (moss) on coarse ._ .----v

1

69 -c,

'14328g

Lit

'19'4' ,

301452 -; .. -

gravpl -Ranunculus (water buttercup)WatercressElodea,(waterweed)

*Selected from Tarzwell 1937

To be productive of aquatic life, a -stream -must provide adequate nutrients,light, a 1 suitable' temperatu,re:. and timeforgrowth to take place.

1 -YOUthfurstreamS; especially on rock.ore sandSUbstrates are Zoo in-essential

's-mutrients. Temperatures in moun-tainous regions are usually low,' and .due to thp steep,gradient, time for.g.xlowth is rt. Although amplelight is a,ilable; growth of trueplankt is thus greatly llmited. 9

s

4,


Y2 As the stream flows toward a more

"mature condition, nutrients tend to. accumulate, and. gradient diminishes

,+ and So time of flow increases, fern;perature tends to increase, andplankton floUrish.

.

_Should a heavy load of-inert siltdevelop on the other hand, theturbidity would reduce the lightpenetration and consequently thegeneral plankton prctduction would

3 As the streaniapproaches base/lev el(old age) and the, time 'available for'plankton growth increases, the cbalance between to nutiie tlevels; and temper ture and therseasonal conditions, determines theoverall productivity.

C -Factors Affecting the Productivity oflakes (See Tape 2)

.

1 The size, shape, and 4epth of thelake basin. Shallow water is moreproductive than- deeper Water" since 1

more light will.reach the bottom tostimulate rooted plant grOwth, Asa corollary, lakes,.with ThOre shore-line, having more shallow water, '

are in general more productive./ Broad shallow lakes and reservoirs

have the greateit production potential(and hence should be avoided for

'Nwater supplies). s

TABLE 2.a

EFFECT OF .SUBSERA TEON LAKE PRODUCTIVITY

productivity of sand-bottcims is taken- s 1)1 .0.

BottomMateHal Relative Productivity, .

Sand.-Pebbles 'Flay ... °

Flat rubble .`Block rubbleShelving,r9ck.

1.

A

'

...

'Ism,1

4

1177

4

,°

--..\

* Selected frolinTarzwell 1937

t9

Aquatic Environment

\

2 °Hard,waters are generally more .

'prOductite than soft waters as thereare more plant nutrientavailable.- This is often greatly in-fluenced byl.the character of the soiland rocks in the watershed'and thequality and quantity of ground waterentering Ze lake. In general; pHranges'oP6.8 to =$. 2 appear to be

^ most 'productive.

, 3 Turbidity reduces productivity asis reduced.

0

4 The presence or absence of thermalstratification with its .semi-annualturnovers affects productivity bydistributing nutrients throughout thewater mass.

5 Glimate, temperature, prevalence ofOf courseice and snow,

important.

D Facto /ale ctini theReservoirs

uctivity of

,\

IV CULT RA EUTROPHICATION1A. The genleral processes of natural

eutroph cation, or natural enrichment .4 and pro uctivity have been briefly out-

lined aboye.

B When the°\activities of 'man speed up. .

these enr chfnent Processes by intro-ducing atural quantities of nutrients

tc. ) the result is often calledcultural e tro hication. This term is

o often exte ded beyond its original usage, to include the enrichment (pollution) of

streams, /estuaries, and even oceans, aswell as likes.

C

V CLASSIFICATION OF LAKES ANDRESE VOIRS .

A The productivity of lakes and impoand-ments is such a conspicuous feature thatit is often used as a convenient of

:44 :classification.

1 The productivity of reservoirs isgoverned by much the same principlesas that of lakes, withoike differencethat the water level'is much moreunder the control of man. Fluctuationsin water level can be used tb de-liberately increase o'rdecreaseproductivity. This can ,be aemOnstated

'CNby a comparison.of the TVA reservoirs

- which practice a summaidrawdownwith some of those in, the 'west where

,a winter drawdown is the rule. 4

2 The level at which watif is removedfrom a reservoir is important to the.productivity of the stream below. ,

'The hypolimnion may. be anaerobicwhile the epilimnion is aerobic, .for

...example, or...the epilimnion is poor innutrients. While the hypolitnnion isrelatively rich,_

4..

3 Reservoir discharges also Profoundlyaffect the DO, temperature, andturbidity in the.stream below a'dam.Too much fluctuationin flow may'°permit sections of he stream, to dry, °or proVide inadequate dilution for

..toxic waste.

.

1 Oligotrozhic lakes are the younger,less productive lakes, which are deep,have clear water, and usually supportSaimonoid fishes in their deeper waters.

2 Eutroyhic lakes' are more mature,more turbid, and richer. They areusually shallower. They are richerin dissolved solids; N, P, and Ca areabundant.. Plankton is abundant andther'e is often a rich bottom fauna.

;;

R

Dy strophic lakes, such as bog lakes,are low in Ph, water yellow to brown,dispolved solids, N, 1', and Ca scantybut hun4c materials abundant, bottomfauna and plankton poor, and fishspecies 13.re limited.

eservoirs lay also be class ified asorage, an run of the river.

orage reserNirs have a largevolume in relation to their inflow.

2 Run'of the river reservoirs have aiarge.flow-thrOZh in relation to theirstorage value:

.

, The Aquatic Environment

C According to 'location, lakes andreservoirs may be classified as polar,temperate, or tropical. Differences in .

climatic .and geographic conditionsxesult in,sdifferences in theiebiology.

VI SUMMARY

A A body of water such as a lake, stream, l,or estuary represents an intricatelybalanced-system in a state of dynamiceqnilibrium. Modification imposed atone point in the system automaticallyresults in compensatory adjustments atassociated'

B The mca4 th rough our knowledge of theentire system the better we can judgewhere to ivnpo e control, measures toachieve a desir d result.

REFERENCES

1 Chamberlin, Thomas C. and Salisburg,Rollin1). Geological Processes andTheir ResUlts. Geology 1: ppand 1-654. Henry Holt and Company.New York. 1904.

2 Hutcheson, George E. A-Treatise, onLimnology Vol. I \Geography, Physicsand Chemistry. 197. Vol. II.Introduction to Lake Biology and theLimnoplankton. 11'l5 pp. 1967.John Wiley CO.

e3 Hynes, H. B. N. The Ecology of Running

Waters. Univ. Toronto Press.555 pp. 1970,

4 De Santo, Robert S. Concepts ofApplied EpolOgy. Spinger-Verlag. 1978.

1

to,

MO 0 .4.

A

0

I

a

40.

.1 -2

4-

Part 4. The Marine Environment

INTRODUCTION,

A The marine emlir onmerit is arbitrarilydefined as the water massextendingbeyond the continental land masse1/4.,including the plants and animals harbored'.therein. This water`mass is large and

'i'deep, coveleing 'about 70 percent of theearth's surface and being as deep as7 miles. The salt content averages. ..about 35 parts per.thousand. Life extends

and its Mile in the Total Aquatic EhAronment. I

.to all depths'.

:B The general nature of the water cycle onearth is well knOwn. Because the largest'portion of the; surface area of the earthis covered-with water, roughly 70 percent .of the earth's rainfall is on the seas.(Figure 1)

TIgmr, 1. TNE WATER CYCLE

Since roughly one third of the .

. rain which falls on the land is againrecycled through the atmospiiere.

1414,' -(see Figure 1 again), the total amountof water washing.Over-Ithei.p.rth's surface,is significantly greaterethan one third cthe total world rainfall. It is thusnotsurprising to note that the rivers whichfinally empty into the sell carry'adisproportionate burden of dissolved and

lTABLE 1 '1 k.e ..PERCENTAGE.COMPOSITION OF THE MAJOg IONS

OF TWO STREAMS AND SEA WATER .-' (Data from Cthrk, F.W., 1924, '"The CoinpOsition of ffiverx

and Lake Waters of the United Stales", U.S. . Geal. Surv,.., , Prof. Paper'No. 135; Harvey, H. W. 1957. "The Chemistry 4

and Fertility of Sea Waters", Cambridge ljniversitrPretx, ?. ,

Cambridge) ..., V'''' " 41 '1 r

- Ioik:,1,-.

-,.DeIaware River

,-.rat _.1,ando.,-,..4-)Alo Grande "", . at '' -

l'ilr.as#

, - i.e.!0e4-41Vajec***-.' -4 -

Na

Ca - ,

Mg

Cl. ,

SO4

CO3,5"

. 1

6.70

"-81.46'17.49 -

4.81. .

h44.23

17.49

32.99 .,

'

-.

e

ire-1.

.

.

1:143,0 :ti: :85i-4-13.I2 41-

.03 / 45

2t. 65 1.

30.10.

11.55

,.:,;30.°4

!\.1-,,,1r

1:16

3.7o55.2

7.7-...

+HCO3 0.35

C 'or this presentation, the marine.environment will be (1) described usingan ecological, approach, (2) characterizedecolOgically by comparing it with fresh-,water and estuarine environments, and(3) considered as a functional: eqological"systerh (ecosystem). .

I FRESHWAT&, ESTUARINE)._ ANDRINE ENVIRONMENTS, 7'

Distinct differences are found in phgsicare.--.--7\chemtical, dr kci biotic factors in going from

a fr,enhwater to an oceanic environment.In general, environmental factors are moreconstant in freswater iriVers) and oceanic'

',.-environments than in the highly variableand harsh environments of estuarine ancoastal waters. (Figure 2) ,

physical and Ch7nical Factors

RiVers, estfaaries, and oceans are..14dompared.in Figure 2 with referenee to

--, the relative instability (or variation) ofseveral important parameters: ',In theoceans, J will be noted, very little changeoccurs in any,parametei.. In fivers, while '"Salinity" (usually, referred to as udissalbed -

-:salids") and teMperature,(accepting normalseasokelvariation,$) changtlittle'3,the otherfour pars: ters'vary c nsiderphly. Inestuattes, they.all chan e.

suspended`golids picked up from theThe chemical.composition of this burdendepends on the 'cOmpoditibn of the rocksand soils through vi,./hIchihe river flMvs,'the'Pr'oximity of an ocean; the directionof prevailing winds, and otherfacars.This latho substance of geological eroSiOn.(Table_ 1) 4f

1 23AK

-The Aquatic Environment -

Type of environmentageneral directionarwater movement

Degree of instability . Avail-

-abilityoS.,00-

nutrients(degree)

TurbiditySalinity-

Temperature

s

Waterelevation

.VerticalStrati- -fication

Riverine

_

scene

.

-

I,

-

I.

,

. .

c--

.

,

.

.

.

I.

Oceanic

Figure 2 .

'B Biotid Factors

1-24

RELATIVE 'VALUES OF VARIOUS PHYSICAL AND CHEMICAL FACTORSFOR RIVER, ESTUARINE, AND OCEANIC ENVIRONMENTS

.A4r-

A complex of physical and ch(micalfactors determine the biotic composi-tion of an environment. In general,the number of species in a rigorous,highly variable environment tends to Ikeless than the number in a More stableenvironment (Hedgpeth, 1966).

The dominant animal species (interms.of total biomass) which occur 4*in estuaries ke often transient,.pending only N. part of their lives in

estuaries. This results in betterutilization of a rich environment.

I

C Zones of the Sea

The nearshore environment is oftenclassified -in relation to tide level andwater depth. The nearshore and offshoreoceanic regions isagether, are oftenclassified with reference to light penetra-tion and water depth. (Figure 3)

1 Neritic - Relatively shallow-waterzone which extends froin the high- \tide mark to the edge of the -7

continental shelf.

27

The Aquatic Environment'

*MARINE ECOLOGY

PEL A 6/C

SwmSuomi 1 T/C

L it !pot(Intertidal)

PELAGIC (WM')Ntelhe,GIctone

EPNI0Pe'Ineptly&.54.1ltyptlaveAtforvilopr

8ENTHIC (Bottom)SuproItttoralLittoral (IntHIJol)SOldtatel

Inns.Oyler

BathyalAbliSta

Sittilittorol

0.

0 C EA N I Copf P opic

tee

wecop logic.1),

.r.z_r_r_rx://xxx e7-7

°we

ACV Mocaor CA)14a NV

tl scret civotle of P.*hVIC.

6.

BolOrpologic

13-ENTHIC

e ' 4/././C'2Atisse0,109,C

txeJhe,

N:N:\

FIGURE 3Classikition of marine environments

..sow ..1) Physical factors fluctuate

less than in the neritic zone.a Stability of physical factors is

intermediate between estuarine00 and oceanic environments.

b Phytoplankters are the dominantproducers but in some locationsattached algae are also importantAs producers.

c The animal consumers arezooplankton, nektoil; and benthicforms.

2 Oceanic - The region of the ocean .beyond the continental shelf. Dividedinto three parts, all relativelypoorly populated compared to theneritic zone. ,

a EuEihotic zone - Waters into whichsunlight penetrates (often to thebottom, in the neritic zone). Thezone of primary productivity oftenextend's to 600 feet below the surface.

011

'70

$0.

14

gra

Kee

Xt.

Xt.

ON*

2) Producers are the phyto-plankton and consumers arethe zooplankton and nekton.

b Bathyal zone - From the bottomof the euptiotic zone to about2000 meters.

1) _Physical factors relativelyconstant but light is absent.

2.) Producers are abSent andconsumers are scarce.

c Abyssal zone - All the sea belowthe bathyal zone,

1) Physical factors'more con-stant than in bathyal zone:.

2) Producers absent and consumersven less abundant than in thebathyal 'zone .

1-25 .

The A uatic Environment

III SEA WATER AND THE BODY FLUIDS

A Sea water is a remarkably suitableenvironment for living cells,, as itcontains 11 of the chemical elementsessential to thesgrowthand maintenance. .of plant and animalS. The ratio and 'often the concentration of the majorsalts of sea water are strikingly similarin the cytoplasm and body fluids ofmarine organisms. This similarity isalso evident; although modified somewhatin the body fluids of fresh water andterrestrial animals. For example,sterile sea water may be usedinemergencies as a substitute for bloodplasma in man.

B Since marine organisms have an internalsalt content similar to that of theirsurrounding medium (isotonic condition)osmoregulation poses no problem. On theother hand, fresh water organisms arehypertonic (osmotic pressure of bodyands is higher than that of the surround-ing water). Hence, fresh water animalsmust constantly expend more energy tokeep water out (i. e., high osmoticpressure fluids contain more salts, theaction being then to dilute this concen-tration with more water).

1 Generally, marine invertebrates arenarrowly 2oildlosmotic, i.e., the saltconcentration of the body fluids chatngeswith that of the external medium. Thishas 'special significance in estuarinesituations where salt concentrationsof the water often vary considerablyin short periods of time.

2 Marine bony fish (teldosts4ave loWersalt content internally than aite externalenvironment (hypotonic).' In order toprevent dehydration, water is ingestedand salts are excreted through specialcells in the gills.

'

N FACTORS AFFECTING THE DISTRI-. BUTION OF MARINE AND ESTUARINE

ORGANISMS

A Salinity. Salinity is the single mostAtconstant and controlling facto n the '

marine environment, probabl bllowedby temperature. It ranges around35, 000 mg. per liter, or "35 parts per . .

thousand" (symbol: 35 %) in the languageof the oceanographer. While variationsin the open ocean= are relatively small,salinity decreases rapidly as oneapproaches shore'and prpceeds throughthe estuary and up into fresh water witha salinity of "0 Too (see -Figure 2)

B Salinity and temperature as limiting. factors in ecological distribution.

1 Organisms differ in the salinitiesand temperatures in which theyprefer to live, and in the variabilitiesofthese parameters which they cant olerate. These preferences andtolerances often change with successivelife history stages, and in turn oftendictate where the organisms live:.their "distribution.' ,

`*,r)

1

2 These requirements or preferencesoften lead to extensive migrationsof various Species for breeding,feeding, mild growing stages. Onevery important-resnkof this is thatan estuarine environment is anabsolute necessity for over half ofall coastal commercial and sportrelated speciesof fishes and invertebrates,' e.

for eitherall or certain portions of theirlife histories. (Part V; figure 8)

3' The Greek word roots "eury"(meaning wide) and "steno '-(ineaningnarrow) are customarily combined,with such Words as "haline" for salt,and "thermal" for temperature, togive us "euryhaline" as an adjectiveto characterize an organism able totolerate a wide range of salinity, forexample; or "stenothermal" meaningone which cahnot stand -much changein temperature. "Meso-" is a prefixindicating an intermediate capacity.

29

ti

C Marine, estuarine, and fresh waterorganisms. (See-Figure 4)

0 Salinity ",; ca. 35

Figure 4. Salinity Tolefance of Organisms

1 Offshore marine organisms are, ingeneral, both stenohaline andstenothermal unless, as noted above,they have certain life history require-ments for estuarine conditions.

Fresh water organisms are alsostenohaline, and (except for seasonaladaptation) meso- or steno ,ermal.(Figure 2)

3 Indi us or native estuarine speciesat normally spend their entire lives

in the estuary are relatively few innumber. (See Figure 5). Tpey aregenerally meso- or euryhaline andmeso- or eurythermal,

t.

10 15au

20 25 30 35bnityFigure 5. DISTRIBUTION OF

ORGANISMS IN AN ESTUARY

tEuryhalirie, freshwaterb Indigenous, estuarine, (mesohaline)c Euryhaline, marine


4 Some will knoWn and interestingexamples of migratory species whichchange their environmental preferemeswith the-life history stage include theshrimp (mentioned above), striped bass,,many herrings and relatives, thesalmons, and many others. -None aremore dramatics than the salmon hordeswhich hatch in headwater streams;migrate far out to feed and grow,then return to the mountain stream.where they hatched 'to lay their owneggs before dying.

5 Among euryhaline animals landlocked(trapped), populations,living in loweredNlinitieskoften have a smaller maximumsize than individuals of the same speciesliving in more saline waters. Forexample, the lamprey (Petromyzonmarinus) attains a length of 30 - 56"in the sea, while in the Great Lakesthe length is 18 -.24".

Usually the larvae of aquatic organismsare more sensitive to changes insalinity than are thesodults. Thischaracteristic both limits and dictatesthe distribution and size of populations.

D The effects of tides on organisms.

1 Tidal fluctuations probably subjectthe benthic or intertidal populationsto the most extreme and rapid variationsof environmental stress encounteredin any aquatic habitat. Highly specializedcommunities have developed in thiszone, some adapted to the rocky surfzones of the open coast, others to themuddy inlets of protected estuaries.Tidal reaches of fresh water rivers,,sandy beaches, coral reefs andmangrove swamps in the tropics; allhave their own floras and faunas. Allmust emerge and flourish when whateverwater there is rises and povers.ortears at them, all must collapse orretract to endure drying, blazingtropical sun, or freezing arctic iceduring the low tide interval. Such acommunity is depicted in Figure 6.

1-27 ,

the Aquatic Environment

O.

SNAILS

Littorina neritoidesL. rudisI.. obtusataL. littoreaP,A dNA CLES

0!) Chthamalus stellatus0 Balanus balanoides

13. pe Hoist uS

Figure 6Zonation of plants, snails, and barnacles on a rocky shore, Whilethis diagram is based on the situation on the southwest coast ofEngland, the general idea of zonation may be applied to any temper- -

att. rocky ocean shore, though the species will differ, The grayzone consists largely of lichens. At the left is the zonation of rockswith exposure too extreme to pport algae; at the right, on a lessexposed situation, the animals re mostly obscured by the algae.Figures at the right hand margin refer to the percent of time thatthe zone is exposed.to the air, i.e., the time that the tide is out.'Three major zones can be recognized: the L4ttorina zone (above thegray zone); the Balan.oid zone (between the gray zone and thelaminarias); and the Laininaria zone, a. Pelvetia cankliculata;b. Fucus spiralis; c. Ascophyllum nodosum; d. Fucus serratus;e.Laminaria digitata. (Based on Stephenson)

.

0

..-/

31

The Auatic Environment

V FACTORS AFFECTING THEPRODUCTIVITY OF THE MARINEENVIRONMENT

A The sea is in continuous circulation. ,With-'on circulation, nutrients' of the ocea.n wouldeventually become a part of the bottom andbiological Production would cease. Generally,in all oceans there.exists a warm surfacelayer which overlies the dolder'water andforms a two-layer system of persistentstability. Nutrient concentration is-usuallygreatest in the lower zone. Wherever amixing or disturbance of these two layersoccurs biological production is greatest.

B The estuaries are also a mixing zone ofenormous importance. Here the fertilitywashed off the lanclis mingled with thenutrient capacity of seawater, and manyof the would's most productive watersresult.

C When man adds his cultural contributionsof sewage, fertilizer, silt or toxic waste,it is no wonder that the dynamic. equilibriuof the ages is rudely upset, and theenvironmentalist. cries; "See what manhath wrought "!

ACKNOWLEDGEMENT':'

This outline contains selected materialfrom other outlines prepared by C. M.Tarzwell, Charles L. Brown, Jr.$C. G. Gunnerson, W. Lee Trent, W. 13.Cooke, 13. H. Ketekum, J. McNulty,'J. L. Taylor, R. M. Sinclair,. and others.

REFERENCES

1 Harvey, H. W. The Chemistry andFertility of Sea Water (2nd Ed. ).,Cambridge Univ. Press, New York.234 pp. 1957.

2 Wickstead,, John . Marine ZooplanktonStudies in Bio gy,no. 62. The Instituteof Biology. 78.

Part 5: Wetlands

I INTRODUCTION. ..,

A Broadly defined, wetlands are areaswhich are "to wetto plough but toothick to flow." The soil tends to besaturated with water, salt of fresh,and numerous channels or ponds ofshaliow or open water are common.Due to ecological features too numerous

,. lid variable to list here, they comprise-,general ta rigorous (highly stressed)

habitat, occupied by a small relativelyspecialized indigenous (native) floraand fauna. ,i

B They are prodigiously productivehowever, and many constitute anabsolutely essential habitat for someportion of the life history of animalforms generally recognized as residentsof other habitats (Figure ti). This isparticularly true of tidal marshes asmentioned below.

(C Wetlands intoto comprise a remarkably

large proportion of the earth's surface,and the total organic carbon bound in

. their mass constitutes an enormous1 sink of energy.

D Since, our main concern here is withthe "aquatic" eironment, primaryemphasis will be directed toward a

rdesc t ption of*vetlands as the transitionalzone' etween the waters and the land, andhow their desecration by human culturespreads degradation in both direction,.

II TIDAL MARSHES AND THE. ESTUARY.

A "There is no other case in nature, savein the coral reefs, where the adjustmentof or anie'relations to physicardttlitiona see in such a. beautiful way as thebalan e between the growing marshepand the tidal streams by which thePareat once nourished and worn awe*:(Shale , ).886):

ON

B Estuarine pollution studies are usuallydevoted to the dynamicsul.the'circulattng .

water, its chemical, physical, andbiological parameters, bottom, deposits, etc.

C It is easy to overlook the intimate relation-ships which exist between the borderingmarshland, the moving waters, the tidalflats, subtidal deposition, and sestonwhether of locale oceanic, or riverineorigin.

D The tidal marsh (some inland areas alsohave salt marshes) is generally consideredtb be the marginal areas or estuaries andcoasts in the intertidal zone, 4hich aredominated by emergent vegetation. Theygenerally extend inland to the farthestpoint reached by the spring tides, wherethey merge into freshwater swamps andmarshes 4-Figure 1). They may range inwidth from nonexistent on rocky coasts tomany kilometers.

Channel LMud Flat Tidal MarshFreshwater

Marsh

tuin Mara

?it- - :::

47 pat 0- Blue Clay Substrate0 *P...ert

rtgurcl. Zonation in e positive New England estuary. 1. Sprtng tide level. 2. Mean higlyide.S. Mean low tide, 4. Dog hole. S. leo eleavap pool, 4. Chunk of Spartina turf deposited by lee.T. Organic oose with associated community. O. pipes* (Zostei ra). 2; Ribbed mussels froodiolusl-

clam QM); mud snail Meal community. 10. lettur vai

The Aquatic, Envir6timent

III MARSH ORIGINS AND 'STRUCTURES

A In general, marsh substrates are high inorganic content, relatively low in mineralsand trace elements. The upper layersbound together with living roots calledturf, underlaid by more compacted peattype material.

1 Rising or eroding coastlines mayexpose peat from ancient marshgrowth to wave action which cutsinto the soft peat rapidly (Figure 2).

1132

Terrestrial turf

Salt marsh eat

Substrate

A1)(11,1( h

7-:;:Figure 2. gran.--13ta matic se ion of.erodlng prat cliff

.

Such banks are likely to be cliff-like,and are often undercut. Chunks of,peat are oen found lying about onharder sViattrate below high tide line.If face of cliff is well above high water,overlying vegetation is likely to betypically terrestrial Of the area.Marsh type vegetation is probably'.absent.

2 Low lying deltaic, or sinking coast=lines, or those with low energy waveaction are likely to have active marshformation in progress. Sand dunes

.. are also common in such areas(Figure 34. General coastalconfiguration is a factor.

Figure 3

Development pi a Massachusetts Marsh since .1300 BC. involving an.

18 foot rise in water level. Shaded area indicates sand dunes. Note

i meandering marsh tidal drainage. A: 1300 BC, Br 1950 AD.

34

O

0

.

I

...The Acpiatic EnVironment

a Rugged or precipitous coasts orslowly rising coasts, typicallyeghibit narrow shelves; sea/cliffs,fjords, massive beaches, and .relatiyely less marsh area (Figure 4).An Alaskan fjord subject to recentcatastrophic subsidoce and rapid -

deposition of glaciflour ShowseVidencelif the recent encToachmentof saline waters in the presence of`recently buried trees and otherterrestrial vegetation, 'exposure .

of layers of salt marsh peat 4ongthe edges of channels, .and.a poorlycompacted younglmarsh turf developingat the. new high water level (Figure 5).

b

Figure 4 A River Mouth on a Slowly Rising Coast. Note abseimeof deltaic development' and relatively marshlandalthough mud flats stippled are extensive,

Shifting flats Tidal marsh I 'Terre striajilt°03

0 Et',

2

8 . 4

it \ /..:,4 ,. .: a ,e

';. g

. . '

. ../Vv\A

.11,

Figure 5 Some genial relationships in 4 northern fjord witla rising water level. 1. ,mean low'water,' 2, maximum high tide, 3, Bedrock, 4. Glacial flour to depths in excess or-b() meters, 5. Shifting flats and channels, 6. Channel against bedrock, 7. Buriedterrestrial vegetation, 8. Outcroppings of salt marsh peat.-

b Lowslying coastal plains tend to befringed by barrier islands, broad c.estuaries and deltas, and broadassqciated marshlands (Figure 3).

F (Vcs

°Deep tidal' channels fan but throughirmumerablebranching and oftenInfer-connecting rivulets: Theizitervening. grassy plains are _

essentially, at mean high tide level.

A

1 -33

-

C.

F_The Aquatic Environment

c Tropical and subtropical regionssuch as Florida, the Gulf Coast,and Central America, are _frequented

"'by mangrove swamps. This uniquetype of growth is able to establishitself in shallow water and move outinto progressively' deeper areas(Figure 6). The strong deeplyembedded roots enable the-mangroveto resist considerable wave actionat times, and the tangle of rootsquickly accumulates a deep layer oforganic sediment...Mangrovesin the south may-be considered tobe roughly the equivalen,t of theSpartina marsh grassin the northas a land builder. Whenfullydeveloped, a mangrove swamp is animpenetrable thicket of roots overthe tidal flat affording shelter to anassortment of semi-aquatic organismssuch as various molluscs aridcrustaceans,' and providing accessfrom the nearby land to predaceousbir'dsa.reptiles, add mammals. ,

Mprigrov.es are not restricted toestuaries, buernay develop out intoshallow oceanic lagoons, orupstreaminto relatively fresh waters.-

'

WACO. Tit=taruS,4ASSCOCS SALtiiIrrerASSOCAOI

:resitIN0001

attiosvoraCOMOOLS

Is=an 1.40.4111

Figure 6warauravasor

Diagrammatic transect of a mangrove swampshowing transition from marine to terrestrialhabitat. ° ,

0

4O

N PRODUCTIVITY OF WETLANDS

A Measuring the productivity of grasslandsnot_easy,tbecause.today,grass is seldpm

used directly 4isisuch by man. Itis thus- usually 'expiessed-as production of meat,

milk, or bythe case of salt marshes, 'thetotal crop'of animals that obtain food perunit of area. '4he.prfirnary producer in a:.

61-14

. .

tidal mar sh is the mar.sh grass, but ver34little of it is use& flreman as grass.(11ab1, 1)

_The nutritionaLanalysis of several.marsh grasses as compared to airy land"hay is shown in Table 2.

..

TABLE 1,, General Order's of Magnitude &Gross Primary Productivity in Termsof ,Dry Weight of Organic Matter Pitad Annually

gms/M

.,/year

Ec9system (grarns/awkre meters/year) lbs/acre/year

Land desekts, deep sceane v Tens

Grass4ds, forests, cutrophic Hundredslakes, ordthary agriculture

Estuaries, deltas, coral reefs, Thousandsintensive agriculture (sugarcane% rice)

#. TABLE

T/A

Dry Wt;

Hundreds

Ttiousends

Tenthousands

2. Analyses of Some Tidal Marsh Grasses

li'ercentage Composition

Protein r Eat . Fiber Water Ash Nfree Extract

Divichln spicata (Ore stand, dry)2.8 ' 5.3 1.7. 32.4 8.1 6.7 4S.5

Short Spartina alternillora and Sabccenia curopaea In standing water)1.2 7.7 2.5 31.1 - 6.6 12.0 37.7

Spartina alterndlora (tall, pure stand in standing water)3.S 7.6 ' 2.0 . 29.0 8.3 15.5 37.3

Sparrinavalftns sport stand, dry) .:. .. .

3.2 , 6,0 ,.. 2.1 30.0 8.1 9.0

Sparrina a/remit/4ra and Sp.iffifla patens (mixed stand, wet)3.4 - 6.6 1.9.4 .29.5 8.1 10.4

Spariniakernillura (short, wet)Z2 13.0 . 2.4 30.4 81 13.3 36.3

Comparable Analyses for Hal 0El 0

1st rut 6.0 2.5 ° 36.2 ",,0 6.7 4.2 44.9

a2,,,roil 11.0 3.7 25.5 10.4 5.9 30.5

AnalrieeperfOrMed by Roland W.. Gilbert, Deptrtmentb. of Agricultural Chemistry, U. R. I.

.

44.5

42.8 o

o

Ow

.

.:.

3C

'I


11

B The actual utjlizaiion'of marsh grass isaccomplighed primarily by its decom-position-and ingestion by micro organ&ns.(Figure 7) A small quantity of seeds andsolids is consumed directly by hirds.

-1

0

,Figure 7 The nutritive composition ofsuccessive stages of decomposition of -Spartina marsh grass, showing increasein protein and decrease pt carbohydratewith increasing age and decreasing size,of detritus particles.

1=

1 The quantity of micro invertebrateswhich thrive on this wealth of decayingmarsh has not been 'estimated, nor hasthe actual production'of small indigenousfishes and invertebrates such as thetop:minn.ows (Fundulus), or the mud-snaili (Nesse), and others.

2 Many forms of oceanic life migrateinto the estuaries, especially the 'marsh areas, for important portiohsof their life histories as is mentionedelsewhere (Figure 8)." It has been .

estimated that in exgess of. 60% of themarine commercia1:4nd sport fisherIe4eare estuarine or marsh dependent in

.some way.

04/0,

',-

0

11;"1.

VrTZtleYOUNG

ISOUNJ23

N OCEAN

.eli4106A ekADULT EGGS

Figure 8 Diagram of the life cycle- .of white shrimp (after Anderson.and

.Lunz .1965).

3. An effort to make an indirectestimate,ofprogiuctivity in a ithOde

^ Island marsh was made on a singledad by recording the numbers

and kinds oebirdirthat fed on arelatively -small area (FigureBetwepn 700 and 1000 Wild oirdsof12 species, ranging from 100 leastsandpipers to uncountable numbersof seagulls were counted. One foodrequirement estimate for three-pound poultry in the confined inactivityof a poultry yard is approximately oneounce per pound of bird per day.

-40;:

.Figure 9

444

Great blue heron

SomeCommon Marsh Birds

4

V.

;

a

Ps.

Greater Yellow legs (left) -and black duck . Nr\

1-35-

ft

t

to


One-hundred black bellied ploversat approximately ten ounces eachwould weigh on the order of sixtypounds, At4the same rate of foodconsumption, this would indicatenearly fotfr pbunds of food requiredfor this species Lone. The muchgreater activity of the wild birdswould obviously greatly increase theirfood requirements, as would their

relatiSely smaller size.-

-Considering the range of foods con-sumed, the sizes of the birds, and thefact that at certain seasons, thousandsof migrating ducks and others pauseto feed here, the enormous productivity,of such a rbarsh can be better under-stood.

V

A

-

INLAND BOAS AND MARSHES

Much of what has been said of tidalmarshes also applies to inland wetlands.As was mentioned earlier, not all inlandswamps are salt-free, any more than allmarshes affected by tidal rythms aresaline.

- ..13 The specificity of specialtz'ed floras to

particula types'Of wetlands is perhapsmore spectacular in freshwater wetlandsthan in the marine, where Juncus,Spartina, and Mangroves tend to dominate.

1 Sphagnum, or peat mass,--is'probably one of the most widespe dand abundant wetland plants on eaDeevey Q.958) quotes an estimate taithere is pt:obably upwards of 223billions (dry weight) of tons of peatin ti( world today, derived duringrecent geologic time, from Sphagnumbogs. Particularly in the northernregions, eat moss t nds to overgrowponds An shallow de fissions, eventuallyforming the vast turd lains-and-moores of the nort

°

2 Long lists of other bog and marsh plantsMight be cited; each with its ownspecial requirements, topographical,

r

and geographic distribution, etc.Included would be the familiar cattails,spike rushes, cotton grasses, sedges,trefoils, alders, and many, many'.Others.

C Types of inland wetlands.

1 As noted above (Cf: Figure 1)tidal marshes often merge intofreshwater marshes and bayous.Deltaic tidal swamps and marshesare often saline in the seawardportion, and fresh in the landward

. . areas.

2 River bottom wetlands differ from-those formed from lakes, since wideflood plains subject to periodicinundation are the final stages ofthe erosion of river valleys, whereaslakes in general tend to be eliminatedbyhe geologic proceSses of naturaleutrophication often involvingSphagnum and peat formation.Riverbottom marshes in the southernUnited States, with favorable climates,have luxurient growths such as thecanebrake of the lower Mississippi,or a characteristic timber growthsuch as cypress.

Aithoug\bird lift is theconspicuoUs animal elener in thefauna (Cf:- izigure 9), many mammals,such as muskrats, beavers, otters,and others areal marsh-oriented.Figure 12) - ,

o. .2 ,,

3

33ft.

4

S

4

VI POLLUTION-

A No "Single statement can summarize theof pollution on Marshlands'as

distinct from effects ,noted elsewhere onother habitats.

B .Reduction of Primary Productivity

The primary producers in most wetlandsare the grasses and peat mosses.Production may be reduced or eliminatedby:

1' Changes iri thewater level broughtabout by flooding or drainage.

a Marshland areas are s5metimesdiked and flooded t9 produce frewater ponds. This may bq for'aesthetic reasons, to ,suppress the

ingrowth of noxious marsh inhabitatingsects -such as mosquitoes or biting

midges, to constructan industrialwaste }folding pond; a thermal or asewagg stabilization pond, a"convenient" result of highway'causeway construction, or otherreason., The result is the elim-ination of an area of marsh. A a

sitallcompensating border ofmarsh may or may nor develop.

b r-High i'dal marshes were oftenditche and drained in former daysto stab ze the sod for salt hay or"thatch' barvesting which was highlysought a sr in colonial days. This

,inevitably changed the characterof the marsh, but it remained asessentially marshland. COnversionto outright agricultural land hasbeen less widespread because of thenecessity of diking to exclude theperiodic floods or tidal incursions,..and carefully timed drainage toeliminate excess precipitation.Mechanical tidal marsheshas_not been econo 'cal in thiscountry, althoUgh the succesb ofthe Dutch andOthers in this regardis well known.

The AcLuatiC Environment

2 Marsh grasRes.may alsO be eliminatedby smothering as, for example, bydeposition of dredge spoils, or thespill or discharge of sewage sludge.

3 Considerable marsh.area has beeneliminated by industrial constructionactivity, such as wharf and dock con-struction, oil well construction andoperation, and the discharge of toxicbrines and other chemicals.

C Consumer production (animal life) hasbeen dr a st Joan:), 're'auised by th'..dr. liberated'distribution of-pesticides. .lesome cases,this has been aimed at nearby agricultural,. ,lands for economic crop pest control, inother cases the marshes have been sprayedor dusted directly to control noxiousinsects.

The results have been universallydisadtrous for the marshes, and thebene \ to the human community oftenquestio able.

.`

.2 Pesticides designed to kill nuisance'insects, are atso toxic to otherarthropods sb that in addition to the'target species, such forage staples asthe various scuds (amphipods), fiddlercrabs, and other macroinvertebratehave either been drastically reducedor entirely eliminated in many placesFor example, one familiar with fiddlecrabs can traverse miles of marshmargins, still riddled with their burrows,)without seeing a single live crab.

a, DDT and related compouna have been'"eaten up the food chain" (biologicalMagnification effect) until fidh eatingand other predatory birds such as heronsand egrets (Figure 9), have been virtuallyeliminated from vast areas, and the /

-accumulation of DDT in .nian himselfis only too well known.

4

The Aquatic Environmen

1'

.1?D Most serious of marsh enemies is

man himself. In his quest for "lebensraum"near the water, he has all but killed thewater he strives fo approach.. Thus up totwenty percent of the marshestuarinearea in various pails of the country hasalready beentutterly destroyed by cut and

. fill real estate development's (Figures,`10, 11).

E Swimming birds uch as duckd, loons,cormorants, pelicans, and many othervsare severely jeopardizedby floating

- pollutants such as oil..

4 rigure 10.

S

Yt

Diagrammatic representation of cut-and-Sill forreal estatexclevelopment mlw = m ean 'low. water

i. .

Figure 11. Tracings of portion of map of a southern1, : .. city. showing, extent of cut-and-fill realj.,, -f estate development. t .

0 z) .

ii

t

- .

6,4

et 6. The Aquatic Environment r

4.0

VII SUMMARY

A "Wetlands comprise the marshes, swamps,bogs, and tundra areas of the world.They are, essential to the well-being ofour surface waters and ground waters.They are essential to aquatic life ofall types living in the open waters. Theyare essential as habitat for all forms of,

wildlife. _

B The tidal marsh is the area of emergent 0= vegetation bordea.igthe,..ocean er ash, estuary.

C Marshes are highly productive areas,essential to the maintenance of a wellrounded community of aquatic life.

D Wetlands may be destroyed by:

1

Aft,

Degradation of the life forms ofwhich it is composed in the name ofnuisance control.

2 Physical destruction by cut-and-fillto create more land area.

REFERENCES

0

Anderson, VP. W. The Shrimp anti theShrimp Fishery of the Southern "United States. TJSDI, FWS, BCF.Fiihery Leaflet 5,89. 1966. °

2 ,,DeeVey,- E. S. ,. Jr. ,BogS. Sci.' Am. Vol.199(4):115-122. October 1958.

3 "Emery, K. 0. and Stevenson. Estuariesand Lagoons. Part II, Biological

0 Aspects by J.W. Hedgepeth, pp: -693-"M. in:. Treatise on Marine Ecologyand Pleoecology. Geol. Soc. Am.Mem. 6 ?. Washington, DC.' 1957.

4 Hesse, R.;; W. C. Allee, and K. P.Schmidt. Ecological AnimalGeOgraphyw John Wiley; & Sons. 1937.

6

Oft

5 Morgan, .J. P. Ephemeral Estuaries ofthe Deltaic Environnient in :'Estuaries;pp. 115-120. Publ. No. 83, Am.Assoc. =Adv. Sol, Washington, DC. 1967.

6 Odum, E.P. and Dela Crug, A.A.Particulate Organic Detritus in'aGeorgia Salt Marsh - EstuarineEcosystem. 'in: Estuaries, pp. 383-388, Publ. No. 83, Am. Assoc. Adv.Sci. Washington, DC. 1957.

7 Redfield, A. C. The .Ontogeny of a Salt.Marsh Estuary. in: Estuaries, pp.108-114. Pulil. No. 83, Arn, Assoc.Adv. Sci. Washington, QC. 1967.

8 Stuckey, 0. H. Measuring the Productivityof Salt Marshes: Maritimes (GradSchool of Ocean. , U. R. I. ) Vol. 14(1):9-11, February 1970.

Williams, -11.B. _CompartmentalAnalysis of Production and Decayof Juncus reomerianus. Prog.Report, Radiobiol. Lab., Beaufort, NC,Fiscal Year 1968, USDI, BCF, pp, 10-12. ,

This outline was prepared by H. W. Jackson,former Chief Biologist, National Training.Center, and revised by R. M._ Sinclair, AquaticBiologist, National Training Center, MOTD,IOWPO, EPA, Cincinnati, OH 45268.

Descriptors: Aquatic Environment, BiologicalEstuarine Environment, Lento Enrironment,Lotic Environment, Currents, Marshes,Limnology, Magnification, Water Properties

( 1-39

A**

CLASSIFICATIONOF COMMUNITIES, ECOSYSTEMS, AND TROPHIC LEVELS -

I A COMMUNITY is an ashmblige ofpopulations of plants, animals, bacteria,and fungi that live in an environmental andinteract with one another, forming togethera destinctive living system with its owncomposition, structure, environmentalrelations, development, and function.

,

II An ECOSYSTEM is a community and itsenvironment treated together as a functionalsystem of complementary relationships,and transfer and circulation of energy and IVmatter. (A delightful litte essajr.on theodyssey of atoms X and Y through an.ecosystem is in Leopold's, A Sand CountyAlmanac. )

III TROPHIC levels are a convenient meansof classify ing organisms according tonutrition, or food and feeding. (See-Figure 1. )'

A PRODUCER, the photosynthetic plant orfirst organism on the food chain sequence. VFossil fuels were produced photosynthe-

-

B

C

D.

E

F

H

I

1 Collectors strain, filter, or otherwisecollect fine particulate organic matterfrom the passing current.

2 Shredders feed on leaves, detritustand coarse particulate organic matter.

3 Grazers feed on attached growths.

4 Predators feed on other organisms.'

Taxonomic Groupings

A TAXOCENES, a specific group of organisms.Ex. midges. For obvious reasons mostsystematists (..xonomists) can specialize-in only one group of organisms. This factis difficult for the non-biologist to grasp:

B Size, which is often dictated by the inves-tigator's `sampling equipment and specificinterests.

Herbivore or primary CONSUMER, thefirst animal which feeds on plant food..

First carnivore or secondary CONSUMER,an animal feeding on a plant-eating animal.

Second carnivore or tertiary CONSUMERfeeding on the preceding.

Tertiary carnivore.

Quaternary carnivore.

DECOMPOSERS OR REDUCERS, bacteriawhich break down the above. organisms.Often called the middlemen or stokers ofthe furnace of photosynthesis.,

Saprovores-ox! DETRITIVORES which feedon bacteria and/or fungi.

....--

Macroinvertebrates have been subdiVidedinto trophic levels according to feedinghabits ,(See Figure 1 from Cummints);

(

BI. ECO, 25a. 3.8042

Arbitrary due to organism habitat prefer-ences, available 'sampling devices,personal preference of the investigator,and mesh sizes of nets and sieves.

A PLANKTON, organisms suspended in abody of water and at the mercy of currents.This group has been subject to numerousdivisional schemes. Plants are PHYTO-PLANKTON, and animals, ZOOPLANKTON.Those retained by nets are obviously, METPLANKTON. Those passing thru even the-finest meshed nets are NANNOPLANKTON:

B PERIPHYTON, the community of micro-organisms which grow on submergedobjects (substrates). -Literal meaning"to grow around plants", howeverstandard glass microslides are sub-mersed in the aquatic habitat tostandardize results.

C BENTHat. is often used to meanMACROINVERTEBRATES; although thereare tenthic organisms in other Plant,ariiinal, and protist grolips. Benthicrefers strictly to the bottom substrates oflakes, - streams, and other water bodies.

2-1

j

Classification of Communities, Ecosystems and Trophic Levels

0.5' METERS)MICROSES.

46.. 2 (1-2 METERS),

Az4!remootNin5PERIPWCTON)

Cr

3 (4-6 METERS)(VASCULAR

HYDROPHYTE0

CPOM

(404

P/R°<I

MICROIE

401W 4 (10 METERS)

0

5

6 (50-75 METE

CPOM

00.

PiR

PRODUCERS(PERIPHYTON)

EDATORS

MicROSES COLLECTORS

4. P/R < 1PROODCERS

(PHYTOPeasicToN)

I I

- 12 METERS)

2-2

OLLEcTORS(COMPLANKTON)

I

. FIGURE 1

43

ORS

Classification of Communities, Ecosystems andTrophic Levels

D ./MACROINVEATEBRATES, are animalsretained on a No. 30 mesh screen (approx-imately 0.5 mm) and thus visible to the;naked eye.

E MACROPHYTES,. the larger aquatic plantswhich are divided into emersed, floating,and submersed communities. Usuallyvascular-plants but may inclUde the largeralgae and "primitive" plants. These haveposed tremendous economic problems inthe large man-made lakes, especiallyin tropical areas.

F NEKTON, in freshwater, essentially fish,salamanders, and the larger crustacea.In contrast to PLANKTON, 4\ese organismsare not at the mercy of the-current.

G NEUSTON, or PLEUSTON, -are inhabitantsofthe surfaCe film (meniscus organisms), -

either supported by It hanging from, or "

breaking through. it. Other organismsare trapped by this neat little barrier ofnature. The micro members of this areeasily sampled by placing a clean cover.slip on top of the surface film then eitherleaving it a specified time or examining:it immediately,under_the microscope. '

H DRIFT, macroinvertebrates which driftwith the streams current either periodically(diel or 24 hour), behaviorally, catastro-phically or incidentally.

BIOLOCIAL FLOCS, are suspended'microorganisms that are formed by , .various means. In wastewater treatmentplants they are encouraged in concrete aeraeration basins 'using diffused air oroxygen (the,heart of the, activated sludgeprocess).

1

.

A

J MANIPULATED SUBSTRATE COMMUNI-TIES. Like the preceding community,theses are manipulated by man. Placingartificial or natural substrates in a bodyof water will cause these communitiesto appear thereon.

K We will again emphasize ARBITRARY,because organisms confound our neatlittle schemes to classify them. Manymove from one community to anotherfor variousreasims. However, allthese basic scheme do have intrinsicvalue, provided they are used with

- reason:

REFERENCES

1 Cummins, Kenneth W. The Ecologyof Running Waters, Theory andPractice in Proc. Sandusky RiverSymposium. International JointCommission. 1975.

2 Leopold, Aldo. A Sand County Almanac.Oxford University Press. 1966.

3 Peters, Robert Henry. The UnpredictableProblems of Tropho-dynamics.Env. Biol. Fish 2:97-101. 1977.

This outline was prepared by R. M. Sinclair,Aquatic Biologist, National Training andOperational Technology Center, MOTD,OWPO, USEPA, Cincinnati, Ohio 45268.

Descriptors: Biological Communities. ,

44,2 -3

LIMNOLOGY AND ECOLOGY OF PLANKTON

I INTRODUCTION

A Most Interference _Organisms are4ap Small.

B Small Organisms generally haveShort Life Histories.

.0 POpulations of Organisms withShort Life Histories may FluctuateRapidly in Response to Key Environ-

, mental Changes.

D Small Organisms are Relativelyat the Mercy of the Elements

E The Following Discussion willAnalyze the Nature of These'Ele-

. ments with Reference to the Res-. ponse of Important Organisms;

TI PHYSICAL FACTORS OF THE ENVIRONrMENT

A Light is a Fundamlental SoUrce.ofEnergy for Life and Heat,"

1 Insolation-is affected by geo-graphical location and mete-orological factors.

2 Light penetration in water isaffected by'angle of .incidence(geographicalY, turbidity, andcolor. 'he. propOrtion of lightreflected depends on the angle )of incidence, the temperature,color, and other qualities ofthe water. In general, as thedepth increases arithmetically,the light tends to decrease geo-:metrically, Blues greens, andyellows tend to penetrate mostdeeply while ultraviolet, vio-lets, and orange-reds are mostquickly absorbed. On the orderof 90% of the total illuminationwhich penetrates the surfacefilm es absorbed in the first10 meters of even the clearestwitdr.

3 TuXidity may originate within

45..

4'B

br outside of a lake.

a That which comes in fromoutside (allochthonous) ispredominately inert solids(ttripton):

b That of internal origin (auto-chthonous) tends totbe bio-1ogieal in nature.

Heat and Temperature Phenomenaare Important in Aquatic Ecology.

1 The total 4uantityof heat avail-. able to a body of water per yearcan be calculated and is knownas the heat budget.

2. Heat is derived directly from in-solation; alsO by transfer fromair, internal friction, and othersources.

Density Phenomena

'1 Density And viscosity affect thefloatation and locomotion ofmicroorganisms.

a Pure fresh water achievesits maximum density. at 4 °Cand its maximum viscosityat 0 °C.

b The "rate of change of densityincreases with the temperature.

2 Density stratification affectaquatic life and water uses.

a In summer, a mass of warmsurface water, the epilimnion,is usually present and separatedfrom a cool deeper mass, thehypolimnion, bYa relativelythin layer known as thethermocline.

'41

b Ice cover and annualspringand fall overturns are due tosuccessive seasonal changesin the relative densities ofthe epilimnion and the hypo-

3,1/

3-1

Limno loley and EcologyAokPlankton

limnion, profoundly influ-enced by prevailing meteoro7logical conditions.

c The sudden exchange ofwater masses having differ-ent chemical characteris-tics may have catastrophiceffects on bertain biota, maycause others to bloom.

d Silt laden waters may seekcertain levels, dependingon their own specific gravityin relation to existing layersalready present.

Saline waters will alsostratify according to therelatiye densities of theVarious layers.

osity of water is greatertemperatures.

a This is important not Onlyin situations involving thecontrol of flowing water as ina.sand filter, but also sinceovercoming resistance to

. flow generates eat, it issignificant in the heating

- of water by internal frictionfrom waive and current ac-tion and many delay theestablishment of anchorice under critical conditions.

3 Theoriseat lower

\

,

b It is easier for planktonto remain suspended in coldviscous (and also dense)water than in less Viscouswarm water. This is re-flected in differences in theappearance of winter vssummer forms of life (alsoarctic vs tropical).

D Shore development, depth, inflow -outflow pattern, and topographicfeatures affect the behavior of the water.

E Water movements that may affect organ-isms include such phenomena as waves,currents, tides, seicheS, floods, andothers.

1 Waves or rhythmic movement

4.

The best luiolAn are travelingwaves. These are effectiveonly against objects.nearthe surface.. They have littleeffect on the movement oflarge masses of water.

tsb Standing waves or seiches

ocdur, in all lakes but areseldom-large enough to be

observed. An "internal seich"is an oscillation in a densitymass within a lake with nosurface manifestation maycause considerable watermoVement.

4G

ly

2 Currents

a Currents are arhythmicwater movements which havehad major study only in ocean-ography. They primarilyare concerned with the trans-location of water masses.They may be generated inter-nally by virtue of density .

changes, or externally bywing or runoff..

Turbulence phenomena oreddy currents are largely re-sponsible for lateral mixingin a current. These are offar more importance in theeconomy of a body of water,than mere laminar flow.

4c Tides, or'rathetidalcurrents, are reversible(or oscillatory) on a relative-ly long and iiredictable period.They are closely allied toseiches. For all practicalpurposes, they are restrictedto oceanic (especially-coastal)waters.

If there is no freshwaterinflow involved, tidal currentsare basically "in and out;"if a significant amount offreshwater is added to thesystem at a constant rate, theoutflowing current will in generalexceed the inflow by the amountof freshwater inpvt.

ee.

ti4

Limnology and Ecology of Plankton

There are typically two tidalcycles per lunar day (approx-imately 25 hours), but there iscontinuous gradation from thisto only one cycle per (lunar) dayin some places,

Estuarine plankton populationsare extremely influenced by localtidal patterns.

d Flood waters ranmfrom torren-. tial velocities which tear away

and transport vast masses ofsubstrate to quiet backwaterswhich may inundate normally dryand areas for extended periods

o time. In the former case, ..._

pl ktonic life is flushed awayco pletely; in the latter, a localpla on bloom may develop whichmay of immediate significance,or which may serve as an inoculumfor receding waters.

F Surface Tension and the Surface Film

1 The surface film is the habitat ofthe "neuston", a group of special "-significance.

2* Surface tension lowered by surfactants"may eliminate the neuston.' This canbe, a significant biological observation.

III DISSOLVED SUBSTANCES

A Carbon dioxide is reanimals in respir

eased by plants andon but taken in by

plants in photosynthesis.

Oxygen is the biblogical complement ofcarbon dioxide, 'a.nd'necessary for allanimal life.

C ,Nitrogen and phosphorus are fundamentalnutrients for plant life.

47'

4Occurin great dilution, Concentratedby plants.

3-3

Limnology and Ecology of PlanktOn

The distribution of nitrogencompounds is generally corred with the oxygen curve, espcially in oceans.

1111.01

D Iron, manganese, sulphur, and siliconare other minerals important to aquaticlife which exhibit biological stratification.

.E Many other minerals are present but theirbiological distribution in waters is lesswell known? fluorine, .tin, and vanadium

shave recently been added to the "essential"list, and more may well follow. ,

F Dissolved organic matter is present ineven the purest of lakes.

BIOLOGICAL FACTORS

A Isttritional Classification of Organisms

Holophyt ic or independent or-ganisms, like green plants, pro-duce their own basic food elementsfrom the physical environment. ,

2 Holozoic or dependent organisms,like animals, ingest and digestsolid food particles of organic

1

3. Saprophytic or carrion eatingorganisms, like many fungi andbacteria, digest and assimilatethe dead bodies of other organ--isms or their products.

B The Prey-Predator Relationship isSimply one Organism ating Another.

Toxic and Horm c Relationships

1 Some organisms such as certainblue green algae apd some ar-mored flagellages produce sstances poisonous to others.

2 Antibiotic action in nature is' not well understood but has been

shown,to play a very influentialrole in the economy of nature.

"3-4

ti

4

0V BIOTIC COMMUNITIES (OR ECOSYSTEMS)

,A A biotic community will be defined.here

j as an assemblage of organisms living ina given ecological niche (as definedbelow). Producer (plant - like); consumer(animal-like) and reducer (bacteria andfungi) organisms are usually .included.A source of energy (nutrient, food) mustalso be present. The essential conceptin that each so-called 'community is arelatively independent entity. ArCtuallythis position is only tenable at any giveninstant, as individuals are constantlyshifting from one community to another inresponse to stages in their life cycles,physical conditions, etc. The only onetosbe considered in detail here is theplankton.

B Plankton are the macroscopic andmicroscopic animals, *ants, bacteria,etc. floating free in the open water.Many clog filters,. cause tastes, odors,and other troubles in water supplies.

4P

1 Those that pass !through a planktonnet (No. 25 silk bolting cloth orequivalent) or sand filter are oftenknown as nannopankton (theyusually greatly exceed the "net"plankton in actual quantity).

2 Those less than four microns inlength are sometimes calledultraplankton.

There are many ways in whichplankton may be classified: taxo-nomic,- ecological, industrial:

Mit4 The concentration of plankton varies

markedly in space and time.

Depth, light,. currents, andwater quality profoundly affectplankton distribution.

The relative abundance ofplankton in the; various sea-sons is generally:

0

1 sprjng, 2 fall, 3 summer,4 winter

.40

1

. 1.

Limnology and EColbgy of Plankton

5 Marine plankton include many_larger animal forms than arefound in freshwaters.

C Th benthic community is generallycgixsidered to be the macroscopic lifeliving in or on the bottom.

es,D The peon community might be

defined as the microscopic benthos,except alai they are by no Means confinedto the bottom. Any surface, floating, ornot, is usually covered by 'film of livingorganisms. ,There is frequent exchangebetWeen the periphytdn and planktoncommunities.

E The nekton is the comMunityof larger,free-swimming animals (fishes, shrimps,etc. ), and so is dependent on the othercommunities for basic plant foods.

F "Neuston or Pleuston

This community inhabits the air/waterinterface, and may be suspended aboveor below it or break it. 'Naturally thisinterface is a very critical One, it beingmicro molecular and allowing interchangebetween atmospheric contaminants andthe water medium./ Rich in bacteria,metals, protozoi, pesticides etc.

I.

VI TIIE EVOLUTION OF WATERS _-

A, The history of a body of water determinesits present condition. Naturalswaters hayeevolvedin the course of geologic timeto what we know today..

. ,B In the. course of their evolution, streams

in general pass through four generalstages of development which may be called:birth, ypvth, Maturity, and old age.

.. ,

Estallishment of birth. In an °.extant' stream, .this might bea "dry run's or headwaterstreambed, before it had erodeddown to the level of ground Water.

2 Youthfultreams; when thestream bed is eroded below thegrouhd water level, spring wateresters and the stream becomes

,,.permanent.

3 Mature streams; have widevalleys, a developed flood plain,

ot4

.deeper, more turbid, and usuallywarmer water, sand, mud, silt,or clay,bottorn -materials whichshift with increase in flow.

4 In old age, streams have appi-oa-ched base level. During floodstage they scour their bed and de-posit materials on the flood plainwhich may be very broad an/ flat.During normal flow the etteneris.._'refilled and many shifting bars aredeveloped.

(Under the influence of man thispatternmaygle broken up, or tem-porarily interrupted. Thus as essen-tially "youthful" stream might takeOn some of the characteristics of a.'.'mature" stream following soil -

erosion, organic enrichment, andincreased surface runoff. Correctionof these conditions might likewise befollowed by at least a partial rever-sion to the "original" condition.)

C Lakes have a developmental historywhich somewhat parallels that of streams.

I°1

1 The method of formation greatlyinfluences the character and sub-sequent history of lakes.;

2 Maturing or natural eutrophicationof lakes

e,a If not already present, shoal

areas are developed throughro.?iop. of the shore by wave

-action and undertow.

b Currents produce bars across'bays and thus cut off irregularsareas.

c Silt brought in by tributtrystreams settles eut in the quietlake water.

d Rooted aquatics grow on shoalsand bars, and in doing so cutoff bays and contribute to thefilling of thelake.

. ,e Dissolved carbonates and other

materials 'are precipitated in thedeeper portions of the lake inpart through the action of plants.

Limno logy and of

.

f When filling is well' ad.anced. sphagnum mats extend Nt-

ward from the shore. Ttibse-mats are followed by sedgesand grasses chfinalkyconvert thelak Into amarsh.

3 Extinction of lakes. After lakesA'reach maturity their progresstoward -hp is accelerated.They become extinct through:

'a The downcuitink Of the out,

let.

b° Filling with detritus eroded, 'from the shores or broughtin by tributary streams.

Q

c Fillingby the acca tion of.s.the remains orvegetable-

geowinen the'e itself. 's

a

t(Oftentiro or 'three pro- ."

cesses may act concurrently)o

When mth hastens tie abode : .,"process, leis Often called"cultural eutrophication. "

CI

VII PRODUCTIVITYQ.

A Thebiological resultant of all physicalsand ctlemical factors is the quantity oflife that may actually be present. Theability to produce this "biomass" isoften referred to as the "produCtivity"

1' of a body of water. This-is neither goodnor bad per se. Aater of low producti-vity is a "poor" water. biologically, andalso a relatively "pure" or "clean" water;hence desirable as a water supply. Aproductive water on the other nand maybe a 'nuisance to man or highly desirable.Some of the factors which influence theproductivity of-waters are as ..follows:

B Factors affecting stream productivity.-A*To be productive of plankton, a, streammust provide adequate nutrients, light,a suitable temperature, and time forgrowth to take place.

.>

o

1 .z Youthful streams, especially onrock or sand substrates are lowin essential nutrients. Tempera-tures in mountainous regions areusually low, and due to the steepgradient; time for growth is short.Although ample light is available,growth of true plankton is thusgreatly limited.

2 As the stream flows toward amore 4'mature" condition nutrients

14 .4.4,1%tend tp accumulate, and gradientdiminishes 91 so time of flow ,

increases, temperature tends toincrease, and plankton flourish.

_Should a heavy load of inert siltdevelop on the other hand, theturbidity would reduce the light

' penetration and, consequently thegeneral plankton production would

As the stream approaches baselevel (old age)' and the time avail-able for plankton growth increases, .

. L. the balance between turbidity,nutrient levels, and temperatureand other_seasonal conditions,detdmines-the overall peoduc-tivit37.- '`

C.' F-actors Affecting the Productivityof ,Lakes i

.

'''10" The size, shapeand depth,of the lake basin: Shallow wateris more productive than deeperwater .since more light will reachstile bottom to stimulate rootedplant growth. As a corollary,lakeslvith more shoreline, havingmore shallow water, are in generalmore productive. Broad shallow

Vekes and reservoirs have thesreatesttproduction potential (andhence should_ be- avoided for watersupplies).

2 Hard waters are generally moreprodhdtive than soft waters asthere are more plant nutrientmtperaleavailable. This is often

5d

0.

51

,)a

HumanInfluence

SewageAgriculture

Mining

PrimaryNutritivMaterial

a.O

FACTORS AFFECTING PRODUCTIVITY

Geographic Location

GeologicalPormatign

Competitionof Substrate

.She of Basin

t.

LatitudeLongitude

Altitude

Climate

Drainage Dept"... Area Bottbm Precipitation- Area Conforination

I I a II h 04W.

Wind

Insolation

Nature of Inflow of Trans- --31....Light edt Penetration 4,2 Penetra Develop o Seasonal Cycle

Bottom A llochthon oils pa. r en cy Penetration and Stratification-01.- and Littoral Circulit. StagnationDeposits Materials ptilizatiop Region Growing Season:.

Trophic Nature of a Lake

O

tz1

16'

ro

'Limnology and Ecology of Plankton

WOO

.

greatly influenced by thecharacter' of the soil and rocksiiithe watershed, and the qualityand quantity of ground waterentering thd lake. lir general,pliTanges of 6.8 to 8.2 appearto be most productive.

3 Turbidity'reduces produCtivityas light penetration is reduced.

4 The presence or absence ofthermal stratification with itssemi-;annual turnovers affectProductivity by distributingnutrients throughout the watermass.

5 Climate, temperature, pre -valance of ice and snow., arealso important.,

Di Factors AffeCting the Productivity ofReservoirs

3 Reservoir discharges also pro-formdly affect the DO, temperature,and turbidity lr the stream belowa dam. Too much°fluctuation inflow may permit sections of thestream to dry periodically.

VIII CLASSIFICATION OF LAKES ANDRESERVOIRS

A The productivity. of lakes and impound-ments is such a conspicuous feature ,

that it is often used as a means ofclassification.

a

2

1 The productivity of reservoirsis governed by much the sameprinciple's as that of lakes, withthe difference that the waterlevel is much more under thecontrol of man. Fluctuations

r in water levelican be used todeliberately increase of decreaseproductivity. This can be dem- *onstrated by a comparison ofthe TVA reservoirs which practicea summer drawdown with some ofthose in the west where a winterdrawdown is the rule.

2 The level at whiir water is re-moved from the /reservoir is alsoimportant( The upper epilimnionmay have a high plankton turbi-dity while lower down the planktoncount may be less, but a tasteand odor causer {such as M_all_o-I be present. Theremaybetwo thermoclines, with -a mass of muddy water flr3wingbetween-a clear upper epilimnionand a clear hypolimnion. Othercombinations ad infinitum mayoccur.

.11

1 Oligotroxhic lakes are thegeologically younger, less produc- °

tive lakes, which are deep, haveclear water, and usually support 6

Salmonoid fishes.

Mesotroac lakes are generallyintermediate between oligotrophicandeutrophic lakes. They aremoderately productive, yetpleasant.to be around.

3 Eutrophic lakes gre more mature;more turbid, and richer. Theyare -usually shallower. They arericher in dissolved solids; N; p,4;ap d Ca are abundant. Plankton is-abundant and there is often a-ricli bottom'fauna. Nuisanceconditions often appear.

4 Dystrophic lakes - bog lake?:low in pH, wate yellow to brown,

I dissolved solids; N, P, and Cascanty but humic materials abun,

. dant; bottom fauna and planktonpoor,

,and fish species are limited.

dr.

B Reservoirs may be classified as storage,or run of the rider.

1 Storage reservoirs have a large4 volume in relation to .their inflow.

4 ).

2 'Run of thel river reservoirs havea large flow throUgh in relation,to their storage value.

A.

.,,

53,

1

S.

Irmamman........

'According to location, lakes andreservoirs may be classified as polar,temperate, or tropical. Differencesinclimatic and geographic conditionsresult in aiffvences in their biology.

rDC THE MANAGEMENT OR CONTROL-OF...

ENVIRONMENTAL FACTORS

A Liebig's Law of the Minimum statesthat productivity is limited by thenutrient present iri the least amoungat any given time relative to the.assimilative capacity of-the organism.

B ShelfOrd's Law of T oler.ation:

Minimum Unlitof toleration

--Wange of Optimum'of factor

Maximum limit oftoleration

Absent Greatest abundance -t-ia-DecreasingAbundance

-4-DecreasingAbundance

C The artificial introductio,(sewage pollution or Perttendsto eliminate existingminimums for some specieintolerable maximums for o

1

of nutrientszer) thus

itingand createher species.

Known limiting min umsimaysometimes be delib ratelymaintained.

2 As the total available ensupply is increased,

; tends to increase.

3 As productivity, increases,, thewhole character o; the watermay be changed from a Meagerly

. productive clear water lake(oligotrophic).to a highly pro-ductive and Usually turbid lake

.,(eutrophia..11

. 4 Eutrophecation leads to tesatmenttroutRes.

gyductivity

a

0 N.11A

Limnology and Ecology of Plankton

to maintain minimum limiting nu-tritional factors.

2 Shading out the energy ofinsola-tion by roofing or inert turbidity;suppresses photosynthesis.

3 Introduction of substances toxicto some fundamental patt of thefood chain (such as copper sul-phate) tends to temporarily inhibitproductivity..

X .SUIVIMARY

A A body of water such as a lake rep-resents an Intricately balanced systemin a state of dyhainic equilibrium. `Modification imposed at one point inthe system automatically results in

' compensatory adjustments at associatedpoints.

*orB The more thorough our knowledge ofthe entire systeM, the better we canjudge where to impòfe control measuresto abhieve a desired result.

REFERENCES .

1 Chamberlin, Thomas C., and SalisburgiO4.Rollin P., Geology Vol. 1, "Geological Processes:and Their Results",pp irzlix, 1.nd 1-654, Henry Holt and_Company, , 904.

meat. ..r.aperties ofOrdinary Water - SUbstario .

Reinhold Publ. Corp., 17ew York.pp: 1-673. 1940.

D . Control of eutrophication may be accom-.plisbed by various means

- Watershed management, ade-quate preparation of .reservoir.sites, and Pollution control tend.'-

.

13 Hutchesoni-George E. A Treatise onLirnnOlogy. John Wiley Company.

4 Wuttner, Franz. Fundamentals ofLimnology. University of Tprontq

"Press. pp. 1-242. 1953.

5 Tarzwell, Clarence M. ExPerimezitalEvidence on the Value of Trout 1937Stmettm Improvement in Michigan.

. American Fisheries Society Trans,661177-187. 1936.

. .1 .:317..

Limnology and EcOlogy of Plankton

,.6 US DHEW. PHS. Algae and MetropolitanWastes, transactions of a seminarheld April 2I-29, 1960 at theRobert A,;' Taft Sanitary Engineer-ing Center, Cincinnati, Ohio.No. SEC TR IN61-3.

7 Ward and Whipple. Freshwater Biology(Introduction). JAn WileyCompany. 1918.,

8 Whittaker, 'R. Ir Communities andEcosystems. MacmillanNeW York. 162 pp.' 1970.

9 Zhadin, V. I. and perd, Sr. Faunaand Flora of the lakes and

- Reservoirs of the USSR., Avail-able from the Office of TechnicalSer4ices, U. S. Dept. Commerce,Washington, DC.

o'41

10 Josephs, lifelvin and Sanders, Howard J.

ACS Publications, WashAgton, DC.Chemistry and the Environment

1967.p

11 Sournia, A. (editor), PhytoplanktonManual. UNESCO. 1978. .

,This outlike"was prepared by IL W. Jackson,formai Chief Biologist,- National TrainingCenter, and z;evisect by 'R. M. Sinclair,Aquatic .Biologist,' NationarTrairiing andOperational Technology Center, ()WYO.' 'USEPA, Cincinnati, Ohio 45268.

-40P

Descriptors: Plankton, Ecology,. Limnology

.t

11,

t

BIOLOGY OF ZOOPLANKTON COMMUNITIES

I CLASSIFICATION

A The planktonic cornmanity is composed oforganisms that are relatively independentof the bottom to complete their life history.They inhabit the open water of lakes(pelagic zone). Some Species have inactiveor resting stages that lie ()Ole bottomand carry the species through periods ofstress; e. g. , winter, A few burrow irithe mud and enter the pelagic zone at night,but most live in the open water all thbtime that the species is present in an activeform.

B Compared to the bottom fauna and flora,the plankton, consists of relatively fewkinds of organisms that are consistentlyand abundantly present. Two major 'cat-egories are often called phytoplankton(plants) and zooplanktonAanimals), butthis is based on an outmoded classificationof living things. The modern tendency isto identify groupings according to theirfunction in the ecosystem: Primary pro-.ducers (photosynthetic organisms), consum(zooplankton), and decomposers (hetero-trophiC.bacteria and fungi).

C, The primary difference ken is nutritional;phytoplankton use inorganic nutrientelements and solar radiation. Zooplanktonfeed on particles, much of which can bephytoplankton cells, but'can be bacteria orparticles of dead organisms (detritus)originating in the plankton, the shoreregion; or the land surrounding the lake.

'D The swimming powers of .planktonicorganisms is so limited that their hort-zontal distributioniis determined mostlyby movements of water. Some of the

viek animals are able to swim fast enough thatthey can migrate vertically tens of meterseach day, but they are capable of littlehorizontal navigation. At most somespecies of crustaceans show a generalavoidance of%the shore areas during,calmweather when the water is movinernoreslowly than the animals can swim.47Bydefinition, animals that are able to controltheir horizontal location are nekton, notplankton.

BI..Aci.29. 6. 46

E In this presentation, a minimum of clas-sification and taxonomy is used, but itshould be, realized that each group is ,

typified by adaptations of structure onphysiology that are related to the plank-tonic mode of existence. These adapta-tions are reflected in the classification.

II FRESHWATER ZOOPLANKTON

A The, freshwater zooplankton is dominatedby representatives of three groups ofanimals, two of them crustaceans:Copepoda, Cladocera, Rotifera. All havefeeding mechanisins that permit a highdegree of selectivity of food, and two canproduce resting eggs that can withstandsevere environmental conditions. Ingeneral the food of usual zooplankton pop-\ ulations ranges from bacteria and, smallalgae to small animals.

B The C.opepoda reproduce by a normalbiparental process, and the females layfertilized eggs in groups which are carriedaround in sacs until they hatch. Theimmature animals go through an elaboratedevelopment with many stages. The later'stages have mouthparts that permit themto collect particles. In many cases, theseare in the form of combs which removesmallparticlps by a sort of filtrationprocess. In others, they are modified toform grasping organs by which smallanimals or large algae are captured

'individually.

C The Cladocera (represented by Daphnia)reproduce much of the time by partheno-

iesis, so that only females are present.Eggs are held by the mother in a broodchamber until the young are developed farenough prrfepd for themselves. The newbornanimals look like

seriesand do

not go through an elaborate of,developmental stages in the water as dothe copepods, oaphni.a, has

somefiltering structures on some of its legs ,that act as filters.

,56

ti

-r

Biology of Zooplankton Communities

D Under some environmental conditions-the'development of eggs is affected and malesare produced. Fertilized eggs are producedthat can resist freezing and drying, andthese carry the population thrbughunsatisfactory conditions.

E The Rotifera are small animals.with'aciliated area on the head which createscurrents used both for locomotion, and forbringing food 'particles to the mouth. Theytoo reproduce by parthenogenesis duringmuch of the year, but production of malesresults in fertilized, resistant resting eggs.Most rotifers lay eggs one at a time andcarry them until they hatch.

III ZOOPLANKTON POPULATION DYNAMICS,

A In general, zooplankton populations are ata minimum in the cold seasons, althoughsome species flourish in cold water. Specieswith similar food requirements seern'toreproduce at different times of.the year orare segregated in-di#e-nent layers of lakes. ,

B There is no single, simple measurementof activity for the zooplankton as a wholethat can be used as an.index of productionas can the uptake of radioactive carbon forthe phytoplankton. However, it is possibleto find the rate of reproduction of species'that carry their eggs. The basis of themethod is that the number of-eggs in asample taken a #a given time representsthe number 9f animals that will be addedto the population during an interval thatis equal to the length of time it takes theeggs to develop. Thus the potential growth'rate of.the populations can be determined.The-actual growth rate, determined bysuccessive samplings and counting, is lessthan the potential, and the difference is ameasure of the death rate..

C Such measurements of birth and death ratespermits a more penetrating analysis to bemade of the causes of population changethan if data were available for populationsize alone.

D Following is an indication of the majorenvironmental factors in the control ofzooplankton.

t-o

1 Temperature has an obvious effect inits general control of rates. ,In addition,the production and hatching of restingeggs may be affected.

4-2

2 Inorganic materials

Freshwater lakes vary in the contentof dissolved solids according to thegeological situation. Tile total salinityand proportion of different dissolvedmaterials in water cartIffect the pop-ulation. Some species are limited tosoft water, others to saline iaterl, asthe brine shrimp.. The maxim.= pop-ulation size developed may be relatedto salinity, but thisris probably'an.indirect effect working through `theabundance of nutrients and productionof food.

3 Food supply

Very strong corrylations have beenfound between reproduction and foodsupply as measured by abundance ofphytoplankton. The rate of food supplycan affect almost.,all aspects of pop -ulation .biology including rate of indi-vidual growth-time of maturity, rateof reproduction and length of life.

4 Apparently in freshwater, dissolvedorganic materials are of little nutri-tional significance, although somespecies can be kept -if the concentration.of dissolved material is high enough.Some species require definite vitaminsin the food.

5 Effect of predation on populations

The kind, quantity and relative pro-portions of species strongly affectedby grazing by vertebrate and inverte-brate predators. The death rate ofDaphnia is correlated with the abun-dance of a predator. Planktivorousfish (alewives) selectively feed onlarger species, so a lake with alewivesis dominated by the smaller species ofcrustaceads and large ones are scarceor absent.

6 Other aspects of zooplankton

Many species migrate vertically con-siderable distances each day. Typically,migrating species spend the daylighthours deep in the lake and rite 'towardthe surface in late afternoon and early'evening.

Some species go through a seasonalchange of form (cyclomorphosis) whichis not fully understood. It may have aneffect 4n reducing predation.

57

REFERENCES

1 Baker, A. L. An Inexpensive Micro-sampler. Limnol. and Oceanogr.15(5) :. .158-160. 1970.

.

2 Brooks, J. L. and DOdson, S. I.Predation, ,Body, Size, and Corn-pqSition of Plankton. Science 150:28 -35. 1965.

3 Dodson, Stanley I, ComplementaryFeeding Niches Sustained by Size.-Selective Pl-edation, Limnologyand Ocenography 15(1): 131-137.

4 Hutchinson, G. E. 1967. A Treatiseon Limnology. Vol. II. IitrodUctionto Lake Biology and the Limnoplankton.xi + 1115. John Wiley & Sons, Inc.New York.

5. Jossi, Jack W. Annotated Bibliographyof Zooplankton Sampling peyices.USFWS. Spec, Sci. : Rep. -Fisheries.609. 90 pp. 1970.

6 Likens, Gene E. and Gilbert, John J.Notes on Quantitativ,e Sampling ofNatural Populations of PlanktonicRotifers. Limnol. and Oceanogr.15(5): 816-820. .

Biolo&Srof Zooplankton Communikies

7 Lund, J. W. G. 1965. The Ecology ofthe Freshwater Plankton. BiologicalReviews, 40:231-293.

8 UNESCO. Zoolplankton Sampling.UNESCO Monogr. Oceanogr. Methodol.2. 17.4 pp. 1968. (UNESCO. -Placede Fortenoy, 75, Paris 7e France).

'This outline was prepared by W. T. Edmondson,Professor of Zoology, University ofWashington, Seattle, Washington.

Descriptor: Zooplankton

53 4-3

L

4

Biology of Zoopl nkton Communities 1111

,

FIGURE 1 SEASONAL CHANGES OF ZOOPLANKTON IN LAKE ERKEN, S-WEDBN-

Keratella hiemali

Kellicottia longispi

Polyarthra vulgaris

Daphnia longispina

"144.1Ceriodaphnia quadrangula

Bosmina coregoni

a a a a a a

J P M

a

M -J J A S 0 N D

1957

Each panel shows the abundan4 of a species of animal.- Each

mark on the vertical axis represents 10 individuals/liter.Nadwerck, A. 1963.11 Die Beziehungen zwischen Zooplankton andPhytoplankton im See Erken. Symbolae Botanicae Upsaltensis, 1,7:1-163.

59

CSI

FIGURE- 2 REPRODUCTIVE RATE OF ZOOPLANKTON AS A FUNCTION OF ABUNDANCE OF FOOD

B

0.20,

a0

000

0.15

0.10

.

co

ae

0.05

'00

Abundance

.. .

*

7 .

.

. 7 .9

..

1

.,. Temperature

more than 10o

...

.

., Temperatureless thanI00

: .

.

.

.

.

200

of food

400 600

organisms ugm /1, dry weight

4Mean rate.of'laying eggs by the planktonicrotifer Keratella cochlearis in natural

populations ap a function of abundance offood'organisms and temperature. W. T.

Edmonton. 1965. ReprOducEive rate ofplanktonic rotifers as related to food

and temperature in nature. Ecol. Mmogr.

35: 61-111.

Young perBrood

:Total young

223,

-177

132

76

Number of young producedin each brood by Daphnia living infour different concentrations of food organisms, rewewed

daily. The total numbetaproduced during the life of a 'mother is shown by the numbers at the right. The Daphnia

at the two lowest concentrations produced their first batch

of eggs on the same day as the others, but the eggs degen-erated, and the first viable eggs were released two days

later. Richman, S. 1958. The transformation of energy by

Daphnia pulex. Ecol. Monogr. 28: 273-291.

1

61

fi

Biology of Zooplankton Commuriities

<,

`Stentor

PROTOZOA.

9,

-

Difflugia

Amoebae

Co-donella

Ciliates

ft.

Epistyks7.

I

.-....-.-

Cladocera

0

1414."


ROT I FERA

Polygarthra

. .

ARTHROPODA

crustaCea

Brachionus

Copepoda

Nauplius larva of copepod

Ins'ecta Chaoborus

ON

. ,

..,. . . . ..- a .

El!,

4

0

I44.

*I.*, .":1

,


PLANKTONIC BIV VE RVAE

380p.

1,1

spinet (fin nttached)..

r

2

864.

3°

sim le (gill attached)

Glochidia '(Unionidae) Fish Parasites-' (1-3)

0

0

s

1

veliger

Veliger Larva6 (Corbiculidae) Free LiArAng Planktonica . ..(4-5)

6.

Pedivpliger' attaches byssus lines)

61

fr/S-a

4 ?*i^

b.

I OPTICS

OPTICS AND n-1E-MICROSCOPE

An understanding of elementary optics isessential to theproper use of the microscope.The microscopist will find th4t unusual pro-blems in illumination and photomicrographycan be handled much more effectively bncethe,underlying ideas in physical opti7 areunderstood.

A Reflection

A food place to begin is-with reflection ata 'surface or interface. Specular (onregular) reflection results when d beamof light leaves a surface at-the same angleat which itreached it. This type ofreflection occurs with highly, polishedsmooth surfaces. It Is stated more pre-cisely as Snell's Law, is e. ,'the angle ofincidence, i, ,is equal to the angle .ofreflection, r (Figure1). Diffuse (or-scattered) reflection results when abeamof light strikes a rough or irregular sur-face and different portions of the incidentlight are reflected from the surface atdifferent 'angles. The light reflected froma piece of white paper or a ground glass isan'example of diffuse reflection.

. Figure 1

SPECULAR REFLECTION SINTELLIS

LAW,., -

,BI. MIC. 18..2. 79

1'

4

Strictly, speaking, of course, all reflectedlight, even diffuse, obey/4 Snell's Law.Diffuse reflected light is made up 8l manyspecularly .reflected rays, each from aa tiny element of surface, and appearsdiffuse when the reflecting elements are'very numerous and very small. The termsdiffuse and .specular, referring to reflection,describe not so much a difference in thenature of the reflection but rather a differ-ence in the type of surface. A polished sur-face gives specular reflection; a roughsurface gives diffuse reflection.

It is also important to note and rememberthat specularly reflected light tends to'be

.strongly polarized in the plane of the reflect-ing surface. This is due to the fact thatthose rays whose vibration directions lieclosest to the plane of the reflection surfaceare most strongly reflected. This effect isstrongest when the angle of incidence issuch that the tangent of the angle is equalto the refractive index of the reflecting sur-face. This particular angle of incidence iscalled the Brewster angle.

B Image Formation on Reflection.

Considering reflection by mirrors, we find(Figure 2) that a plane mirror forms avirtual image behind the mirror, reveredright to left but of the same size as theobject: The word virtual means that the

,image appears to be in a Oren plane butthat a ground glass screen or a photographicfilm placed in that plane would stiOw noimage.' The converse of a virtuariniage isa real image. '

Spherical 'mirrors are either convex or con-cave with the surface of the mirror repre-senting a portion of the surface of a sphere.The center of Curvature is the...center of theSphere, part orwhose surface forms Themirror: The Tocus,lies halfway bb'tween the

' -center of curvature 1144:14he milk9F surface..7,

5-1

,

Opttr..5 and the Microscope

°bloat

4.

Mirror

t*Figure 2

IMAGE FORMATION BY PLANE MIRROR

itI

VirtualImago

Construction of an image by a concavemirror follows from. the ,two premisesgiven below (Figure 3):

<I)

c_ ( Figure -3

IMAGE FORMATION BY CONCAVE MIRROR.

111

1 A ray of light parallel to the axis ofthe mirror must pass through thd'foCus after: reflection.

2 A ray of light which passes through thecenter of. curvature mast return alongthe sarrie pat11.-

11.

A corollary of the firstpremiseis:N

<

,

5-2

a

,<

3 A ray of light which passes through thefocus is reflected parallel to the axisof the mirror. .

- The image from an object can tie locatedusing the familiar lens formula:

+I 1

where p = distanCe from the object tothe mirror

q = distance from the image tothe mirror

f = focal length

C Spherical Aberration

No spheyical surface can be paeftct in itsimage-forming ability. The most seriousof the imperfections,' spherical aberration,occurs in spherical mirrors of largeaperture (Figure 4). The rays of light

,making up an image point from the outerzone of a sphericd.1 mirror do not passthrough the same point as the more centralrays. This type of aberration is reduced byblocking the outer zone rays from the imagearea or by-using aspheric'surfaces.

Figure 4

SPHERICAL ABERRATION BYSPHERICAL MIRROR

66.

- .

O

Optics and the Micrpscope

D Refiaction Of Light

Turning now.to lenses rather than mirrorswe find that the inostìninortant t liaicte-istic is refraction. Reratilni releys tothe change of dirlction and/or velocity Oflight as it passed from one 111..diutil toanother. The ratio of the veloc it) in air

, (or more corrcu.tly in a vacuum) to, thevelocity ip-th«.-; 111 rdit1111 i, called theLte frac ti've index. Some.. typical values ofrefractive index measured with mono-

into focus and the new micrometer readingis taken. Finally, the microscope is re-focused until the surface of. the liquid appearsin sharp focus. The micrometer reading.,is taken again and, with this information, .

the 'refractive index may be calculated fromthe simplified equation:

active index actual depthapparent depth

, chromatic' light (sodium I) line) are listedin Table 1.

.

.Refraction causes an object inimer,s(el in

'Table I REFRACTIVE INDICES OE COMMONMATERIALS MEASURED WITH SODIUM LIGHT

a medium of higher refractive index than V a't uu m 1.0,000000 Crown glass. 1.48 to 1.61air to appear Olosert.to the surface thah it Air 1.00.029 i 8 Rock salt)oirs:.1.5443actually is (Figure 5). This effect CO2,

WateL0004498

'I. 3330Diamond 2.417Lead sulfide 1E3.912

re

Actual-.depth ,

Apparentdepth

.33

.Figuse 5

REFRACTION OFfrL,H.T41' INTEAFACE

,

Air

Medhim

'TOO

Ob 4ject-.0.

11 4 s. F 1.

be used to dete'rimine th,e,gpfractive indexor a.liquid:with the nlivipscope. A flat

'. ;,t . vial`with a. scHtcl).eiti Otte bottom cipside)cr ,. is Placed,ern the' stage of the mic"rbscope.e.

.. The' microscope iS fo'cubed on ale stratchf and the fine adjustment micrometer reading

. 1. .. is noted. A small amount of the unknotArn.

liquid is added: the scratch is again brought ., .., . ,

.., . , ...

-... .

When the situation' is reversed, add a rayof light from a medium of high refractiveindex p'a4es through the interfaCe of amedium of lower index, the ray is refracteduntil a critical angle.is reached beyond whichall pf the ,light is reflected frOm the interface(Figure 6). This critical angle, C, has thefolldwing relationship to the refracti've irdicesof the two Media:

sin C= n9n i

, where\n2 <n1.

When the second medium is air, the formulabecomes:

'six; C = nj

Figure.4

-4 . REFLECTIONAT CRITICAL ANGLEa,

6

67A I

Ks ,t

t .3

C

`' .3

a,


iss

E Dispersion

Dispersiodis another important propertyof transparent materials. This is thevariation of refraetive index with color,(or wavelength)of light. When white'lightpasses` thro6gh a glass prism, the lightrays are-reffraCted'hy different amountsand separated into the colors of the .-,-spectrum.' This spreading of light intoitg component, colors is due to disperseswhich,. in turn' is due to the fact that the

.refractive index of transparent substances,. liquids and solids, is lower for long wave-

lengths than for short wavelength.

BeCause of dispersion, determination ofthe refractive index of a substance re-quires designation of the particula, ye-length'used. Light from a sodium lamphas a strong, closely spaced doublet withan average wavelength of 5893A, calledthe D line, which is commonly used as areference wavelength. Table 2 illustratesthe change, of refractive index with wave -'length for a few common substrCes.

Table 2. DISPERSION (iF REFRACTIVINDICES OF SEVERAL COMMON MAT

0

F Lenses.16

There are two cPasses of lenses, con-.verg ing and diverging, called also convexand concave., respectively. The focalpoint of a converging lens is defined asthe pqint at which a bundle of light rays

...parallel to the axis of the lens appearA to'converge aftter,passing through the lens.The focal lerigth of the lens is the distancefrom the lens to the focal point (Figure 7):,

f . '

. .

IALS : , .,

, Figure 7.

CONVERGENCE OFLIGI-IT AT FOCAL POINT0 Fefractive indexF line D line C line.blue (yellow) '(red)

, 4861A 5893A 6563A

Carbondisulfide 1.6521 1,6276 1.6182

Crown glass 1.5240 '1%5172 -.1.5145

Flint glass 1.:6391 1.6210 \..1.t221/

Water 1. 3372.1.'3330 1. 3312

mr - .

The dispersion Qt a material can be defined tquanittativel§ as:

n (yellow) - 1v dispersion z-b1 n (blue)-- n (red)

.

n (593mii) - 1'n (486mv) - n(656mv)

where, n is the refractive index, of Thematerial at the par+icular wavelengthnoted( in the parentheses.

G Image FoAnation by Refraction

Image formation by lenses (Figure 8).follows rules analogous to those alreadygiven above for mirrors:

1 Light traveling parallel to the axis ofthe lens Will he refracted so as to passthrough the foeus of the lens.

Light traveling through the geometricalcenter of the lens will be unrefracted.

he,pos ition of the, image c e determined'by remembering that a light r y passingthrough itcus,',F, will be parallel tothe axis of the lens.ori the opposite "sine' ofthe lens and that a 'ray passing through thegeometrical. center of the lenswill beunrefracted. w

°Optics and.the Microscope

Figure 8

IMAGE FORMATION BY A CONVEX LENS

The magnification,.object,prothiced byrelationship:

.M =image sizeobject size

M, of an image° of ana lens is given by the

image distance qobject digtance p

rom image to lensfrom object to lens.

where q = distance.;and p =. distance

H Aberrations of Lendm

Lenses have aberrations of several typeswhich, unless corrected, cause loss ofdetail in the image. Spherical aberrationappears in lenses with-spherical surfaces.Reduction bf spheriCal aberration caneaccomplished by.diaphragming the outerzones of the lens ,or by designing specialasplferical surfaces in the lens system.

Chromatic aberration is -a lifienomkwrioncaused by the vaiiation of refractiVe. indexwith wavelength (dispersion). Thus' a lensreceiving white light from an object willfarm a violet image closer to the lens' anda red one father away. AchroznA'CiC'lenses arb,employed to minimize this .

effect...The lenses ar4"cornliiratizins oftwo or more lens elements made tip.of,materials having different dispersivetiters: The use of monochromatic light-fs. obvious way of eliminatingchromatic aberration.. , , .

Aistigmatism-is a third aberration ofspherical lens. syStems. It occurs' when

.$

a.

object points are riot located on the opticalaxis of the lens and results in the formation-of an indistinct image. The simplestremedy for astigmatism is to place ?tie '

objey.t close to the dxis of the lens system.

I IntCrft.rence Phenomena

Interference and diffraction are two phe=-nomena which are due *to thq wave character-isties of light. The superposition of two .

light rays arriving simultaneously At a givenpoint will give rise td interference effects,whereby the intensity at that point will varyfrom. dark to bright depending on the, phasedifferent es between the two light rays..

The first requirement for interferencethat the light must come from a singlesource. The light may be split into anynumber of-paths but must originate fromthe same poipt (or .coherent source). Twolight waves from a coherent source arriv-ing at .a.'point in phase agreement willreinforce each other (Figure 9a). Twolight waves,from a coherent source arriv-ing at a point in opposite phase will canceleach other (Figure 9b).

is

Figure

9

.

9a. Twolight rays, I and .2, ofthe same:frequency'but dif-ferent amplitudes, are in phaiein the upper diagram. in thelower diagram, rays l 'and 2interfere constructively to givea' single wave of the same fre-quency aricP.with an, amplitudeequal to the summation of thetwo former waves.,

.44.t

.1

,

ak

'Optics and the Microscope

a

otb

Figure 9b. Rays 1 and 2 are no.v 180°-"out of phase and interferedestructively. The resultant,in the bottom diagram, is ofthe same frequency but is ofreduced amplitude (a isnegative and is subtractedfrom b).

The reflection of a monochromatic lightbeam by a thin film results in two beanis,one reflected from the top surface and onefrom the bottom surface. The distance.traveled by the latter beam in excess ofthe first is twice the ,thickness of the film

. and equivalent air path is:

2 nt

where n is.the refractive index andt is'the thickness-of the film.

The second beam, however, upon reflectionat the bottom surface, undergoes a halfwavelength shift and now the total retaril-.

tapon of the second beam with respect to°the-first is given as:

retardation = 2 nt +4

Where k is the wavelength of the lightbeam. e 4

When .retardation is_ exactly an *odd -Timberof half wavelengths, destructive interfer-ence takes place resulting in darkaess. 'When it is zero di an even number-of halfwavelengths,, constructive interferenceresults in brightness (Figure 10).

Figur,e 19

INTERFERENCE IN A THIN FILM

A simple interferometer can be made,bypartially silvering a microscope slide andcover slip. A preparation between the twopartially silvered surfaces will show inter-ference fringes, when viewed with mono-chromatic light, either transmitted or byvertical illuminator. The fringes will beclose together with a wedge-shaped prep-aration and will reflect-refractive indexdifferences due to Temperature variations,concentration differences, different solidphases, etc. The method has been used tomeasure quantitatively the concsrgration ofsolute around a growing crystalui(Figure 11).

50% Morrom/..N

--Cover s-p -

Specimen

0000

b,

Figure 11MICROSCOPICAL METHOD QF VIEWING

INTERFERENCE IMAGESa Examination is by transmitted light.

Light ray undergoes multiple .

reflections and produces dark andlight frifiges in the-field. A speci-men introduces a phase 'shift andChanges the fringe pattern.

b Illumination is froni the top. Theprinciple is the same but fringesshow greater contrast.

Each dark band represents an equivalentair thickness of an odd number of halfwavelengths. Conversely, each brightband is the result of an` even number' ofhalf wavelengths. '

Witlyinterference illumination, the effectof a transparent object of different re-fractive index than the medium in themicroscope fittia is: '4

1 a change of light intensity of the objectif the background is uniformly illumi-nated (parallel cover slip), or

2 a shift of the ipterference bands withinthe object if the background consistsof bands ( tilted cover slip).

The relationship of refractive indices ofthe surrounding medium and the object isas folloWs:

=

es

exnm( 360t

where ns = refractive index of thespecimen

= refractive index of thesurrounding_medium

= phase shift of thostwobeams, degrees

X = wavelength of the lightt = thickness of the specimen.

nITI

J Diffraction'In geometrical optics; it is assumed that,light travels in straight lines. This is notalways true. We note that a beam.passingthrough a slit toward a screen creates abright band wider than the slit with alter-nate bright and dark bands appearing oneither side of the central bright Band,decreasing in intensity as a function of :a

the distance from the center. Diffractiondescribes this phenomenon and, as one ofits-practical'consequences, limits thelens in its ability to reproduce an image.For example, the image of a pin point oflight produced by a-lens is not a pin pointbut ,is revealed to be a somewhat largerpatch of light surrounded by dark andbright rings. The diameter, d, of thisdiffraction disc ( to the first dark ring)is given as:

g4k,


2 44 f Xd

D

where f is the focal length of thelens,X the wavelength, and D the., diameteof the lens.

It is seen that in order to maintainsmall diffraction disc at a given wave-length, the diameter of the lens shouldbe as large as possible with respect tothe focal length. It should be noted,also, that a shorter wavelength producesa smaller disc.If two pin points of light are to be distin-guished in an image, their diffraction discsmust not overlap more than one 'half theirdiameters. The ability to distingvish suchimage points is called resolving power andis expressed, as one half of theprecedingexpression:

resolving power = 1.22 fD

II THE.°C.OMPOUND MICI:PSCOk'E

The compound m'principle ofhe

Ccoscope is an extension inimple magnifying glass;

hence it is e ntial-toutidefstand "fully theproperties of this simple lens system.

A Image Formation by the Simple Magnifier

The apparent size of an object is determined'by the angle that is, formed at the eye by theextreme rays of the object. By bringing the

4 object closer to the eye, that angle (calledthe visual angle) is increased. This_alsoincreases the apparent size. HoWever alimit of accommodation of the eye is reached,at which distance the eye can no longer focus.This limiting distance is about 10-inches or 25

. centimeters. It is at this distance that themagnification of an object observed, by theunaided eye is said:to be unity. Theeye can,of course, be focused at shorter distances butnot usually in a relaxed condition,

A positive, or converging, lens can be'usedto'permit placing an object closer than 10inches to the eye (Figure 12). By this meansthe visual angle of the object is increased(as is its apparent size) while the image of

...

71

5 -7°


t'---1r

4 IMO.. ObjeCI

4Ilast414,

1. 111060

EV,

Figure 12

VIRTUAL IMAGE FORMATION BYCONVEX LENS

the object appears to be 10 inches fromthe eye, where it is best accommodated.

B Magnification by a Single Lens System

The magnification, M, of a simple magni-fyinass is given Py:

25 ,M = + 1

where f = focal length of the lenscentimeters.

in

Theoretically the magnification can beincreage-d-with-shorterlocal-lengtirienses7However such lenses require placing theeye very close to the lens surface and

have much 'mage distortion and otheroptical aber ations. The practical limitfor a simple magnifying glass is about20X.

In order to go to magnifications higherthan 20X, the compound microscope isrequired. Two lens systems are usedto form an enlarged image of an object

..,.(Figure 13). This is accamplished intwo steps, the first by a lehs called theobjective and the second by a lens knownas the eyepiece (or ocplar).

C The Objective

The objective is the lens or lens system)closestito the object. Its function' is -to.reproduce an enlarged image of tht objectin the body tube of the Microscope..Objectives are available in various focal

5 -8

Eye

%

Imaget

ob,.... 111

4

Object t

Eyepiece

v 'tug Image

Figure 13

IMAGE FORMA ION INCOMPOUND MIC OSCOPE

lengths to give different agnifications(Table 3). The magn cation is calculatedfrom the focal 1 by dividing the latterinto the tube length, usually 160 mm.

The numerical aperture (N. A.) is a measuz4of the ability of an objective to resolve detail''This is more fully discussed in the nextsection. The working distance is in the freespace between the objective and the coverslip and varies slightly for objectives of thesame focal length depending upgn the degreeof correction and the manufacturer.

There are three basic classifitations ofobjectives: achromats, fluorites andapochromats, listed in the order of theircomplexity. The achromats are good fOrroutine work' while the fluorites and apo-chromats offer additional optical correctionsto compensate for spherical, chromatic and'other aberrations.

72

.

a

Optics and the Microscope,

Table 3. NOMINAL CHARACTERISTICS OF USUAI. MICROSCOPE OBJECTIVES

Nominal ' NominalWorking Depth

focal length N. A. distance focusmagnif.mm mm ii

Diam. of,fieldmm.

Resolvingpower, white

light, p.

Maximum Eyepieceuseful for niax,magnif., useful magnif.

56 2. 5X 0.08 .40 50 8.5' 4..4 80X 30X

32 4 1 4... 'O. 10 ,25* 16 5 3.9 el )X 20X

16 \ 10 0.25 7 8- 2. 1.4 250X 25X

8 20 0.50 1.3 2 1 0.7 500X 25X

4 \ 43 . 0.66 0.7 1 0.5 0.4 660X 15X

4 45, / 0.85 0,5 1 0.4 0.35 850X 20X

1.8 90 1.30 0.2 , 0.4 0.2 0.21 1250X 12X

Another tem of objectives employsrefytti surfaces in the shape of concaveand vex Reflectionx mirrors. Re optics,

...1.--eSuse they have no refracting elements,do not suffer from chromatic aberrationsas ordinary refraction objectives do. Based

. entirely on reflection, reflecting objectivesare extremely useful in the infrared and

'ultraviolet regions of the spectrums Theyalso have a much longer working distancethan the refracting objectives.

The body tube of the microscope supportsthe objective at the bogom (over the object)and the eyepiece at the top. The tubelength i's maintained at 160 mm except forLeitz instruments, which have a 170-mmtube length.

The objective support may be of two kinds,an objective clutch changer or a rotatingnosepiece:

1 The objective clutch changer ("quick-change" holder) permits the mbuntingof only one objective at a time on themicroscope. It has a tentering arrange-nient, so that each objective need becentered only once with respect to thestage rotation. _.The changing of objec-tives with this system is somewhatawkward compared with the rotatingnosepiece.,

2' The revolving nosepiece allows mountingthree or four objectiV,es on the microscope

at one time (there are some nosepiecesthat accept five and even six objectives).In this system, the objectives areusually nccncenterable and the stage iscenterable. Several manufacturers pro-vide centerable objective mounts so.thateach objective on the nosepiece need becentered only'once to the fixed rotatingstage. The insides of objectives arebetter protected from dust by the rotatingnosepiece. This,yas well as the incon-venience of the so-called "quick-change"objective holder, makes it worthwhileto have one's microscope fitted withrotating nosepiece.

.D The Ocular

The eyepiece, or ocular, is necessary inthe second step of the magnification process.The eyepiece functions as a simple magni-fier viewing the image formed by theobjective.

There are three classes of eyepieces incommon use: huyghenian,,compensatingand flat-field. The huyghenian (or huyghens)eyepiece is designed4to, be used withachromats while the compensating type isused with fluorite and apochromaticobjectives. Flat-field eyepieces, as thename implies, are employed in photo--micrography or projection and can be usedwith most objectives, It is best to followthe recoramendations ofthe manufactureras to the proper combination of objectiveand eyepiece. - .

5-9

.

.1


The usual magnification available inoculars run from abouttBiC up to 25 or30X. The 6X is generally too lo;,v to be ofany real value while the ?5 and 30X ocularshave sligh y poorer imagery than mediumpowers an have avery low eyepoint. Themost useful eyepieces lie in the 10 to 20Xmagnification range.

E Magnification of the Microscope

The total magnification of the objective-eyepiece combination is simply the productof the two individual magnifications. Aconvenient working rule to assist in theproper choice of eyepiecesl states that themaximum useful magnification (MUM) forthe microscope is 1,000 times the numeri-cal apei-ture (N. A.) of the objective.The MUM is related to resolving powerin that magnification in excess of MUMgives little or no additional resolvingpower and results in what is termed emptymagnification. Table 4 shows the resultsof such combinations and a comparisonwith the 1000X N.A. rule. The under:.lined figure Shows the magnification near-estto the MUM and the eyepiece requiredwith each objective to achieve the MUM.From this table it is al_parent that onlyhigher power eyepieces can give full useof the resolving power of the objectives.It is obvious that a 10X, or even a 15X,

eyepiece gives insufficient magnificationfor the eye to see detail actually resolvedby the objective. ,

F Focusing the Microscope

The coarse adjustment is*used to roughlypositron the body tube (in some newermicroscopes, the stage) to bring the imageinto focus. The fine adjustment is usedafter the coarse adjustment to bring theimage into perfect focus and to maintainthe focus as the slide is moved across thestage. Most microscope objectives areparfocal so that once they are focused anyother objective can be swung into positionwithout the necessity of refocusing except,with the fine adjustment.

The student of the microscope should firstlearn to focus in the following fashion, toprevent damage to a specimen ox objective:

1 Raise the body tqbe and place the speci-men on the stage.

2 Never focus the body tube down,(or thestage up) while observing the fieldthrough. the eyepiece.

3 Lower the body tube (or raise the stage)with the coarse adjustment while care-* fully observing the space between the

Table 4. MICROSCOPE MAGNIFICATION CALCULATEDFOR VARIOUS OBJECTIVE-EYEPIECE COMBINATIONS

ObjectiveFocal Magni-length fication

56mm

32

16

8

4

5X 10X

3X 1.5X 30X

5 25X 50X

10 50X 100X

20 . 200X

40 . 200X 400X

1.8 90 . 450X 900X

Eyepiece MU Ma(1000 NA)15X 28X 25X

46X .-60X 75X 80X

75X 100X 125X 100X .

150X -200/ 250X 250X

. 300X 400RI .500X 500X

OOX 800X 1000X 660X

135')X , 1800X 22504 1250X

'aMUM= maximum useful magnificati9p

74

objective, and slide and permitting thetwo to come close together wiothouttouching.

4 Looking through the microscope andturning the fine adjustment in such away as to move the objective away fromthe specimen, bring the image intosharp focus.

The fine adjustment is VR ua11y calibratedin one- or two-micron steps to indicate,th vertical movement of the body tube.ThN feature is useful in making depthmeasurements but should not be reliedupon for accuracy.

G The Substage Condenser

The substage holds the condenser andpolarizer. It can usually be focused in avertical direction so that the condenser canbe brought intethe.correct position withrespect to the specimen for proper

'illumination. In some models, the conden-ser is centerable so that it ay be setexactly in the axis of rotati of the stage,otherwise. it will have been centered atthe factory and should be permanent.

H The Microscope Stage

The stage of the microscope supports thespecimen between the condenser andobjective, and may offer a mechanical stageas an attachment to provide a means ofmoving the slide methodically during obser-vation. The_polarizipg mica °scope isfitted with a circular rotating stage towhich a mechanical stage may be added.The rotating stage, which is used fowbjectorientation to observe opticakeffects, willhave centering screws if the objectives arenot centerable, or vice versa. It is un-desirable to have both objectives and stagecenterable as this does not provide a fixedreference axis.

i The Polarizing Elements

A polarizer is fitted to,tlie`condenser of allpolarizing microscopes. In routine instru-ments, the polarizer is fixed with itsvibration direction oriented north-south(east-vr-st for most European instruments)


while in research microscopes, thepoWrizer can be rotated. - Modern instru-A,Inints have larizing filters (such asPblaroid) eplacing the older calciteprisms. Polarizing fgters are preferred

.,N beause they:

I are low-cost;

2 require no maintenance;

3 permit use of the full condenseraperture.

An analyzer, of the same construction asthe polarizer, is fitted in the body tube ofthe microscope on a slider So that it may

- be easily removed from the optical path.It is oriented with its plane of vibrationperpendicular to the corresponding directionof the polarizer.

J The Bertrand Lens

The Bertrand lens is usually found only onthe polarizing microscope although somemanufacturers are beginning to include iton phase microscopes. It is located in the-body tube above the analyzer on a slider(or pivot) to permit quick removal fromthe optical path. The Bertrand lens is usedto observe the back fooal plane of the objective.It is convenient for checking cgickly the typeand quality of illumination, for observinginterference figures of crystals, for adjust-ing t phase annuli in phase microscopyand fo adjusting the annular and centralstops in aspersion staining.

K The Compensator Slot

The compensator slot receives compensators(quarter:wave, first-order red and quartz-wedge) for observation of the optical prop-erties Of crystalline materials. It is usuallyplaced at the lower end of the body tube justabove the objective mount, and is oriented45° from the vibration directions of thepolarizer and analyzer.

L The Stereoscopic Microscope

The stereoscopic microscope, also calledthe binocular, wide-field, dissecting or

541

*.,T4


-Greenough binocular microscope, is inreality a combination of two separatecompound microscopes. The two micro-scopes,_ usually mounted in one body, havitheir optical axes inclined from the verticalby about 7° and from each other by twice'this angle. When an object is placed onthestage of a stereoscopic microscope, theoptical systems view it from slightlydifferent angles, presenting a stereoscopicpair of images to the eyes, which fuse thetwo into a single three-dimensional image.,,

1

,The objectives are supplied in pairs, eitheras separate units to be mounted on themicroscope or, as in,the"new instrtments,,built into a rotating drum. Bausch andLomb was the first manufacturer to have azooni Iens system which gives a continuousChange in magnification over the full range.Objectives for the stereomicroscope runfrom about 0.4X to 12X, well below themagnification range of objectives availablefor single-objective microscopes.

The eyepieces supplied with stereoscopicmicroscopes run from 10 to 25Xand have

fAt.wider fields than their counterparts in thesingle-objective microscopes.

Because of mechanical limitations, thestereomicroscope is, limited to about 200Xmagnification and usually does not permitmore than about 120X. It is most usefulat rela,tively low powers in observingshape and surface texture, relegating thestudy of greater detail to the monocularmicroscope. The .stereomicroscope isalso helpful in manipulating small samples,separating ingredients of mixtures, pre-paring specimens for detailed study athigher magnifications and performingvarious mechanical operations under micro-scopical observation, e. g. micromanipulation.

HI ILLUMINATION AND RESOLVING POWER41110

-Good resolving power and optimum specimencontrast are prerequisites for .good microscopy.Assuming the availability of suitable optics(ocular, Objectives and substage condensei)it is still of paramount importance to useproper illumination. The requirement for a

5-12.

good illumination system for the microscope. is to have uniform intensity of illumination,

over the entire field of view with independent .

control of intensity and of the angular apertureof the illuminating cone.

A Basic Types of. Illumination

There are three types 'of ilfuminatiop(Table 5) used generally:

1 Critical. This is7used when high levelsof illumination intensity are necessaryfor"oil immersion, darkfield, fluores-cence, low birefringence or photo-micrographic studies. Since the lampfilament is imadd in the plane of the'specimen, a ribbon filament or arclamp is required. The lamp must befocusable and have an iris diaphragm;the position of the filament must alsobe adjustable in all directions.

2 KOhler. Also useful for intense illumi-nation, Kohler.illumination may beobtained with any lamp not fitted with'aground glass.' The illuminator musthowever, be focusable, it.must have anadjustable field diaphragm (iris) and thelamp filament position must b\adjust-able in all directions.

3 "Poor man's". So-called because a low-priced illuminator may be used, thismethod gives illumination of high qualityalthough of lower intensity becauseqf thepresence of a ground glass in the system.No adjustments are necessary on theilluminator or lamp filament althoughan adjustable diaphragm on the illuminatoris helpful.

All three types of illumination require thatthe microscope substage condenser focusthe image of the illuminator aperture in theplane of the specimen. In each case, then,the lamp iris acts as a field diaphragm andshould be closed to just illuminate the fieldof view. The difference in these threetypes o'f illumination lie the adjustmentof the lamp condensing le . With poorman's illumination there is'no lamp conden-ser,.hence no adjustment. The lamp shouldbe placed close to the microscope so that

76

0

Table 5. COMPARISON OF CRITICAL,OHLER AND POOR MAN'S ILLUMINATION

Optics and the Microscope°

Critical Kohler Poor man's

Lamp filamnent

.Lamp condensing lerisLamp( iris °

Ground `glass at lamp

Image of light source

Image of field iris

IMage.of substage iris

ribbon filament

requiredrequirednone

t.

in object plane. °

nedr objectplane

back focal Planeof objective

any type any type °required nonerequired useful.none . presentat substage none

irisin object

plane

back focal' planeof objective

neap objectcplane

back focal planeof objective

the entire field of view is alwaysilluminated. If the surface structure of theground glass becomes apparent in he fieldof view the substage condenser veryslightly defocused:

Critical Illumination

Withcritical illumination the lamp conden-;ser is focused to give parhllel rays; focuring the lamp filament on a far wathis.sufficient. Aimed, then,. at the substageMirror, the substage condenser will focusthe lamp filament in the.object plane. 'Thesubstage condenser iris will now be foundimaged in the back focal plane of the ob-jective; it serves as a control over con-v'ergence.of, the illumination. Althdligh thesubstage4ris also affects the light intensityover the field of view it should most decid-edly not be used for this purpose. Theintensity of illumination may be varied bythe use of neutral density filters and, unlesscolor photomicrography is anticipated, bythe use of variable voltage on the lampfilament.

Kohler illumination (Figure 14) differsfrom critical ilhitmination in the use of thelamp condenser. \With'critical illuminationthe lamp condenser focuse§ the lampfilament at infinity; with Kohler illuminationthe lamp filament-ts focused in the plane of

77O

the substage condenser iris (also coincidentwith the anterior Meal plahe of the substage.condenser). The functions of the lampcondenser iris and the substage condenseriris in controlling, respectively, the area .

of the illuminated field of view and theangular apertureof the illuminating coneare precisely alike for all three types ofillumination.

Critical illumination is seldom used becauseit requires a special-lamp filament and be-cause, wtren used, it,phows no advantageover well-adjusted-Kohler illumination.8 IIKohler Illumination

To arrange the microscope and illuminato.°for Kohler illumination it is well to proceedthrough the following steps:

a ,Removelffediffusers and filtersfrom the lamp:

b Turn the lamp on and aim at-a con-venient wall'or vertical screen about

'19' inches away. Open the lampdiaphragiri.

c By moving the lamp condenser, focusa sharp image,of the filament. Itshould be of such a size as to fill,not necessarily evenly, the microscope

5r13

a

O

,Optics and the Microscope

Eye

Eyepoint

Ocular

Focal plane

Focal plane

Objective

Preparation

"Critical

Substagecondenser

Poor man's

/NJ

.1)

'I/

Substage' ,IriS

Lainp irisLamp

condenser

Light source,

5-14

'I

O

O.

1

I

Optic's and the Microscope

substage ,condenser opening. If itdoes note -move the lamp away ("vonthe wall to enlarge the filament image;refocus.

d Turn the lamp zuurain it attlie micro-scope mirror-so as,to maintain th.:same, 18 inches (oradjuided lampdistance).`

e Place a specimen oil' the iirospe.`stage and focus sharp1.wilh.0 I ti -mm(lox) objective. Open fully theaperture diaphragm in the substagecondenser. Irthe light is too bright,temporarily place a neutral densityfilter or a diffuser in the lamp.

f Close the lamp diiphragm, or field.diaphragm, to about a 1-cm operringiRack the Microscope substage con-denser up and down to focus thefield diaphragm sharply in the sameplane as the.speeimen..

g. Adjust the mirror to center the fielddiaphragm in The field Of view.

solro h Remove the 16-misr,,objectiveandreplace with.a4-mm objective, Movethe specimen sb that a clear area isunder observation, -Place the

au ;

Bertrand lens in the optical path, orremove the eyepiece and insert anauxiliary telescope (sold with phasecontrast accessories) in itefilace,orkremove the eyepiece and observe,theback aperture of the objectivedirectly. Remove any ground glassdiffusers from the limp. Nowobserve the lamp filament throughthe microscope. .

.i If the filament does not appear to becentered,'-swing the lamp housing ina horizontil,arc whoire center is at-Effie field-diaphragm. The putposeis.to maintain the field diaphragm on

"the lamp in_jti3.centered position, Ifa -.vertical movement of the filamentis required, loblieri the bulb base andslide it up or down. If the base is'.fixed,7-2tilt-theTlamP housing'vertical arc with`thefieI8diaphragm

k

. .as the center of movement (againendeavoring to keep the lamp dia-phragm in the cienivred position),If you have mastered this step, you

4 have Zieromplishd the must difficultportion. (}letter tilierosopd-laritshave adjustments to move the bulb,

'indepindently of the lamp housing tosimplify this step,.)

Put. -

the specinulri in place, replacethe eyepiece and the desired olSjec-live and refocus.

Open orlose. tIm.rfield diaphragmuntil it jtist disappears fi:om.the field.

-c

Observe -the back aperture of theobjective, preferably with the Bertrandlbns or the auxiliary telescope,, andclose the apeiture diaphragm on thesubstage condenser until it is aboutfour-fifths the diameter of the backaperture-. This fs the best positionfor the aperture diaphragm, a posi-tion which minimizes glare and maxi-,mizes:the resolving power. It isinstruttiVe to°.vary the aperturedia-phragm'and observe the image criti--cally during the manipulation:

m If the illumination is too great, : 'insert an appropriate neutradennityfilter between the illuminator and .

the condenser. Do not use the con.-denser aperture d.iaphragm or .thelamp field diaphrAgm to controLtheintensity of illumination.

Poor Man's Illumination

Both critical ana 'Kohler illumination re-quireexpens illuminatorg' with adjust-able focus, lamp 'iris and adjustable lamp

© mounts. Poor man's illumination requiresa cheap illuminator although an expensiveilluminator may be used if its expensivefeatures are negated by inserting a groundglash diffuser or by rising arosted bulb.Admittedly an iris diaphragm on the lampwould be a help though it is not necessary.;, -

a The illuminator must have a kOstedbulb

,or a ground glass diffuser.

'7'

79

5-15 ta

APF

Orlties.and the Miernope, r

0

.

should be possible to.direet it inthe general direction of the substagemirror, very close thereto or inplace thereof.

b Focus on ahy preparation aftertilting the mirror to illumin9te the'field.

c RemOve the top lens of the condenserand, by racking the condenser pp*,maze often, down, bring into focus(in the same plane as the specimen)a finger, iclneil or other object placedin the same general region as thegroundtlass diffuser on the lamp.The glass surface itself can ern befocused in the plane of the specimr.n.

.d Ideally the grotknd glass surface will ,

just fill `the field of view when centered'by the substage mirror; adjustmentmay be made by moving the lampcloser to or farther from the micro-scope (the position 'Might be markedfor each objective used) or by tuttingpaper diaphragms of fixed aperture(one for each objective used). In thisinstance a lamp iris would be usejul.

e .Low 9i condenser just sufficientlyto defocus the ground glass surfaceand render the field of illumhtationeven.

f,' Observe the back aperture of theobjective and open the substage con-denser iris about 75-percent of theway... The final adjustment of thesubstage iris is made 'while observingthe preparation; the iris should beopen as far as possible, still givinggood contrast.

g The intensity of illumination should-be adjusted °WI, with neutral densityfilters or by changing the lamp voltage.

Proper illumination is one of the most im-portant operations in microscopy. It iseasy to.41:dge microscopist's ability bya glance at his field of view and the objec-tive back lens.

0

4

(li ItcsoWing Power

The resolving pow r of the microscope isits ability to distinguish ,separate detailsof closoly tpaced microscopic structures..The theoretical limit of resolving two' 'dim:rely points, a. Ilistance X apart,',is:

X1. 22 k2 N. A. ;,

wIn.re = waveleniftlmtf light, used toilluminate °the specimen

N, A. = nktnericalaperture of theobjective ?

Substituting a wavelength of 4,500Angstroms an a numerical aperture of1. 3, about est that can be done withvisible light, we find that two points about2, 000A (or 0.2 fnicron) apart can be=seenas two separate points. Further increasein resolving power can be achieved far thelight microscope by using light or shorterwavelength. Ultraviolet light near 2; 000,Anqtroms lowers the- limit to about 0.1micron, the lower limit for the lightmicroscope.

The.numerical aperture of an objective isusually engraved on the objective and isrelated to the angular aperture, AA(Figure 15), by the formula:

AAN.A. r n sin2

where n = the lowest index in the spacpbetween the object and theobjective.

Figure 15

ANGULARAPERTURE OFMICROSCOPE OBJECTIVE

so

4

et

s 2"

o

d.

I

Optide and the Microscope

1 Maximum useful magnification.

A helpful rule of thumb is that the use-ful magdificatio'n will not exceed 1,000times the numerical aperture of theobjective (see Tables 3 and 4). Althoughsomewhat higher magnification may beused in specific cases, no additionaldetail will be resolved.

It is curious, considering,the figures. in the table, that most, if ilot all, manu-

facturers of microscopes furnish a 10Xeyepiece as the highest power. 4-10Xeyepiece is useftiltlit anyone interestedin critical work should use a 1-L2SX eye-piece; the 5-10X eyepieces are best forscanning purposes. . ra)

2 Abbe's theory of resolulion

One of the ,mostcdgent theories,bfresolution is due to Ernst Abbe, whosuggested that mtroscopic objects act`like diffraction gratings (figure 16) andthat the angle of diffraction, therefore,increases' withthe fineness of the detail.He proposed,tiltit a given microscopeobjective Would resolve a particulardetail if at least two or three' transmittedrays (one direct and two diffracted rays)

"°.entered the objective; In Figure )6 thedetail shown would be resolved in A andC but not in B. This theory, which canbe,borne ougy simple experiment, is'useful in shoWing how to improve resolu-tion. Since shoker wavelengths willgive a smaller diffraction angle, thereis more chance of resolving fine detailwith short wavelengths.. Also, sinceonly two.of the transmitted rays areneeded -Oblique lightmand a high N.A.condenser will aid in resolving fine detail.

4 0 33 Improv,ing resolving power

. .

The folToeing list summarize thepractical approaches to higher resolu-tion with the light microscope:

a The specimen should be illuminatedby either critical or Kohler

- illumination.

'4,

e '

10111 1111114,-

a b

Figure 16

ABBE THEORY OF RESOLUTIONe

b The conde.nser shotild be well-corrected and have a numericalaperture as high as the objective tobe used.

(c An apochromatic oil-immersionobjective should be used with a come-pensating eyepiece of at least 15Xmagnification. The immersion oilshould have an index close to 1.515and have proper dispersion for theobjective being used.

(cf--"Immersion oil should be placedbetween the' condenser and slide andbetween cover slip and objective._The preparation itself should besurrounded by a liquid having arefractive index of 1.515 or more.

e The illumination should be reasonablymonochromatic and as short in wave-length as possible. An interferencefilter transmittinga wavelength ofabout 480-500 millimicrons is asuitable answer to this problem.Ideally, of course, ultraviolet lightshould be used to decrease the wave-length still further.

The practical effect of many of thesefactors is critically discussed byLoveland(2) in a paper on the optics ofobject

81

5-17


IV PHOTOMICROGRAPHY

A Introduction

-Photophotograpthrtughgraph is a small photograph; a photomicro-graph is a photograph of a small object.Photomicrography is a valuable tool-illrecording the results of microscopicalstud . It 6nables the microscopist to:

1 describe a 'microscopic field objectively)bithout resorting to written descriptions,

2 record a particular field for futurereference,

3 make particle sl,ze counts and countinganalyses easily and without tying p amicroscope,

curvature of field and depth of field. Thephotographic plate, however, lies in oneplane; hence the greatest care must beused to focus sharply on the subject plane

(gra h , as distinct from micro- of interest and to select optics to givey," is tie art of ta iiii-TAC--tures---------minimtim_a.mounts of field curvature and

microscope. A microphoto- chromatic aberrations.

4 enhance or exaggerate the visual micro-scopic field to bring out or emphasizecertain details not readily apparentvisually, .

5 record images in ultraviolet and infra-red microscopy, which are otherwiseinvisible to the unaidecl-eye.

There are two general approaches to photo-micrography; one requires only a plate orfilm holder supported above the eyepieceof the microscope with a light -tight be,illows;the other utilizes any ordinary camera withits own lens system, supported with a light-tight adaor above the eyepiece. It isbest, in the latter case, to use a reflex'camera so that the image can be carefullyfoCused on the ground glass. photomi-crography of this type can be regardedsimply as replacing the eye with the cameralens system. The camera should be focusedat infinity, just as the eye is for visualobservation, and it shoal be positionedclose to and over the eyepiece.

.

. the requirements for photomicrography;hOivever, are more rigorous than thosefor visual work. The eye can norinallycompensate for varying light intensities;

5.48

With black and white film, color filtersmay be used to enhance the contrast of

\some portions of the specimen -while mini-mizing chrdmatic aberrations of the lenses.In color work, however, filters cannotusually be used for this purpose and betteroptics may be,required.

Photomicrographic cameras which fitdirectly onto the microscope are available .

in 35-min or up to 3-1/4 X 4-1/4 inch sizes.Others are made which accommodate largerfilm sizes and which have their own supportindependent of microscope: The former,however, are preferred for ease of handlingand lower cost. The latter system is-pre-ferred for greater flexibility and versatilityand lack of vibration. The Polaroid camerahas-many applications in Microscopy andcan be ilised on the microscope directly but,becauSe of its weight, only when the micro-scope his a vertically moving stage forfocusing rader than a focusing body tube.

B Determination of COrrect Exposure

;Correct exposure .gitterntinatisin can beaccomplished by tfial and error, by. relatingnew conditions to previously used successful

,conditions and by photometry.

With the trial and error method a series oftrial exposures is made, noting the type ofsubject, illumination, filters, objective,eyepiece, magnification, 'film and. shutter/speed. The best 'exposure is Selected. The.fillowing parameters can be changed'andThe exposure time-adjusted accordingly:

1 Magnification. Exposure time.vviesas the square of the magnification.

Example: Good exposure was obtained.; With a 1/19-second exposure.

and a Inagnification of 100)1.If the'magnification is now:

'OP

824 '

C°,

r-

i Optics and the Microscope '

° o ...1 2004, the' correct exposure,

__ ..._ I__ is calculated as follows.,,.,;,....

. -.ifew exposure time= 'old exposure time

new ma gnification0 (20012 -old magnification ' 100'

4/10 or say,' 1/2''sRcond.`°

1 should-hem- .hOWever, that theabove calculation carl:be made only whenthere has been no change IQ the illumi-nation system including the condenserorAthe objective. Only changes in magni-fication due to changing eyepieces or

bellows extension distance can hand-led in the abovg manner..

*4.- 'Numerical aperture:- Exposure time, \varies inversely as the square ofm:the-%smallest for king _hume B id ai!:"apof the condenser and objective.

..,

Example; Good exposure wad obtainedat 1/10 'geeorld wit)fi tide,

lt° objective, N.A. 0.25, '°4t-0o full aperture., With a 204,r

t objective, N,A. 0.25, at. . full aperture and the same ;

final tnagnificatioV what is-the correct expcialire time?

new exposure time = old e. pure time

;old N.A. 1 ..°1/19 11'40;or,new N. A.' O. 0/ , I

shy, .1450 second.,It it -Oeen that moreeliglit reaches the -photographiF filttkitytth highat humeri- P

CP cal apertures at tlic pagnemagnifiCation..1

3 Film. Etpbsure time varies inversely..1. with the Aifierican Standard's Association

'Co

Kodachrome II-Type AProfessional is 40.

4

new exposure time = old exposuie time

A. S. A. of old film 0.

X 11100(400/40)A. S. A. of new film _

° .`

10/100 or 1/10 second.

4 Other parameters may be varied but theprediction of exposure time cannot bemade readily. Experience and photo-electric devices at'e the best guides to'the proper exposur'e. ' .

;!'a Photoelectric devices are excellent for

determining correct,exposure. Sincebrdinary photographic.exposure meters

'Vre not sensitive enough for photomi-crography, more sensitive instruments,having a galvanometer'or electronic .

4amplifyined'ircuit, are required. Somephotosensitive cells are inserted in thebody tithe in place of the eyepiece forlight intensity roadings. This has the

advantage of delecting the light level at apoint of high intensity but &les not takeiqo account the eyepiece, the distance tothalm or the filar speed.

4The cell May be placed just above the eye-iece so that it. egisters 'the total amountP 5

alight leav,i0Vthe eyepliCk. 'Again, .theeffec,ts of film stgaetoatiinte44eidti*rojection .distake are no "'r!..., The-04ns ip.11dial,vbet.Fk with the total _light -.1:=,%- ,measurihk-photomefer'is ip

,e difficulty' of ,.

taking into account the arkaof field covered.: -!`,,' .

ake, for example, a.bright fieldin Mich , 1 ,,,t .°

.

4

speed index of the film..

Exainple : A'gdeid.picture was obtainedwith Edstman'Tri-XfilrrYat

second. What is the'Correct ext)sureforEastman Kodachrome ..Type A. .The A. S.A. speedfor Tri-X it 400 and for

.

t8

ly few crystalsappear,t perhaps 1 per-derhf of the light entering the field, of view isscatter0,03/ the crystals and the photomete..y : ,. '"shows closAto a ma9imum reading!' Now - .

asstime that evetrythihg r_e_2mains_constant- ,t-the hunib-Ei. of crYitals and, conse-

.quently-,,the amount of light.-scattered:' 'The'pt6torpe'ter rdadini couId easily drop Iby 501:lerdent., yet ttiii° proper,,xposure isunchanged.. The.,:;ituation forphotoTrdb.rograp y wi *th crb s,ed polarp since,

. ' the photometer reading depends. oh thei. intensity -of lildrninationvOn the bire-

.

fringence andstlicckness of the,,crystals and. ;". ./ ,.14;

t

't44,

°


on the number and size of the crystals infield or, alternatively, on the area of

the field covered by birefringent crystals.One of the best solutions to this problemis to measure the photometer reading withno preparation on the stage: A first-orderred compensator or,xqirartz wedge is in-serted when crosoRfOlars are being used

illuminate -the entire field.-

S

An alternative to place the cell on theground,glass ere the film will belocated. ever, although all variablesexcept m speed are now taken intoaccount, measurements in the image planehave the disadvantage of requiring a moresensiOve electronic photoelectric apparatus.

No matter what- m ?thod is used for placing'the photocell, the exposure time can ,,W$determined by the general formula:

exposure time meter reading

The constant k will'depend on the physicalarranerdnt and film used.; To determine.'k for any rticular system, fillst set upthe microsccr totake a picture.- Recordthe meter' reading anci take a series oftrial exposures. Pick out the best exposure

. and calcylate k.- Then the k which wasdetermaned holds as Iongas no change ismade lit* right pathpeyond the photocell,-e. g: chAiging to a faster film or changing

'thF668atiOridratance.,. Thus the, objective,'condenser position or illuminator may bechanged without affectinili if the cell is

;,used as deicribed above.

Example: With one particular arrange-ment of photocell and

. found too be 40. A.serfes of photographs

arelakensat 1/2, 1/5, 1/10,.IL2,5 and 1/N secon *. The.ph9toinicrograpil taken at 1/5second is judged to be. the best;

',hence k is calculAea as follows:

W4nieter.reading' X exposuretime = 40X 1/5,..= 8.

Assume,,novithat. a new picture.is to be takenat.sinot)iermagnification (hut witty-the

r -* t

-4

4t

er,

.4.;,t

5--20

e

same film and projectiondistance) and that the newmeter reading is-16; therefore:

expVsure time = k/meterreading = 8/16 = 1/2 second.

.V MICROMETRY

A Particle Size Determination

Linear distances and-areas can bemeasured with the microscope. Thispermits determination of particle sizeand qUantitative analysis of physicalmixtures. The usual unit of length for ,

microscopical measurements is to micron(1 X 10-3mm or about 4 X 10-5incffl.Measuring partici% in electron microscopyrequires an even smaller unit the milli-micron (1 X 10-3 micron or 10 Angstromunits). Table 6 shows 'the approximateaverage size of a few common airbornematerials:'

Table 6.. APPROXIMATE PARTICLE SIZE OFSEVERAL COMMON PART,IcULATtS

N

Ragweed pollen

Fc.:,g droplets

PnW Pr- plant flyash(after prIcipitators)Tobacco smoke

Foundry fumes ..

25 micr ons

20 microns

5 rriic rons

0.2 micron(200.millimicrons)

6

' 0.1 - 1 micron--(IUO =-1b00 rfaiiiinicrons)

The practical lower Unlit of acaurateparticle size measurement withthe 4ght,microscope is about 0.5,micron: Themeasurementeasurement of a particle,smaller thanthis with the light microscope leads toerrors which, under the best cirertm-stances, increase to about + 100 percent,

,(usually +). 7

Qne of th .e.princijIal-uses res.oltringpower is in the precise measurement of

-

*fr

Optics and 'the Microscope

partide size.. There are, 'however, avariety otapproximategand useful proce-dures -as well.

1 Methocid of particle size measurement

a Knowing he Magnification of themicroscope (product of the magni.-fication of objective and ey'epicce),the siie -of.particles-can be esti- -mated. For example, with a.10Xeyepieceind a 16-mm (or 10X10X)'. objective', the, total magnificationis 100X. A particle that appears tobe 10-mm at 10 inches from the eyehas an actual.size of ,l0 mm dividedby 100 or 0.10 mm or 100 microns.This is in no sense an accuratemethod, but it does permit quickestimation of partible size; the errorin this estimation is usually 10-25percent.

An9ther approxitrote method i's alsobased on the use of known data'. If.we know approximately the diameterof the microscopelield, we can

.6- estimate the percentage of'thediameter occupied by the object tobe measured and calctlate from -

these figures the approximate sizeAof the object. The size of the micro-scope field depends on both the objec-tive and the ocular although the latteris. a minor influence. The size ofthe field should be determined witha millimeter scale for each objective

.lend ocular. If this is done, eiti-tiori .0.fizes-by-comparigiiii with-entire field diameter carbe quite

( accurate (5-10%).

p.

b'

V

c The movement of a graduated mechan-ical stage can also be used for roughMeasurement of diameters of largt

larticles. Stages are usually gradaid (with vernier) tO'read to 0.1mittimeter, or 100 microns. In

..,practb-b,' the leading 'edge of theparticlelis brought to one of the linesof the cross hair in.thp eyepiece anda.reading iSe,t4100 bf the AtAkeposition.

Then the particle if mimed across thefieldby moving the mOf tartirCailttage'

\\'' ot_ `

''op

Po'

in an'appropriate direction until thesecond trailing edge just touches thecross-hair line. A second reading istaken- and the difference in the two`readings is the distah-Ce mov- r thesize of the particle. This method is.especially useful when the particleis larger than the field, or when theoptics give a distorted image near theedge of the field.

d The above method can be extended toprojection or photography. The image

.of the particles can be projected on ascreen with a suitable light source orthey may be photographed. The finalmagnification, M, on the projectionsurface (or film plane) is given approxi-mately' by

M = D X O. M. X E. /25

where 0. M. = objective magnific4onE. M. =eyepiece magnifica ion

D = projection distanC0from the' eyepiece'inCentimeters.

The image detail can theft be measured,' in centimeters and the actual size com;)

puted by dividing by M. This methodsis usually accurate to within 2-5 percentdepending on the size range of the detailmeasured.

e The stated magnifications and/or focallengths of the microscopeoiltics_a.renominal and bit from objectiveto objective or eyepiece to eyepiece.To obtain accurate measurements, .astage micrometer is used to calibrateeach Combination of eyepiece antobjective. T1* stage micrometer isa glass microscope slide that.has,accurately engraved in the center, ascale., usually 2 millimeter slong,divided into goo parts, each part repre-senting 0.01 millimeter. Thus 'when .

this scale is observed, projected.or,..,photbgraPhed, the exact image inagni-.

fication can be:determined. For'example, if 5 spaces of theg,stage micro-meter measure 6 millimeters when '

'projected, the'actual magnification is

O .

4

vo'

'4 .4

tics and the Microsc

65 (0. 01)

120 times.

This magnification figure canused to improve the accuracy ofmethod 4 above.

The simplest px.iXedure and the mostaccurate is based. on the use of a

_ micrometer eyepiece.: Since theeyepiece, magnifies a real imagefrom the objective, it is possibleto place a transparent scale in thesame plane as the image from theobjective and thus have a scalesuperimposed over the image. Thisis done by first piecing an eyepiecemicrometer scale disc in the eyepiece.The eyepiece micrometer has anarbitrary scale and mast be cali-brated with each objective used. Thesimplest way to do this.is to place .the stake micrometer -on the stageand note a convenient whole numberof eyepiec4 micrometer divisions.,The value inmicrons for each eye-piece micrometer division is theneasily computed.. When the stagemicrometer is removed and replacedby the specimen; the ,superimposedieyepiece scale can be used for accu-

, rate measurement of any feature inthe spec:igen by direct observation,photogrFWor projection. 4

2 Calibration of eyepiece micrometry .

Each znicgometer 'stage scale hasdivisions 100g (0. 1mm) apa t; qneor two of these are gusuall subdividedinto '10g (0. 01-mm) div ions. Theseform the standard against which thearbitiary divisions in the micrometereyepiece are to be calibrated. Eachobjective must be "calibrated-separatelyby noting the correspondence betweenthe stage scale and the eyepiece scale.Starting with the lowest power objectivefocus -on the stage/scale, arrange/ thetwo scales parallel- and in goodldQus.It should be pcishible to determine thenumber of eyepiece Azisions exactly,equal to some whole umber ofdivisibns of the stage scale, a distancereadily expressed in microns.

5-22

e

a

The calibration consists, then, ofcalculating. the number of microns pereyepiece Scale division. To make thecomparison as accurate as possible': alarge part of each scale must be used(see Figure 17). Let's assume thatwith the-low power 16-mm objective.6 large divisions of the stage scale(s.m.d.)-are equal to 38'divisions ofthe eyepiece scale. This means that38 eyepiece micrometer divisions (e.m..are equivalent to 600 microns. Hence:

1 e. m. d. = 600/38= 15. 8g.

Step litcrentieler Scale

14./4,14.A.16L4.1.1 .1.4.1411,

0Eyepiece blieremeise lids .

Figure 17

COMPARISON OF STAGE MICROMETERSCALE WITH EYEPIECE MICROMETER SCALE

t'Thus tvhen that micrometer eyepieceis ased.with that 1.6 -nim objective eachdivision of the eyepiece scale is eqUivalentto 15.8µ, and it can be used to 'make.anaccurate measurement of any object onthe microscope stage. A particle, forexample, 'observed with the 16-mm objec-tive and measuring 8.5 divisions do theeyepiece scale is 8.5 (15.8) or 135g indiameter,

Each objective on your microscope angstbe calibrated in tis manner. 0.

A- convenient way to record_the. necessary:data and tct calinlate g/emd is by. meansof a table.

Optics ancrtlie Microscope

Table

Objective nb. emd- no. emd 1 emd*

32-mm 18 = 44 , 1800 = 44 40.9F.,

16-mm 6 = 38 600 = 38 15.. 8Fi

4-mm 1 = 30 100 30 3.33µ. _

3 Determination of particle size.distribution

The measurement of particle size canvary in complexity pending on parti-cle shape'. Ttfe ze of a sphere ma edenoted bycube may be'expressed by engt

4n edge or diagonal. Be d these twoconfigurations, the p- cle "size" mustinclude information' out the shape ofthe particle in quest on, and theexpression'ofthis s ape .takes a morecomplicated form. t

Martin'S diameter is e simplest meansof measuring and expr ssing the dia-meters of irregular pa icles and issufficlVtly accurate wb n averaged fora largeltAnber of partic es. In thismethodi the horizontal east-westdimension of each icle which _dividesthe projected,area into halves-is taken asMartin's diameter (Figure 18):

4

1-74 I 1----4 1i 1 1 1

t1

1 II i

Figure 189

. MARTIN'S DIAMETER ,

87

0

The more pafticles counted, the moreaccurate will be the average particlesize. Platelike and needlelike particlesshould have a correctioir factor appliedto account for the third dimension sinceall such particles ale restricted in theirorientation on the rrTicroscope slide.Whet; particle site is report11, the'general shape of the particles as well asthe metboditied-to-delefrifine- the"diameter" should be noted.

Particle size distribution is determinedroutinely by moving a preparation ofparticles past an eyepiece micrometerscale in such a way that their Martin's

tdiameter-.can be tallied. All particleswhose centers fall within two fixeddivisions on the scale are tallied. Move-Ment of the preparation is usuallyaccomplished by means of a mechanicalstage but may be carried out by rotation ,of an off-center rotatingstage. A sampletabtabulation rn rTea .obnloe s8. sTohe ear -piece

.at least-six, but not more than tweive,,size classes are required and sufficieritpArticies are counted to give a smoothcurve. The actual number tallied 200

000) depends on particle shapere laOty and the' range of sizes. The

ze tallied fqr each Particle is thatnumber of eyepiece micrometer divisionsmost closely approximating Martin'sdiameter for that par rticle.

4 C alculation of.size1

averages

The size dat'a maybe treated in a varietyof ways; one simple, straightforwardtreatment .is shown in Table 9. For amore complete discussion of the treat-ment of particle size data see Chamotand Masdn;s, bybook of Chemical/Vlicroscopy(aage 26.

The averages with resperugiiier,7:11-, surface, d3; and weight or volume,d4, are calculated as follows for thedata in Table 9.

0

re

5 -23 ,

,;1;

,-

4


4

Tags S. PARTICLE SIZE TALLY FORA SAMPLE OF STARCH 9RA1NS

&so class. (sods) Numb** of particles ° Total

S

4

!

1 r1=4.1

-r-t-s4rt-1-4

re-44r-1-44

r-r44 rt.4.4

rt44 214..1rt.4.4

rt-i4 1-14.4rt-44 2444

-2-1-4.1

1-1-14 rt-4-1

rt-s4 rt.44

11-14

ft4.1#1'S-1.1

ri--1.4

11-441-t-4-1

rt-44

1$

9$r-144re-4.4

r-24.42-14.1

r144

11,1

1-144

ri-441-1-44

1"-11-4.1

rt-4-4

11-541

lit -4.l

rt-4.1.

110

1S7

.7-1

45

04

r 1-4.1

r1-4.311

ri-44rt441

rtZ1.1

7 t71-1..1 I-I-14 14-3-1 -1

.

11

*find eiralide microrneteL dizialeas

di = En'd/En = 1758/470 ,

= 3. 74 emd X 2. 82* =7. 10. 5p.

.713 = End3/ End2 = 37440 /7662

= 4. 89 emd X 2. 82 = 13. 81.1.

d4 = End4/End8 = 1991941 37440

= 5: 32 emd X 2. 82 = 15. OR

*2:82 micronSper emd(determined by calibration of the

.eyepiece - objective combingtionused for the determination).

Cumulative percents by number,surface and weight (or volume) may beplotted' frOm the data in Table 9. Thecalculated percentages, -e.g.

5-24

5

.

for the. cumulative;wei4ht or volumecurve, are plotted against d. Finally,the.specific surface, Sm, .in squaremeters per gram, m, maykbe calculatedif thedensity, D, is known; the surfaceaverage ;53, Is used.

-If D 1, Sm = 6 /d3D = 6/13. 8(1. 1)

= 0, 395m2ig.

83

Optics and the Microscopen.

/Table 9. CALCULATIONS FOR PARTICLE SIZE AVERAGE '

d(Aver. diam. n

in emd)nd nd2 nd2 nd4

1 16 16 ^ 16 16 16

98 196, 392 784 . 1588

3 -.1.10 330. 990 2970 . 8910

4 '107 428 1712 6848 27392

5 355 1775 8875 "'"44375

,1' 6 45 270 -..41620, "\ , 58320

7. 2f 147 1029 '''7203. 50421

8 2 16 128 1024 8192

-470 1758 7662 37440. 199194,".

B Counting Analysis,

\of .Mixtures of patteulates can often bequantitatively aniprzed by counting thetotal number of particulates from eachcomponent in a' representative sample. 'The calculations are,' however, comPli.-cated by three factors: average particle

,size, particle shape and the densityof the componints: If all of tht compon-ents were equivalent in particle size,shape and density then the weight.per-centage would'be identical tothe number_perceritige. llsally, however, it isnecessary to determine correction factorsto account for the differences.

When pxoperly applied, this method calbe-accurate to within + 1 pirced and,in special cases, even better. It is oftenapplied to e analysis of fibefmixturesand is then ually called a dpt-countbecause the y of fibers is kept aathepreparation is oved past a point or dotin the epiece.

A variety of methods n be used tosimplify recognition of the differentComponents.-----Theseinclude chemicalstaineor dyes and enhancementOrOptical- ---differences such as refracWe indices,dispersion or color. Often, however, one

.relled.cc.the differences, in morphology, ,

4

89

c4 .e..g.seounting the percent of raxon fibers

ip a sample of "silk".

Example 1:"A dot-count of,a mixture ,offiberglass and nylcin shows:

Therefore:

nylon 262fiberglass . 168

% nylon = 262/(262 + 168) X= 60.9i by number.

-However, although both fibers are smoothcylinders, they do have different densitiesand usually differegt diqieters. Tocorrect for diameter one must measure. -

the average diameter of each type of fiberand calctilatethe volume of a unit length

aver. diam. volume of4 1-p, slice, p.3

nylon 185' 268

fiberglass 13.2 117

"le percent by volume is, then:

% nylon " 2620( 26,8 X 100(262 X 268)+(168 X 1 .17)

= 7; 1%-by-volume. _ _

Still we must take into account the density ofeach in order to calculate the weight percent.

,

5-25


If the densities are 4.6 for nylon and 2.2efor glass then the percent by weight is:

262 X 268 X.]. 6.% nylon

:-. N L X 100,(462 268 X 4)+0 >c68 ,417 TEI3

= 72% by weight.,

ElamP11%.2: A count of quartz andgypsumshows:

_

gypsum."283

dypsum. 467I

To calculate the percent by weight we musttake into account the average particle size,the shape and the-density'of

The average particle size with respect toweight, d4,. must be measured for eachand the shape factor must be determined.Since gypsum is more platelike than quartzeach particle of gypsuni is thinner. Theshape factor can be approximated or can beroughly calculated by measuring the actual,thickness of a number of particles. Wemight find, for exaMple, that gypsum parti-cles average 80% of the volttme of the aver-

- age-quartz particle; this is our shape factor.The final equation for the weight percent is:

283 X itc14/6 X Dq% quartz = 238 X it-a4/6 X f)q.+ 467 X it cit/6 X 0. 80 XIDg

where Dq and Dg are the densities of quartzand gypsum respelcitrely;--0 : 80 is the shape,.

-.factor and d4 and d4, are the average parti-cle sizes with respect to weight for quaytzand gypsum respectively..

-

/ACMIOWLEZIGNIEN. T: This ogtlih'e was 2preparicl.by the U.S. Public Health Service.Departmenfjctf-- Health. Education and Welfare,'for use in its Training Program. 3

lEhmn. C. W. Crystal Grovrth from

X ipo

Loveland. R. P.. J. Roy. Micros. Sc.'O79, 59. (1980).

Chamot. Emile.Monnin and Mason.Clyde Walter, Handbook of Chemical .Microscopy, Vol. 1, third ed.John Wiley and Sons. New York (1959).

Solution. Discussions of the Faraday DESCRIPTORS: Microscope and Opticaleociety No.5. 132. Glittery and Jackson. propertieP

- g_

. 5-26.4,

1

.

5.

I INTRODUCTION

atr

STRUCTURE AND FUNCTION .0E- CELLS 5-11-.

What are cells? Cells may. be defined as thebasic structural units of life. The dell hasmany different parts which'carry on thevarious functions of cell-life.- These arecalled organelles ("little organs")..' .

A The branch of biology which deals With theform and structure of plants and animalsis called "MorPho loci." The study of thearrangement of their several parts iscalled "anatomy", and the study ,pf cells,is called "cytology','..

B "There is no "typical" cell, for cells differfrom ,each'other in detail, and thesedifferences are in part respdhAble for thevariety of life that exists on the earth..

.1

II FUND'AMENTAI;S OF CELL STRUCTURE

A tiow do, we recognize a structure as a cell?We must look for certain characteristi-ea.--and/or -structures which have been foundto occur in cells. The cell is composedof a variety of substanc.es and structures,some of which result from cellularactivities. These include both.living andnon-living materialS. ,

Non-living components incltide:

a. A "cell Wall" composed of cellulosemay be found as the outermostcovering a many plant cells.

b t "Vacuoles" are chambers in .the .

protoplatm which contain.fluidS ofdifferent 'densities (i. e., *differentfrom the surrounding protoplasm).

*7

2 The "living" partii of the cell areicsilled"protoplasm.", The following structuresare, included: .

a. A thin "cell Membrane" is locatedjust inside the ref wall. This

BI. CEL. 6. 76

s.

membrane may be thought of as theoutermost layer f protoplasm.

b In plant cells the most-conspicuous-protoplasmic structures are, the .

"chloroplasts", which contain-highly organized menibeane systemsbearing the photosynthetic pigments(chlorophylls, carotenoids, andxanthophylls) and enzymes.

c The. "nucleus" is a spherical bodywhicli regulates cell function bycontrolling enzyme synthesis.

d "Granules". structures of smallsize and. be "living" ornon-living' material.

e, "Flagella are whip-like structures'found in both plant and animal cells.Thy flagella are used for locomotion,or to circulate the surroundingmedium.

f "Cilia" resemble short flagella, foundalmost exclusively-on animal cells.In the lower animals, tilts are usedfor,Aocomotion and food gathering.

-g The "pseudopod", or false foot, is

an extension of the protoplasm ofcertain protozoa, in' which the -

colloidal state of the protoplasmalternates from a "soh' to a "jel"condition from titne-to time tofacilitate cell movement.

.

h "Ribosomesuare,protoplaarnic bodieswhich are the site of protein ;synthesis. They are too small(150 A in diameter)ito be seen witha light microscope.' .

i "Mitochondria" are small. mem-branoUs structures containing .

enzymes that oxidize food to producelienergy' transfer cdmppunds, (A TP )

'

yG7,

6-1

k

4

. -Structure and Function of Cells

B how basic strutture is expressed in somemajor types of organisms. '

We call better visualize the variety Of cellstrUCture'by considering several spedifipcells-.

1. li,4Cieria have fevrorganelle's;_ and areso minute that under the light -

, MicroscOPe only general morpholOgicaltypes (ire., the three basic shapes;rods, spheres-, and spirals) can berecognized: The following structureshave been defined: :

r-a The. "capsul e'',*s a thick protectivecove0ng,of the cell ekterior., con-sisting of pArsacci-bifi, cie orpolypeptide.

b The dell watt and plasma membrane"- are present..

.

c Although no well.defined nucleus isvisible in bacterial cells, theelectron microscope has revealedareas pfdeoxyribone nucleic acid(DNA).con8entration.' This sub-stance is piese,nt within the nucleus ofof higher cellk, and is'the geneticor hereditary material: ;

d Some types -Of pa-cttriaTkynt.iinspecial type of chlorophyll 1(bacterioc rophyll ) and carry onitotosyn esis,

2 ue-green algae are similar to thebactbria. in structure,. but contain the

0

pho ospithgtic pigment chlorophyll) a.

a t ke the bacteria, these forms alsotlackan organized nucleus (the

nuclear region is not bounded by amerhbrane). 7

41.

+he Chlorophyll,b,ea.ring membranesaide not localized in distinct bodies

. .

IChloroplasts), but are diaperded-throughout the. cell. .

.... '. .

Gas -filled structures called"pse'Udovacuoles" are found iii sometype g.of blue-greens:,

/ '

o

1

I

3 Thp green algae as a group include a.great variety of .Structural types, .ranging from singleq-celled non-motileforms to large motile colonies. Some,types are large enough to resemblehigher aquatic plants.

a

I... att'

a. The' chloroplasts.aremoditied irkbosA variety of: shapes and ire loomedin different positions, Examples ,of dh3.oropla.st shape and position.aret

. sokt,

; .3)

1) Parietal - located c5xrtheperiphery of the cell;' usuallypup- shaped and may extendcompletely around the innersurface of the plasnia membrane.

2) Discoid - also located oq theperiphery of the cell; but are.plate-shaped; usually many percell:

Axial - lying in the central axis-of the cell; may be ribbon-likepr star-shaped.

. .*-

4) Radial - have-os or processesthat extend' outwa-i'.rpd from the

scenter of the cell (radiate),er" reaching the plasma melnlYrane..

.5) Reticulate - a mesh -like network _

that extends throughout Volumeofthe cell.

b Located in the chloroplasts may bedense, proteinaceous, starch.forming bodies call "pyrenoids".

4 The flagellated algae .pbasess dne-to-, eight flagella per cell., The chloro-

plants may contain brown and/or redpigments in addition to chlorophyll.

4,1

a ReserVe food may be stored asstarch (Chlamydomonas)paramylon(EUglena), Or as oil. 4

5 The protozoa are single-celledarlimals whidh exhibit a variety ofcell structure.

.92It

/.

a

,;

,

44

*ift

a The amoebae move.by means ofpseudopodia, as described-,

- previously.r-

b The flagellated prbtozoa(Mastigophora) possess one or more

c The ciliates are the Most highly-friodifiedprotozdans. The cilia may,be more or less evenly distributed,bver:the entire surface of the cell,

.or mapbeelocalized. ,

Structure and Function of Cells

III FUNCTIONS OF CELLS

.: 4 .a higher concentration of a substance"(Such as, phosphate) inside the cell thanis found outside. Algae are able tosynthesis 'organic matter from inorganicraw.materials (carbon dioxide and ,

water), with the aid of energy derivedfrom light; whereW3 animal cells mustobtain their organic matterready- .made" by consuming other organisms, ".organic debris,. or dissolved organics.

.. _

What are the functions of Cells and theirstructural components? Cellular function,is called "life", and life is difficult to define.Ltfe..is charadterized1:89moccsse ommonlyreferred to as rejarbduction, growth, photo-syntflegis, etc. °

IV SUMMARY

The cell is made up of many highly special-,izedgilbstructures. The types of sub-structures present, and then. appearance(shape, color, etc, ) are very imnortant, inunderstandinethe role of the organisth inthe aquatic comMunity4 and in classification.

%.-

.\REFERENCES - °

.'A" Microorganisms living in surface waters -1. Bold, H. C. Cytology of algae. In: G.M:.

are subjected to corratant fluctuations in 4 4 - Smith, (ed. ), Manual of Phyvlogy.the physical and chemical characteristic , Rona ld Press. 1951. . r .

. . 41

of thet h e e n v i r ondAt, and mus t constantlyIm O d ifr t h e i r activities. " 2 B. otn.e) Geoff fry H ed° : , Cytology aid

.The. cell requtres'oe. source of chemical,energy to, carry On life proceies andsuccessfullZoompete with other-,

° oFgarnsms. Plant cells may, obtain1" this energy from light, ,which is,.

absorbed by chlorOphyll and convertedinto ATP or food reserves, 'dr from , -the pxidation of food s tiffs. Animalcells obtain energy on y from the

o°oxidation oflood.,-

0 0 0eF.' , ,. ,

ells. must obtain *ravio5ixaci eriars, fromthe *envirourpent In:ordeF to grovrandcarryout otheilifeyfilnctiOris: Inorgaiitc,and'orgailic..inatelials may, be to:keit-ups

C

Cll Physiology. .3rd ed.,' AAcademic', .

"Press. 1964:- o; . .

4 ,.. ... .. °'

3 :lara_chet, Jean. The Living Cell:' Scientific American. 205(3): n61.

.,..4r

.4 Corliss, John O. Ciliated Protozoa. ,

' Pegamon. 1961. -...

i'rittch; Fre- The: structure and .treproduction.of the algae. CambridgeUnik( Press. , 1965, . ,P

6

t`6 Frobisher, .M. I'luilamentals ofitliepdbiology .7th edition. W.B.

. Saunders Philaalphia; ;1962.0...by passive diffusion' or by . 44, . .* ;-:.

.'7 Round, F'. E. The biology of-the algae.

p St. Martin's .Prese. 'New York. 1999*.. .

transport". Irl,theaFter process,energy Is used to,laulc1 up and maintain.

a .. `.14'4; .4 4 ; ' Ili.. re ,, o o .1 ,. ca., '-; ... Tlis°outline,originally.prepared by ttilhaer4.. :

.° s .4- 9' E. Bencier;Biologist, /orfnierlyth

0 ,,;:. , c 9 . 4.1

.0 . Training ACtivities; FWPCA, SEC. andrevised by Cornelius I. Weber, March 19_700.- $ '' ..

A

;- ' '44

4.04o Descriptor: °Cytological Studies

9

I INTRODUCTION -

.,

BACTERIA AND PRO'TOZOAAS TOXICOLOGICAL INDICATORS

The. WINOGRAD,SKy COLENIN is an excellentsimple classroom experiment as wklfas aminiature_ecosystem which yields a varietyof-photosynthetic and other prbtista -especiallythe bacterial forms important in photosynthesis°research. These photosynthetic bacteria, aspointed out by Dr. Hui r, are ubiquitous inwet soils and natural waters but ordinarilyescape notice.

II PREPARING THE COLUMN

The materials needed are simple laboratoryitems. \

A The'ino6ulurit la back sludge)*may beeasily obtained From a lOcal'sewagetreatment plant or the bottom of a pondoil lake. Because the USPHS document,contairiing Dr. Hutner's paper, is out ofprint\it reproduced, ere.

0B Directqls for preparing the cofiren,and

other'useful inforination are given in that,paper.

01/4.12r. Buttesufficient f

. information:

III ECOSYSTEM DEVELOPMENTi

tit -

1s*bittliography Should ber these who wish" more

.

A yactoi' such as the substrate used, theinoculum, the overlying supernatant water,end labOrafory, condiUsi- as temperature

light: all;ittfltred440 p&rticul Ir.type of biota forfaing successive layersor zones.. The.4.cdtsmpanying figure isthier efore 'generalized, and is not intended '"°

.

, to be absolute. ,

B Some. representative forms are listed for.'general inforination, The numberscorrespondtO those,on the figur.

. -Inorganic substrate on towe/ing.-f.,

o

a.9.72

.1

2 Green photOsynthetic bacteriaMicvochloris, Chlorobium, andChlorochromatum; methane bacteria;and SO4 reducers.

13 Photosynthetic pUrple sulfur bacterfa -Thiopedia and Thiosarcina.

4 Filamentous sulfur bacteria,Beggiatoa.

5 Non-sulfur photosynthetic bacteria,Rhodopseudomonas.

- ,

6 Blue-green algae, Schizothrix andOscillatoria, .

.

Diatoms, Nitzschia.

8 Coccoid green algae, Ankitrodesmus;and flagellate greens, Chlamy'domonas;

, similar to a stabilization'pond flora,

9 Filamentous green algae, Ulothrix.

C Besides the phbtosynthetic bacteria and'ather.piotista there will be a variety of °peotozoa found in the aobic levels(app. 6 through,9). Many of,theseprotozoans are typical fauna of activatedsludge.

°

D The possibilities are endless for furtherexperimentation. These ecosystems ax'salso conyeniehf and inexpensiye seurcesfor protozoa and,ottterltiodsta for class,arid laboratory instruction. ° .

, 4 o.,MICRIDAQUARIA

4 ,

II

A Fenchel describes a microlquariunc(1.5 X 54u) which may be observed undera .compounernicroscope. (Figure 2y-

Be, The development of cpmrnunities of.- 'orkanisMs is, quits? similar-to the

Winogradsky Column..,(Figure 3)o"

0 1. , 7:1

, Bacteria -aildIProtozoa as Toxicological Indifators

C The basic media consists of one liter ofnatural water 10 g CaSO4, k g glucose,1 g of peptone utoclaved and stored atSo C;

\ Befo*re use agar is boiled with the media.White hot7 the media is introduced intoone end of the "micr6 aquarium" with a

_pip et . _After. tiv agar _congeals, naturalwater samples are added. Duringincubation and When not being observed,the-micro aquaria are kept_in a moistchamber..

E Although Fenchel used a 'seawater mediumand inoculum, freshwater sources wouldbe equally useful.

F In these microaquaria simple ecosystemsdevelop when they are kept in complete.darkness. Complex photosyntheticcommunities develop when they areilluminated, A natural ecosystem isfigured by FenChel (Figure 4), ald arelated food web is shown in Figure-5(both from Fenchel).

. .

G Microaquaria Using PAitic Petri Dishe'se.

Sessile ciliates have been successfullycollected, cultured, and used for bioassay \'using the' same petri dish (membrane filter 'type, with tight fitting lids). 3-, t

,. H. IVIicro*Aluartia,usindisilltorfe.cement ringssillies,. ,,

, 0,,*hilh Allow;diffiision of 'gases throtth the-' pi/icohe: cultures win $12.erfbyflemp.in .....

active ipdelinirgy. 4 . % . . s.

. 4 . . %tti '., ,

, g 0 ....

SO '. ADDr l'IONAL REFERENCES ... I. - ,-

r Hutner, S.H. i?rotoxoa as.Toxicokgical(roofs: The Jour. of Frotozoology.11(1):1:6.' l9B4. :' ',, ft', ;

, ers

4

3 Burbanck, W. D. and Spoon, D. M.The Use of Sessile Ciliates Collectedin Plastic Petri Dishes for RapidAssessment of Water Pollution.Jour, of Protozoology. 14(4):739-744.1967.

4 Curds, C.R. and Cockburn, A.Protozoa in Biological Sewage Treat-

, Pnent Processes. I. A Survey of theProtozoan Fauna of BritiSh Percolating

. Filtermand Activated Sludge Plants.Water Research 4:224-236. 1970.

5 Curds, C.R. and Cockburn, A.II. Protozoa as Indicators in theActivated Sludge Process. WaterResearch 4:237-249. 1970.

6 Curds, C. R. An Illustrated Key to theBritish Freshwater Ciliated ProtozoaCommonly Found in Activated Sludge.Water Poll. Research Laboratory.Stevenage, England. 90 mi..

.7 Fenchelk Tom. The Ecology of Marine-- 4 "I

Microbenthom' IV. Structure and..1i4=1._..onof the nenthic EeosAtem,its, Chemical and Phisical Factorsand the`Micrpfauna Communities withSpecial Reference to the CiliatedProtozoa, 01:Melia 6:1-182. 1969.*

. Botanical Gardens and. # Hol-izong in Ald3.1.Rese,.arcIA. In Challenge,

for Survival. Pierre )5anseteau*ed.Columbia.University Press. -1970. , .

. ..., .G .

9-. Hutner, S.11. ; Baker, H. ;,031nColi, D..

4,t

2 Spoor; D. M. and.lierpanc It, , W. DrA New Method X9r Collecting SessileCiliates in Plastic Petri Dishes withTight - Fitting bids. Jour, ofProtozoology J4(10:735:730. 1967.'

- ,Water Pi'ograms Operations, EPA, Cincinnati,,.

,'7 -2

.Nutrition and Metabolism ro ozoa.Chapter in ictlogy *pp: 85-177.

Adited rprganton.35*ress.197.2. ' 15'

i :A

*". 4?* "*" .-*At :10 Hu S. H. The Urban.tiot°431ical:Giffae*A°

A cadeknis Wildlife,.B..vesekirct. ,bardenn. 19L2V7,-40: 1969.wpw .** '4

4;

. Thisilyntlinevran prepared by Ralph M. Sindlair,4quati4 Biologist, National Training Center,

01# 45268.-a

6

.95

41.

*1.

O

J

.

is

1

o

,

Bacteria and Protgzoa Toxicolngical Indicators .

o

du 1

.

.WINOGRADSKY :COUMN

S.

GENgRALI2ED.

green

green

I -iipt1.01041%, 74.

r m'clge.no

a 4,

rest'

-gyttja

a

.,

o. 4.

... .. tl ' 0 a

' ',40%.

A

' ' ^7. %

FILAMENTOUS GREEN 'ALGAE, ,..

4 . .411. vtt ,--.. t

c , S.

IS

SUPERNATE , .7 sis - °

41' a° 4 1 ' .` ' s. °

.' CPCCOID GREEN' akbdA,E '.41/4 )V ! ... '''' .' /6- : ,tt / "

6 . 14*. ..; ..i e' - , +. A' A . . . ' ;

.: C % i 4 O

k 0 ' .. 6

.4)4.1.016AS;

..I.J_EpREEN ALGAE =P.

N01.LLLFUt PHOTO,----FIL-KM EN:TO-USSU L-FUR

0PURPLE_SULFUR.B*CTERIA

04

PREEN PHOTO. ,..

A

J

,FIGEIRE 1 .

Did

Aft'411W

4

$INORGAtildSUBP'RATE,

45

4

4

o,V

Bacteria and Protozoa aS Toxicological Indicators'

c

t f

FIGURE 2

A micro aquarium fitted with electrodes and the redox conditions through 17 days.

ic

4

_i , ,

/T c I ,---, . , j:,,..4.-- .., .,,,, ,, `2

.. ?":-.-,:i,i 1.ezt',_'r/-A-,-4,1- r- r

"

,1.-4; ),) / ". is-'----,1

C.' . i _...)

-......., (< r---1.,V\--,to 221 c"' :'t t %

/ t1,,,,..,-,,r,, - "---

-----,__. ,, .2, ,-- 4,j,/.-.....,-::-. :.--/'4.'d , -,i

\>1. -";- \

4

ul

FIGURE 3.

brawing made by tracing a micrograpirof the bacterial plate in a micro aquarium(same experiment as shown pn Figure t.) Most conspicuous are the filaments ofBeggiatoa and thg ciliates byclidium citrpllus, Euplotes elegans and HolostichaBp: Below the Oscillatoria filament (lower left) a Plagiogogon loricatus is seen.Eafteria (exctpt Beggiatoa) 'arse not shoWn,

, .

Bacteria and Protozoa as Tokicolosica.1 Indicators

..

v.

-t

300 it

r

FIGURE 4

The microflora and fauna in the surface of the Beggiatoa patches. (Oscillatoria,Beggiatoa, Thiovolum, diatoms, euglenoids, nematode, Tracheloraphis sp.,Frontonia marina, Diophrys scutum, Trochiloides recta).

4.1ACaff LA S.

1;ONO' (9STom S.

F...tv-7nusetczettu LA.& Kt.tAlt.

/11c34.4/24./0_2Psf_

FKKONIA ,I.ONO°6 .

etaramAti2

M'AA MU A S.

F ht),0*.4AV

ealre.S&...61121P5AMtatayjiczpauyi'

isSrf PO, sPe.

RN Li. T VADLOSTSn ClEvA

=4=S LifMf.n220.11±.M

The food relationships ofthe most common ciliates in "estuarine" sediMents and in sulphureta.

93fi

5

,

,

ENVIRONMENTAL REQUIREMENTS OF FRESH- WATER INVERTEBRATES

L. A. Chambers, *Chairman

Bacteria:- Protozoa as Toxicological Indicators in Purifying Water

S. H. Hutner,, 'Herman Bakers S. Aaronson and A. C. Zahalsky+

There is a cynical a ge that all travelersbecome entomologi ts. But now with DDTand detergents, t 4Velars and stay-at-homes

_ alike are beco 4 g toxicologists. We havean immediate in Brest in pollution problems:Our laboratory receiiies, like the Eait Riyerand adjoining United Nations buildings, agenerous sootfall from a nearby power plant.Also, we have'seen a superb fishing ground,Jamaica Bay in New York City, become asewer. (Jainaica Bay is, however, beingrestored to its pristine cleanlines§but notthe U. N. 'read. )" We take our'themeneverthel ss not from aesthetics but fromstateme s by Berger (1961): {1) It is anexpensive, timq- consuming project". .. tpredict with confidence -a'new waste'sprobable iinpact on certain important' down-

! strewn water uses." And (2) "Thetoxicological.phase,octie study is p1rhapsits Mod perplexing aspect' The specializedServices and cost necessary for determiningthe effect of repeated upoSure -to low con-centrations of the waste forlong periods of-dine would inevitably plaCe this job out ofreach of most public agencies. Equallydiscouraging, ,perhaps, is the probabilitythat the, toxicological study may take as long'as two years."

As describtd here, advances in niicrobiology,offer Hopes, of lightening this burden. Thefirst question is: What kind of microcosmwilAserve'for toxicological surveys, especiallyforipredictinithe pOisOning of biological meansof waste disposal? The second is: Cap theprotozoa of this microcosm predict toxicityto highei aninials?,,

,*Iiirect6r,' Allan Hancock Foundation and Head, Bio. Dept.-, Univ. South. Calif.+Haskins Laboratories, 305 E. 43rd St., New York 17, K. y.,; a/id Seton Hall Collegeof 1Viedidine and Dentistry, Jersey City, N.J. Pharmacological work from HaskinsLabdrafoPies discussed here vitas assisted by grant R6-9103, Div. of General Medical.Sciences, of the Nationtillnstitutes of Health. Paperresented by Hutner.

The food -chain pyramids of sewage plantsand polluted waters haverbeen adequatelydescribed (Hynes, 196e; Hawkes, 1960).A problem treated here is hoW to scale thosemicrocosms to experimentally manipulablemicrocosms yielding dependable predictionsfor the behavior of sewage-plant microcosms.

THE WINOGRADSKY COLUMN AS 'SOURCEOF INOCULA FOR MINERALIgATIONS ANDAS TOXICOLOGICAL INDICATOR SYSTEM

Total toxicity °depends on intrinsic toxicity- k

persistence relationship,. Techniques fortesting the degradability, of organic compounds-and so their persistenoe in soil arilwater--are' still haphazard. The enrichment culturetechnique, in which one seeks microbes thatuse the compound in.question as sole substrate- -hence degrades it and even "mineralizes"it - -was developed by the Dutch school ofmicrobiologists. Enrichment cultures are/used routinely by biochemits wishing to.

work out the microbial catabolic metabolicpathway for a compound of biochemicalinterest. Since the compounds dealt with bybiochemists are of biological origin, micro-bial degradability 'can be assumed. Still,finding a microbe to degrade a rare biochemicalis not always easy: Dubos, in a classicalhunt for a microbe able to live off the capsulesof pneumococci, found the bacterium only,after a long search which ended in a cranberrybog. Stkh.difficulty in finding mierobes thatdegrade raz biochernrcale implies an evengreater difficulty in finding microbes thatdegrade many products of the synthetic

7-6O

'.99

I

't1,

,Environmental Requirements of Fresh-Water Invertebrates

organic chemicals indu"Stry, since suchcompounds may embody4oiochemically rareor biochemically nonexistent linkages,.Intimations of the frnpTaante of inoculumabound in the literature, e.g., Ross andSheppard (19561 could not at first obtain .

phenol oxidizerslfrom ordinary inocula(presumably soil and water); but manure

A-and a trickling filter from a diemicar_plant______proved abundant sources of active-bacteria.- \Onelvonders how extensive a study underlies \the statement mloted by Actixander (1961)tilat

Z "soils treated witI 2,4, 5-T (trichlorophenoxy-acetic acid) still retain the pesticide Mengafter all vestiges of toxicity due to equivalentquantities of 2, 4-D have disappeared.','

What then is a reasonable inoculum for testinga compound's susceptibility to microbial

...attack? The size range is wide: from thetraditional crumb or gram of soil or mud tothe scow-load ot activated sludge.contributedby New York City to inaugurate the Yonkers,

..)_ sewage-disppsal plant. We suggest that to' strike a practical mean in getting a profile

of Soil, mud, or sludge to bye used as inoculumthe uses of the Winogradsky column should beexplored. Directions for Winogradsky columnand bacteriological enrichments have beendetailed (Hutner, 1962) and so only an outline'is given hei.e. The column is prepared byputting a paste of shredded:paper, CaCO3,and CaS0 at ine bottom of a hydrometer jar,Ming the4jar with mud smelling H2S, coveving

with a..shallow layer of water, and illuminatingfrom the side With an'incandescent lamp. In2 or 3 weeks sharp zones, appear: a green-and-black anaerobic zone at,the bottom(a mixture of green photosynth 'tics bacteriaalong with SO4-reducers, meth roducers,and the like); over tha0. red zonesk '....sf

(predominantly photosinthetic bacteria);above this garnet or magenta spots or zone(pre dominantly non- sulfur ,photo syntheticbacteria); above this a layer rich in blue-green algae (the transition to the aerobiczones); above this aerobic bacteria along withgreen algae, diatoms, other algae, and an-assortment of protozoa. This makes anexcellent siTple classroom experiment todefhonstrate the different kinds of photo-synthetic organisnis, especially the bacterialforms that are important in photosynthesisresearch and that ordinarily escape noticeyet are ubiquitous in wet soils and naturalwaters. .

As-pointed out by our colleague, Dr. L. ,

Provasoli41961), the heterotrophic, ,capacities of algae are very imperfectlyknown. This is underscored by recentstudies of the green flagellate Chlamydomonasmundane. as a dominant in sewage lagoons inthe Imperial Valley of California (Eppley andMaciasR,' 1962); other than that it prefersacetate among the few substrates tried; itsheterotrophic capacities are unknown.More unexpectedly, some- strains of thephotosynthetic bacterium Rhodopseudomonapalustris use benzoic acid arferobically asthe reduclant in photosynthesce--(SchF,_Scher,and Hiitner, 1962) narrowing the gap betweenthe photosynthetic pseudomonads and theubiquitous pseu,domonada-so often represented ,

among bacteria attacking resistant substrates(e.g.', hydrocarbons) as well as highly_vulnerable subtra s. For the widelystudied, strongly h erotrophic photosyntheticflagellates Euglena. gracilia'aind E. viridis,./common in sewage, no speceic enrichmentprocedure is known, .meaning that theirecological niches are unknon but labotatoiydata provide hints. , ,i

The increasing use of oxidation ponds wouldin any case urge a greater use of Sealed-down ecological systems in which developmentof none of the photosynthesizers present inthe original inoculum was suppressed;Conceivably, some of the rare microbesattacklikg rare substrates- -and such micrObesare likely to represent a sourcei of degiadersof resistant non-biochemicals--are specialistsin attacking products of photosyntheticorganisms.

Traditionally, the inoculum for a Winogradskycolumn is a ma.rincorgbrackish mud (asNew Yorkers we Would be partial to mud fromflats of the Harlem River). Little is knownabout the effectiveness as inocula of freshwatermuds or water-logged soils, It would bevaluable to know how complete a cohimn

'',coUld develop from material from a tricklingfilter or an activated-sludge plant.' A practicaliattue is: Might the poisoning of 'a sewageaoxidation system be paralleled, by the poisoningof a Winogradsky column}-whgre'the poisonWas Mixed with the inoculum for the column?Might tile, variously 5t111 photosyntlietic-zones of the column and the.'aerobic populationon top procride sensitive indicators for the

-

4

0 Environmental Requirements of Fresh -Water InvertebratesI

,performance of a sewage-oxidation systemsubjected to chern,ical wastes?

If a,particular compoundmixed with inoculationmud Fr sludge suppressed development of thelull Winogradsky pattern, one might assumethat the compound at the test concentrationwas poisonous and persisteirt. Biological Nv/,destruction of such poisons, if at all possible,might demand a long hunt for Suitable micro-organisms, tHexi buildup of the culture to apActical scale. This might best be done withilluminateti shake or aerated' cultures, withthe inoculg coming from a variety of environ-ments.. Optimism that microbes can be foundcapable Of breaking almost all the linkages oforganic chemistry is fostered by the study ofantibiotics, which include a wealth ofpreviously "unphysiological" linkages--azocompounds, oximes, N-oxides, aliphatic and

. aromatic nitro and halogen compounds, andstrang9 heterocyclic ring systems. Somenatural heterocycles, e. g,, pulcherriminicacid and 2-n-nony1-4-hydroxyquinoline, havea disquieting resemblancelto the potentcarcinogen 4-nitroquinoline

@

N-oxide.

PROTOZOA AS TOXICOLOGICAL TOOLS

A difficult pfoblerri is one mentioned earlier:persistence joined with lr-grade toxicity tohigher animals. Recent developments in theuse of protozoa as pharmacological tools showthat protozoa can serve as sensitive detectorsof metabolic lesions ("side actions " ?) of awide assortment of "safe" drugs. The listincludes the "antich,olesterol" triparanol(MER/29). TriParanol toxicity manifested

. itself with severalprotozoa, includingOchromonas danica (Aaronson et al., 1962)and Tetral1mm (Holz et al., 1962).Tkparanolwas not acting simply as an anti-.

cholesterol for its obvious toxicity to protozoawas annulled by fatty,acids as well as' bysterols. The connection between the protozoanresults and the-ttside actions" of triparanol--baldness, impotence, and cataracts- -are ofcourse Unclear, but protozoal toxicity mightserve as an initial warning That it might not,be as harmless as assumed from short-termexperiments with higher animals.

A

. 7-8

The.anticonvulsant primidone provides aclekr indication of how protozoa can be usedto pinpoint the location of a metabolic lesion.Primidpne had been known to cause folic acid-.responsive-anemias. It is therefore ea+tofind that with joint use of a thymine-dependent .

Escherichia coli and the flagellate Crithidiafasciculata reversal of growth inhibition byfolic acid and related compounds permittedthe charting of interferences with the inter-connected folic acid, biopterin, and'DNAfunction (Baker et al., 1962), which amplyaccounted for the megaloblasic anemias.Lactiohacithis casei, a bacterium much usedin chemotherapeutic research, was unaffectedb§ the drug.

In another instance, Where the mode pf actionof the drug in higher animals was Unknown,growth inhibition property of the anticancercompound 1-aminocyclopentane-l-carboxylicacid was. reyersed for Ochromonas danicaby L-alanine and glycine, as wag-th-e'rnhibitionproperty of 1-amino-3-methyl-cyclohexane-1-carboxylic acid by L-leucine (Aaronsonand Bensky, 1962).

Growth irklvdbition of Euglena bythe, potentcarcinogen 4-nitroquinoline N-oxide wasannulled by a combination of tryptophan,.tyrosine, nicotinic acid, phenylalanine,uracil (Zahalsky et al, ; .,1962) and, in morerecent experiments, the vitamin K relativephthiocol. These N-oxides are of interestbecause of recent work indicating that perhapsthe main way in which:the body converts suchcompounds as the amino hydrocarbons to theactual carcinogens m4y be by an initialby oxylation of the nitrogen, e. g., workb Miller et al., (19610. Whether thepe oxides in photo - chemical smogs of theLos geles type action hydrocarbons toprodu e carcinogen' N-oxides is entirelyunkno n. Leighton ( 961) ,lists, an array ofperoxy reactions pro uced by sunlight inpolluted ail..

Our aforementioned experience with primidone,a ketonic, heterocycle, le us to test thesedative thalidomide. It as toxic forOchromonas danica O. m. emensis andTetrahymenapyriformis; this toxicity was

ry 101

Environmental Requirements of Fresh-Water Invertebrtes

annulled by nicotinicacid (or nico inam de)or vitamin K (menadione) (Frank et al, 1963).We do not know whether a similar protectioncould have been conferred on human embryosor polynedritis in the adult.

Many widely used herbicides of the dinitrophenoltype are powttrful poisons for higher animals.We do not know how sensitive protdzoa wouldbe Rix- detecting their persigtence. However,since somewhat similar thyro-,active com-pounds can be Sensitively detected by their-exaggeration of the B12 requirement ofOchromonas malhamensis (Baker of al, 1961),this flagellate might be a useful test objectfor dinitrophenols and congeners.

The PararneCium (ai-id perhaps too the.Tetrahymena) test for polynuclear benzenoidcarcinogens has remarkable sensitivity andspedificite (Epstein and Btirroughs, 1962; Hull,1962): This test depends on the carcinogen-sensitized phot9dynamic destruction ofparamecia by ultraviolet light. This test isapproaching practicality for air, and there is.noTeason to suppose it cannot be applied to;benzene extracts of foodstuffs and water.

CONCLUSIONS

We have suggested here new procedures forexamining the intrinsic toxicity-persistencerelationship, using as test organisms the,protists represented conspicuously in aWinogradsky column. The new field ofmicro - toxicology is virtually undeveloped.Th4rgent need for detecting chronic, low-grade toxicities is evident from many sides.This is not the place for a detailed discussion..,of the medical implications of phis area ofresearch, but it should be eplphasiied thatchronic toxicities and carcinogenesis are'related. Conversely, Umezawa (1961) hasremarked that most antitumor subStance'shave chronic toxicities and that elaboratetesting procedures for toxicity are requiredtq, fix the daily,tolerable dose; appa'rently thisproblem is a central theme in medical as wellas pollution research. Inhibition of girth ofanarray of protozoa is now in practiCal useUsi a means of detecting anticancer substancesin antibiotic beers (Johnson et al, 1962).

.

4.

1 f . 102.

0

Since the embryos appear to lack thedetoxication mechanisms of the adult animal(Brodie, 1962), toxicity for protozoa (which _-presumably lack these detoxicationmechanisms) should be compared with thatfOr the embryo, not the adult, as emphasizedby the thalidomide disaster.

There 'are further limitations on the use ofmicrobes as detectors of toxicity. High-molecular toxins seem inert to microbes,and antihormOnes (with the exception of

anti-thyroid compounds) are generally inert.'The main usefulness of microbial indices bftoxicity would-appear, then, to be for detectinglow-molecular poisons acting on cellilartargets rather than on cell` systOms and

, organs. These are precisely the poisonslike to'put out of: business a pollution-contr 1 installation primarily dependent onmicrobial activity.

REFERENCESfr

Aaronson, S. and Bens*, B. 1962._protozoological: studies of the cellular'action of drugs. I. Effect of1-aminocyclopentane-1- carboxylic acidand 1-4amino -3 -methylcyclo-hexanel-1carboxylic acid on the phytoflagellateOchromonas danica. Biochem.` Pharmacol.11:983-6.

Aaronson; S., Bensky, B., Shifrine, M. andBaker, -H. 1962. Effect of hypotholes-teremic agents on protozoa. Proc. Soc.Exptl. Biol. Med. 109:130=2.

IAleXander, M. 1961. "Introduction to Soil

Microbiologir", John Wiley & Sons, N. Y. .'(see p. 240).

Bhker, H., Frank, O., Hutrier, S. H.,Aaronson, S., Ziffer, H. and Sobotka,. 1t.1962. Lesions in foljc acid ,metabolisminduced by sprimidone 'Ekperientia18:224-6.

Baker, H., Frank, O., Pasher, , Ziffer, H.,Hutner, S.H. and Sobotka, H, 1961.Growth inhibition of microorganisms bythyroid hormones. Proc. Soc. Exptl. .

Bio. Med. 107:965-8.

7-0

.)

I.

5Berger, B.B. +1961. Research needs in

quality conservation. In "Algae and'MetriSpOlitan Wastes", Trans. 1960Seminar, Robt. A. 'itrftSanitazy Eng.Ceriter, Cincinnati 26, Ohio, p. 156-9.

Enviromtental Requirements of Fresh-Water Invertebrates

Brodie, B. D, 1102. Drug Metabolism-subcellular mechanisms. In "Enzymesand Drug Action", J. L. Mongar andA. V. S. de Reuck, eds., Ciba FoundationSymposium, J. & A. Churchill Ltd.,p. 317-40.

Eppley; .R. W. and MaciasR, F. M. 19.62.Rapid growth of sewage lagoonChlamydomonas with acetate. Physiol.Pls.ntarum 15:72-9.

ter

Epstein, S. S. and Burroughs., M. ' 1962.Some factors influencing the photodynamicresponse of Paramecium caudatum to3,4-benzypyrene. Nature 193:337-8.

Frank, 0.4 Baker, H., Ziffer, H.,- - --Aaronson, S., and Hutner, H. 1963.Metabolic deficiencies in protozoa inducedb-§ Thalidomide. Science 139:110-1:

Hawkes, IL A. 1960. Ecology of activatedsludge and bacteria beds. In "Waste .

Treatment ", P. C. G. Isaac, ed.,Pergamon, Nr. Y., Oxford, etc., p.'52 -97;discussion p. 97-8.

Holz, G. . Jr., Erwin, J., Rosenblum, LT.and Aar son, S. 1962. Triparanol 'Inhibition of Tetrahymena and its preventionof lipids. Arch. Biochem.. Biophys.98:312-22.

Hull, R.W. 1962. Using the "Parameciumassay' to screen carcinogenic hydrocarbons.J. Protozool. (Suppl) 9:18.

Hutner; S. H. .1962. Nutrition'of protists.In "This is Life: Essays in ModernBiology", W. I. Johnson and W, C. Steere,eds., Holt, Rinehart and Winston, N.Y.P. 109-37.

Hynes, H. B. N. 1960. The Biology of 13olltitedWaters. Liverpool Univ. Press.

1-10

Johnson, I. S. , Simpson, P..J, and Cline, 'J. C.1962. Comparative studies Aithchemotherapeutic agents in biologicallydiverse in vitro cell systems. CancerResearch 22:617-26.

Leighton, P.A. 1961. "Photochemistry ofAir Pollution ", Academic Press, N.Y.

Miller, J. A. , Wyatt, C. S. , Miller, E. C.and Hartman, -H. A,. 1961. TheN-hydroxylaiion of 4- acetylamino-bifheyl by the rat and dog and thes g carcinogenicity of N-hydroxy-4:cety ,sminobiphenyl in the rat, Cancer

Research 21:1465-73.

Provasoll, L. 1961. Micronutrients f4mdheterotrophy as possible factors in bloomproduction in natural waters. In "Algae1an Metropolitan. Wastes", Trans. 1960Seminar, .Robt. A. Taft Sanitary Eng.Center, 'Cincinnati 26; Ohio, p. 48-56.

.Ross, W. K. and Sheppard, A . A. 1956.Biological oxidation of petroleum phenolicwastewaters. In "Biological Treatmentof Sewage-4nd IndUstrial Wastes", J.McCabe and W. W. Eckenfelder, Jr., eds.,1:376=8. Reinhold Publ.. Co., N. Y..

Scher, S., Scher, B. and hutner, S. H. 1963.Notes on the natural history of ,RhodOpseudomonas galustris. In"Symposium on Marine Microbiology",ed. C.H. Oppenheimer, C: C. Thomas,Springfield, Ill., p. '580-7.

Umezawa, M. 1961. Test methods for- antitumor substances. Sci. Repts. 1st.

Super-.- Sanita 1:437-38 4.

Zahalsky,. A. C., -Keane, K., 'Hutner', H.,Kittrell, M, and Amsterdam, D. I962. '-Protozoan response to anticarCinogenicheterocyclic N-oxides: toxicity andtemporary bleaching. J. Protozool.(Suppl.) 9:42.

-"Biological Problems in Water Pollution"(Third Seminar 1962) U.S. Public HealthService-Pub. No. 999-WP-25, Cincinnati,OH . 1965, II

.5

%.

110 3

-\ I /INTRODUCTION

FILAMENTOUS BACTERIA

1

There are a number of types of filamentousbacteria that occur in the aquatic environment.They include the sheathed sulfur and ironbacteria such as B_ esgiatoa, Crenothrix andSphaerotilus, the actinomycetes-which areunicellular microorganisms that form chainsof cells with special branchings, andGallionellae a unicellular organisrli thatsecretes a long twisted ribbon-like stalk.These filamentous forms have at timescreated serious problems in rivers,reservoirs, wells, and water distributionsystems.

II BEGGIATOA

Beggiatoa is a sheathed bacterium that growsas a long filamentous form. The flexiblefilaments may beias-large as 25 microns wideand 100 microns long: (Figure 1)

Begglatoa albk

,=,13.A.: 8a. 5.80

151.1 X

up to 1.000pi,

104

Transverse separations within the sheathindicate that a row of 'cells is included inone sheath. The sheath may be clearlyvisible or so slight that only special-stainingwill indicate that it is present.

The organism grovis as a ii,hite slimy or.felted cover on the, surface of various objectsundergoing decompdsition or on the surfaceof stagnant areas of a stream receivingsewage. It has also been observed.on thebase of a trickling filter and in contactaerators.

It is most commonly found in sulfur stringsor polluted waters where H2S is present.3eggiatoa is distinguished by its ability todeposit sulfur within its cells; the sulfurdeposits appear as large retractile globules.(Figure 2)

Figure 2

Filaments-of Beggiatoacontaihing granules ofsulphur.

t

When H2S is no longer present in the environ-ment, the sulfur deposits disappear.Dr. Pringsheim of Germany has recentlyproved that the organisin can groiv as a trueStitotroph obtaining all its energy from theoxidation of H 2S and using.this energy to fixCO,,' into alkmaterial. It can also use .

certain Materials.if they are presentalong with.

the H2S.

,1Faust and k\rolfe, and Scotten and Stokeshave grownpe organism in pure culture. inthis country'... Beigiatoa exhibits a motilitythat is quite different,from the typicalflagellated motility of most bacteria; thefilaments have a flexible gliding motion.

8-1

, _

Filamentous Bacteria

The only major nuisance effect of Beggiatoaknown has been overgrowth on.trickling filtersreceiving waste waters rich in H2S. Thenormalinicroflora of the filter was suppressedand the filter failed to give good treatment.Redi Oval of the H 2S from the water by blbwingair through the water before it reached thefilters caused the slow decline of theBeggiatoa and a recovery of the normal ,microflora. Beggiatoa usually indicatespolluted conditions with the presence of H2Srather than being a direct nuisance.

III A CTINOMYCETES! AND EARTHY ODORSIN WATER

Actinomycetes are unicellular microorganisms,lsmicron in diameter, filamentous, non-sheathed, branching monopodially, andreproduced by fission or by meanstof specialconiqia.... (Figure 3)

1,

Figure 3

-Their filamentous habit and method ofsporulation is reminiscent of fungi. However,-their size, chemical cbrigiosition,4and other.characteristics are more similar to,bacteria.(Figure 4)

Filamente. of Actlnomycetes.

-

du

Figure 4

Egg albumin isolation plate.'A' an actinomycete colony.and 'B'.a bacterial colony

`11

4viofAppearance:

II and powdery /smooth. and MUcOld

These organisMs may be considered as agroup intermediate between the fungi andthe bacteria. They require organic matterfor growth but can use a wide variety ofsubstances and are widely distributed.

Actinomycetes have been implicated as thecause of earthy-odors in some drinlqgwaters (Romano and Safferman, Sily and.Roach) and in earthy smelling substance hasbeen isolated from several members of thegroup by Gerber and Lechevalier. Saffermanand Morris have reported on a method for the"Isolation and Enumeration of ActinomycetesRelated toyWateeSupplies. " But the actino-mycetes are primarily soilinici-oorganismsand often grow in field's or on the banks of ariver or lake used for the water supply.Although residual. chlorination will kill theorganisms in the treatment plant or distribqion

105

o

1

.r

. Filamentous 'Bacteria

system, the odors often are present-beforethe water enters the plant. Use of perman-ganate oxidation and activated carbon filters

shave been most successful of the methodstried to rJmove the odors from the water.control procedures to prevent the odorousma rial from being washed into the watersup151 rains or to prevent possible develop-men of t e actinomycetes in water rich indeca ing organic matter is still'needed.

IV FILAMENTOUS IRON BACTERIA

The filamentous iron bacteria of theSehaerotilue- Lept othrix group, Crenothrix,and Gallionella have the ability, to eitheroxidize manganous or ferrous ions to innicor ferricsalts-or are able toprecipitates of these compounds within thesheaths of the organisms. Extensive growthsor accumulations of the empty, metallicencrusted sheaths devoid of cells, havecreated much trouble in well's or water dis-tribution systems. Pumps and back-surgevalves have been clogged with masses ofmaterial, taste and odor problems haveoccurred, and rust colored masses ofmaterial have spoiled products in contactwith water.

Crenothrix polyspora has only been examinedunder the microscope as we have never beenable to grow it in the laboratory. The orga-nism is 'easily recognized by its specialmorphology. Dr. Wolfe of-the University-ofIllinois has published photomicrographs ofthe organism. (Figure 5)

Orgapisms of the S haerotilus-Leptothrixgroup have been extensivel studied by manyinvestigators (Dondero . al., Dondero,Stokes,- Weitz and 1,0. ey, Mulder and van,Veen, and Amberg an Cormack.) Under

jdiffereht ,environmental conditions the mok-,

.phological se- ance of the organism varies,, -The usual farm f in polluted streams or

bulke vated sludge is Sphaerotilus natans;(Fly re 6)

..10G.

Figure.5Crenothrix polyspora

cells are very variable insize ttom small cocci orpolyspores ta cells 3x12/.4

Figure 6

Sphaerotllus natans

3-8 x 1.2 1.8fp. cells

8-3

Filamentous Bacteriae

This is a sheathed bacterium consisting of ,

long, unbranched filaments', .whereby individualrod- shaped bacterial cells are enclosed in alinear order within the sheath. The individual

,cells. are .31 microns long and 1.2-1.8microns wide. Sphaerotilus grows in greatmasses; at times in streams or rivers thatreceive wastes from pulp mills,' sugarrefineries; distilleries, slaughterhouses,or ilk prOoessing plants. In these conditions,it ap ars as large masses or tufts attachedto rocks, twigs, or other projections and -themasses may vary In color from light grey to

,reddish brOwn. In sortie rivers large tnassesof , Sphaerotilus break loose and clog watetintake pipes or foul fishing nets. When thecells 'die, taste and odor proble4 may alsooccur in the wa ,

Amberg, Cormack, and Rivers and McKeanhave reported on methods to try to limit 'thedevelopment of Sphaerotilus in rivers byintermittant discharge of wastes. Adequatecontrol will probably only be achieved once

.the wastes are treated before discharge tosuch an extent that the growth of Sphaerotilusis no longer favored in the river. Sphaerotilusgrows welt at cool temperatures-and slightlylow DO levels in streams receiving thesewastes and domestic sewage. Growth is slowwhere the only nitrogen present is inorganicnitrogen; peptones and proteins are utilizedpreferentially.

Gallionella is an iron bacterium which appearsas a kidney-shaped cell with a 'twisted ribbon-like stalk emanating from the concavity of thecell. Gallionella obtains its energrbyoxidizing ferrous ironlnerric iron and usesonly CO2 and inorg)cnic salts to forth all- ofthe. cell material; it is an antotroph. Large.masses of Gallionella may clause problemsin wells or accumulate in low-flow how=pressure water mains. Super chlorination(up..to-p0, ppm of sodium hypochlorite for48 hours) followed by flushingill oftenrer hove tile masses of growth and'periodictreatmen will preirentthe nuislince,effeCtsof the extensive masses1/4& Gallionella.(Figure 7)

4

Figure 7Galionella furrugfnea

OA X PI -1.1pCells

r- Faust, L. and Wolfe; R. $. Enrichment.*; and Cultivation of Beggiatoa Alba.

Jour. Bact., 81:99-106. 1961.

2 Scotterrr-H. L. and Stokes, J. L.Isdlation and. Propertifs of Beiatoa.Arch Fur. Microbiol. 42:3537368.

. .1962.

3 -Kowallitk, U. and Pringsheim, .B. G.The Oxidation of Hydrogen Sulfide by

a Beggiatoa: Amer. Jour. of Botany:53:801-805. 1966, .

Actinomycetes and Earthy Odors.,.4

5

REFERE ES

Beggiatoi.

Silvey, J.,K. G: et al. A ctinomycetes andCommon Tastes and Odors. JAWWA,42:1018-1026. 1950.

Safferman, R. 5; and Morris, M. E%A Method for the Isolation and,Enunieration of ActinomyCetes Relatedto Water Supplies. Robert A. TaftSanitary Engineering Center Tech.Report W-62-10. 1962.

1Q7

^1.

00

Filamentous Bacteria

.6 Gerber, N. N. and Lebhevaliert, H.A._j Geosmin, an Earthy-Smelling Substance

Isolated from Actinomycetee. Appl.Microbiol. 13:935-938. 1965.

.

re-

°Filamektous Iron Bacteria

7 Wolfe, R. S. Cultivation, Morphology, andClassification of the Iron Bacteria.:JAWWA. 50 :1241 -1249. 1958.

.E1 Kucera, S. and Wolfe, R. S. A SelectiveEnrichment Method foi Gallionellaferruginea. Jour. Bacteriol. 74 :344-349. 1957.

9 Wolfe, R. S. Observations and Studiesof Crenothrix polyspora. JAWWA,52:915-918. 1960.

10 Wolfe, R. S., Microbibl. Concentrationof Iron and Manganese in Water withLow Concentrations of theiefrElements.JpWAr. 521335=1337. 1860.

11 Stokes,c4.,L. Studies on the FilamentousSheathed Iron Baetiriurn Sphaferotilusnatans. Jour; Bacteriol. 67:278-291.

41954.

1Z, 'Waltz, S. and Lackey; J. B. Kerphologicftiand Biochemical Siudies on the _

OrIganismSphaerotilus natans. arf.Joy. Fla. Acad. -Sci. 24(4)131958.

"

13' Dondero, N.C., Philips, R.A. anr .

Henkelkian, H. Isolatidn andPreservation of Cultures of SohaerotillAppl. 9:2191-227% 1961.

14 Eikelboom D. H. , Identification-QS__Filamentous Organisms in BulkingActivated Sludge.- Frog. WaterTe0.,h. 8(8):155-161.' . 4

Pergamon Press.

<

15 Donlero, N. C. Sphaeretilust Its Natiireand Economic Signifirianee. AdvancesAppl. Microbiol. 3:717-107. 1961.

,16 Mulder', E. G. akid van Veen, W. L.,

Investigations on the SphaerotilUs' Leptothrix Group. Antonie vanLeewenhoek 29:121-153. 963.

kip). .r .-17 Amberg, H.R. and Cormac , J. F. _

Factors Affecting Slime Growth inthe Lowers Columbia 'River andEvaluation of Some Possible ContrOl.Mea'sures. Pulp and Paper Mag. ofCanada. 61.;T70:-T80. 1960.

18 Amberg, H.R. , Cormabk, J. F. andRivers, M. R. Slifne. Growth Controlby Intermittant Discharge of SpentSulfite Liquor.. Tappi. 45':770-779.1962.

.

19 McKeown, 4. J. - The Sonttol ofSphaerotii.us natans. Watchrand Wastes, 47:137 19-22 and8:(4)30-33'. 1963.

-20 CtiPtis, ,L J. C:, Sco,a3e Fun-,:us;: Nature.and Effects. Wat. Res.

.3:289.4311. 1969,

21 Lechevalier, Hubert A., P:ctinomycefesof Sewage 'treatment Plants. Envir.Prot. Tech. Series USEPA,'600/r-75-031. 1975.Io °

This outline was prepared by R. F. 'LewisBacteriologist, Advanc aste TreatmentResearchLaboratory, NER,C, 'USEPA,Ciricinniliz-OhiO 45268.

Descriptors: Aquatic Bacteria, Sphs.erotiluslActinomycetes, Nocardia

j .

I INTRODUCTION

I

FUNGI AND TUE SEWAGE FUNGUS" COMMUNITY

\st III ECOLOGY

Descri tion

Fungi a e heterotrophicachylorophyllousplant-like organiSms which possess-truenuclei with nuclear membranes andnu-cleoli. Dependent upon the species andin some instances the environmentalconditions, the body of the fungus, thethallus, varies from a microscopicsingle cell` to an extensive plasmodiumor mycelium. Numerous forms produce r

. ,macroscopic fruiting bodies.

B Life Cycle

Thelife cycles of fungi vary from simpleto complex and may include sexual andasexual stages with varying spore typesas the reproductive units'.

C ClairSifica.tion -

Traditiorially, true fungi are classifiedwithin the Division Etnnycotina of thePhylum Mycota of the plant kingdom.Some authorities consider the fungi anessentially monophyletic group distinctfrom the classical plant and animalkingdoms.

U AC' IVITY

I gen eral, -fungi possess:btoad enzymaticapacities; Various species are able- to

a.ctiOely, degrade such compounds as.complex polysaccharides (e. g., cellulose,chitin, and glycogen), proteins (casein,albumin, keratin), hydrocarbons (kerosene)and pesticides. 'Most species, possess anoxidative or microaerophilic metahq4111,bit anaerobic catabolism is not uncommon.A fespeCie.s.iihow anaerobic metabolismand growth.

BI. FU. 6a. 5. 80 109

A Distribution

Fungi are ubiquitous in nature and mem-bers of all classes may occur in largenumb aquatic habitats. Sparrow(1968) has riefly reviewed the ecology

of fungi in reshwaters with particularemphasis on the zoosporic phycomycetes.The occurrence a.p.d ecology of fungi in'marine and estuarine waters has beenexamined recently by a number of in-vestigators (Johnson and Sparrow, 1961;Johnson, 1968; Myers,1968; van Udenand Fell, 1968).

B -Relation to Pollution

Wm; Br'cllxe Cooke, in a series of in;vesti ions (Cooke, 1965), has estab-

, lished that fungi other than phycomycetes_occur in high numbers 'in sewage and

pollutedAters. His reports on organicpollution of streams (eooke, 1961; 1967)show that the variety of the Deuteromy-cete flora is decreased at the .immediatesites a pollution, but dramatically in-creaseddownstream from these regions.

Yeasts,- in particular, have been foundinnarge numbers in organically enrichedwaters (Cooke, et al., 1960;- Cooke and.

.Matsuura, 1963; 'Cooke, 1965b; Ahearn,et al., 1968). > Centain yeasts are ofspecial interest. due-to their potential°use as "indicator" organisms and theirability to degrade or utilize proteins,various hydrocarbons,. straight andbranch, chained alkyl-benzene sulfonates,Tats, .metaphosPhates, and wood sugars.

9-1*

(

Fungi

C "Sewage Fungus" Community (Plate. I)

A few microorganisms have Tong beentermed "sewage fungi. " The mbstcommon microorganisms includsidinthis group are the iron bcterkSphaerotilus natans and the phycomy-cete- Leptomitus' lacteus.

\1 Sphaerotilus natans is nota fungus;'

rather it'is a sheath bacteritlyn ofthe order -chlamydobacterialts.This polymorphic bactersium occurs

.clommonly in organically enriched'streams where it may produceextensive slimes.

a Morphology

Characteristically, S. natansforms chains of rod shaped

o cells (1. - 2.04 x 2.5 - 174).within 'a clear sheath or tri-clome composed of a protein-poly saccharidae-lipid complex.The rod cells are frequentlymotile upon release from thesheath; the flagella are lophO-trichous. Occasionally tworow.s-of cells may be presentin a single sheath. Single tri-chomes may be several mmin lerigth and bent at variousangles. Empty sheaths, ap-pearirig like thin cellophanestraws, may be present.

- $

13' Attached growths

The trichomes are cementedat one end to solid substratasuch as stone or metal, andtheir cross attachment andbending gives a .superficialsimilarity to true fungal hYphae.The ability to attach firmly tosolid substrates gives S. natansa selective advantage in thepopulation of flowing streams.For more thorough reviews ofS. natans see Prigisheim (1949)and Stokes (1954).

4..

2 Le om. itus lacteus also ,producesex ensi, e slimes and fouling flocs ,In es water's. This speciesiorrnsthal ified by regular constrictions.

a Morphology

Cellulin plugs may be present *near the constrictions and theremay be nurrterous granules inthe cytoplasm. The basal cellof the thallus may possessrhizoids.

b Reproduction

The segments delimited by the..partial constrictions are con-verted basipetally to sporangia.The zoospo'res are diplanetic(i.e., dimorphic) and eachpossesses one whiplash and onetinsel flagelluth. No sexualstage has been demonstratedfor this species.

. c Distribution

For further information on thedistribution and systematicsof L. lacteus see Sparrow (1960),Yerkes I1h66) and Emerson andWeston (1967). Both S. natansand L. lacteus appear to thrive -

in organically enriched coldwaters (59-22°C) and both seemincapable of extensive growth attemperatures of about 30°C.

d Gross morphology

Their metabolism is oxidativeand growth of both specie ayappear as reddish brown flocsor stringy.slimes of 30 cm ormore in length.

e NUtritive requirements

Sphaerotilus natans is able toutilize a wide variety of organiccompounds, whereas L. lacteusdoes not assimilate simple

14.

Fungi .

PLATE, I

"SEWAGE FUNGUS" COMMUNITY OR "SLIME GROWTHS"

(Attached "filamentous" and slime growths)

Sphaetotilus natans

9

Begeatoa alba

3

BACTERIA

:

.

Fusarium aqueductum Leptomitus lacteus

1--I44-

FUNGI

7

9

CarchesiuM

ro

OpercUlaria

PROTOZOA

Fungi

.° PLATE IIREPRESENTATIVE FUNGI

Figure

Fusarium aquaeductuum(Radlmacher andRabenhorst) Saccardo

Microconidia (A) producedfrom phialides as in Cephalo-IPOriUM. remaining in slimeballs. Mac:soma& (B), withone to several cross walls,produced from collared phial.ides. Drawn from culture.

Figure

Leptomitus lacteus (RAgardh

Cells of theqy hae show.ing_constrictionVivith cellulmplugs. In ,one/cell large zoo.spores hive been deliraid.

. Redrawn from Coker, 1

Figure 4).-

Zoophagus in:idiomSommerstorff

Mycelium with hyphal pegs(A) on which rotifers willbecome impaled; gemmae (B)produced as conidis on shorthyphal branches; and rotiferimpaled on hyphal peg (C)from which hyphae have

grown into the rotifer whosethell will be discarded afterthe contents are duisumed.Drawn from culture.

Figure .3Geouichum candulumLink ex Persoon

Mycelium with short cellsand arthrospores. Young bypha (A) ; and mature arthro-spores (B). Drawn from culture.

Figure Snichlya americana Humphrey

Oeogonium with three oo-spores (A); young rmosier-121011M with delimited Zoo-spores (R) and zeosporangia(C) with released zoosporesthat remain encysted in dus-ters at the month of the dis-charge tube. Drawn from ma-ture-

r wrier/dub rolm M11111111111

A

FIGUR); 7 Ilapinsporideum ccWale. spots;

Figures 1 th;ough 5 from Cpoke; FigUres 6 and 7 from Galtsoff.

Fungi

sugars and grows most lu,iniriantly:inthe pres1,-nce of organio nitrogenouswastes.

3 Ecological roles

Although the "sewage fUgg?"_oltoccasion attain visdally notic.eableconcentrations,,the less obitiouspopulations. of cfeuteromycetes maybe more important in the ecology ofthe aquatic habitat. Investigations ofthe past decade indicate that numerousfungi and of primary importance in themineralization of organic wastes; theoverall significance and exact roles offungi in this process are yet to beestablished.

D Predacious Fungi

1 Zoophagus insidians

(Plate H, Figure 4) has been observed. to impair functioning of laboratoryactivated sludge units (see Cooke and'Ludzack). ,

2 Arthrobotrys is usually found alongwith Zoophagus in laboratory activatedsludge units. This fungus is pretiacioua

, upon nematode's. Loops rather than"pegs" are used in snaring nematodes.

PLATE II (Figure 4)

. 113

IV CLASSIFICATION4,.

In recent classification schemes, classesof fungi are distinguished primarily onthebasis of the morphology of the sexual andzoosporic stages. In practical schematics, 'however, numerous fungi do not dernonstVethese sta s, -.Classification must thereforebe based nlhe sum total of the morphologicalarid/or physiological characteristics. Theextensive review by Cooke (1963) on methodsof isolation and classification of fungi fromsewage and polluted waters precludes theneed herein of extensive keys and species-illustrations. A brief synopsis key of thefungi adapted in part from Alexopholous(1962) j.s presented on the following pages.

1,58

This outline was prepared by Dr. Dontld G.'Ahearn, Professor of Biology, Georgia StateCollege; 'Atlanta, Georgia 30303.

Des Cr iPtbr Aquatic Fungi41,

,

-1

9-5

t

e,a

'Fungi

.KEY TO THE MAJOR TAXA OF FUNGI.A.

Definite cell walls lacking. somatic phase a free li;)ing Plasmod ium : :$41=n

Sub-phylum Myxomycotina . (true slime cnolcfs)..Class Myxomycete'Cell walls usuallyiwell defined, somatic phase not a free-living Plasvmodikm. a

(true fungi) Sub-phylum Eumycotina . ..... .2 t.

2 Hyphal filaments usually coenoctytic, rarely septate, sex sElls when pciesent farmingoospores or zygospores, aquatic species propagating asexually by zoospores, terrestrialspecies by zoospores, sporangiospores conidia or conidia-like sporangia .VPhycomycetes".: . 3

The phycomycetes, are generally considered to include the most primitive of the truefungi. As a whole, they encompass S wide diversity of forms with some ,showing relation-ships to the flagellates, while others closely resemble colorless algae, and still othersare true molds. The Vegetative body (thallus) maybe non-specialized and entirely con-verted into a reproductive. organ (holocarpic), or it may bear tapering rhizoids, or bemycelial and v?ry extensive. The outstanding characteristics of the thallus is a tendencyto be nonseptate and in most groups, multinuNiate; cross walls are laid down in vigorouslygrowing material only to delimit the rAfeorductive organs. The spore unit of nonsexual re-production is borne in a siporangium, and, in aquatic and semiaquatic orders, is providedwith a single postdrioror anterior flagellum or two laterally attached ones. Sexual activityin the phycomycetes characteristically results in the formation of resting spores.

2.t1'1 Hyphal filaments when present sefitate. without zoospores. with or without sporangia.usually with conida; sexual reproduction absent pr culminating in the formation of ascior basiaia 8

3 (2) Flagellated cells characteristically produced 43' Flagellated cells lacking or rarely produced .7

4 (3) Motile cells uniflagellate 5

. 4'. Motile cells biflagellate 6

.

5 (4) Zoospores posteriorly uniflagellate, formed inside the sporangium class .Chytridiomycetes

The Chytridiomycetesoroduce asexual zoospores with a single posterior whiplashflagellum. The thallus is highly variable, the Most primitive forms are unicellular andholocarpic and in their early stages of development are plasmodial (lack cell walls). moreadvanced forms develop rhizoids and with further evollikonary progress develop myceliumThe principle chemical 'component of the cell wall is chitin, but cellulose is also present.Chytrids are typically an atic organisms but may be found in other habitats. Some speciesare re chitinolytic and/or k ratinolytic. Chytrids may be isolated from nature by baiting (et. g.hemp seeds or-pine pollen) ytrids occur both in marine and fresh water habitats and areof some economic importance due to their parasitism of algae and animals. The genusDermocystidium may be provisionally grouped with the chytrids. Species Of this genuscause serious epidemics of oysters and marine and freshzater fish,

5' Zoospores anteriorly uniflage4late, formed inside or outside the spbrangium. classHyphochyt r iciiomycetes

.These fungi are aquatic-(fresh water or marine) Chytfid-like force whbtZe motile cellsposses; a single anterior flagellum of the tinseThYpe (feather-like). They are parasitic onalgae and fungi or may be saprobic. Cell walls contain chitin with some species also demon-strating cellulosecontent. Little information is available on the biology of this class andat present it is limited to less than 20 species.

a ',. ,re' t!16.

' 6 (4!) Flagella nearly equal, poe whiplash the other tinsel claws... ...Oomycetes

. .a.,,... A number of representatives of the Oomycete; cAve been shown tp have cellulosic cell.

* rtwalls. The mycelium is coenocytic, branched and well doieloped in most cases, The sexualprocess results in the formation of a resting spot4 of the.00gamous type, i:e., a type offertilization.in which two heterogametangia cqme fri contact'and fuse their contents through...,a po4s or tube. Tke-thalli in this class r e from unicellular to profusely branchedfilamentous types. Most forms arc eucarpi zoospores arc produced throughput the classexcept in the more highly advanced species Certain species are of economic ithportance due

'ec: their destruction of food crops (potatoes and grapes) while others cause serious diseases offish (e. g. Saprolegina parasitical. Members of the family Saprolegniaceae are the common

6

4

water molds and are among the most ubiquitous fungi in nature, The order Lagenidta esincludes only a few species which are parasitic on algae, small'inimal, and other aqua islife. The somatic structures of this taxon are holocarpic and endobiotic. The sewage f ngi4are classified in'the order Leptomitales. Fungi of this order are ch:aracte rued by theformation of retractile constrictions. cellulin plugs ' occur throughout the thalli or, at 1 ast,at, the bases o( hyphae or to cut off reproductive structures. Leptomitus lacteus mayproduce rather extensive fouling floes or slimes in organically enriched waters.

II

Flagella,of unequal size, both whiplash ...... class....Pfasmodiophoromycetes

Members of this cla e obligate endoparasites of vascular plants, algae. and fungiThe thalls consists of a pla chum which develops within the host cells. Nuclear divisionat some stages of the life cycle of a type found in no other fungi but known to occur inprotozoa. Zoosporangia which ari e directly from the plasmodium bear zoospores with two"unequal anterior falgella. The cell waifs of the fungi apparently lack cellulose.

7 13') Mainly .sapro,bic. sex cell when present a zygospore.. ..... class . 7%vomycetes

This class has well developed mycelium with septa developed in portions of theolder hyphae, actively growing hyphae are normally non-septate. The asexual spore,. at Enon-motile sporangioipores (aplanospore,$). Such spores lack flagella and are usuallyaerialy disseminated. Sexual reproduction is initiated by the fusion of two gametangiawith resultant formation of a thick-walled, resting spore, the zmospore. In the moreadvanced species. the sporangia or the sporangiospores are conidia -like Many of the ,

Zygomycetes are of economic importance due to their ability to synthesize commerciallyvaluable organic acids and alcofiols. to transform steroids such as cortisone, and toparasitize and destroy food crops. A few species are capable of causing disease in manavid animals (zygomycosis)

Obligate commensals of arthropods, zygospores usually lacking chss ...Trichomycetes

The Trichomycetes are an ill-studied group of fudgi which appeao be obligatecommensals of arthropods. The trichomycete are associated with a wide variety of insecta.

2111iplopods, and crirstacea of terrestrial and aquitic (fresh and marine) habitats. None ofthe members of this class have been cultured in vitro for continued periods of times with anysuccess. Asexual reproduction is by means of sporangiospores Zygospures have beenobserved in species of several orders.

8.(2'1 Sexual spores borne in asci class ..fikscOmycetes

In the Ascomycetes the products'of meiosis; 'the ascospores, are borree in sac.like structures termed asci. The ascus usually contains eight ascospores. but the numberproduced may vary,with the species or strain. Most sp6cles produce extensive sepiatemycelium. This large class is divided into two subclasses on the presence or absenceof an.ascocarp. The Hemiasconcetidae lack arLascocarp and do not produce ascogenoushy'phae; this subclass includes the true yeasts. The Euascomycetidae usually are dividedinto'three series (Plectomycetes, Pyrenomycetes, and Discomycetes) on the basis ofascocarp structure.

8' Sexual spores borne on basidia class...4113asidiomycetes.

The Basidiomycetes generally are considered the most highly evolved of the fungi.Karyogamy and meiosis occur in the basidium which bears sexual exogenous spores,basidiospores. The mushrooms. toadstools, rusts, and smuts are included in this class.

_Sexual stage.14cking . .Form clarri.(Fungilmperfecti).Deuteromycetes

The Deuteromycetes is a form class for those fungi (with orphological affinitiesto the Ascomycetes or Basidiomycetes) which have,not demon trated a sexual stage.The generally employed classification scheme for these fung is based on the morphology

'and color of the asexual reproductive etages. This scheme is briefly outlined be'lowNehver concepts of the classification based on conidium development after the classicalatoirk_of S. J. Hughes (1953) may eventually replace the gross morphology system (seeBarron 1968).

Fungi

iY

7A

14.F

r

-J

,

- KEY TO THE,FORM-ORDERS OF THE FUNGI IMPERFEC1i

1 . Reproduction by means of conidia, oidia, or by budding 2

1' No reproductive structures present Mycelia Sterilia

1

8

2 (1) Reproduction by means of conidia borne in pycnidia Sphae rolisidales2' Conidia, when formed, not in cycnidia 3 ,

.'3 (2') Conidia borne in acervuli t;',. ,. , . Melanconiales 1

3' Conidia borne otherwise, or rep'roduction by oidia or by budding, Moniliales.-.i

KEY TO THE FORM - FAMILIES OF THE MONILIALES

1 Reproduction mainly by unicellular budding, yekst-like; mycelia' phase, if present,secondary, arthrospores occasionally produced, manifest nielanin pigmentation lacking 2

Manus mainly filamentous; dark melanin pigments sometimes produced 3

2 (1) Ballistospores produced2' No ballistospores

.'SporobolornycetceaeCryptococeaceae

'3 COnidiophore*, if present, not united into 8porodochia or synnernata 4

3' Sporodochia present Tuberculariaceae3" Synnemata present Stilbellaceae

4 (3) Conidia and conidiophores or oidia hyaline or brightly colored Monies liacCae

4' Conidia and/or conitliophores, containing dark melanin pigment Dematiaceae

444

O 4

0e

Fungi and the Sewage Fungus Community'

SELECTED REFERENCES ,I.

Ahearn, D. G., Roth,F. J. Jr . ; Meyers; S.P.Ecology and Charact erization Of Yeastsfrom Aquatic Regions of South Florida.Marine Biology. 1:291-308. 1968

Alexopoulos J. C.' Introductory Mycology.2nd ed. John Wileyand Sons, New York,613 pp. 1962

Harron, G. L. The Genera of Hyphomycetesfrbm Soil. Williams and Wilkins Co.,Baltimore. 364 pp. 1968

Cooke, W. B. Population Effects on theFungus Population of a Stream.Ecology 42:1-18. 1961

. A Laboratory Guide to Fungi inPolluted Waters, Sewage, and SewageTreatmept Systems. U. S. Dept: ofHealth, Education and Welfare, Cincinnati,132 pp. 1963 1

Fungi in Sludge Digesters.\ Purdue Univ. Proc. 20th Industrial

Waste Conference, pp 6-17.. 1965a

4The Enumeration oreastPopulations in a Sewage Treatment' }dart._Mycologia 57:6 6-103. 1965b

. Fungto Pollution ofUtah Acad. Pr

1 Populations in. Relationhe Bear River,:ldaho-Utah.c. 44(1):298-315. 1967,

and Matsuura, George S. A Studyof Yeast Populations in a Waste StabilizatipnPond System. protoplasm 57:163-187..19%3

, Phaff, Miller, M.W.,Shifrine, M., and Knapp, E. Yeastsin Polluted ater and Sewage:Mycologia 5:210 -230. 1960

EmersonTlialpTrandVe'ston,. W; H.:Aqualinderella fermentans Gen. et Sp.

Nov., A PhyComyceie Adapted to

4:4

Stagnant. waters. I. Morphology andOccurrence in Nature. Arher. J.Botany '54:702-719." 1967 t.

Hughes, S. J. Conidiophores, Conidia andClassification. Can. J. Bot. 3 r:577 -659. 1953

Johnson, T.W., Jr. Saprobic Marine Fungi.plzt-:-.95-104. In Ainsworth, G. C. andSussman, A. The Fungi, III.

_Academic Press, New York. 1968-

and Sparrow, F. K., Jr. Fungiin ceans and Estuaries. Weinheim,Germany. 668 pp. 1961

/#

Meyers, .S.P. Observations on the Physio-7'1 logicalltcology of Marine Fungi. Bull.

Misaki Mr. Biol. Inst. 12:207-.228.'1968. ,

Prigshei, E.G. Iron Bacteria. Biol. Revs.Cambridge Phil: Soc. 24:200-245. 1949

Sparrow, F. K., Jr. A quatic Phypomycetes.2nd'ed. lUniv. Mich. press, AnnArbor.

. Ecology of Fr_Fshwatertungi:pp. 41 -93k. Ainsworth G.C. andSusSman, A.S. The Fungi., III. Acad

Jress,'New York. 1968'

Stokes, J.L. Studies on thetFilamentousSheathed Iron BaCterium Sphaerotilusnatans. J. Bacterio1.67:278-291. 1954

van'Uden, N. a,rid Fell, J.W. Marine Yeasts.167 -201. In Droop, M.R. and Wood,

E. J_. F. Advan-cealn-Microbiology ofthe Sea, Academic Press, New York.1968-

1187 pp: 1960.

Yerkes, D. Observations on an Occurrenceof-Leptomitus lacteus in Wisconsin.Mycorogis. 58:976-978. 1966

4.,Cooke-,---William B. and Ludzack, F. J.Predac s Behavior in

c hated Sludge Systems. Jour. WaterPoll. Cont. Fed.s30(12):1490-1495. 1958.

. Fungi and the Sewage Fungus Corn pity

SELECTED REFERENCES (t ontinued)

Curtis, E. J. C. Review Paper'SewageFungus:_ Its Nature and Effects.Wet. Res. B:289.-311 1969.

. Curtis, E. J. C. and, Curds, C. R.Sewage Fungus in Rivers in theUnited Kingdom: The SlimeCommunity and It&ConsituentOrganisms. Wafer Res,'5:1147-1159. 1971.

O

.'PE;r1).1

-PROTOZOA, NEMATODES, AND ROTIFERS

A

I GENERAL CONSLDE4ATIONS

A Microbial quality ?onstitutes only oneaspect of water sanitation; microchemicalsand radionuclides are attracting increasingamount of attenticn

bi Microbes considered:here include bacteria,protozoa, and microscopic tnetazoa; algaeand fungi excluded..

C Of the free-living fOrmi, some aremembers of the flora and fauna, of surfacewaters; others washed into the water fromair and soil; still others of wastewaterorigin; nematodes most commonly fromsewage effluent.

D Hard to separate "native" from "foreign"free-litring 'microbes, due to closeassociation of water with soil and otherenvironments; generally speaking, bacteriaadapted to water are those that can growon very low concentrations of nutrientand zoomicrobes adapted to water arethose that feed on algae, and nematodes,especially bacteria eaters, are uncommon

yin water but in large numbers in sewageeffluent.

E More species and lower densities of.mic`robes in clean water and fewer speciesand higher densities in polluted water.

F Pollution-tolerance or nontolerance ofmicrobes closely related to the DO level,required in respiration.

G From pollution viewpoint, the followinggroups of microbes ate of importance:Bacteria, Protozoa, Nemateda, andRotifera;

II BACTERIA ,

A No ideal,method for studying distributionand ecology of bacteria in freshwater,

. _B According to Collins, (9)Pseudomonas,

Achrombacter, Alcaliguies Chromobac-Tterium, Fla..7/212usteriurn, and,Micrococcus

'" are the most widely distributed and may be- ,. W. BA. 45c. 5.80

4

considered as indigenous to naturalwaters. Sulfur and iron bacteria aremore common in the bottom mud.

C,Actinomycetes,' Bacillus sp. Aeroaressp., and nitrogen-fixation bacteria areprimarily soil dwellers and may be washedinto the 'water by runoffs.

a

E Nematodes are usually of aerobic sewagetreatment origin..

D E. coli, streptococci, and Cl..perfr iugeaare True indicators of fecarpollyon..

III PROTOZOA

A Classification

119

1 Single- cell animals in the mostprimitive phylum (Protozoa) in theanimal kingdom.i

2 A separate kingdom, Protista, Aci in-elide protozoa, algae, 'fungi, and

'bacteria proposed in thp 2nd edition'ofWard-Whipple's Fresh-Water2.3.191210,) 10) 14;

Four subphyla or classes:3

MattigdPhora (flagellates)- SubclassPhytomastigina dealt with underalgae; only subclass Zoomastig aincluded here; 4 orders:'

1) Rhizdmastigina - with flagell mor flagella and psetidopodia

2) Protomonadine. -. With 1 to 2flagelmostly free-living manapamsttic

3) 'Polymastigina - with 3.to 8flagella;..mostly parasitic Inelementary tract 9f animalsand man

4) Hyperinaktigina - 1 inh itants, of-alimentary 'tract o sects.

.

9:

10-1

Protozoa, Nematodes) anti Rotifers

4

0

b Ciliophora-or Infusoria (ciliates)no pigmented members; 2 classes;

1) Ciliate - cilia present during they_,whole trophic life; containingmajority of the ciliates

2) Suctoria - cilia present whileyoung and tentacles during p-ophic

c Sarcodina (amoebae) - Pseudopodia(faise feet) for locomotion andfood-capturingi 2 subclasses:

1) Rhizopoda - Pseudopodia withoutaxial filaments; 5 orders:

a) Proteornyxa - with radiatingpseudopodia; without test orshell

b) Mycetozo f ming plasmodium;resembilin in sporangiumformatIon

c) Amoebina - true amoebaforming lobopodis2

d) Testacea amoeba with singletest or shell of chitinausmaterial

e) Foraminifera amoeba with 1or more shells of calcareousnature; practically all marineforms

d Sporozoa - no organ of locomotion;amoeboid in asexual-ohase; allparasitic

B General' Morphology

1 Zoomastigina:'

.Relatively small size (5 to 40 p);*with- the exceptionOUthizomastigina, the

body has a definite shape (oval, leaf-like, pear-like, .etc.); common memberswith 1 or 2 flagella and some ,with 3, 4,or more; few formpit colonies; ..;:torne

present in many for feeding. .

2 Ciliophora:

Most highly' developed protozoa; withfew exceptions, 'a macro and a micro-

._ nucleus; adoral zone of membranellae,-F mouth, .and groove usually present in

swimmingand crawling forms, somewith conspicuous ciliation of a disc-likeAnterior region and little or no bodycilia (stalked and shelled forMs);Suctoria nonmotile (attached) and with-out cytostome cysts formed in most.

Sarcodine:

Cytoplasmic membrane but no cell wall;endoplasm and ectoplasm distinct or i-,distinct; nucleus with small or fargenucleolus; some with test or shell;

eying by protruding pseudopodia; fewapabIe of flagella transformation; fresh-

water actinopods usually sperical withmany radiating axopodia; some Testaceacontaining symbiotic algae and mistaken.

N. for pigmented amoebae; cysts with single\or double wall and 1 or 2 nuclei.

4 Sporozoa: to be mentioned later'.

...

C General Physiology

1 Zoomastigina:Free-living forms normally holozic;.food supply 'mostly-bacteria in growth 's

film on surfaces'or clumps relativelyaerobic, th -refore the first protozoa todisappear -in naerobic conditions andre-appearing at recovery; reproduction

. by sim-Aelission or occasionally bys,r,--,_budding. ',/,

2 Ciliophora:

Holozoic; trt# ciliate's concentratingfood particles by ciliary movement.around the mouth part; suctoria suckingthrough teycles; bacteria and small -

120 1

(

4

algae and protozoecon titute mainfood under natural condi 'ons; soshown in laboratory to th ixe otyfleadorganic matter and serum protein; not asaerobic as flagellates -.some survivingunder highly, anaerobic conditions, suchas Metopus; reproduction by simplefission, conjugation or encystment.

3 Sarcodina:

Holozoic; feeding through engulfing bypseudopodia; food essentially same asfor ciliates; DO requirement somewhatsimilar to Ciliates - the small amoebaeand Testacea frequently present in largenumbers in sewage effluent and pollutedwater; reproduction by simple fissionand encystation.

1

IV NEMATODES

A Classification

1 All in'the phylumffiemata (non'egment-ed round worms); subdivided by someauthors into two classes:

Secernentea - 3 orders:(phasmids)

Tylenchida, Rhabditida, Strongylida,a Teratocephalida; with papillae onmale tail, caiudal glands absent.

Adenophora - 6 orders:( aphasmids)

Araeolaimida, Dorylaimida,Chromdorida, Monhysterida, Enoplida,and Trichosyringida no papillae onmale caudal glands absent.

2 Orders encountered in water and sewagetreatment - Free-likring fot6s inhabitat-ing. sewage treatment plants are usuallybacteria-feeders and those,feeding onother nematodes; those inhabitating clean'waters feeding on plant matters; theyfall into the following orders:

Protozoa, Nematodes, and Rotifers

3 Tylenchida - Stylet in-mouth; mostlyplant parasites; some feed onnematodes, such as Aphelenchoides.

4 Rhabditida - No stylet in Mouth or caudalglands in tail; mostly bacteria-feeders;common genera:

MRhabiditistDiplogaster,

DiplospAte...:2ALest MT.hoides Peisferat1.fa.12..a.......ellus, and Turbatrix.

4%.

5' Dorylaimida - 'Relatively large nematodes;stylet in mouth; feeding on other nematodes,algae and probably zoomicrobes; Dorylaimuscommon genus.

6 Chrom dorida - Many marine forms;some f, e'shwater dwellers feeding onalgae; ch acterized b strong orna-

tio of knobs, bristles or!punc1 atio s in cuticle.

M nhysteridi. - Freshwater dwellers;e ophago-intestinal valve spherical toelongated; ovaries, single or paired,usually straight; common genus inwater - Monhystera.

8 Enoplida - Head usually with a numberof setae; Cobb reported one genus, .

Mononchulus, in sand filters inWashington, D. C..

.

B General Morphology

Round, slender, n nsegmented (transversemarkings. in.cuticle of some) worms;some small (about a mm long, as Tri-

o cephalobus), ma 1 to 2 mm long(Rhabditis, Dinlogaster, and tiplogasteriodeefor instance), and some large (2 to 7. nnn,such as Dorylaimus)rsex separated but fewparthenogeristic complete alimentary; canal; -with elaborate mouth parts with or without__stylet; complete. reproductive system ineach sex; no circulatory or restiratorysystem; complex nervous system withconspicuous nerve ring across oesophagus.

AK,

C General Physiology

1 Feeding - Most sewage treatment plantdwellers feeding bn bacteria; otherspreying on protozoa, nematodes, .rotifers,

10-3


. etc., clean-water species apgarentlyvegetarians; those with stylet in mouthuse the latter to pierce the body of/anithal-or plant and suck contents; metabolicwadte mostly liquid containing ammoniiunc rbonate or bicarbonate; entericpa ogens swallowed randomly withsuspending fluid, hence remote possixtaffy of sewage effluent-borne nematodesbeing pathogen-carriers.

2 ,OxYge n requirement - DO, amiarentlydiffused through cuticle into body; DOrequirement somewhat simgar toprotozoa; Rhabditis tolerating reducedDO better than other Rhabditida members;,all disappear under sepsis in liquid; somethrive in drying sludge. ,

3 Reproduction Normal life cycle requiresmating, egg with embryo formation,

- hatching of eggs inside or outside femals,4-larval Stages, and adult; few repro-duce in the Operice of males. ,

,- V ROTIFERS -

A Classificatiolii

- 1 Classified either as a crass of the 'phylumAschelminthes (various forms of worms)or as a separatel phylum (fiotifera); cbm-,monly called wfirelanimalcules, onaccount df apparent circular movement ofcilia arbund head (corona); corona con-tracted when 244a4ing or swimming andexpan. dedwithenched to catch food.

2 Of the 3 classes,-.2 (Selsooidea andBdelloidea) rollped by some authorsunder Digon8nta (i °Varies) and theother beibOlonogimont (1 ovary);eisonideacontaining rn stly marine

forms. /-3 Class Digononta containing 1 order

(Bdelloida) with 4. families, 'Philodinedae- --being the most important.

4 °Class Monogononta comprising 3 ordetst-:'Notommatida (mouth not near cente -Ofcorona) with 14 families, FloscularidaMelicertida (corona with two wreaths ofcilia and furrow between them) with 3 .

families; most import .genera includedin the order Notommatida: Brachionus,

10.4__ -

Keratella, Monostyla, richocerca,Asplanchna, Polyarthrta, Synchaeta,Midrocodon; common genera under theorder Ffosculariaceae: Floscularia,and Atrochus. Common genera underorder Melicertida: Limnias andConochilus.

5 Unfortunately orders and families( ofA rotifers partly based on character of -

corona and trophi (chewing organ),which are difficult to study, -esp. thelatter; the foot and cuticle much ea.4ier

,to study.

General Morphology and Physiology

1 Body weakly differentiated into head,.neck, trunk,- and foot, separated byfolds; in some, these regions aremerely gradual changes in diameterof body and without a separate neck;Segmentation external only.

2 Head withccirona, dosal antenna, andventral mouth; mastax, a chewing organ,located in head and neck, connected to /mouttLanteriorly by a ciliated/gullet anposteriorly to a large stoma.01 occupy

,mucli of the trunk..

3 Common rotifers reproducing partheno-genetically by diploid eggs; eggs laid inwater, cemented to plants, or carriedon female until hatching. b

g

4 Foot, a prolongation of body, usually'with 2 toes; some with one toe; some °

with one toe and an extra toe-like_structure (dorsal spur).

ome, like Phlilodina, concentratingbacteria ariciother microbes and minuteparticulate organic matter by ciliarymovement on corona larger-microbeschewed by mastax; some such orsMonostyla feeding on clumped matter,such as Cacterial growth, fungal massed,etc. at bottom; virus generally notingested - apparently undetected bycilia.

6 DO requirement somewhat simil ar toprotozoailome disappe ring underreduced DO, others, lik Philodina,surviving at as little as ppm DO,

122

.1,

eia0111=0.

VI SANITARY SIGNIFICANCE

A Pollution tolerant and pollution non-toledigat species - hard to differentiate -re/411Eing specialist training in protozoa,

- nematodes, and rotifers.

B Significant quantitative difference in cleanand polluted waters - clean waters con-taining large variety of genera and speciesbut quite low in densities.

C Aerobic sewage treatment processes(trickling filters and activated sludge

'processes, even primary settling) idealbreeding grounds for those that feed onbacteria, fungi, and mintztelLotozoa andpresent in very large numbers; effluentsfrom-sticifiCocesses carrying large num-bers of the-fie zoomicrobes; natural watersreceiving such effluents show.ing significantincrease in all 3 categories.

D Possible Pathogen and Pathogeri Carriers

1 Naeleria causing swimming associatedmeningoencephalitis and Acanthameobacausing,nonswimming associated cases.

2 Amoebae. and nematodes grownpathogenic enteric bacteria in lab;.tionealive in amoebic cysts; very few alivein nematodes after 2 days after ingestion;,virus demonstrated in nematodes onlywhen very high virus concentrations... 41*present; some"TreeliNiing amoebaepalasitizing humans.

5\ Swimming ciliates and some rotifers(Concentrating food by corona) ingestinglarge numbers of pathogenic enerticbacteria, but digestioh rapid; no .

evidence of concentrating virus; crawlingciliates and flagellateei feeding on- clumpedorganisms,' ,

,4 Nematodes concentrated from sewageeffluent in Cincinnati area showinglive E. coli and streptococci, but nothuman enertic pathogens.

VII EXAMINATION OF WATER FOR MICROBES

A Bacteria - not dealt here.

ProtozolLIImatodes.1_and Rotifers°

B Protozoa and rotifers 4 should be includedin examination for planktonic microbes.

C Nematodes

D Laboratory Apparatus(3)

1 Sample Battles - One-galron glass orplastic bottles with metal or plasticscrew caps, thoroughly washed andrinsed three times with distilled water.

2 Capiplor Pipettes and Rubber Bulbs -Long (9 in.) Paaeur capillary pipettesand rubber bulbs of 2 ml capacity.

3 Filtration Unit - Any filter holderassembly usetd bacteriologicalexamination. 'The funnel should beat least 650 ml and the filter flask atleast 2 liter capacity.

j 4. Filter Membranes - Millepore SS (SSiiirgiM) type membranes or equivalent.

5 MicrOsc..._me - Binocular microscope,with 10X eyepiece, 4X, 10X, and 43Xobjectives, and mechanical stage.

E Collection of Water Samples

Samples are collected in the same manneroas those for bacteriological examination,except that a dechlorinating agent is not:needed. One-half to one gallon samples arecollected from raw water and one-gallonsamples from lap water. Refrigeration isnot essential and samples may be transportedwithout it unless examination is to be delayedfor more than'five days.

F Concentration of Sainples\

1 One gallon of tap water can usually befiltered through a single 8-u membranewithin 15 minutes unless the water hashigh turbidity: -"At least one gallon ofsample should be used in 'a single examina-tion. Immediately after the last of thewater is disappearing from the membrane,the suction line is disconnected and themembrane placed on the wall of a clean50 to 100 ml beaker and flushed repeatedlywith about 2-5 ml of sterile distilled water

10-5

Pr'otozoa, Nemato4s and Rotifers

with the aid of acapillary pipette and arubber bulb. The concentrate is thenpipetted into a clean Sedgewick-RafterCounting Cell and is ready for examina-tion.

2 1p concentration of raw water sampleshaving visible turbidity, tem to :our87rriicron membrapes. may be requiredper sample, with filtration through eachmellibrane being limited to not morethan 30 minutes. Samples ranging from500 ml to 21iters may be filtered Withone membrane, depending on degree ofturbidity. After filtration the membranesare placed M the walls of separatedbeakers and washed as above. Toprevent the particulates from obscuringthe nematodes, the washing from eachfilter is examinedsin a separate countingchamber.

G 'Direct Microscopic Examination,L,

Each counting chamber containing thefilter concentrate is first examined undera 4X objective. Unless the concentratecontains more than 100 wormso they wholeFell area is surveyed for nematodes, withrespect to number, develbpmental stage,and motility. When an object having anoutline. resembling that of a nematode isobserve it is re; examined under a ipxobjective for anatomical structures, uklessthe object exhibitg typical nematode move-ment, which is sufficient' for identifying theobject as a nematode. When the concentratecontains more than 100 worms, the wormdensity can.be estimated by counting thenumber ofworms in representative Micro-scopic fields and multiplying the averagenumber of worms per field by the numberof fields in the cell'trea. The nematodedensity may be expressed as number ofworms, per gallon with or without differenti-ation as to adult or larval stages or as toviability.

H' General Identificatiotrof Nernt.todes

1 While actively motile nematodes can bereadily recognized by any person whohas some general concept of micro-scopic animals, the nonmotile or

10-6

VIII

sluggishly motile nematodes may beconfused with root fibers, plant fila-ments of various types,' elongatedciliates such as Homalozooji vernii-culare, or segments of appendages_ofsmall crustacea. To facilitate a,geAral identification of nematodes, thegross morphology of three of the free-living nematodes that are frequentlyfound in water supplies is sh9wn in theattached drawing. The drawing providesnot only the gene ral anatomy for recogni-tion of nematodes but also most of theessential structures for guidance to thosewho want to use the "Key to Genera" inchapter No. 15 on Nemata by B. G.Chitwood and M. W. Allen in the book,Fresh Water Biorogy. (10)

2 Under normal conditions, practicallyall nematodes seen in samples offinished water are in various larvalstages and will range from 100 to 500microneiri length and 10 to 40 micronsin width. Except in the fourth (last)stage, the larvae have no sexual organsbut show other structural characteristics.

3 If identification of genera -i's desired,the filter washings are 'centrifuged at500 rpm for a few minutes. Thesupernate id-discarded, excapt a fewdrops, and the sediment is resuspendedin the remaining water. A drop of thefinal suspension is examined under both10X and 43X objectives for anatomicalcharacteristics, without staining, and forsupplementary study of structures therest is fixed in 5% formarin or otherfixation fluid and stained according toinstructions given in Chitwood and

,Chapter 'on Nemata, (7):Goodey's Soil and Freshwater Nema-todes(11) or other bboks on nematology.

USE OF ZOOMICROBES ASPOLLUTION INDEX

A Idea not new, protozoa suggested long ago;many considered impractical because ofthe need of identifying pollution - intolerantand poIlution-tolerant specie's.- proto-zoologist required. Method'also timeconsuming. . .

M

r

B Can use them on a quantitative basisnematodes, and nonpigmentedprotozoa present in small numbers inclean water. Numbers greatly increasedwhen polluted with effluent from aerobictreatment -plant or recovering from sewagepollution; no significanrerror introduced

"when clean-water members included in theenumeration if a suitable method of com-puting the pollution index developed.

C Most practical method involves theequatibn; A + B + 1000 C = Z. P. I. ,

A

whereA = number of pigmented protozoa,B = non pigmented protozoa, andC = nematodes in a unit volume of sample,azd Z, P. I. = zoological pollution index.For relatively clean water, the value ofZ.P.I. close to 1; the larger the valueabove 1, the greater the pollution by aerobic'effluent (see attached report on zoomicrobialindicator of water pollution).

CONTROL

A _-Chlorination of effluent

B Prolongation of detention time of of

C Elimination of slow sand filters innematode control.

LIST OF COMMON ZOOLOGICAL ORGANISMSFOUND IN AEROBIC SEWAGE TREATMENTPROCESS

PROTOZOA

Sarcodina - Amoebae

Amoeba proteus; A radiosa

Hartmanne lla

Amelia VulgarisNoegleria gruberiA ctinophrys

125

Protozoa; Nematodes, and Rotifers

FLAGELLATA

Bodo caudatus

Pleuromonas jaculans

Oikomonas termo

Cercomonas longicauda

Peranema trichophorum

Swimming type

Ciliophora:

Colpidium colpoda

Colpoda cuculus

Glaucoma pyriformisParamecium candatum; P bursaria

,r0

Stalked type

a2rcularia sp. (short stalk dichotomous)

Vorticella sp. (stalk single and contractile)

aistylis plicatilis (like opercularia, morecolonial, stalk not contractile);

Carchesium sp. (like vorticella bat colonial,individual zooids contractile)

foothamnium sp., I entire colony contracts)

Aspicisca costataEuplotes patella

Stylonychia mylitus

UrOstyla sp.

Oxytricha sp.

Crawling type

NEMATODA

Dipluaster sp. Doryhrr...itis sp.

Manochoides sp. Chlindrocorpus sp.

2klorosteroides sp. agtloblis sp.

Rhabditis sp. Rhabditolaimus sp.

Pelodera,sp. NIonhystera.sp.

AphelelichoideS sp. r-..24lolaus sp.

imp*,

10-1

Protozoa,. Nematodes, °s.nd Rotifers

ROTATORIA

21..glena

,

Polarthra

Philodina

Ke rate lla

Brachionus

OLIGOCHAETA (bristle worms)

Aelosomh hempachi..

,Aulophonts limosa

Tubifex tubifex

Lumbricillus lineatus17=11D 111 all.aDiISEC LARVAE

Chironomus

3 Chang, S. L. Interactions between AnimalViruses and Higher Forins of Microbes.Proc. Am. Soc. Civ. Eng. J1. San. Eng.96:151. 1970

4 Chang, S. L. Zoomicrobial Indicators° of Water Pollution presented at the ---

Annual Meeting of Am. Soc. Microbial,Philadelphia, April 23 -28, 1972.

5 Chang, S. L. Pathogenic Free-LivingAmoebae and Recreational Waters.Presented at 6th International Confer-

, en%e of Water Pollution ResearchAssociation, Jerusalem, Israel,.June 19-24, 1972.

> 6 Chang, S. L. Proposed Method forExamination of Water for Free-LivingNematodes. 5.A.W.W.A. 52:695-698.1960.

7 Chang, S. L., et al. Survival and ProtectionAgainst Chlorination of Human EntericPathogens in Free-LivingNematodesIsolated from Water Supplies. Am. Jour.

e Trop. Med. and Hyg. 9:136-142. 1960.

8° Chang, S. L. Growth of Small Free-LiiiingPsychoda sp. (trickling filter fly) Amoebae in Bacterial and Bacteria-FreeARTHRoPODA Cultiires 4. Can. J. Microbial.

6:397-40 s'1960. ''f-,Lessertia sp

9 ChaneSs. L. and fabler, /1. W. tree- LivingNematodes in Aerobic Aeatment PlantPorrhomma sp. Effluents. J.W.P.O.F. 34:1256-2161. 4

Achoratua subuiata s ('collembola). 1963.

10 Chitwood, B. G. and Chitwood, M. B. AnFolsomia sp. (collerhhola) Introduction to'Nernatolozz. Section IAnatomy: 1st e71. Monumental PrintingTomocerus sp. (collembola) Co. Baltimore. 1950. pp 8-9.

11 Cobb, N.A. Contributions to the Science ofNematsology VIt. "Williams and Wilkins Co.Baltimore. 1918.

12 Collins, V. G. The Distribution and Ecologyof Bacteria in Freshwater, Pts. I &

REFERENCES

1 American Public Health Association, '4merican Water Work's Association andWater Pollution Control Federation.Standard Methods for the Examinationof Water a Warftewater, 13th ed.New York. 1971.

2 Chang, S. L., et al. SurveY of Free-,Living Nematodes and Amoeba inMuicipal Supplies. J. A. W. W. A.. 52:613 -618. .

Proc: Soc. for Water Treatment and Exam.12:40-73. 1963: (England)

13 Edmondson, W. T., et al.. Ward-Whipple'sFresh Water Biolm. 2nd ed. JohnWiley & Sons, New York. 1959. pp 368-401.


14 Goodey, T. Soil \and FreshwaterNematodes.' (A Monograph) 1st ed.Methuen and Co. Ltd.' London. 1951.

9

tI

This outline wa prepared by S. L. Chang,Chief, Etiology, Criteria DevelopmentBranch, Water Supply Research Laboratory,NERC, USEPA, Cincinnati, Ohio 45268.

Descriptors: Protozoa, Ne atodes, Rotifers

k

127

:%"=.!

.1

109

AV

Piotozoa, Nematodes, and tifers

Effluent

;1 TR RRFILTERS

AERATIONTANKS

TIT FILTERS

j

InsectsA

OligochaeteS-&insect larvae-

Nematodes& rotifers

Nonpimentedprotozoa

I It AHeterotrophic

bacteria

1-7-4- Raw Sewage

\.\

Suspe) oed ank matter

Fungi

Algae

(by h crolysis)

01=11

DisSolved organic matter

(respiration,de amination,decarboxylation, etc.)

Inorganic C, P, N,S comp.

Autott'ophic bacteria

Pathogenic organiSms

(Nitrification, sulfur& iron bacteria)

Food Chain in Aerobic.Sewage Treatnient Processes

The same sequences and processes occurin polluted streams and streami'receivingtreated wastes.

1 28

ACTIVATED SLUDGEPROTOZOA.

.IMO

Larggr animals (worms, snails, fly larvae, 1

etc. ) dominate trickling filters. Why arethese always absent from the activatedsludge process?

Why are there numerous micro- speciescommon to both trickling filters andactivated sludge?

"2

What Organisms besides protozoans andanimals are present in activated sludge?

3

What is (he advantage(s) of microscopicexamination of activated sludge?

4

One sampling site would be the one of choice 5 .in sampling Ni activated sludge plant formicroscopic analysis. Why? Where?

What organisms predominate in activated 6

sludge?a 4.

Why are photosynthetic green plants (incontrast to animals) basically absent fromthe activated sludge process in general andmixed liquor specifically?

7

hy are the same identical species of protozoa 8

f d in activated sludge plants all over theworld?

What is, the significance of a microscopic 9

examination of mixed..)liquor?

Define and characterize: 10

Activated Sludge.Mixed LiquorFlocs

What is the relation between bacterial) .

protozoan populations in activated sludgeand the proces,s itself?

11

At what total magnificatiori wereyou able to 12believe the smallest cells observed.w.ere infact bacteria?

Activated sludge is a dynamic (although 13

man-manipulated) ecosystem. How-does,it differ from a natural ecosystem?

Activated-Sludge PiOtozoa

What is the greatest problem(s) with a wetmount slide preparation? . I

How do you overcome these disadvantages?

How do yoU slow down fast moving protozoanson a wet mount?

Why are quantitative counts of protozoa (likenumber/ ml) generally meaningless?

What' is the significance of proportionalcounts?

Scanning a slide (in making a count) shouldgenerally be done at X. (Totalmagnification) .

The iris diaphrUm on thenlicroscope isused to adjust light intensity (true-false).

Why sample the surface film of thesettleometer?

Why is the thinnest film most ideal for awet mount?

What did you learn from the microscopicexamination' of the activated sludge?

What is the physical nature of the flocsobsakved?

What filamentous organisms wereobserved?

Why are "rare" species of no practicalsignificance in microscop analyses ofactivated sludge?

Why are there no protozoan indicator speciesof process efficiency in activatedillludee?

Activated sludge-biological communitiesare temporal in contrast to biological.communities in trickling filters whichare spatial-(TRUE/FALSE).

"Seeding" a newly started activated sludgeplant with cultures or material from otherplants is only a. wasted effort (TRUE/FALSE).Justify,y.our answer. .

e

14

a

C-

-AO

15

16

17

18

19

20

21-

2263

239

24

25°

26

o

27

.0 4

28 oak

29

13

O

co

4-. .

.1 .

if a wet mount slide` of mixed liquor is 30prepared and placed in 'a petri dish with awet blotter underneath and allowed to sitfor sgveralhours, what will be the distri-bution of the protozoa under the cover slip?

ti

Activated Sludge Protozoa

In a mixed liquor sample nearly all of the 31stalke,d ciliates have""broken off. the stalksand are frerswimmineas utelotrcichs."What does'this indicate? tto

What are Monags? And are they good, tad, 32

4or indifferent in activated sludge?

What are hypotrichs or crawling ciliates, 33and are they good, bad or indifferent iiactivated sludge? A %. Q

What are swimming ciliates, and are they 34 _

goodbad or indifferent in activated sludge?

What are flagellAtes, and are they good, bad '35 .

or indifferent in activated sludge?o

What are amoebae, and are they good, bad, 36

A or indifferent in activated sludge?

What are the ideal characteristics of a wet 't 37mount slide preparation?

Why does total community givea- betfbr 38indication of process efficiency in activated ..sludge?

O

In observing and identifying protozoa one looks 39for what, characteristics of an individualorganism?

What is the role of bacteAa in activated a ;10

sludge?

Vihatsis the role or,protozoa in activated 41;sludge?

Microscopic analysis ofAmbeed liquor 42 ot, 4&)

.sample can be very quick, simple, and -.

meaningful (TRUE /FALSE). .

11-3

0

Activated Sludge Protozoa f

Protozoan communities present in activated 43'sludge reveal:

a. Plant efficiencyb. Settlfabil4c. ROD removald. Solids removalei.; Plant ling

..(Circle applicable .descriptions)

o

Pr.otozob.n-communities in activated sludge 44c. °reveal complete-and instantaneous to,nditions;

average of physical and_chemical-conditions;-egtremes of chemical and physical conditions.(Draw a line through phrases true.)

Rank in increasing plant efficiency the 45°following ptotoioan group which would pre-,dominate.

RotifersStalked ciliates ,AmoebaeSwimming ciliatt s

9 Crawling ciliateslagellates

(For exa ple, use number 1-6. One would bestartup co ditiops or least efficient, and sixwould be t e.most efficient.) ..Identification is usually done at X and 46sometimes requires X.

Immersion oil should be used sparingly at 47what two points on a slide?

'''''''4:9 Which comes first in microscopic examina- 48lion; s anning at low power to pick out.unkno ns or higher power to identify ?,

°

1<

In making proportional count, which total. 49number to count would be better; total of ten

_organisms or a total of 100 organisms? Why?

Why not kill the organisms so you canidentify and count them'on the slide?

50

,What simple chemical solutions are usefulto immobiliie protozoa if methyl celluloseor polyvinyl alcohol is hot-available?'

51

t4

Activated Sludge Protozoa

Initially the° wet mount slide should be 52racked up close to the low power objective:by your eye on the eyepiece through thescope; or by glancing at the actual distance .

with the naked eye While you rotate thecoarse adjustment knob. (Underline which)

What are par -focal objectives on the 53microscope? ,

Why should water on the microscope and all 54its parts be carefUlly avoided?

lj

If activated sludge is a mah manipulaiied 55system, 6re there comparable,na.tuPal,ecosystems? Exainple?

.What is the " community '; concept in exami- 56nation of aptivated sludge?

What a're the appliCations of direct micro- 57scopist examination of activated sludge?

What are rotifers, and are they good, bad - 58* or indifferent in activated sludge? . o*.

List the five kingdomS of organismsnd give 59a specific example for each.

What techniques are most useful in 60identifying an unknown Organism, and whyis -correct identification Important?

Scanning and counting is done at * X 61magnification. Identification of mostPROTOZOA usually requires . X-magnification and occassionally X

- magnification. -5' ,

OBJ. TOTAL MAG. USE

43.5 X

10 X

40 X ".

boo° x

(1,0 X eyepiece's)

62

I.

11-5

hand is constantly operating the 63

hand issconstantly, operating the

The microscope is manually operated andrequires skill and understanding on the partof the operator. A microscope, no matterhow costly is only as good as the micro-scopist operating it.

Activated Sladge Protozoa

List the basic skills required in utilizing the 64 .7optimum capabNty of your microscope.

NÀ h,

, _0

This outlinerv/as prepared Isr,R. M. Sinclair,.fiational Training tenter, MOTD, OWPO,USEPA, Cincinnati, Ohio 45268.

4

Descriptors: Zlicroorganisms, Protozoa;.... .Rotifers Activated Sludge. Riota

C

.

*I

.? 1,

". 3

13401

A

ea.

FREE- LIVING AMOEBAE AND -NEMATODES

I FREE - LIVING AMOEBAE

Importance of Recognizing Small,Ftee- Living Amoebae in WaterSupplies

1 Commonly found in soil, aerobic is

sewage effluent and natural, freshwaters - hence,- frequently en-countered in examination of rawwatek. '

stenot infrequently found inwits]. al supplies - not pathogenarrie- s.

1

\ 1:-.lage 41,e-a,moebae Naeolerla

involved-140 some cases ofmeningoencephalitis, about half .in the U:S.; associated withswimming in small warm lakes.Acanthamoeba rhisodes parasitizinghyman throats and causing (3 cases)nonswimming-associated meningo-encephalitis. .

4 Cysts not to be confused with thoseof Endamoeba histoljtica in water -borne. epidemics.

.81

B Classification of Small, Free-LivingAmoebae

1 Recognized classification basedon characteristics in mitosis.

2 k Common species fall into thefollowing families and genera:

Family Schizopyrenidae: ,Genera_

Naegleriaj Didascalusi andSchizopyrenu.s.- first-two beingflagellate amoebae.

Family Hartmannellidae: GeneraHartmanella (Acanthamoeba)

3 H9w to prepare material's forstudying mitosis Feulgen stain

BI. AQ."14b. 6. 76 I

C Morphological Characteristics ofSmall, Free-Living Amoebae

.1 Morphology of Trophozoites'-Ectoplasm and endoplasm usually,dIst ot; mar:dens-with large nucleolus.

2 Morphology of cysts - Single ordouble wall with or without pores

D Cultural Characte ristics of Small;Free - Living- Amoebae

1 How to cultivate these amoebae -.plates with bacteria; cell cultures,axenre culture.

2 Growth characteristics on plate,cell, and axenic culture

,3 Complex growth requirementsfor most of these amoebae

E Resistance of Amoebic Cysts toPhysical andChemical Agents

II FREE- LIVING NEMATODES

A Classification of Those CommonlyFound in Water Supplies

42>

1 Phasmidia (Secerneutes):Genera Rhabditis, piploga.ster,Dipl.oacqeroidesi CheilolmsPanalalgromus

Aphasmidia (Adenophoro): GeneramoL2-il st1., Aehelenchusr. Turbatibc..(vinegar eel), l2c2D22.imusj andRhabdolaimus ,

B "MorphOlogical F4tures

1 Phasnaids: papilla on tail 'at males,mouth 'adapted to feed on bacteria,few exceptions.

2 Aphasmids: no papilla on male ts.- -

glandular cells in male.

2-1

V

'!$Amoebae aid IsTematodes in Water Ues

4C Life Cycle

1 MetIods Of mating

2 Stages of developthent

3 Parthenogenesis

b Cultivation

1 Bacteria -fed, cultures

2 Axenic' cultures,,>;

E Occurren-ie in Water Supplies

1. Relationship between their_.---appearance in finished,waerand that in raw water.

S'

2 r Frequency of occurrence in f

diffetent types of'raw water,,,and sources.

3 Survival of human enteric path-ogenic bac't'eria and viruses in

'- nematodes.

4 Protection of human entericpathogenic bacteria and virusesin lematode-carriers.

F Control.

1 Chlorination of sitwa.ge--eltu-entA

Floqdulation and sedimentationof A4ater

3 Chlorination of water

4 Other methods of destruction

REFERENCES

Amoebae

1_ Sinkh, B. N. , "Nuclear Division in NineSpecies of Small, Free-Living Amoe-bae and. its Bearing on the Classifica-tion of the -Order Amoebida",Trans; RcrialSoc. London; Series B,-236:405-461;, 1952. ,

'11

(2.

we, 4.

2 Chang, S. L., et al. IsSurvey of Free -Living Nematddes and Amoebas' in

' Municipal Supplies". J. A. W . W. A.52.613-618, 1960. ,

3 Chang, S. L., "Growth of Small Free-Living Amoebae in Various Bacterialand in Bacteria-Free Cultures".., Can,Jour. -.Microbiol:. 6 :397 -405, 1960.

Nematodes

1 Goodey, T., "Freshwater Nematodes",1st. Edition, Methuen & Co., LondyA,1951.

2 Edmondson, W. T. ,. Ed., Ward & Whipple' s'''Fresh-Water Biology" 10th Edition,page 397, 1955.

3 Chang, S. L., et al. , "Occurrence of a'Nematode Worm in a City-Water Supply ".J.A.-W.W;A., 51:671:676, 1959.

4 Chang, S. L., et al., "Suzvival, 3-11AProtection Against Chlorination, of ;HumanEnteric Pathogens in Free- '

Living Nematodes Isolated From. WaterSupplies". Am. Jour. Trop. Medicine& Hygiene, 9:136-142, 1960.--,

5 Chang, S. L., et al., "Survey of Free-Living Nematodes and Amoebas inMunicipal Supplies". J. A. W. W. A. ,52:613-618, 1980.

6 Chang, Si L. , "Proposed Method forExa-nination'of Water for Free-LivingNematodes". J. A W. W. A. , 52:695-638,-1950->7 -4*

Ch, g, S. L.; "Viruses, Amoebas,a d Nematodes and Public WaterS pplies". J.A9rW.W.A., 53 :288 -296, ,1961.

8 Chang, \S. L. and Kabler, P. W., "Free4Living Nematddes 4n Sewage Effluentf tom erobic Treatment Plants": To

ub shed. -

This outline as prepared by ShT,ii L7Charig,M. D: , Chief, iology, Criteria Lievelopment.Branch, Water Supply Research Laboratory,

_NERC, EPA, C cinnati. OH 45268.

Descriptons; Amebae, Nematodes

1 3

Subphylum:

SUGGESTED, CLASSIFICATION OF SMALL AMOEBAE

Sarcodina Hertwig and Lesser

ss: ,Rhizopoda von Siebold

Subclass: Amoebaea Butschli

'Order: Amoebida Calkins and Ehrenberg to

Superfamiky: Amoebaceae - free-living(Endarooebaceae parasitic in animals)

Family: Schizopyrenidae - active limax form common; transcient° flagellates present or absent; nucleonus-origin of

polar masses; polar caps and interzonal bodip-presentabsent

Genus: Schizopyrenus - no transcient flagellates; single-walledcysts; no polar caps or interzonal bodies in mitosis

Species: S. erzthaenu,3a - reddish orange pigment formed In agarcultures with gram-negative bacillary bacteria

S. russelli - no pigment produced in agarcultures

Genus: Didascalus - morphology and cytology similar toSchizopresmsbuFsmall 'numbers' of transcient flagellates fdrmed at times

Species: D. thorntoni - only species described by Singh (1952)

Genus: %aerie. Alexeieff - double-walled cysts; transcientfla,gellatcs formed readily; polar caps and interzonalbodies present in mitosis

Species: N. gr.utieri (Schardin-ger) - only species established;Singh (1952) disclaimed the N. soli he described in 1,951

Family: _Hartmiannellidae - no trans dent flagellate formed; motilitysluggish; no limax form; nucleolus disappearing, probably

Vh., forming spindle in mitosis; no polar caps or masses, aster,and centrosome not known

Genus: Hartmannella - ectoplasm clear or less granular than. endoplasm; single-walled cysti3; single vacuole

Species: H. fasbae - clear ectoplasm,agricola.- ectoplasm less granular than endoplasm

Genus: Acanthamoeba - filamentous processes from ecto- orendoplasm; growing axenically in fluid bacteriological

a/Arne dta

137 12-3'

I

Su p, estecassification of Small Amoebae ..

Air

. , Species: A. rhLsodea, .

Genus: Sinehella -,double-walled cysts; ecto- and endoplasmindistinguishable; many vacuoles

Species: Singilella latocherrius

x

/

* ,...tr,

i

C5

I.

0s

0

t

. ...

61> /z,

. .. :

13L

.-'

4

ANIMAL PLANKTON

,I INTRODUCTION

A Planktonic animals or zooplankton arefound in nearly every major group ofanimals.

1 Truly pla:hktonic species (euplankton)spend all or most of their active life

,Cycle- suspended in the ;water. Threegroups are predominantly involved in

. fresh water; the protozoa, rotifers,"and microcru.stotea.

2 Transient planktonic phases such asfloating eggs and cysts, and larvalstages occur in many Other groups..

B Many forms are strictly seasonal inoccurrence.

C Certain rare forms occur in great numbersat unpredictable intervals.

D Techniques of collection, prese'rvation,and identification strongly influence the

, species reported.

E In oceanographic work, the zoop ?Acton isconsidered to include many relath.vely largeanimals such as siphonophores, ctenoplores,phepteropods, pteropods, arrowworms, andetiphausid shrimp,

F The plant-like or phytoplankton on theother hand are essentially similar in allwaters, and are the nutritional foundationfor the animal community..

II PHYLUM PROTOZOA

A The three typically free living classes,Mastigophora, Rhizopoda, and Ciliophora,all have planktonic representatives. Asa group however, the majority of the phylumis benthic or bottom-loving:, Nearly anyofothe,benthic forms may occasionally bew.ashe'd up into the overlying waters andthus be collected along with the euplankton.

B Class mastigophora, the nonpigmentedzooflagellates.

These have frequently been confused withthe phytomastigina or plant-like flagellates.The distinction-is made here on the basisof the presence or absence of chlorophyllas puggested.by Earner 'and Ingram 1955.-

139.

(Note Figure: Nonpigmehted, Non - OxygenProducing Protozoan Flagellates in theoutline Oxygen Relationships.)

1 Commonly encountered genera

Bodo

.Peranema

2 Frequently associated with eutrophicconditions

C Class Rhizopoda amoebOi.dprotozoans

1 Forms commonly encountered asplankton:

Chaos

Arceila

Difflusia

Euglyphe.

fi4noeba)

Centropyxis77Fieliozoa

2 Cysts of some types may be encounteredin water plants bedistributiori systems;rarely in plahkton of open lakes orreservoirs.

D Class Ciliophora

1 Certain "attached" forms often roundfloating freely with plankton:

Vorticella

2 Naked, unattached ciliates, Halteriaone of commonest in this grouirTa7Rousheavily ciliated forms (holotrichs) maYoccur from time to time ailch asColpidium, Enchel, etc.

3 Ciliates protected by a shell or test(testaceous) are most often recordedfrom preserved samples. Particularlycommon inthe experience of the National'Water Quality Sampling Network are:

Codonella fluviatile

Codonella cratera

a

Tintinnidium (usually with organic matter)'4..

TintinnoRgis

13 -1

Animal PlanktA

III PHYLUM ROTIFERA,,

.i/N Some forms such as .Anuraea cochlearisand Asplanchna pridorat all tames of the year. Others such aslc,Totholca striate, N. Ion spina and Poly-arthra platypTera are repor e to be essen-tially winter torms..

B Species in approximate order of descendingfrequency currently recorded by NationalWater. Quality Sampling Network are:

Keratella cochlearis

Polyarthra vulgaris

Synchaeta pectinate

Brachionus quadridentata

Trichocerca longiseta

Rotariesp.

Filinia longiseta.

Kellicottia longispina

Pompholp sp. a

C Benthic species almost without number maybe collected witb4e plankton from time totime.

IV PHYLUM ARTHROPOD.A

A Claes Crustacea

1 The Class Crustacea includes the largercommon freshwater euplankton. Theyare also the greatest planktonic consum-ers of basic nutrients in the form ofphytoplankton, and are themselves thegreatest planktonic contribution to thefood of fishes. Most'of them are herb-ivorous. Two. groups, the cladoceraand the copepods are most'conspicuous.

2 Cladocera,(Subclass Branchiopoda,Order Cladocerarar Water Fleas

a Life History

1) During most of the year, 'eggswhich will develop without fertil-ization (parthenogenetic) aredeposited by the female in a dorsalbrood chamber. Here they hatchir}to minature adults which escapeand mini away. ,

4

2) As unfavorable conditions develop,males- appear, -and- thick walledsexual eggs are enclosed in egg'cases called ephippia which canoften endure freezing and drying.

3) Sexual reproduction may occurat different seasons in differentspecies.

4) Individuals of a great range of *sizes, and even ephippia, arethus encountered in the plankton,but there is no "Larval" form.

b Seasonal variation - Considerablevariation may occur between winterand'summer forms of the samespecies in some cases. Similarvariation also occurs between arcticand tropical. situations.

c Forms commonly encountered asopen water plankton include:

Bosmina longkostrisCand others

Daphnia galeata ai.14.-erthers

Other less common genera are:

Diaphanosoma, Chydorus, Sida,7croperus, Creriodaphnia, Byt 'ho-trephes, analTeEataVorousreYEZ'ara and Polyphemus.

d :Heavy blooms of Cladocerans maybuild up in eutrophic waters.

3 The copepods (Order Copepoda) are theperennial microc'ustacea of open waters,both fresh and marine. They are themost ubiquitous of animal plankton.

a Cyclomis the genus Most oftenrdiirarby the National Water QualitySampling Network activities. Eucr.clops, Paracyclops, Diaptomue,UrriffiocaraptirCiiiiiscTirra, anLimnocIlanirrare other formsFFIWe7-1-5.-We planktonic.

b Copepods 'hatch into a minute char-acteristic larvae called a nailiuswhich, differs considefably from theadults. After five r six moults, thecopepodid stage is j'eached, and after.-six more moults, the adult. Theselarval stages are often encounteredarfd are difficult to identify.

I-

i

Insecta1 Only a few kinds of insect that can be

ranked as a truly planktonic. Mainly. the phantom midge fly Chaborus:

2 The larva of this insect has hydrostaticorgans that enable it to remain suspendcdin the water, or make vertical asaens ii:thewate column.

3 It is usually benthic .during the daytime,but ascends to the surface at night.

V. OCCASIONAL VLANKTERS

A While the protozoa, rotifers, and micro-crustacea make up the bulk of the plankton,there are many other groups as mentionedabove that may also occur. Locally orperiodically these may be of major impoit-

- ance. Examples are given below. 4.

B Phylum Coelenterate

T Polts of the genus Hydra may becomedetached and float aHourhadiging fromthe surface film or floating detritus.

. 2 The freshwater medusa Craspedacustaoccasionally appears in Tale% or reser-voirs in great numbers.

C Phylum Platyhelminthes

1 Minute Turbellaria (relatives of thewell known Planaria) are sometimestaken with tbe 'plankton in eutrophic--conditions. They are often confusedwith ciliate protozoa.

2 Cercaria larvae of Trematodes (flukes)parasitic on certain wild animals,frequently appear in great numbers.When trapped in the droplets of wateron a swimmer's skin,' they attempt tobore in. Man not being their naturalhost, they fail. The. resultant irritationis called "swimmer's itch". Some canberidentified, but many unidentifiablespecies may be found.

3 In many areas of the world, _cerparialarvae of -human parasites such as theblood fliAce-Schistosorna japonicum mayJive as plan orkr7-'1-fT-"IpEngfiirf. Fia-humahskin,directly on contact.

141tr

Animal Plankton

yluirt-Nymathelminthes

1 N atodeslbr nemas) or roundwormsappr h the bacteria and the blue-greenalgae in, iquity. Thevare found inthe soil and in tke water, and in the airas dust. 'In both marine and fresh watersand from,the Arctic tb the tropics.

2 Although the majority are free living,some occur as parasites of plants,animals, and Man, and some of these

. parasites are among our most serious.

3 With this dfstribution, it is obvious thatthey. will occasionallz be encountered as'plankton. A more complete discussionof nematodes and their public healthimplications in water supplies will befound elsewhere (Chang, S. L. ).

E Additions.' 1 crustacean groups ,sporadicallymet with in the plankton include the folloWing:

1 Or er Anostraca or fairy shrimps, merly included with the two

f lowing orders in the Euphyllopoda)primarily planktonic in nature.

a Extremely local and sporadic, butwhen present, may be dominatihg.

b Artemia, the brine shrimp,cantolerate very high salinitie.

c Iiery widely distributed, poorlyunderstood., -

2 Order Notostraca, the tadpole shrimps.Essentially southern and western indistribution.

3 Order Conchostraca, the clam shrimps:Widely distributed, sporadic in occur-vrence.' Many local species.

4 Subclass Ostracoda, the seed shrimps.Up to 3 in. in length. Essentiallybenthic but certain species of C ris,and Notodromas may occur in consi -erabTerniribiTs as plankton at certaintimes of the year.

5 Certain members of the large subclassMalacostraca are limnetic, and thus,planktonic to some extent.

a The scuds, (odder Amphipoda) areessentially benthic but are sometimescollected in plankton samples around

13-3;0

'Animal Plankton

weed beds or near shore. Nekto-planktonic forms include Pontoporeiaand some species of Gammarus.

b The mysid, or opossum-shrimps arerepresented among the plankton byMysis-relicta, which occurs in thedeeper waters,, large lakes, as farnorth as the Arctic Ocean. '

_ .

F The Class Archnoidea, Order Hydracarina(or A cari) the mites. Frequent in planktontows near shore although Unionicola crass-ipes has been reported to-be yirtuallyplanktonic.

G The phylum Mollusca is but scantilyrepresented in the freshwater plankton,in contrast to the marine situation.Glochidia (ciliated) larvae are occasion-ally collected, and snails now and then .

glide out bn a.,quiel surface film and aretaken in a plankton net. An exoticbivalve Corbicula heave planktonicveliger stage.

H Eggs and other reproductive structuresof many forms including fish, insects, androtifers may tpe found in plankton samples.Special-reproductive structures such asthe statoblasts of bryozoa and sponges,and the ephippia of cla.doCerans-may alsobe ineltided.

I Adveptitious and Accidental Plankters°

Many shallow ,water benthic organismsmay become accidentally and temporarilyincorporated into the plankton. Many ofthose in the preceding section might belisted here, in addition to such forms ascertain free living nematodes' smalloligochaetes, and tardigrades,. Collembolaand other surface film livers are also,taken at times but should not be ,Mistaken-for plankton. Fragments and molt. skins' ,from a variety of arthrOpods are usuallyobserved.

Pollen from terrestrial or aquatic plantsis often unrecognized, or confused witone, of the above. Leaf hairs fromtexrestrial plants are also confusing to

the uninitiated, they are sometimesmistaken for fungi or other organisms(and vice versa).

In flowing waters, normally, benthic(bottom living) organisms are often found 41kPdrifting freely in the stream. Thisphenomenon may be constant or periodic.When included in plankton collections,theyamust be reported, but recognizedfor what they are.

S,uria..t. films are especially rich inmicro "biological garbage" and theseenrich the plankton.

REFERENCES

1 Edmondson, W. F., ed. Ward andWhipples'aFreshwater Biology, 2ndEdition, Wiley & Sons) Inc., New York.1959.- ,

2 Hutchinson, G. Evelyn. A Treatise onLimnology. Vol. 2. Introduction to

. Lake Biology and the Limnopla.nkton.Wiley. 1115 pp. 1967.

3 Lackey, J. B. Quality and Quant y of. Plankton in the South End of ke

Michigan in 1952. JAWW36:669-74. 1944.

4 McGauhey, P. H. , Eich, H. F. , Jackson,H. W., and Henderson, C. A Studyof the Stream Pollution Problem in theRoanoke, Virginia, Metropolitan ,

District. Virginia Polytech.Engr. Expt. Sta.

5 Needham, J. G. and Lloyd, J, T. TheLife of Inland Waters. Ithaca, NewYork, Comstock Publishing Co., Inc.

j 1937..

6 Newell, G.E. and Newell, R. C.Marine Plankton. Ilutchinson duc.Ltd. London. 221 pp. 1963.

7 Palmer, C. M. and Irlgram, W M.Suggested Classification of Algae andProtozoa in Sanitary Science.Sew; &''Ind. Wastes. 27:1183-88.1955. ," ,

142

Animal Plankton

8 Pennak, R. W. Freshwater Invertebratesof the United States. The Ronald Press,New York. 1953.

9 Sverdrup, IL W. , Johnson, M. W. , andFleming, R. H. The Oceans, TheirPhysics, Chemistry and General,Biology. Prentice-Hall, Inc., New York.

1 1942.

1

10 Welch, P. S. Limnology, McGraw-HillBook C'o., Inc., New York. 1935.

This outline was prepared by-H. W. Jackson,former Chief Biologist, National TrainingCenter, and revised by R. M. Sinclair,Aquatic Biologist, National Training CenterMOTD. OWPO, ,USEPA,_ Cincinnati, Ohio45268.

Descriptor: Zooplankton

A

re

t

6

Anim1 Plankton

3/4 -PhylumPROTOZOL

Free Living Representative.'

I. Flagellated Protozoa, -C sae Mistigophora

Antholihysis.

Pollution tollerant6,u

Pollution tollerant19 A..

II. Amsboid Protozoa; Class Saroodina

LColony of PoteriodeidronPollution tollerant, 35jz

.t;$

to

DimastizamoehaPollution tollerant

1067,50)u

tiuolearia,reportedto be intollerant ofpollution,-451u.

_III. Ciyied-Protozoa, Class'Ciliophora

I

DiffluxiaPollution tollerant

601563.14

Coliod;Pollution tollerant

20-120)4

6/'

Nolovhrva,reportodto'be intollerant ofpollution, 35,E

4.

Aaitvlis, pollutiontollerant. Colonies often,

maprosoopio.

H.W.Jaokson

Ahimal Plankton

PLA NKTONIC PROTO ZOA

Peranema trichophorum

,

A r cellavu garis

."11.%:..

J, A

VorticeilaGodonellacratera

A ctinosphad r him

_Tintnidium

fluviatile

(.,

;

Animal Plankton

a

$4,

a 11.4'

a

*

PLANKTONIC ROTIFERS

.'Various Forn of Keratella cechlearis

Synchaetapectknata

b sPolyarthrAvulgaris

Brachionusquadridentata I

Rotaria sp0

4

Animal Plankton

4

1

SOME PLANKTONIC CRUSTACEANS

, 14,

Copepod; Cyclops, Order Copepoda

2=.3 mm .

CRUSTACEA

_ A NatiPtiis larva of a Copepod

1-5 mm

Water Flea;-At

Daphnia2-3 mm'

OSTRA CODE

Order Cladocera

Left: Shell closed Right: Appendages extended

1-2 mm

147

a

st

- Aninial Plankton

O

10

PLANKTONIC 4FiTHR OP ODA

A mysid shrimp - crustacean.

A water mite - arachnid

Chaoborus midge larva - Insect174,"

...

a

S.

/- Aspects in the life cycle of the human tapeworm,Diphyllobothrium Tatum, Class Cestoda. Aintestine; B. procercoid larva in copepod;larva in flesh of pickerel (X-ray view).

. 9

. adult as in human /C. plerocercoid

143

H. W. Jackson

D

PREPARATION AND ENUMERATION OF PLANKTON IN THE LABORATORY

Ì RECEPTION AND PREPARATION OF,SAMPLES

A Preliminary-sampling arid analysis is an .

essential preliminarS, to the establish-ment of a permanent or- semi-permanent'

'program.

B Concentration or sedimentation of pre-served samples 'may precede analysis.

1 Batch centrifuge

2 Continuous centrifuge

3 Sedimentation

C Unpreserved (living) samples should beanalyzed at once or refrigerated fofuture analysis.

II PREPARATION OF MERTHIOLATEPRESERVATIVE

A The Vater Pollution Surveillance Systemof the FWPCA haS developed a, modifiedmerthiolate preservative. (Williams,1967) Sufficient stook to "make an approx-imately 3. 5% solution in the bottle whenfilled is placed in the sample bottle inthe laboratory. `The bottle is then_ filled'witli water in the \isteld and returned tothe laboratory for nalysis,.

B Preparation of Mertbiolare Preservative

Merthiolate is availablelfrom manychemical laborator supply houses;one should specify he water 'solublesodium salt.

9

The amount of sodium borate andmerthiolate may be vAried slightly toadjust to different waiters, climates,ana organic, contents.'

3 "Prepae a saturated aqueous. Lugol''Ssolution- as follows:

a Add 60 grams of potassium iodide(IP) and 40 grams of iodine crystalsto 1 liter of distilled water.

, 4 Prepare the preservative solution by1adding approximately 1. 0 ml of theLugol's solution to 1 liter of merthi-.olate stock.

III SAMPLE ANALYSIS

A Microsc'opic examination is most frequentlyemployed in the laboratory to determine 7

what plankton organisms are-present and -NI_

bow many there .are:

2 Merthiolate stock: d, -solve approxi--mately 1.5 dram of sodium borate(borax atd approximat ly 1 gram ofmerthiolate in 1 liter o distilled water.

MIC. enu. 15f. 6.76

1-Optical equipment need not be elaboratebut should include.

a Compound microscope with thefollowing equipment: #

1) Mechanical stage2)- Ocular: 10X, with :Whipple type

counting eyepiece or reticuleObjectives:

approx. 10X( 16mm)aaprox, 20X(8 mm).approx. 40X(4 mm)

.

approx. 95X( 1. 8 mm)( optional)

3)

A 40X objective with a workingdistance of 12,8 mm and an erect .image may be obtained as specialeqUipment. A water, immersionobjective (in addition to oil) .mightbe considered for use with watermounts. k

S

Binocular eyepieces are 'optional.

14-4_

L

40Preparation and Enumeration of Planktonr

-At,

0

Stage micrometer (this may beborrowed, if necessary, as it isusually used only once, when the-equipment .is calibrated) .

.

Inverted microscopes offer certain ,

advantages but a r efoot widelyavailable. The same is true ofsome of the newer optical systems,such as phase contrast microscopy.These are often excellent but ex-pensive for routine plant use.

2 Precision made counting chambers 4--

are required for quantitative -work withliquid mounts.

a Sedgwick- Rafter cells' (hereafterreferred to as S-R 'cell) are usedfor routine counts of medium andlarger forms.

b Extremely small forms or "nanno-plankton" may be counted by use ofthe nannOplankton (or Palmer) cell,a Fisher- Littman cell, a hemacytp-.

meter, the Lackey drop method, orby use of an inverted microscope.

g Previous to starting serious analyticalwork, the microscope should be cali-brated as described elsewhere. Di-mensions of the S-R cell should alsobe checked, -e ecially th depth/

C

Z

3 Direct 'counting of the unconcentrated'sample eliminates manipulation, savestime, and reduces error. If frequencyof organis is low, more area mayneed to be xamined or concentrationof the, samp

4 Conventional techniques emplpyingconcentration of the sample provide moreorganisms for observation, but becausethey involve more m nipulations,

moreadditional error and fake moretime.

may be in ortfe'r.

Several methods of counting plankton arein 'general use.

1 The anumerical or clump count isregarded as tile simplest.

a Every organism observad must be'enumerated. If it cannot be identi-fied, assign a symbol or numberand make a sketch of it on the backof the record sheet.

b Filaments, colonies and otherassociations' Of cells are countedas units., equVr to single isolatedcells. _Their identity as indicatedon the record sheet is the key tothe significance' of such a count.

Individual cell 'count. In this method,every cell of every colony or clump .of organisms is- counted, as well aseach individual single- lled organism.

4 Autom is rticle cou ters may beuseful for c9ccoid organisms,

B Quantitative Plank-Ion Counts

1 All quantitative eounting techniquesinvolve the filling of a standard cellof known dimension's with eitherstraight -sample or a concentrate ordilution .theieof.

2 The organisms in a predeterminednumber. of microscope fields or otherknown area are then observed, and bymeans of a' suitable series of multi-plier factors, projected to a number orquantity per ml gallon, etc.'

14 -2

The_ areal standard unit method rscertain technical advantages, but alsinvolves certaininherrent difficultie

a An areal standard unit is 400 squaremicrons. This is the area of oneof the smallest subdivided squaresin the center of the Shipple eyepieceat a magnification of 100:X.

In .operation, the nurnber f arealrunits of each species is recorded on

the record. sheet rather than'thenumber of indiviiduals. Averagear=eas of the common spe'cies are

b

O

t Preparation And Epuinberation of Plankton

are sometimes printed on recordsheets for a particular plant toobviate the necessity of estimatingthe area .of each cell- observedindividually.

c The advantage of they method lies inthe cognizance taken of the relativemasses of the various species as-indicated by the area presented tothe viewer. These arel.s., however,are often very difficult to estimate.

....4 The ,cubia standard unit method is

logical extension of the areal met od,but ha§ achievedness acceptance.

5 Separate field "count

a In counting separate fields, the-'question always .arisee as to howto count Organisms touching orcrossed by the edge of the Whipplefield. Some 'workers *estimate the

e proportion of, the organism lyinginside the, field as compared to -thatoutside. °nit), those which are overhalf way inside are counted. 'Y./

b Another system is to select twoadjacent sides of the square forreference,. such as the top and leftboundaries. Organisms touchingthese lines in any degree, fromoutsid e or inside, are then counted,while organisms touching the oppositesides fire ignored. It is importantto adopt some such system andadhere to it consistently.

k

c It is suggested that if separate101P 'microscopic fields are examined,,

a standard number of ten be adopted.These should be evenly spaced in tworows* about one-third of the distance .

down from the top and one-third ofthe distance lip frc he bottom ofthe S-R cell.

6 Multiple area count., This is an ex- 'tension of the separate field count.A Considerable increase in accuracyhas recently been shown to accrue byentptying and refining the S-R sell

ti

after each group of fields are countedand making up to 5 additional suchcounts. This may not be practical withhigh counts.

7 The strip count. When a rectangularslide such as the S-R cell is used, a,.strip (or strips) the entire length (orknown portion Thereof) of the cell maybe counted instead of separate isolatedfields. Markinahe bottom of the cell.by evenly, spaced cross lines as ex-plained elsewhere greatly facilitatescounting. .

JJ

a When the count obtained is multi-plied by the ratio of the width ofthe strip counted to the width ofthe cell, the product is the esti-mated number of organisms in thecell, or per ml.

b Whenthe material in the cell isunconcentrated sample water, thiscount- represents the condition ofthe water being evaluated withobtfurther calculation,

8 Survey count. A survey count is anexamination of the entire area of avolumetric cell .using a wide field lowpower microscope. The objective isto locate and record the larger ,forms,especially zooPlankton such as copepor large rotifers whi2h may be presentin size. Special large capacity cellsare often employed for this purpose.For still larger marine forms; numerousspecial devices have been created. .

9 Once a procedure for "concentrationand/or counting is adopted by a plantor other organization it should beused consistently from then on sothat results from year to year can becompared.

rz Differential or qualitative "counts" areessentially lists of the kinds of organisms.found.

Preparation and. Enumeration of Plankton1.

Jr

E It9portional or relative counts of specialgroups are often very us%itilth .For ex-ample, diatoms'. is b alwayscount a standard numbers of cells.

F Plankton are sometimes measured bymeans other than microscopic counts.

1 Settled volume of killed plankton in anIrnhoff-,cone may be 'observed after astandard length of time. This willevaluate primarily only the llsher forms.

2 A gravimetric-method employs dryingat 60° C for 24 hours followed by ep

ashing at 6000 C for 30 minutes. Thisis particularly useful for chemicaland radiochemical analysis.

3 Chemical and phySical evaluation of.plankton pdpulations employing variousinstrumental techniques are coming - tobe widely used. Both biomass andproductivity rates can be measured.Such determinations probably achievetheir greatest utility when coordinatedwith microscopic examination.

4 The membrane (molecular) filter hasa great- potential, but a generallyacceptable technique has yet to beperfected.

a Bacteriological techniques forcoliform determination arewidely accepted.

'b Nematodes and larger organismscan readily be washed off of themembrane after filtration.

c 'els also being used to measureultraplankton that pass treatmentplant operations. -

.

d Membranes can -be cleared andorganisms depbsited thereoni';observed directly, although 4.3accessory staining is desirable.

-C.

e Difficulties include a predilec-tion of extremely fine membranes-to clog rapidly with silt or .

increase in plankton counts, andthe difficulty of making observationson individual cells when theorganisms are piled on top of eachother. It is sometimes necessaryto dilute a sample to obtain suitabledistribution.

IV SUMMARY AND CONCLUSIONS

A

B

The field' sampling program should becarefully planned to evaluate all significantlocation in the reservoir or stream,giving e consideration to the capacityof the la ratory.

Adequate records and notes- should bemade of field conditions and associatedlit h the laboratory analyses in apermanent file.

C Optical equipmentbe calibrated.

in the -laboratory should

D Once a procedure for processing planktonis adopted, it should be used exclusivelyby all workers at the plant..

E Such a procedure should enable the waterplant operator to prevent plankton troublesor at least to anticipate them and havecorrective materials or equipment stockpiled.

REFERENCES

1-

2

3

Ely Lilly Company. Merthiolate as aPreservative. Ely Lilly & Co.Indianapolis 6, Indiana.

Gardiner, A. C! Measurement of ,Phytoplankton PopulationWY-the.Pigment Extraction Method. Jour.Marine Biol. Assoc. 25(4):739-744. 1943.

Goldberg, E. D. , Baker, 1VI. , and Fox, D. L.Microfiltration in OceanographicResearch Sears. Foundation. 'Jour:Mar. Res. 11:194-204. 1952.-

'D

152

Preparation and Enumeration of Plankton

4' Ingram, W. M., and Palmer, C. M.Simplified Procedures for Collecting,Examining, and Recording Planktonin Water. Jour. AWWA; 44(7):617-6'24. 1952.

5 Jacksti, H.W Biological Examination(of plankton) Part III in SimplifiedProcedures for Water Examination.

,AWWA Manual M 12. Am. WaterWorks Assoc. N.Y. 1964.

6 Lund, J. W. G., and Talling, J. F.Botanical Limnological Methods withSpecial References to the AlgaeBotanical Teview. 23(8&9):489-583;October, 1957.

'7 Weber, C. I. The Preservation ofPlankton Grab Samples.- WaterPollution Surveillance Systems,Applications and Develppment ReportNo. 26, USDI, FWPCA, Cincinnati,

° Ohio. 1967,

8 Williams, L. G. Plankton PopulationDynamics. National Water QualityNetwork Supplement 2. U. S. PublicHealth Sy.vice Publ. No. .663. 1962

9 Wohlschag, D.- D., and Hasler, As'. D.Some Quantitative ASpects of AlgalGrowth in Lake Mendota. Ecology.32(4):581-593. 1951

This outline was prepared by W. Jackson,former Chief Biologist, National TrainingCenter, and revised by R. M. Sinclair,Aquatic IJologist, National Training.Center,,MOTD,OWPO, USEPA-, Cincinnati, Ohio 45268

153

Descriptor: Plankton

14 -5

.r.A-T-TACHED GROWTHS(Periphyton or AUfwuchs)

I The cOmmunitS of attached microscopicplants and animals is frequently investigated .durtng water quality studies. The attachedgroWth community (periphyton) and suspendedgrowth community. (plankton) are the principalprimary producers in waterwaysthey con-vert nutrients to organic living materials andstore light originating energy through theprocesses of photosynthesis.. In extensivedeep waters, plankton is probably the pre-dominant primary- producer. In shallow lakes,ponds, and rivers, periphyton is the predominantprimary producer. During the past twodecades, 'investigators of microscopicorganisms have increasingly placed emphasison periph)tic growths because of inherentadvantages over the plankton when interpretingdata from surveys on flowing waters:

A Blum (1956) ". 4 workers are geninallyagreed that no distipctive association ofphytoplanktdn is found in streams, althoughthere is some e% idence of this for individualzooplankters (animals) and for a fewindividual algae and bacteria. Plankton,organisms are often introduced into thecurrent from impoundments, backWater

areas or stagnanf arms of the streain....Rivers whose plankton is not dominated byspecies from upstream lakes or ponds arelikely .to exhibit a majority; of forms whichhave been derived from the stream bottomdirectly and which are thus merelyfacultative or opportunistic plankters, "

i"B "The transitory nature of stream planktonmakes it nearly impossible to ascertatwhich point upstream agents producinglchanges in the algal population wereintroduced, and whether the changesoccurred at the sampling site or at someunknown point upgtream. In contrast,ibottom algae (periphyton) are true com-ponents of the stream biota. Their ,

sessile-attached mode of life subjectsthem to the quality of water continuouslyflowing,,over them. -By.observing thelongitudinal distribution of bottom algaewithin a stream, the sources of the agentsproducing the change can be traced(back- 'tracked)" (Keup, 1966).

BI. IVIIC. enu. 19b. 5.80 154.

II TERMINOLOGY

A Two terms are equally valid and commonlyin use to describe the attached communityof organisms. Periphyton literally ,means"around plants" such as the growths over-growing pond-weeds; through usage thisterm means the attached film of growthsthat rely on substrates as a "place-to-.,grow" within a waterway. The componentsof this growth assemblage consists ofplant's, animals, bacteria, etc. Aufwuchsis an equally acceptable term (probablyoriginally proposed by Seligo (1905)] .

Aufwuchs is a German noun w ithoutequivalent english translation; it isessentiall) a ,collective term equivalentto the above American (Latin root) termPeriphyton. (For convenience, only,PERIPHYTON, with its liberal modernmeaning will be used in this outline,. )

13 Other terms, some rarely encountered inthe literature, that are essentially ^

synonymous with periphyton or describeimportant and dominant components of theperiphytk community are: Ne.eiden,Bewuehs, Laison, Belag, Besatz, attached,sessile, sessile-attached, sedentary,seeded-bn, attached mterials, slimes,slime-growths, and coatings...

The academic community occasionallyemploys terminology based on the natureof the substrates the periphyton grows on(Table 1).

TABLE 1

Periphyton Terminology Basedon Substrate Occupied

Adjectiveepiholitic, nereiditic, sessileepiphyticepizooicepidenditic, epixylonicepilithic

[ After Srarneck-Husek (1946) and is Slacleckuva(1962)j Most above listed latin-root adjectivesarse derivatives of nouns Such as ethihola,epiphyton,

Substratevariousplantsanimalswoodrock

Attached Growths (Periphyton or Aufwuchs)

III Periphyton, as with all other componentso'f the environment, can be sampled quell.-tatively (what is pregent) and quantitatively(how much or many are present).

A Qualitative sampling can be performed bymany methods and may extend from directexamination of the growths attached to asubstrate to unique "cuttings" or scrapings.It may also be a portion of quantitativesampling.

B Quantitative sampling is difficult becauseit is nearly impossible to remove theentire community from a standardized orunit area of substrate.

1 Areas scraped cannot be determinedprecisely enough when the areas apeamorphous plants, rocks or logs thatserve as the principal periphytonsubstrates.

111

ARTIFICIAL SUBSTRATETLACEMENT

A Position or Orientation

1 Horizontal - Includes effects of%'-ettledmaterials.

2 Vertical - Eliminates many effects ofsettled materials.

B Depth (light) - A substrate placed in lightedwaters may not -reflect conditions in awaterway if much of the na ural substrate(bottom) does not receive li receiveslight at reduced intensity. (Both excessivelight and a ,shortage of light can inhibitgrowths and influence the kinds of organismspresent. )

C Current is Important

**2 Collection of the entire community withina standard area usually destroys individualspecimens thereby making identificationdifficult (careful'scraping can providesufficient intact individuals of the speciespresent to make qualitative determinations); VIor the process of collection adds sufficient'foreign materials (i. e. detritus, sub-strate, etc. ) so that some commonlyemployed quantitative procedures arenot applicable.

dsa,a-

IV Artificial substrates are a technique,designed to overcome the problems of directsampling. They serve their purpose, butcannot be used without discretion. They areobjects standardized as to surface area,texture, position, etc. that are plaCed in thewaterway for pre-selected time periods duringwhich periphytic growths accumulate. They -

are usually made of inert materials, glassbeing most common withzlastiesisecond infreqUency. Over fifty various devices andmethods of suppoil or suspension of the., A

substrates have been devised (Sladeokova,:--.-,,"1962) (Weber, 1966) (Thomas, 1968).

A

4

1 It can prevent the settling of smotheringmaterials.

2 flushes metabolic wastes away andintroduces nutrient s.to the colony.

THE LENGTH OF TIME THE SUBSTRATEIS EXPOSED IS IMPORTANT.

The growths need time to colonize anddevelop on the recently introducedsubstrate.

Established growths may intermittentlybreak-away from the substrate because .

of current or weight induced stresses, or"over-growth" may "choke" the attachmentlayers (nutrient, light, etc. restrictions)which then weaken or die allowing releaseof the mass.

_C A minimum of about ten days is requiredto produce ,sufficient growths on anartificial substrate; exposures exceedinga longer time than 4-6 weelss,may produce"erratic result" because of:sloughing orthe accumulation of senile growths insituations where the substrate isartificially protected from predation andother environmental stresses.

155

VII Determining the variety of growths present:is presently only practical with microscopicexaminatioh. ,(A few,micro-chemical pro-

. ,cedures for differentiatiOn'show promise,but, are only irrthe early, stages of development. )

VIII DETERMINING THE QUANTITY OF.GROWTH(S)

A Direct enumeration of the growths whileattached to the substrate can be used, but

A is-restricted to the larger organismsbecause (1) the problem' of maintainingmaterial in-an acceptable condition underthe short working distances of thecobjectivdlenses on' compound microscopes, and(2) transmitted iight is not adequate,because of either opaque substrates a /orthe density of the colonial growths.

B Most frequently, periphyton is scrapedfrom the substrate and then processedaccording to several available procedures,the selection being based on theneecneed,use of the dttta.4 .

1 Aliquots of the sample maybe countedusing methods frequently. employed iri`plankton analysis.

4 Number of organisms

b Standardized units

c V.olumetric units

2 Gravimetric

a Total dry weight of scrapings

b Ash-free dry weight (eliminatesinorganic sediment)

c A comparison of total and ash-freedry, weights

3 Volumetric, involVing centrifugation ofthe scrapings to det(ermirie a packedbiomass volume.

15G

Attached Growths (Periphyton or Aufwuchs)

r

4 Nutrient analyses serve as indices of4" the biomass by measuring the quantity

of nutrient incorporated.

a Carbon

1) Total organic carbon

2) Carbon equivalents (COD"'

b Organic nitrogen

c Phosphorus - Has limitationsbecause cells can stare excessabove immediate needs.

d Other

5 Chlolophyll and other bio -pigmentextractions.

-_ Carbon-14 uptake

Oxygen production, or respiratory1 oxygen demand

IX EXPRESSION OF 11:OULTS

A Qualitative

1 Forms found

2 Ratios of number per group found

3 Frequency distribution of varietiesfound

ACM

4 Autotrophic index (Weber)

5 Pigment diversity index (Odum)

B Quantitative,',1 Areal basisquantity per square

inchrfoot, centimeter, or meter.Fcir example:a 16 mgs/sq.',inch

b 16, 000 cells/sq. inch. . ,

2 Rate basis. For example: /Z..a .2 mgs/day, of biom accumulation

b 1 mg 02/mg of owth /hour

15,3

*4

Attached Growths (Perinhyton or Aufwuchs)

REFERENCES

1 Blum, J. L. The Ecology of River Algae.Botanical Review. 22:5:291. 1956.

2 Dumont, H. J. A Quantitative Method forthe Study' of Periphyton. Limnol.Oceanogr. 14(2):584-595.

3 Keup, L. E. Stream Biology for Assessing .

Sewage Treatment Plant EfficiencY.Water and Sewage Works. 113:11-411.1966.

4 Seligo, A. Uberden Ursprung derFischnahrung. Mitt. d. Westpr.Fisch. V. 17:4:52. 1905.

5 Sladeckova,. A. Limnological Investigation-Methods for the Periphytoh Community.Botanical Review. 28:2:286. 1962.

6 ,Srameck-Husek. (On the UniformClassification-of Animal and Plant,Communities in our Waters).Sbornik MAP.20:3:2,13. Orig. inCzech. 046.

4.

I

7 Thomas, N.A. Method for SlideAttachment in Periphyton Studies.

-..e\slylanuscript. 1968,

8 Weber, C.I. Methods of Collection andAnalysis of Plankton and PeriphytonSamples in the Water PollutionSurveillance System. Water PollutionSurveillance System Applications andDevelopment Report No. 19; FWPCA,

- Cincinnati. 19 +pp. (multilith). 1966.

9 Weber, C. I. "Annual BibliographyMidwest Benthological Society.Periphyton. ,1014 Broadway,Cincinnati, OH 4520'2.

10 Hynes, H. B,N. The Ecology of RunningWaters. Univ. Toronto Press.555 pp. 1870: .0

11 Spoon, Donald M. Microbial Communitieso the Upper Potomac Estuary: TheAqfwuchs in: The Potomac Estuary,Biological Resources, Trends andOptions. 1CP1113 Tech Pub. No. 76-2,1976.

12 Whitton, B.A. "Plants as Indicatorsof River Water Quality. " (Chapter 5in Biological Indicators of WaterQuality. Ed. 5, James, A. andEvison, Lilian Wiley. 1979

This outline was prepared by Lowell Keup,Chief, Technical Studies Branch, Divisionof Technical Support, EPA, Washington,DC 20242

Descriptor: Periphyton

.157

I.

J

J

EFFECT OF WASTEWATER TREATMENT, PLANTEFFLUENT ON SMALL STREAMS

I -*BENTHOS A,i ORGANISMS GROWINGON Oh ASS° ATED PRINCIPALLYWITH THE OTTOM OF WATERWAYS

Benthos is the noun.

Benthonic, benthal and benthic areadjectives./

II THE BENTHIC COMMUNITY

A Composed of a wide variety of lifeforms that are related because theyoccupy "common ground"--substratesof oceans, lakes, streams, etc.They may be attached, burrowing, ormove on theinterface.1 Bacteria

A wide .variety of decomposers workon organic materials, breaking themdown to elemental or simple corn-pounds.

2 Algae

Photosynthetic 'plants having no trueroots. stems, and leaves. The basicproducers of food that nurtures theanimal components'of the community.

3 Flowering Aquatic Plants, (Riverweeds,yondweeds)

The largest flora, composed.of .

complex and differentiated tissues.May be emersed, floating, or sub-mersed according to habit.

4 Microfauna

Animals that pass through a U. S.Standard Series No. 30 sieve, butare retainedon a No. 100 sieve.E'Xamples are rotiferS.and micro-crustaceans. Some forms haveorgans for attachment to substrates,while others burrow into soft-niateri.Ior occupy the interstices between rocks,flfral or faunal materials.

BI. MET. fm. 8j.. 5.80. 153

t

5 .Meioiauna

Meiofauna occupy the interstitial zone(like between sand grains) in benthicanti hyporheic habitats'. They are inter-mediate in size between the micrOfauna(protozoa and rotifers) and the macro -fauna (insects, etc. ). They passa No. 30sieve (0.-5 mm approximately). In fresh-water they include nematodes, copepods,tardigrades, naiad worms, and some flatworms. -They are usually ignores in fresh-water studies, since they pass the standardsieve and/or sampling devices.

6 Macrofauna (macroinvertebrates)tAnimals that are retained on a No. 30 mesh

sieve (0. 5 mm approximately). This groupincludes the insects, worms, molluscs, andoccasionally fish. Fish are not normallyconsidered as benthos, though there are bottomdwellers such as sculpins, settlesdarters, and madtotns.

B;The ,benthos is a self-contained community,though there is interchange with othercommunities. For example: Planktonsettles to it, fish prey on it and lay theireggs there, terrestrial detritus and eavesare added to it, and many aquatic insectsmigrate from it to the terrestrial environ-ment for their mating cycles.

It is an in-situ water quality monitor. Thelow mobility of the biotic components requiresthat they "live with" they:quality changes of the

-overrpassing waters. ChanOs imposed in thelong-lived components remain visible forextended periods, even after the cause hasbeen eliminated. Only time will allow a curefor the community by drift, reproduction, and re-cruitment from the hyporheic zone; .

D Between the benthic zone (substrate/waterinterface) and the underground water tableis the hyporheic zone. There is considerableinterchange from one zone ,to another. ,

III HISTORY OF BENTHIC OBSERVATIONS

C

A Ancient literature records the vermin associ-,ated with fouled waters.

16-1

Effect of Wastewater Treatment Plant Effluent on Small Streams

B 500 -year- oldlishing lit rature refersto animal forms that are fish food andused as bait.

C The scientific literature associatingbiota:to water pollution problems isover 100 years old (Mackenthun andIngram, 1964).

, D Early this century, applied biological-investigations were initiated.

1. The entrance of state boards of healthinto water pollution control activities.

-

2 Creation of state conservation agencies.

3 Industrialization and urbanization.

4 Growth of limnological programs,at universities.

E A decided increase in benthic studiesoccurred in the 1950's anifinuch oftoday's activities are strongly influencedby .1evelopmental work conducted duringthis period. Some of the reasopS for thisare: .

1 Movement of the universities from"academic biology" to applikdpollution programs. ,

2 Entrance of the federal governmentinto enforcement aspectS of waterpollution control.

3 A rising economy and the developmentof federal grant systems.

4 Environmental Protection Programsare a current stimulus.

IV WHY THE BENTHOS?..

A t is a natural monitor .

B The' community contains all of thecomponents of an ecosystem.

1 Reducersa bacteriab fungi -

2 Producers (plants)

vot

- .3 Consumers

a Detritivores and bacterial feeders

b Herbivores

c Predators

C Economy of Survey

1 Manpower

2 Time

3 Equipment

D Extensive Supporting Literature

E Advaniages of the Macrobenthos

.1 Relatively sessile

2 Life history length

3 Fish food organisms

4 Reliability'of Sampling

5 Dollars/information

6 Predictability

17 Universality8 Sensitivity to.perturbation

-.F "For subtle chemicalchanges,

unequivocal_,data, and observationssuited,th some statistical4evaluatipn willbe needed. This requirement favors themacrofauna as' a parameter. Macro-invertebrateiare easier to samplereproductively that) other organisms,numerical estimates are possible andtaxonomy needed,for synoptic invest-gatioris is withirithe reach of a non-specialist." (Wuhrmann)

G "It is self-evident that for a multitude ofnon-identifiable.though biologically activechanges of chemical conditions in rivers,small organisms.with high physiologicaldifferentiation are most responsive.Thus. the small macroinvertebrates(e. g. insects) are doubtlessly the mostsensitive organisms for demonstrating

259

, 4

Af r,

Effect of Wastewater-Treatment Plant Effluent on S 11 Streams

unspecified changes of waterchemistry called 'pollution'Progress in knowledge on useful,autecological properties oforganisms or of transfer of suchknowledge 'into bioassay:practicehasbeen very small in the past.Thus, the bioassay concept(relation of organisms in astream-to water quality) inWater chemistry has'brought notmuch more than visual demon-stration of a few overall chemicaleffects. Our capability to derivechemical conditions- from biOlogicalobservations is, therefore, almoston the same level as fifty years ago.In the author's opinion it is idle toexpect much more in the future becauseof the limitations inherent to natural bio-assay systems (relation of organismsin a stream to water quality)." (Wuhrmann)

4,

V REACTIONS OF-THE BENTHIC NINCRO-INVERTEBRATE COMMUNITY TOPERTURBATION

A Destruction of Orgariism Types

_1 Beginning, with the most sensitiveforms, pollutantskill in order ofsensitivity until the most tolerant formis the last survivor. This results in areduction of variety or diversity oforganisms.

2 The generalized order of macro-inyertebra- te disappearance on asensitivity soale,below pollutionsources is shoign in Figure 2.

WaterQualityDeteriorating

;

Stoneflies aterMayflies alityCaddisflies mprovingAmphipods-IsopodsMidgesOligochaete s

160

As water quality improves, thesereappear inlhe same order.

B The Number of Survivors Increase

1 Competition and predation are reducedbetw.pen different species.

2 When the pollutant is a food (plant's,fertilizers, animals, organic 'materials).

C The Number of Survivors Decrease

The m tevial added is toxic or has nofood va ue.

2 The material added p'roduces toxicconditions as a byproduct of decom-position (e.g., large organic loadingsproduce angnaerobicevironmentresulting in the production of toxicsulfides, methanes, etc. )

D The Effects May be Manifest in Com-binations

1 Of pollutants and their effects.

2 Vary with longitudinal distributionin a stream. (Figure 1)

E Tolerance to Enrichment Grouping(Figure 2) -

Flexibility must be maintained in theestablishment of tolerance lists basedon the response of organisms to theenvironment because of complex relation-ships among varying environmentalconditions. , some general tolerance=.patterns can be established. Stoneflyand mayfly nymphs, hellgrammites.,and caddisfiy larvae represent a grouping(sensitive or intolerant)'that is generallyquite sensitive to environmental.changes. Blackfly larvae, scuds, sow -bugs, snails, fingernail clams, dragon-flyskana damselfly naiads, and mostkinds of midge larvae are facultative(or intermediate) in tolerance.Sludge-worms, some-kinds of midgelarvae (bloodworms)t and some leeches.t

16-3


cctua)

z

DIRECTION Of FLOW 7*C.

D.

I-

TIME OR DISTANCE

.NUMBER OF KINDS,NUMBER OF ORGANISMS,SLUDGE DEPOSITS

,Fciur basic responsei of bottom animals to pollution.

A. Organic wastes eliminate the sensitive bottom animalsand provide; ood in the form of sliidges for the surviving toldr-ant, forms. B. Large quantities of decomposing organic wasteseliminate sensitive bottom animals and the excessive qucmtities of byproducts of organic decomposition inhibit the tolerantforms: in time, with natural stream purification, water qualityimproves so that the tolerant forms can flourish, utilizing thiisludges as food. C. Toxic materials eliminate the sensitivebottom animals; sludge is absent and food Is restricted to thatnaturally occurring in the stream. which linos the number oftolerant surviving forms. Very toxic materials may eliminateall organisms below a waste source. D. Organic sludges withtoxic materials reduce the number Of kinds- by eliminatingsensitive forms. Tolerant 'survivors do not utilize the organicsludges because the toxicity restricts their growth.

Figure 1

16 -4

J

are tolerant to comparatively wavy loadsof organic pollutants. Sewagp mosquitoesand rat-tailed maggots are tolerant ofanaerobic environments for they areessentially air-breathers.

F Structural Limitations

1 The morphological structure of aspecies limits the type of environmentit may occupy.

a Species with complex appendagesand exposed complicated respiratorystructures, such as stoneflynymphs, mayfly nymphs, andcaddisfly larvae, that are subjectedto a constant deluge of settep.bleparticulate matter soon abandonthe_polluted area because of theconstant preening required to main-tain mobility or respiratory func-tions; otherwise, they are soonsmothered.

b Benthic animals in depositing zonesmay also be burdened by "sewagefungus" growths including stalkedprotozoans. Many of,these stalkedprotozoans are host specific.

2 Species without complicated externalstructures, such as bloodworms andsludgeworms, are not so limited inadaptability.

a A sludgeworm, for example, canbarrow in a deluge of particulateorganic matter and flourish on theabundance of "manna."

b Morphology also determines thespecies that are found in riffles, onvegetation, on the bottom of pools,or in bottom deposits.

A

e

(

Effect of Wastewater Treatmen$ Plant Effluent op Small Streams

VI SAMPLING PROCEDURES

A Faufta.

1 Qualitative 'sampling determines thevariety of species occupying an area.Samples may betaken by any methodthat will capture representatives of thespecies present. Collections fromsuch samplings indicate changes in theenvironment, but generally ,do notaccurately reflect the degree of change.Mayflies, for example, maybe re-duced from 100 to 1 per square meter.Qualitative data would indicate thepresence of both speciest .but might notnecessarily .delineate the change in pre-dominance from mayflies to sludge-worms. The stop net or kick samplingtechnique is often used.

2 Quantitative sampling is performed toobserve changes in predominanc-e.The most common quantitative samplingtools are the Petersen, Ekrnan, and Pottergrabs and the Surber stream bottom orsquare-foot sampler. Of these, the,

_ Petersen grab samples the Widest varietyof substrates. The Ekman grab is limited

- to fine-textured and soft substrates, suchas silt and sludge, unless hydraulicallyoperated: .-

f

The Surber sampler is designed forsampling riffle areas; it reqtkiresmoving water to transport dislodgedorganisms into its net and is limitedto depths of two feet or less.

Kick samples of one minute duration willusually yield'around 1,000 macroinyert,ebrates per square meter L10.5 X a oneminute kick= organisms/mh).

3 Manipulated substrates (often referred toas "artificial substrates") are.placed in a stream and left for a specifictime period. I3enthicjcroinvertebratesreadily colonize these forming a manipu-lated community.. Substrates may be con-structed-of natural materials or synthetic;may be placed in a natural situation orunnatural; and may or may not resemble, I,'

the normal stream community. Thepoint being that a great number, of envi-ronmental variables are standardized andthus upstream and downstream stationsmay be legitimately compared in. terms ofwater quality of the moving water column.They naturally do not evaluate wliat mayor may not !e happening to the substratebeneath said monitor. The latter couldeasily be the more important.

REPREStNTATIVE,BOTTOM-DWELLING MACROANIMALS

Drawings from Geckler, J. K. M. Mackenthun and W.M. Ingram, 1963.

Glossary of Commonly Used Biological and Related Terms in Water andWaste Water Control, DHEW, PHS, Cincinnati, Ohio, pub. No. 999cWP -2.

A Stonefly nymphB Mayfly nymphC Hellgrammite or

Dobsonfly larvaeD "Caddisfly larvaeE Black fly larvaeF ScudG Aquatic sowbugH Snail

(Plecoptera)(Ephemeroptera)

(Megaloptera)(Trichoptera)(Simuliidae)(A mphipetla)(Isopoda)(Gastropocla)

I Fingernail Clam (Sphaeriidae)J Damselfly naiad (Zygoptera)K Dragonfly, naiad (A nisopt e'ra )L..,Bloodworm or midge

f ly larvae. (Chironomidae)M LeeCh (Hirudinea)N Sludgeworm (Tubificidac)O Sewage fly larvae (Psyc4odidae)P Rat-tailed maggot (Tubifera-Eristali

KEY TO FIGURE 2

v

O

\

SENSITIVE

G

INTERMEDIATE

ti

1",`

e,6

110Effect op-Wastewater Treatment -Paant Effluent on Small, Streams

Invertebrates which are=: part obenthos, but under certain cobecome Carried downStrearn inappreciable numbers, are, knownDrift.

z2411

MOP,'

Groups which have members forminga4conspicjous partvf the driftinclude the insect orders Ephemeropterab,Trichoptera, Plecoptera and thecrustacean order Amphipoda. .

Drift net studies are widely used andhave a proven validity in streamwater quality studies.

5 The collected sample is screened tha standard sieve to-concentrate theorganiSms;_these are separated fromsubstrate and debris, and the numberof each kirisVot'Organism determined.Data area` adjusted to number perunit ar,4,7els!Vy to numberof bottomorganisztil per ii:guar meten.-

6 Indequananal

ndently74itheCqualitative ot 'ati,ye data- sufftee for fhor gh,ofAn'Viron6ental conditions. VII, ANALYSES OF MICROFLO_FiA

'''eitarnifiaion to, detect damagea r be made4fth either method; but- .,

a co bination of the two gives a morepi-ec e determination. If a choice mustbe de, Tiantitative sampling wouldbe est, because it incorporates apartial qualitative sample.

,7 '7 'Studies have shown-that a significant -

number and variety of.ma'croinve0e-bl-ates inhabit the hyporheic zone in streams.As much r6: 80% of the macrolfiverte-

' brdtts may be below 5 cm in tipshyporheic zone, Mos't samples andsampling techniques do not penetratethe substrate below the 5 cm depth.All quantitative studies must take this-and otter substrate factord'into account

hen absolute figures are presented onanding crop and numbers per square

ter, etc.

. rF

ora 3.B, Fl

1

:-Direct quaptitative sampling of natu-

rally growing bottom algie is difficult.Ills basically one of collecting algae'from a standard or uniform area of the'bottom substrates without disturbingthe delicate growths and thereby dis -'tort the,eample. Indirect quantitativesampling is the best available method.--

2' Manipulated substrates, such as Aood-blocks,f, glass or plexiglassbricks, etc., are placed in a stream.Bottom-attached _algae will grow onthese artificial substrates. _After twoor more weeks, the artificial sub-strates are removed fo? analysis.Algal growths are scraped from, thesubstrates,and the quantity measured.Since the exposed substrate area andexposure periods are equal at all otthe sampling sites,. differences in thequantity of algae can be related tochanges in the quality of water flcaving.over the substrates.

0

1 64

A Enumeration

1 The quantity of:algae on, manipulatedsubstrates can be measured in severalways. Microscopic counts of algalcells and dry weight of a algal mater-ial are.long-established-methods. _ _

.4.

2 Mieroscopic counts involve thoroughscraping, mixing and suspension ofthe algal cells. From this mixturean aliquot of-cells is withdrawn forenumeration under a microscope.Dry - weight is determined by dryingand weighing the algal sample. then---Nigniling the sample Ito burn off e

gal materials, leaving iner inorganicmaterials that are again weighed.The difference between initial dry weightand weight afterignilikion is attribilted to, algae.

r 3 Any organic sediments, however,that settle on the substrate along

--with the algae are processed also..i..+.0

-.

0 .4

Effect of Wastewater Treatment_Plant'Effluent-on-Small-Streanis--Y

A.*

Thus, if organic wastes are presentappreciable errors may enter intothis method.

13 Chlorophyll Analysis

s

i During the past decade, chlpro,phyllanalysis has become a popular methodfor estimating algal growth.' Chloro-phyll is extracted from the algae andis used as. an index of the quantity ofalgae present. The advantages ofchlorophyll analysis are rapidity,simplicity, and.ViVid pittoriat results..

9 The algae are scrubbed fr,om theplaced substrate samples, ground,then each ,sample is keened in equalvolumes? 90% aqueous acetone, whichex*acts the Chldrophyll from the algalcells. The chlorophyll extracts maybe compared visually.

3 Because the cholorophyll extracts fadewith time, oolorimetry should be usedfor permanent record For routine'records, simple 'colorimeters will

At high cholorOPhylldensities, interference Vvith colori-metry occurs, which must he correctedthrough seiial dilution of the sampleor with a nomograph.

aututrOpliic

The ChlosoRhyll'content of the periphytonis used to estimate the alkdl Inomas-sandcis :In inditA;",e,t. tmr. Inelont. onfeentfor trophic status) cir"to7i-Icity of the waterunit the taxononne-comPosition of the

: \,- ,community:- Periphytbn growing in sur- ..1.fa.ce water relatively free of organic-nonillion consists largely of algae,which contain approXimately 1 to 2 percentchlorophyll'a by dry 'Weight. If dissolvedOr partiCulate organic matter presentIn high concentratione, large popula;tionsof filamentOut-bacteria, stalked prOtozoa, 'and otlidrli. liorophyll'beai-ing micro-r

viii

organisms ,,yelop and the percentageof chlorophyll is'then reduced.. 4f thehioniass-chlorophyll a relationship

expressed as a ratiolthe autotro-index); yallies'greater than 100

.may result from orgahad'pollution(Weber and 11/IeFarlan,dq 19169; Weber,

16 -$' r

Autotrophic Index""Aish-free Wgt (n m2 )

Chlorophyll a (mg/m2),_.

1

MACROINVERTEBRATE ANALYSES

A Taxonomic

The taxonomic level to whic.l'h animals .areidentified depends on the needs, experience,and available resources. However; thetaxonomic level to which identifications arecarried in each major group should beconstant throughout a, given study.

li Biomass

Macroinvertebrate biomass (weight oforganisms per unit area) ip °a. usefulquantitative estimation of standing crop.

C Reporting Units

Data from quantitative samples may be used_ -to obtain:

1 Total standing crop of individuals, orbiomass, or both per unit area or unitvolume or sample unit, and

2 Nilmbers orbiomass, or both, of individual"'taxa per unit area or unit volume or sample

Data, from devices sampling a unit areaof bottom willbe reported in grams dryweight or ash5free dry weight per squaremeter (gm/ m ), or numbers ofindi-viduals per square meter, or both.-

44 ; Data from multipldte samplers will-bereported in terms of the total surfacearea of the plates in grams dry weightor ash-free dry weight or numbers ofirdividualper square. meter, or both.

5 data fr'Orn rock-filled basket samplerswill be reported as grams dry weightor numbers of individuals per sampler,or both.

165

.s


IX FACTORS INVOLVED IN DATA INTER-PRETATION ,

Two very important factors in data evalua-tibn are a thorough kriowledge of conditionsunder which the data were collected and acritical assessment of the reliability of thedata's representation of the situation.

A Maximum-Minimum Values

The evaluation of physical and chemicaldata to determine their effects on aquaticorganisms is primarily dependent onmaximum and minimum observed values.The mean is useful only when the data arerelatively uniform. The minimum ormaximum values usually create acur-conditions in the environment.

B Identification

, Precise identification of organists tospecies requires a specialist in that.taxonomic groups. Many immatureaquatic forms have not been, associatedwith the adult species. Thbrefore, onewho is certain of th& genus but not thespecies should utilize thenot a potentially incorr,eThe method of interpret logicaldata on the basis of numbers of kindsand numbera of organismavill typically'Suffice.

C Lake and Stream Influence

neric name,Gies name.

Physical tharacteristics of a body ofwater also affect animal populations.Likesor impounded bodies of watersupport Afferent faunal associationsthan rivers, The number of kindspresent in a lake may be less than thatfound -in a stream. because of a moreuniform habitat. *A lake is all pool,but a river is compdsed of'both poolsand riffles.- The nonflowing water oflake exhibits a more complete set-tling of particulate arganic'matter thatnaturally supports a higher populationof detritus consumers. For these

i6

reasons, the bottom fauna of a-lake orimpoundment, or stream pool cannot bedirectly compared with that of a flowing,.stream riffle

D Extrapolation'

How can bottom `-dwelling macrofauna databe extrapolated to other environmentalcomponents? It must be borne in mindthat a component of the total environmentis being sampled. 1-114e sampled com-ponent exhibits cha.nges, then so must theother interdependent components of theenvironment. For example, a clpan streamwith a wide variety of desirable bottomorganisms would'be expected to have awide variety of desirable bottom fishes;when pollution reduces the number of bottomorganisms, a comparable reduction wouldbe expectedin the number of fishes. More-over, it would be logicApto conclude thatany factor that 'eliminates 'all bottom organ-isms,.would eliMinate most other aquaticforms of life. A clean- stream with a widevariety of desirable bottom organismsWould be expected to permit a variety ofrecreational, municipal and -industrial uses.

E Expression of Data

1 Standing crop 'and taxonomic composition

Standing crop and numbe-ts of taxa (q`pesor kinds) in a community are highlysensitive to environmental perturbationsresulting from the introduction of contam-inants. These parameters, particularlystanding crop, may vary considerably inunpolluted habitats, where they may rangefrom the typically high standing $rop oflittoral zones of glacial lakes to thesparse ,fauna of torrential soft-waterstreams. Thus, it is important thatcomparisons areznade only between trulycomparable erOironments.

16-9.

)

Effect of Wastewater 11)eatment Plant Effluenton Small Streams

2 Diversity

Diversity indices are an additional tool .for measuring the quality of the environ-ment and the effect of perturbation onthe structure of a community of macro-invertebrates,: Their use is based onthe generally observed phenomenon thatrelativelPtuidtsturbed environmentssupport communities hiving largenumbers of species with no individualspecies 'present in overwhelmingabundance. If the species in such acommunity are ranked on the basis oftheir numerical abundance, there will ,

--rbe relatively few species with largenumbers of individuals and largenumbers of species represented by onlya few individuals. Perturbation tendsto reduce diversity by making theenvironment unsuitable for some speciesor by giving other species a competitiveadvantage..

Indicator-organism scheme ("rat-tailedmaggot studies")

a-=-For-thi-s-teehnique, the individualtaxa are classified on the basis oftheir tolerance or intolerance to ' -various- levels of putrescible wastes.Taxa are classified acdortling totheir presence or absence ofdifferent environment's as deter-mined by field studies.- Somereduce data based on the presenceor absence of indicator organismsto a simple numerical form for easeirrkesentation.

b Biologiats are engaging in fruit-., less exercise if they intend to make

any decikons about indicatororganisms by operating atthegenerie level of macroinvertebrateidentifications." (Resh and Unzicker)

4 Reference station methods

Comparative'or control station methods, compare the qualitative characteristics

of the fauna in clean water habitats withthose of fauna in habitats subject to stress.Stations are compared on the basis,ofrichnqss bf species.

4

If.adequate background data are avail-able to an experienced investigator,these techniates can prove quite usefulparticularly for the purpose of demon:strating the effects of gross to moderateorganic contamination on the macro -invertebrate community. To detectmore subtle changes in the macroinver-tebrate community, collect quantitativedata on numbers pr biomass of organisms.Data on the presence of tolerant andintolerant taxa and richness of speciesmay be effectively summarized for evalu-ation and presentation by means of linegraphs, bar graphs, pie diagrams,histograms, or pictoral diagrams.

X IMPORTANT ASSOCIATED ANALYSES

A The Chemical Environment

1 Dissolved oxygen.

2 Nutrients

3 Toxic materiels...-'

4 Acidity and alkalinity

5 Etc.

B The Physical Environment

1 Suspended solids

2 Temperature

3 Light penetration

4 Sediment composition

5 Etc.

XI AREAS IN WHICH BENTHIC STUDIESCAN BEST BE APPLIED

A Damage Assessment or Stream Health

If a stream is suffering from abuse thebiota will so indicate. A biologist candetermine damages by looking at the"critter" assemblage in a matter ofminutes. Usually, if damages are notfound, it will not be necessary to alert

'the remainder of the agency's staff,

167

yr1

B

Effect of Wastewatei TreatmentPlant Effluent on Small Strea.rris

pack all the equipment, pay travel and perdiem, and then wait five' days beforeenough data can be assembled to begin.evaluation.

By determining what damages have beendone, the potential cause "list" can bereduced to a few items for emphasis andthe entire "wonderful worlds" of scienceand engineering need not be practiced withthe result^that much data are discardedlater because they were not applicable tothe prOblem being investigated.

C Good benthic data associated with chemi-cal, physical, and engineering data canbe used to predict the direction of futurechanges and to estimate the amount ofpollutants that need to be removed fromthe waterways to make them productiveand useful once more.

D The benthic rnacroinvertebrates are aneasiled index to stream health thatcitizens may use in stream improve-ment programs. "Adopt-a-stream"efforts have successfully used simplemacroinvertebrate indices.

E The potential for restoring biologicalintegrity in our flowing streams usingmacroinvertebrates has barely beentouched.

REFERENCES

1 Freed, J. Randall and Slimak, Michael W.Biological Indices in Water QualityAssessment. - (in The FreshwaterPotomac Acidatic Communities and .Environmenta.1 Stresses). InterstateCommission on the Potomac'RiverBasin. 1978.

2 Hellawell, J. M. , Biblogical SurveillanceOf Rivers. Water Research Center.Stevenage and Medenham. 333 pp.1978.

4

:3 Hynes, .11*B.N. The Ecology of RunningWaters. Univ. Toronto Press. 1970 .

+,

- 163

4 Mackenthun, K. M. The Practice ofWater Pollution Biology. FWQA.281 pp.. 1969.

5 Weber, Cornelius I., Biological Fieldand Laboratory Methods for Measuriftthe Quality of Surface Waters andEffluents. U. S. Environmental Pro-tection Agency, NERC, Cincinnati,OH. Environmental Monitoring Series670/4.73.001 July 1973

6 Keup, L. E. and Stewart, R. K. Effectsof Pollution on Biota of the Pigeon River,NorthCarolina and Tennessee. U. S. EPA,National Field Investigations Center. 35 pp.1966. (Reprinted 1973, National Training

-C enter)

7 Wuhrmann, K., Some Problems andPerspectives irk-Applied LimnologyMitt. Internat. Verein Limol. 20:324-402.1974.

8 Armitag, P. D. , Macliale, Angelu M., andCrisp; Diane C. A Survey of StreamInvertebrates in the Cow Green Basin(Upper Teesdale) Before Inundation.Freshwater Biol. 4:369-398. 197+.

9 Resh, Vincent H.., and Unzicker, John D.- Water Quality Monitoring and Aquatic

Organism's: the JWPCF 47:9-19. 1975.

This outline was prepared by Lowell Keup,Chief, Technical Studies Branch,Division of Technical Support, EPA,Washington, D.C. 20460, and revised byR. M. Sinclair, National Training andOperational Technology Center, OWPO,USEPA, Cincinnati, /Ohio 45268.

Descriptors: Aquatic Life, Benthos, WaterQuality, Degradation, Environmental Effects,Trophic Level, Biological Communities.Ecological Distributions

16-11

4. ECOLOGY OF WASTE STABILIZATION PROCESSES

I ITT >RODUCTION

Living organisms will live where they can live.This holds for treatment plant environmentsjust as it does for streams, impoundments,oceans, dry or weft lands.

A Each species. has certain limits or toler-ances, growth, feeding habits and othercharacteristics that determine its favoredhabitat. .

B The presence of certain organisms withwell defined characteristics in a viablecondition and in significant numbers alsoprovides some inference with respect tothe habitat.

C The indicator organism concept has certainpitfalls. It is not sufficient to base anopinion upon one or more critters whichmay have been there as a result of gasliquid or solid transport. It is necessaryto observe growth patterns, associatedorganisms, environmental conditions, andnutritional characteristics to provideinformation on environmental acceptability.

D Organisms characteristic of wastewatertreatment commonly are those found innature under low DO. conditions. Perform-ance characteristics are related tocertain organism progressionS and assoc-iations that are influenced by food toorganism ratios and pertinent conditions.One single species is unlikely to performall of the functions expected during wastetreatment. Many associated organismscompete in an ecological system bonafavored position. 'The combination includessynergistic, antagonistic, competitive,predative, and other relationships that mayfavor predominance of one group for a timeand other groups under other conditions.

E It is the responsibility of the treatmentplant control team to manage conditionsoftreatMent to favor the best attainableperformance during each hour of.the dayeach day of the year. This outline con-siders certain biological characteristicsand their implications with respect totreatment performance.

*PC 10.1.0.69

.

II TREATMENT PLANT ORGANISMS .

WaStewater is characterized by overfertili-zation from the standpoint of nutritionalelements, by varying-amount of items thatmay not enter the metabolic Vattern but havesome effect upon it, such as. silt, and bymaterials that will interfere with metabolicpatterns. Components vary in availabilityfrom those, that are readily acceptable tothose that persist for long periods of time.Each item has some effect upon the organismresponse to the` mixture.

A Slime forming organisms including certainbacteria, fungi, yeasts; protista moneraand alga tend to grow rapidly, on dissolvednutrients under favorable conditions. Thesegrow rapidly enough to dominate the overallpopulation during early stages of growth.There may be tremendous numbers ofrelatively-few species until availablenutrients have been converted to cell massor other limiting factors check the pop-ulation explosion:

it.B Abundant slime growth favors production

of predator organisms such as amoeba orflagellates. These field upon preformedcell mass. Amoebas-tend to flow_around_particulate materials; flagellates also aterelatively inefficient food gatherers. Theytend to become numerous when the nutrientlevel is high. They are likely to be assoc-iated with floculated masses where foodis more ablirdant.

C Ciliated organisms are more efficient'food gatherers because they have theability to move more readily and mayset up currents in the water to bring foodto them for ingestion. Stalked ciliatesare implicated with well stabiliied effluentsbecause they are capable of sweeping thefine particulates from the water betweenfloc masses While their re§idues tend tobecome associated with the floc.

D Larger organisms tend to become establish-, ed later and serve as scavengers. These.

include Oligocha:etes (worms), Chiobomids(bloodworms and insectilaryae), Isopods(Sow bugs and crustacea), Rotifera andothers.

17-1.

Ecolog_of Waste Stabilization ProoesAes

104,

III TREATMENT OPERATIONAL CONTROL

An established treatment pla-nt is likely to*pntain representative organisms from allgroups of tolerant species. Trickling filters,activated, sludge, or ponds tend to retainpreviously developed organisms in largenumbers relative to the incoming feed. Thenumber and variety avail determine theie.nature, degree and time

rquiredfor partial

oxidation and conversion of pollutants fromliquid to, solid concentrates.

A Proliferation of slime forming organismscharacterize the new unit because they growrapidly on soluble nutrients. Predatorsand scavengers may start growing as soonas cell mass particulates appear but growthrate is slower and numbers and mass lagas compared with slime organisms. Asslime growth slows due to conversion ofsoluble nutrients to cell mass, the slimeformers tend tO associate as agglomeratesor clumps promoting floculation and liquidsolid separation.

13 Overfeeding an established unit encouragesrapid growth of slime organisms as individ-ual cells rather than as flocculated masses.This results in certain characteristicsresembling those of a young, rapidly .

growing system..

Toxic feeds or unfavorable conditionsmaterially reduce the population of exposedsensitive organisms. The net effect is apopulation selection requirihg rapid regrowthto reestablish desired operating character-istics. The system assumes 'new growthcharacteristics to a degree depending uponthe fraction remaining after the toxic effecthas been relieved;h-y dilution, degradation,sorption, or other means.

D Treatment units are characterized bychanges in response to-feed sequence,load ratio,, and physical or chemical,conditions. Response to accutectoxiCity

4.--." may be immediately apparent. Chronicoverloading or mild toxicity may not beapparent for)several days. It may be

, expected. that it will require 1 t ,t3 weeksto restore effective performa e after anymajor upset.' Performance . iteria,maynot indicate a smooth progression towardimprc:qed operation. .

E Observations of the growth characteristicsand populations do not provide quantitativeinformation, but they do indicate trendsand stages of development that are usefulto identify problems. It is not possible toidentify most slime organisms by directobservation. It is posSible to recognizegrowth and flocculation characteriStics.Certain larger organisms are recqgnizableand are useful as indicator organisms tosuggest past or subsequent developments.

IV ILLUSTRATIONS OF ECOLOGICALSIGNIFICANCE

A The first group repreents initial devel-opment of non-flocculent growth. Singlecelled and filamentous growth are shown.Rapid growth shows little evidence offlocculation that is necessary to producea stable, clear effluent.

B The next group of slidesAndicate develop-ment of floc forming tendencies fromfilamentous or non-filamentous growth.Clarification and compaction characteris-tics are relatable to the nature and densityof floc masses.

Organisms likely to be associated withmore stabilized sludges are shown in thethird grntip Scavengers-esseOn-sist of a large alimentary canal withaccessories.

D The last two slides illustrate changes in -appearance after a toxic load. Scavengers,ciliates, etc. have been inactivated. New ,

growth at the edge ot the floc masses arenot apparent. Physical structure indicatesdispesed residue rather than agglomera-tion ndencies. The floc probably contains

- living organisms protected by the surround-ing organic material, but only time andregrowth will reestablish a working flocwith good stabilization and clarificationtendenc %s.

ACKNOWLEDGEMENTSThis,outline contains significant materials fromprevious outlines by H. W. 1Jackson and R. M.Sinclair. Slide illustrations were provided byDr. Jackson.

Thisoutline was prepared by F. J. Ludzack,Chemist, National Training Center,.EPA, Cincinnati, OH 45268.

170

ECOLOGY PRIMER(from Aldo Leopold's A SAND COUNTY ALMANAC)

Ecology is a belated attempt to-con<rertour collective knowledge of biotic materialsinto a collective wisdom of biotic manage-ment.

II The outstanding scientific discovery ofthe twentieth century is not television orradio, but rather the complexity_of theland organism.

LEI One of the penalties Of an ecological edu-cation is that one lives alone in a worldof wounds. Much of the damage inflictedon land is quite invisible to laymen. Anecologist must either harden his shelland make believe that'the consequencesof science are none of his business, orhe must be the doctor who sees the marksof death in a community that believesitself well and doe's not want to be toldOtherwise.

IV Ecosysteins have been sketched out aspyramids, cycle', and-energy circuits.The concept of land as an energy circuitconveys three basic-ideas:

A That land is not merely sail.

B That the native plants and animals keptthe energy circuit open.; others may ormay not.

C That man-made changes are of a different

VI

VII

A thing is right when it tends to preservethe integrity, stability, and beauty of thebiotic community. It is :wrong when ittends otherwise.

Every farm is a textbook on animalecology; every stream is a textbookon aquatic ecology; conservation isthe translation of the book.

VIII There are three spiritual dangers in notowning a farm-

A One is the danger of supposing that break-,,fast comes from the grocery.

B Two is thZt heat comes from the furnace.

C' And three is that gas comes from thepump.

IX in general, the trend of the evidenceindicates that in land, just as in thefishes body, the symptoms'may lie inone organ and the cause in another. The-.practices we now call conservation are,to a large extent local alleviations ofbiotic pain. They are necessary, butthey must not be confused with cures.

order than evolutionary changes, and haveeffects more comprehensive than is intend-ed or foreseen (See figures 1-4).

D To keep every cog and wheel is the firstprecaution of intelligent tinkering.

V The process of alteringthe pyramid forhilman occupation releases Stored energy,and. this Often gives rise, during-thepioneering period, to a deceptive exuber-ance of plant and animal lite, both wildand tame. These releases of bioticcapital tend to becloud or postpone thepenalties of violence. .

BI. ECO. 26b. 9.79

X An Atom at large in the biota is too freeto knokw freedom; an atom back in the seahas folgotten it. For every atom lost tothe seaXthe prairie pulls another out ofthe decaying rocks. The only certaintruth is that its creatures must suckhard. live fast, and die often, lest itslosses exceed its gains.

REFERENCES

1 Leopold, Luna 13. (ed.) Round River.Oxford University Press. 1953.

2 Leopold,' Aldo. A Sand County Almanac.'Oxford University Press. 1966.

3 United States Environmental ProtectionAgency. The Integrity of Water.WashingtOn, D. C. 1977.

. -171 18-1

Ecology Primer (from Aldo Leopold's A Sand County Almanac)

REFERENCES (Continued)

4. United States Geological Survey.ographic Maps"

. Reston, Virginia. 1978.

V--,:ca'

egs--,,,,,......,,,........./ ,,,,.....- .--.,Y. Vi alo#

. ..i.y..1:: } :4 / #

-4. t 141' CC, CC iwt

'n*PPrt,t,:tt t tTaint ...,,,;..:%t,. Ir% 4..1it ett--z.,;,, ....;:e.

./;.:'.7.,:',At.,' '/' .4.7.;14::

e';4:"'S"'",%'?i7A) o'f

.

er geA'v*%-1"f;g4.14:1

.14,t4 b.e:14.144.; {44

41{

t SSt

K,.4%:s4 CI,ts>rGreyvik ;j;Ct'41 .."

t*1- %('

. sJ-7

11 1

Mapped in 1949

figure 1.

-4,./_; ,

m;;I 4, -tio

. 61- - /, 0\-?tt% / ,''. .;

oll Course .7,'A ,;, Landing U.

15,:,i 1 D :;:s70, /0 0:1/,,,/

1

..C'14 g

.1i 'Xe;

I.

16. .

ii .......

I '

I .!

. . '.-"

Golf Course

s

182

1 1 Q06 , I '

.641 it5"1.::44:1.Vtri.k51.15/Ve-t

Photorevised In 1969'

li,igure 3.

... y

': OCean Reef

This oudne was prepared bly R. M. Sincla-o

gational Training and Operd.tional TechnologyCenter, OWPO, USEPA, 'Cincinnati, Ohio45268.

Descriptor: Ecology

.4 104.,'Lancing Field

AY

4teNt

*. ,A* _Ocean Reef

Club 4

S. . 4.

Revised In 1956

. Figure 2.

0

;)errRoll Course

Of().,i//'I"?-00 '1

Phototevised In 1973

Figure 4:

1'2

:1:Ocean Reef'

lub )

04.

I

I

THE LAWS OF ECOLOGY

These so-called Laws of. Ecology havebeen collected and reformulated byDr. Pierre Dansereau.

A They have a broad range of appli-cation. in aquatic as well as terres-trial ecosystems.

B Only the ones which have strictapplication to fresh -water organismswill be discussed.

II The Laws Verbatim

A Physiology of Ectopic Fitness (1-9)

Law of the inoptimum. No speciesencounters in any given habitat theoptimum conditions for all of itsfunctions. .

2 Law of aphasy. "Organid evolu-.tion is slower than environmentalchange on the average,. and hencemigration occurs. "

3 Lavr of tolerance.. A species isconfined, ecologically and geo-graphically, by the extra:ids ofenvironmental adversities that itcan withstand.

Law of valence: In each part ofits area, a given species shows agreater or lesser. amplitude inranging through various 'habitats'(or communities); this is condi-tioned by its. requirements sand,tolerances being satisfied, ornearly bvercome.

5 Law of competition- cooperation.Organisms' of one or more speciesoccupying the same site over agiven period of time, use (andfrequently reuse) the same re-sources thrOugh various sharingprocesses, which allow a greaterportion to the most efficient.

BI. AQ. 35.5 80

6 -Law of the continuum. The gamutof ecological niches, in a regionalunit, permits a gradual' shift" in the

-qualitative and quantitative compo-sition and structure of communities.

7 Law of cornering. The environ-mental gradients upon which speciesand, communities are ordained eithersteepen or smoothen at various timesand places, thereby reducing utterlyor broadening greatly that part ofth5 ecological spectrum which offersthe best opportunity-to organisms of

- -adequate valence.

171

8 Law of persistence. Many species,especially dominants of a community,are capable of surviving and main-taining their spatial position after

- their habitat, and even the climateitself have ceased to favor fullvitality.

9 Law of evolutionary opportunity. Thepresent ecological success of aspecies is compounded of its geo-graphical and ecological breadth, itspopulation structure, and the nature

of its harboring communities. ,

B Strategy of Community Adjustment (10-14)

10 Law of ecesis. The resources of -anunoccupied environment will first beexploited by organisms with high,tolerance and generally with low

, requirementS. -,--

11 Law of succession. The same sitewill not be indefinitely held by thesate plant community, because thephysiographic agents and the plantsthemselves induce changes in thewhole environment, and these. allowother pants, heretofore .unable toinvade, but now more efficient, todisplace the present decupants.

40,101

fork; I

19*k

YIThe Laws of Ecology

Law of regional climax. Theprocesses of succession gothrough a, shift of controls butare not indefinite, for they tendto an equilibrium that allows nofurther relay; the climatic-topo-graphic-edaphic-biological bal-ance of forces results in anultimate pattern which shirtsfrom region to region.

13 Law of factorial control. 'Al:-though living beings react holo-,cenotically (to all ,factors of the /environ ent in their peculiarconjun tion , there frequentlyoccurs a discrepant factor whichhas controlling power throughits excess or deficiency.

14 Law of association segregation.Association of reduced composi-tion and simplified structure havearisen during physiographic orclimatic change and migrationthrough the elimination of somespecies and the loss of ecologicalstatus of others.

C Regional Climatic Response (15-20)

.15 Law of geoecological distribution."The specific topographical dis-tribution (microdistribution) of anecotypic plant species or of aplantrcorninUnity is a parallelfunction of its general g,eograpicaldistribution (macrodistribution),since both are determined by the .

same ecological amplittides andultimately by uniform physiologicalrequirements."

16/, Law of climatic stress. It is atthe level of exchange ,,,,b.04-cen theorganism and the. environment(microbiosphere) that the stressis felt which eventually cannot beovercome and which willAistablisha geographic boundary.

I

17 Law of biological spectra. Life-,form distribution is a characteristicof regional floras which can be

41,

correlated to climatic conditionsof the, present as well as of thepast. 4.

18 Law -of vegetation regime. Undera similar climate, in differentparts of the world, a similarstructural- physiognomic- functionalresponse can be induced in thevegetation, irrespective of floristic.affinities and/or historical con-nections.

19 Law of zonal eqUivalence. Whereclimatic gradients are essentiallysimilar, the latitudinal and altitudinalzonation and cliseral shifts of plantformations also tend to be; wherefloristic history is essentially iden-tical, plant communities will alsobe similar.

20 Law of irreversibility. Someresources Imineral, plant, oranimal) do not renew themselves,because they are the result of aprocess (physical or bilogiCal) which'has ceased to function in a particularhabitat of landscape at the present

r time.

21 Law of specific integrity. Since thelower taxa (species and subordinateunits) cannot be polyphyletic, theirpresence in Widely separated areascan be explained o'nly by formercontinuity or by -migration.

22 Lavi-Of phylogenetiC trends.relative geographical positions,within species (but more often generaand families), of primitive and ad-vanced phylogenetic features are goodindicators of the trends of migration.

The

23- Law -ofmigration. Ge'agraphicalmigration is determined by populationpiessure and/or environmental change.

24 .Law -cif differential evolution. eo,graphic and ecological barriersfavor independent evolution, biz tthedivergence of vicariant pairs is notnecessarily proportionate to the 'gravity,of the barrier or the duration ofisolation.

174Fes-

The Laws of Ecology

.

25 Law of availability. The geo-graphic distribution of plantsand animals is limited in thefirst instance by their placeand time of origin.

26 Law of geological alternation.Since the short revolutioriaryperiods have a strong selectiveforce upon the biota, highlydifferentiated life forms areare more likely to developdUring .those times than duringequable normal periOds.

27 Law of domestication. Plantsand animals whose selectionhas been more or:less domi-nated by man are rarely, ableto survive without his continuedprotection.

III THIENEMANN'S ECOLOPICAhXRINCIPLES

These three Principles apply to streaminvertebrates and will be noted specif-ically during your stream examinationsas you compare aquatic communities.

A The greater the diversity ofthe conditions in a locality thelarger is the number of specieswhich make up the bioticcommunity.

The more the conditions in alocality deviate from normal,and hence from the normaloptima of. most species, the,smaller is the number of species'which 'occur there and thegreater the number of individualsof each of the species which dooccur.

C The longer a locality has beenin the same condition the .

richer I§ its biotic communityand the more stable it is.

'? *

Iv THE LAW OF THE EQUIVALENCE 440

OF WINDOWS (deAssis)

"The wayto compensate for a'Cclosedwindow is to, open another window."

-41P

REFERENCES

Dansereau, P. Ecological Impact and Human'Ecology, in the book Future 4nvironinentlof,North America, edited by F. FraserDarling and John P. Milton; The NaluralHis tory press, division of Doubleday,Company;New York,' 19661970.

dassis, Machado. Epitaph of a Small Winner.Noonday Press. 1966.

Hynes, H. B. N. The Ecology of RunningWaters. University of Toronto Press.1970.

This !outline was prepared by Ralph M.,S,inclair,Aquatic Biologist, National Training Center,MP&ODWPO, USEPA, CinCinnati, OH 45268.

Descriptor: Ecology.

O

V

APPLICATION OF BIOLOGICAL DATA

I ECOLOGICAL DATA HAS TRADITIONA 41.X. III- BEEN DIVIDED INTO TWO GENERAL ,

CLASSES: /.

..

AA Qualitative - dealing with the taxonomic

composition oP communities

B Quantitative - dealing with the population'densityyor rates of processes occurringin the communities

Each kind of data has been useful in its own

II QUA LITA TIV E. DA TA

A Certain species have been identified as:

.1 Clean water (sensitive) or oligotrophic

2 Facultative, or tolerant

3 Preferring polluted regions(see: Fjerdinstad 1964, 1965; Ganfin .

Tarzwell 1956;Palmer 1963, tt,69;'laws-60056; Teiling 1955) *

B Using our knowledge about ecolcigicalrequirements the bioLogist may comparethe species present

At different stations in the same river(Gaufin 1958) or lake (Holland 1968)

In different rivers or lakes (Robertsonand. Powers 1967)

or changes in the species in a rive?: or/lakeover a period of several years. (Carr& Hiltunen 1965; Edmondson & Anderson1956; Fruh, Stewart, Lee & Rohlich 1066;Hasler 1947).

C Untilcomparatively recent times taxonomicdata were not subject to statistical treat-,ment..

F

QUANTITATIVE DATA: TypicalParameters of thity,pe inblude:

°

Counts algae / ml; b enthos/J;fish/net/day

Volume mm3 algae/liter/

C Weight - dry wgt; apt}-free .wgt..

D Chemical content chlorophyll;carbohydrate;'ATP; DNA,; et6.

E caloric equivalents)

Processes - productivity; 0respiration

IV Historically, the chief use of statisticsin treating biological data has been in the .

collection and analysis of sampreS-for-thesep4meters. Recently, many methods havebeen devised to convert taxonomic data intonumerical form to permit:

A Better communication between,thebiologists and other scientific disciplines

B Statistical treatment of taxonomtc data

C In the field of pollution biology thesemethods include:

1 Numerical ratings of organisms on .thebasis of their pollution ,tolerance

(saprobic'valency: Zelinka Sladecek. 1964)

(pollution index: Palmer 1969)

, 2 Use of quotients or ratios of species indifferent taxonomic groups (Nygaard1949).

17G20-1,

AppUcdL± on of BLO1b 4

>.

Q

Simple indices of community diversity:

a Organisms are placed in taxonomicgroups wHichbehave similarly underthe. same ecological conditions. Thenumbe'r of species in these groupsfound at "healthy" stations is corn-parIed to that found at "experirhental"stations. (Patrick 1950)

b A truncated log normal cuveplotted on the basis a the numberof individuals per diatom species.(Patrick Hohn, &.Wallace 1950°.

Sequential comparison irldex.(Cairns, Albough, Busey & Chanay

1968).. In this te,chniqUe,' similarorganism's enco.unteredosequentiallyare grouped intoeruns".

6'

-runsSCI = total Organisms examined

d R.tio-sof° caroterioidiJo chlorophyllin phytoplanktoq populations:

- OD430 /OD 665iNlargalef 1968)

ADD43.5/OD670

(Tanaka} et al 1961)

I

e .The number of diatOm species presentat ac, station ,1s considered indicativeOf water quality, or pollution level.(Williams' 1964) '

numbe'r of specieS (S)-f; number of individuals W)

,--ivriber of- .species (S)Stiar.irelOot. of nuinb&l.of indiYiduals ( N)

fFi°-4- (Menhinick 1964)-log N.

E n i 1) (Simpson 1949)d N (N 1)

where n. = number of individuals1 belonging to the i-th species,

20 -2

and

N = tot tuber of individualss4)

j infor ation theory:

The basic equation used forinformation theory applications was.deyeloped by Margalef (1957).

E

1 N!/.0 = log

N 2 Na ! Nb !' ..Ns !

where, I information/individulal;N ...Ns are the number of

species a, b, . , .

sW4dN is their sum.4740041e4; ,

This equation has' also beefiliSedwith:

1) The fatty, acid content of algae(McIntire, Tinsley, and Lowry

Jew:

2) Algal productivity (Dickman 1968).

3) Benthic biomass (Wilhm 1968)

-REFERENCES

1 " Cairns, J. Jr., Albough, D. W. ,Busey, F, and Chaney, M, D.The sequential comparison index -

- a simplified method for non-biologiststo estimate relative differences inbiological diversity 'in -stream pollutionstudies. J. Water Poll. Contr. Fed.40(9):1607-1613. 1968. 0

2 Carr, J. F. and Hiltunen, J. K. Changesin the bottom fauna of Western Lake'Erie from 1930 to1981. Limnol.OceanOgr. 10(4):551-569.. 1965.

3 Dickman, M. Soine indices of diversity.Eco1ogy49(6):1191-1193. 1968.

1

1

Tto

r-1

. A

Tt"

EStnondson,, Anderson, G. C.Artificiai,EutA ation of LakeWaahingtcin. Oceanogr;1(1):47-53: 195'6.

5. Fjerdingstad, E. 'Pollution's-of Streamsestsimateclby,;5berIthal phytomicro-organisms-. -''.4,%h\ saprobic.,systembased on commtinitios of organismsand ecological facto1:44, Internatil,Rev. Ges.. Hydrobiol. 49P:63-M1.1964.

J.D

6 Fjerdingstad, E. Taxonomy and saProbicvalency of b'enthio phytomicroi,,,,_organisms. Hydrobiol. 50 (4):05-604.

7 Fruh, E. G. , Stewart, K. M , Lee, G.7t: .

And Rohlich, G.A . Measurements-*eutrophication and trends. J. WatW-,.:Pail, Contr. Fed. 38(8):1237-1258.

A liCation of`BiolO ical Data

15 Menhinick, E. F. A comparison of s omespecies - individualsdiver'sity indicesapplied to samples of field,insects.Ecology 45:p59. 1964.

16 Nygaard, G. Hydrobiological studies in -some ponds and lakes: II. The .quotient hypothesis and some new orlittle-known phytoplankton organisms. ,

Mg. Danske Vidensk. Selskl. Biol.Skrifter 7:1-293. 1949.

17 Patten, B. C. Species diversity in netplankton& Raritan Bay., J. Mar.Res. 20:57-75. 1962. -

iB Palmer, C. M. The effect of polltifion onriver algae. Ann. New York A.Cad.Sci.' 108:389-395. 1963.

'19 Palmer, C. M. A compOsite rating ofalgae tolerating organic pollution.,J. Phycol. 5(1):78-82. 1969.8. Gaufin, IVA. Effects Of Pollution on

nkidAyeSthrnstream, Ohio J. Sci.40i:197-208. 1958.

9 Gaufin,1 A.R. and Tarewell, C. M.,macroinvertebrate comindicators of organic pCreek. Sew. Ind. Ira-St924. 1956. "

AquatiOunities aslution,in Lytle

s. 213(1):906-

10/ Hasler, :-g__titrophication of lakes bydomeiti& ralOge.395.1=

11 Holland, R,E. Correlation of Melosiraspecies with trOphic conditions in LakeMichigan. Limnol. Oceanogr. .

13(3):555-557. 1968.

12 Margalef, R, Information theory inecology. Gen. Syst. 3:3Q-71, 1957.

Ecology 28(4)1383-

0

galef, R. Perspectives in ecologicalry. Univ. Chicago Presi, 1968.--t.%

14 McIntire,,

Tinsfey, I. J. and o

Lowry, R. fk Fatty acids in loticperiphyton: 'notheRipreasure ofcommunity, strritcture T Phycol.5:26-32. 19.69.

20 Patrick, R., Hohn, M. H. and Wallace,J.I. -4 A new method for determiningthe pattern of the diatom flora. Not.

Natl.,Acad. Sci., No. 259.,-PhiladePphia'. 1954.

7

21 tRawson, D. S. Algal.indicators of trophic'Jake types. 7 Limnol. Oceanogr,1:18-?5. 1956.

22: Robertson, §. and Powers, C.F.Cdinfijarison,of the distribution oforganie,Irnatitet..in the five Great Lakes.in: J. C. A y e s., an d D. C. Chandler,.eds. Stiftlies on the environment andeutrophigtion of:`:-.L.ake Michigan.Spec. Rpt, No. 364.:Great Lakes Res.Div., Inst. Sci; .8t:,%echn. , Univ.Michigan, AnKAOiar. 1967. .; a

23 Simpson, E. H. Measur ement of iversity.Nature (London-) 1949.

. 24. Tanaka, .0.H., Irie, STlzuka, and Koga, F.Nndainent41 investigation on the

biological productivity*: the Northw estof Kyushu. I. The investigation ofplankton. Rec. Oceanolr. W, Japan,Spec. Rpt. No. 5,

/

I

z.4

'Applicat riof aiologicarData

25 !Ta iling, E. Some mesOtroptric phyto-plankton indicators. Proc. Intern.

- 'Assoc. Limnol. 12:212-215. 1955.....

26 Wilhm, .1, L. 'Comparison of somediversity- indices applied to populationsof ben.thic,macroinvegtebOtes in a

/. stream receiving organic wastes. J._` Water Poll. Conti% .ked, 39(10):1673-1,683,

/967.

27 Wilhm, J. L. Use of biomass units inShannon's formula. Etdogy 49 :153 -156.1968.

1'

t IS

4

a.. .I

-

.

.1

.

1

28 Williams, L. G. Possible relationshipsbetween diatom numbers and water -

quality. 2cology 45(4):810-823. 1964..

29 Zelinka, M. and Sladecelt, V. Hydro-,biology fog water management.State Publ. House for TechnicalLiterature, Prague'. 122 p. 1964.

This outline was,prepared by C. I. Weber, .

Chief, Biological Methods Section, AnalyticalQuality Control Laboratory, NER:C, EPA,Cincinnati, Ohio, 45268

i6

Descriptors: Analytical 'Techniques, Indicators

4

o .

a

,

L.

4

,<-. .

I.

O

Q

I

e'

SIGNIFICANCE OF "LIMITING FACTORS" TO POPULATION VA RIA-TION

I INTROCUCTION

A All aquatic organisms do not react uniformlyto the vario4chemical, -phy-ical and' iolokal features in the,ir environment.Through normal cvolutibliary processesvarious organisms,have.become adapted'totccrtoln combinations of environmentalconditions. The successful development,and maintenance of a population or communitydepend uponitarmonious ecological oalancebetween environmental conditions andtolerance of the organisms to variationsin more of these conditions.

. B A factbr whose presence or absence exertssome restraininit, influence upon a populationthrough incomPat;ibility with speciesrequirements or tolerance 4-: said he a

factor. The principle of limitingfactors is one of th'e major aspects of theenvironmental control of aquatic organisms(FigurEl 1).

II PRINCIPLE OF r:IMITING FACTORS'

This principle rests essentially upon two basicconcepts. One of these relates organisms tothe environmenial supply of materials essentialtor their growth and development. The second

-pertains to the tolerance which orgonismsexhibit toward'. n . it uomental LynoitiOns.

1

/UNLIMITED, GROWTH///

DECREASE IN.." -LTiarATIOMNS

&GUI $RIUM WITHN' ONMENt

1"... INCREASE INs,TIMM/IONS

POPULATION DECLINE

'Figure f.

' TIME4

..

The relationships- of lirnitin, factorslei population,gro.:.th' and dcs,olopment.

13 Eo0. 20a. 7.79

180

v.

A Liebig's-Jaw of the Minimum enunciatesthe first, basic concept. In orner for auorganism to intiatAt a pain( u1a envion-ment, specified levels of tiA ateriaksnecessary for growth and de elopment(nutrients, respiratory gases, etc.) tnust

_be present. If one_of these materials isabs'en"- m lb,. It\ ,runmen pt .

in rnn ma, quantities, a given spe(Ieswill oti..y sui rive in limited numbers, ifat all Ileigurr 2).,Coppe, for example,is essential in trace amounts tomam, spet tes.

r

zo.z

10

OPTIMUM

RITI AL RANLOW - MAGNITUDE OF FACTOR - HIGIH

1

.0

Figure-2'. Relationships of environmentalfa( tor:- and the abundam. c of organisms.

1 The subsidiartv principle of fi, tor!aciton stals that high LootArratiott

or availability of 'sornc sunstance, c.the action of some factor in the environ-ment, may ti,o(lif:, of theminttnuri one. For example:

a Tne uptake of-phospitous oy tilealgal Nitzcnia losterium is influenced

the 0- lati vt pia tit ities of nitrate',and phosphate in the elivirourptlitt;'

,, now .:ver, nitrate ut iftZ'at ion ,hppearsto be unaffec:tecrby th,e phersphate(Reid, 1961),

b Tb as..;imilatio,1 of sonic 'algfie isclosely r ateu to temperature.

c The rate ofpxy.,e utilisation,bymay "e affected try many 19,1-ter U1). e

standes or factors in the environment..

21

ti

aa

a Significance of "Limiting Factors" to oputation Variation

a.

S.

d Where strontium is abundant, mollusks.are able to substitute it, to a partialextent, for ealcium in their shells(Odum, 1959).

2 If a material is present in large amounts,but only a small amount is available.forpse/bv the organism', the amount availableandnot-the -total amount present deter-

:mines whether or not tke particular.1 material is liinkting.(calcium in the form

of CaCO3

B Shelford pointed out in his Law.of Tolerancethat there are maximum as well as minimumvalues of moss environmental factors whichcan be tolerated. Absence or failure of anorganism can be 'controlled by the deficiencyor excess of any factor which may approachthe limits of tolerance for that organism

1 (Figure 3).

CONCENTRATION

Figure 4. Relationship of purely harmfulfactors and the abundance oforgani'sms.

Minimum Limit of Range of Optimum ' Maximum Lunit ofToleratg,n ,_

Vof Factors Toleration

stO!,--1- ....e.^,..

Absent Decreasing Greatest Abundance i Decreasing Absent ,

Abunsance e, Abundance

.

Figure 3. Shelford's Law of Tolerance.

1 Organisms have an ecological minimumand maximum for each environmentalfactor with a range inbetween calledthe 'critical rage which represents themange of tolerance (Figure2).. Theactual range thru which an organism cangrow, develop and reproduce normallyis ususWy much smaller than its totalrange aiK.t,olerance.

2 Purely derZte*.ciOds factors (heavy metals,pesticides, etc.) have a maximum,tolerable valut but no qptimuin (Figure 4).

2

iD

e. V, *`,.

.- .

W

3 Tolerance to environmental factors"varies widely among aquatic organisms.

A species may exhibit a wide range'of tokrance toward ote faCtor and anarrow..Lnge toward anothem. Trout,for instance, have a wide range oftolerance for salinity and a narrowrange lot; temperature.

b All stages in the life,historiof anorganism do not necessarily have thesame ranges of tolerance. Theperiod of reproduction is a criticaltime'in theIife cycle of mostorganisms. .

0c The range of tolerance toward one

factor may be modified by anotherfactor. The toxicity of most sub-stances .increases as 'the temperatureincreases.

d The range of tolerance toward a givenfactor may vary geographically within

'the same species. Organisms Thatadjust to local conditions are calledecotypes.'

-

6

Significance of "Limiting Factors" to Population Variation

e The ran,p of tolerance toward a givenfactor may vary seasonally. In generalorganistend to be more "sensitiveto environdiental changes in summer-than in other seasons. This isprimarily due to the higher surqmer

/ temperatures.'

4 A wide range of distribution of a speciesis usually the result of awide range oftolerances. Organisms with a widerange Of tolerance for all factors arelikely to be the most widely distributed,although their growth rate may varygreatly, ,A one-year old..carp, forinstance, may vary in size'from lessthan an ounce to more than a poilnddepending on the habitat.

5. To express the relative degree oftolerance for a particular environmentalfactor the prefix eury (wide) or steno(narrow) is added to a term for thatfeature (Figure 5).

4,

a

3 .

STENOTHERMAL STENOTHERMAL(OLIGOTHERMAL)""ERRAL (POLYTHERMAL)

AihAFigure 5.

TEMPERATURE

Comparison cd'relativelimits otolerance 'of stenothermal andeurythermal organisms.

C The law of the minimum as it pertains tofactors affecting metabolism: and the lawof tolerance as it relates to density anddistribution, can be combined to form abroad principle of limiting factors.

1 The atundance, 'distribution, 'activity"and growth of a population aredeter-mined by a combination of factors, anyone of which may through scarcity oroverabundance' be limiting.

2 The artificial introduction of varioussubstances into the environment tendsto eliminate limiting minimums forsome species and create intolerablemaximums for others.

3 The biological productivity of any bodyotwater is the end result of interaction

a of the organisms present with thesurrounding environment.

III VA LasND USE OF THE PRINCIPLE OFLIMITIK, FACTORS

A The organism-environnt'ent relationshipis apt to be so complex that not all factorsare of ecfual importance in.a given situation;some links of the chain guiding the organismare weaker than others: Understandingthe broad principle of limiting fa.ctors andthe 'subsidiary principles involved makethe task of ferreting out tire weak link ina given situation much easier and possiblyless time consuming and expensive.

1 If an organism has a ide range oftolerance for a fac or which isrela,tiVely constant in the environmentthat factor is not li ely to be limiting.The factor cannot b completelyeliminated from cons eration, however,bfa'ause of factor interaction.

2' If an organism is known to have narrowlimits of tolerance for a factor which isalso vs/viable in the environnfent, that.factor merits caref. study since it.mht be limitirig. //'

.04

I

Significance of "Limiting Factors" to Population Variation

,

B Because of the compleCity of the aquaticenvironment, it. is riot always easy toisolate the factor in the environment thatis limiting a particular population.Prematfire conclusions may result fromlimited Observations of a particular.situations. Many important factors maybe overlooked unless a.sufficiently longperiod-of-time is covered to permit thefactors to fluctuate within their ranges of .

possible variation. Much time and Moneymay be wasted on control measures withoutthe real limiting factor ever being (As"-

covered or the situation being improved.

C Knowledge of the rinciple of limitingfactors may be u ed to limit the numberof parameters t t need to be measure&orobserved for a articular study. Not allof the numero s physical, chemical andbiological par meters need to be measuredor observed 7.r each study undertaken.The aims of pollution survey are not tomake and ob erve long lists of possiblelimiting facto but tip discovq whichfactors are sign icant, how they bringabout their effects, e source or sourcesof the problem, and w control measures.should be takenif

21 -4

O

.

I

9

D Specific factors in the aquatic environmentdetermine rather precisely what kinds oforgt.nisms will be present In a particulararea. Therefore, organisms present orabsent can be used to indicate envion-mental conditions. Theeciiversitroforganisms provides a better inc/cation ofEnvironmental conditansthan doWS anysingle-ape-cies.- Strong ph3)sio-chemical-limiting factors tend to reduce the diversitywithin a community; more tolerant speciesare then able to undergo population growth.

REFERENCES

1 Odum, Eugene P. :Fundamentals ofEcology, W. B. 'Saunders Company,Philadelphia. (1959)

2 Peid, George K. Ecology of Inland -Watersand Estuaries. Reinhold PublishingCorporation,' New YOrk.(1961)

/403 Rosenthal H. and Alderdice, D. F.

Sublethal gffects of EnvironmentalStressors, Natural and Pollutional,on Marine Fish Eggs and Larvae.J. Fish. Res. Board Can. 33.2047-2065,1976.

4 Thorp, James H. and Qib>rke-r\J.Whitfield (editors), ,Energy andEnvironmental Stress in AquaticSystems. Tech Inform .Ctr,U. S. Dept of Energy. 71978.

This outline was prepared by John E.Matthews, Aquatic Bfologist, Robert S. KerrWateg Research Center, Ada,' Oklahoma.

Descriptors: %Population, Limiting Factors

\)

-I, INTRODUCTION -'

O

ALGAE AND CULTURAL EUTROPHICATION

4

A

This top% covers a wide spectrum_ of itemoften depthiding upon the individual discussing

r. the subject and, the particllar situation orobjectives that fie .is trying to "prove "._Since the -writer is not a-biologist, these*viewpoints arev,"from the 'outside-looking in".Any impreSsion of bias is intentional.

t

A Some Definitions are in Order,to Clarify,Terminology:

Eutrophication - a process or action ofbecoming eutrophic, an enrichment.To me, this,is a dynamic pr"ogressionCharacterized by nutrient enrichment.Like many deitiitions, this one is not''precise; stages of eutrophication areclassified as olig-, meso-, and eutrophicdepending upon increasing degree. Justhow a given body of water may be'classified is open to question. Itdepends upon whether you look at quiet6.1; tip-bulent, water, top or'bottomsamples, season of the year, whetherit is'a.. first impression or seasonedjudgement. It, also-depends upon the,water use in which you areintemated,such as for fietting-tewaste discharge.The tisnsitiOnaf'stageS are the majorproblems -. it is loud and clear to atrout fisherman encountering carp andscum.

2 CultureI

Fostering of plant or animal growth;cultivation. of living material andprOducts of such cultivation, both fit.Some degree of control is implied but,the -control may have limitations as .

well as advantage sL Human culturaldevelopnient has fogii-fr human num-bers successfully, but, has promotedrapid degradation of his natural environ-.,tient. e

BI. ECO. hurn. 3.6. "76. .

3.e

3, Nutrients

A component or element essential tosustain life or living organisms. Thisincludes many different materials,some in gross quantities - others inminor quantities. Deficiency of anyone essential item make livingimpossible. Nutrients needed in largequantities include carbon, hydrogen,oxygen, nitrogen, phosphorus, sulfurand. silica. N andoP frequently areroosely considered as "the"nutrientsbecause of certain solubility, con-version and "known" behavior'characteristics.

,

4 Algae

t ,

A group of nonvascular plants, capableof growth on mineralized nutrients withthe aid of chlorophyll and light enerknown as producer organisms, ,sincethe food chain is. based directly or

°indirectly upon, the organic =materialproduced by algae.

B Now that we havelalkacked Alto' itlewords via 'definitions, some of theramifications of eutrophication, nutrientenrichment,, and cultural behavior arepossible. ,

,..

II NUTRIENTS INTER'R'ELATIONSHIPS

A nutrients are interchangeabletin form,solubility, availability,. etc. There areno "end" productg. We caa9 isolate, cover,convert to gas liquid or-seid,. oxidize,reduce, complex, dilute, etc. sometime, some place,- that nutrient mayredycle p.'s part of cultural behavior.

a1 Water contact is'a major factor in

recycle,dynamics just as waterrepresents two-thirds or more of cell

22-1

a

.

J

Algae and Cultural Eutrophicaan

/mass and appears to be the medium inwhich living forms started. Wastedisposal interrelationships (Figure 1)suggests physical interrelationships ofsoil, air and water., The wet apex ofthis triangle is the basis for life. It'sdifficult to isolate water frog the soilor atmosphere - water contact meanssolution of available nutrients.

, /AS ".1Si'OSALINTER:lc:LAI-IONS:11PS

ATMOSPHERE

:-:"\

FOG DUST

4

tWATER t > SOIL

PAUD;k-:'''

2 Figure 2 takes us Into the'biosphere (1)via the sohible element cycle. Thisrefers 'mainly tophosphorus interchange.Phosphorus of geological origin'may besolubilized in water, used by plants or

- animals and returned to water. Naturalrrfovement is toward the ocean. Lessphosphorus returns by water transport. .Phosphorus does not vaporize; hence,atmospheric transport occurs mainlyas windblown du t. Man and geologicalupheaval, parti reverse the 'flow ofphopshoerus tow rd the ocean sifik. .

ti

SGUIDLE ELEMENT- CUL:7.

ATLIOSPHERE

1

LITHOSPHERE \

p

41:04k.o.

.\

0 4 0.0 0 4BIOSPHERE are4i, SeAs

SS.

NO pa0 #i$N,

r .-

3 The nitrqgen cycle starts with ele-Mental nitro in the atmosphere.It candle 'converted to combinedformby electrical discharge, certainbacteria and algae, some plants andby industrial fixation. Nitrogen gasthus may go directly into plant formor be fixed before entry. Derlitrificationoccurs mainly via saprophytes.

(Figure 3) Industrial fixation is a. relatively new contribution to

eutrophication. '

Of.

11r3St1Uts

MI* MIISIllit1111(11

E

A2S/IIIIC1103113

c111:(1

.171--,M__--1.111111111

.---,411111Cit

i.M4U)

M

iIlnitIn

1iMIT° ial:"1l1(111133$

11113.--NI

111(11$MIS

1** IA, AA 4

4 Carbon Con versions (Figure 4) showmost of the carbon in the form of,.

111% geological carbonate (1) but bi,carhonateand-CO, readiily are c'onyerted to plant

, cell maw -end into other life forms:Note the relatively small fractipn ofcarbon in liting mass.

. CA:13 01 c: ;cZIOSPNEIIE , - _

..., :: tinsmite4.4 IBS

Irlt2111'Vet

[4$11 S...... 411

. It ESIN LAIS ,VITTUA141111 "S""1"

.--ks /2 IINSISII,11111 111

$

4s.

71 TIII1131111 MTI, 'S 411 VS

,,A4 EICSAISt

i i III?;111 ;BMX 1111 1111/114\,...1

IASI'SEA

14.311

III SI

SEA

SIII/Ct .

LAMSSBA

SIM ITS10.11111 III

CI

4

B, Nutrient -.Growth Relationships

Nutrient cycles could go on, but, lifedepends upon a mixture of essentialnutrients under favorable conditions.Too much of any significant item in thewrong place may be considered' aspollution. Since toxicity is related tochemical concentration, time of exposureand organism sensitivity, too-muchbecomes toxic. If it happens to 6e toomuch growth, its a result of'eutrophication.'How much' is generally more importantthan op 'what'? Botli"natunal and manmade

..processes lead to biolbkical conversions,to pollution, to eutrophicatiOn and to

111 toxicifye Man is the only animal that can ,

concentrate, speed up, invent, of otherwisealter these conversions to make a collossalmetes.

1 Life forms hie been...fpirpulated,iriterms of elemental or htitrient com-ponents 'many times. The simplest is;C irp N. A more complex formula?,

5 .2..is C8 0 N Ca Cl P CuF100 76 80 20 6 7 2 - 2

SilftMn2IcNaS21Zzi. This includes

Al e and Cultural Eutto hication

18 elements. More than 30 have been' implicated as essential and they still

Would not, "live", unless they were'correctly assembled. As a nutrientMnemonic H. COPKINS - - Mg(r)-CaFe-MoB does laitly well. It alsoindicates Iodine 7 I, Iron-Fe,MolybdenAm-Mo, and Boron-B thatWere not included earlier.

2 The Law of Distribution states that"Any given habitat tends to favor allsuitable species - any given speciestends to be present in all suitablehabiiats." Selection tends to favor'themost suitable species at a given placeand time.

3 Liebigs Law of the Minimum, statesthat "The,essential material availablein amounts most closely approachingthe critical minimum will tend to bethe limiting growthfactor."

4 Shelfords law recognizes that therewill be some low concentration of anynutrient that will not s nt growth.Some higher concert atio will stimulategrowth. Each nutrient will have somestill higher concentration that will bebacteriostatic or toxic. This has beendiScussed earlier but was considered ina different manner.

HI BIOLOGICAL PROGRESSIONS

The biological "balance" appears to bey. verytransitory condition in cultural behavior,Man favors 'production. A steady slate"balance" does not persist very long unlessenergy of the system is.too low to permitsignificant growth. A progression-of specieswhere each predominent form thrives for atime, then is displaced by another tempgrarilyfavored group is usual. Yearly events in the-lawn start with chickweed, then dandelion,plantain, crab grass, rag weed, etc., in .

successive predominence. Occasionally,more 'desirable grasspg may appear on thelawn. ,,prass is a selected unstable "culture".

1 8C

f

3

...%-"

Algae and CulturalEutre?phication

A Figure 3 shows a biological progression (2)following introduction of wastewater in anunnamed stream. Sewage or slime baCteriaproliferate rapidly at first followed by

6.

o

ciliates, rotifers, etc.

.16

CL 300

3.

2 200CCWa.

foo

.7

COLIFORM(NO.rER MI.)

THE BIOTASEWAGE BACTERIA

NO:PERIM:

c:).CILIATES

NO.PER ml.x1,000. ROTIFERS. &CRUSTACEANS

NO. PER ml.x 25,000

1 3 4 5 6 7 8 9DAYS

24 12 0 12 24 36 48 60 72 84 96 108MILES

Figure 5. qactetio thrive and finally become prey of the ciliates, which in turn are food for the atifers and crustaceans._ .

B Figure 4'shows another'progression ofbottom dwelling larva. Here the sequenceof organisms changes after'wastewaterintroduction from aquatic insects to sludgeworms; midges, sow bugs and then tore7es lishment of insects.

(

4

L

THE BIOTASLUDGE WORMS ,

TIC INSECTS .

ttl_.. DUOS (X 30)

"...FA I: Se 6.

I I 014g I IniS41

4 $ I .1 1 f

11 11 It /4 1MILES li 72 14 14 '

. .

Figure 6. The gonfalon curee,,_44,,FIgure 7 is composed of a 'series of maximaId, Individual species, each math:404 and, dying off as stream conditions spit.t

N.- . . .

A 87. -

..'


C Another progression afier waste .

introduction changes the biota froman algal culture to sewage moulds withlater return to algalpredominence.

,

\CM

A

t

figure ?shortly after sewage discharge, the moulds attilin maximum growthese ore assooted with sludge deposition shown in the fewer curve. The sludge i

decomposed gradually; as conditions clear up, algae gain a foothold and multiply.

.Figures 5, 6, and 7 are shown separatelyonly.,6ecause one visual would be Unreadabl%wiith'all possible progessionson it. There'

eare progresips For fungi, protista, insectlarvae, worms, fish, algae, etc. Each

_SpIt,it,cannot compete-successfully, it willBe replaced by.those that can compete.under prevailing conditiOA at the tit'ne:ConditionS shift rapidly with rapid growth.

4IV :ilhe inter ctio pf bacteria or fungi andalgtOrre 8) are particulgrly ,

sigpffic .to eu.trophicAtionj

A The b cteri,a orFr the saprophytic group"amore hem tend to work on preformedbrga .materials -. pre-existing organicsfro dead or less favored organisms.

. Alg 1 dells produce theyorganics fromlig t ...eArgy chlortppyll and mineralized

. ntlt netts. This is a happy combinationfo both: The algae release the oxygen

,4 use by the bacteria while the bacteria.. . . algae.elease the cOlvneeded by theA.,

4

,

Since the algae also acq ire CO2 fromthe atmosphere, f rbin ewater andfrom geologital sources, it always endsup with moire enrichment of nutrients inthe water - more enrichment means more

ro h and rowingtorganisms eventuallyclump.ansi deposit. The nature of growthlifts om free growth to rooted forths,starting in the shallow,s. Anotherprogression occurs (Figures 9 and 10).

, 183 .

It is this relationship that favors profusenuisance growth of algae below significantwaste discharges. There is a tremendouspool of carbon dioxide available ingeological formations and in the airsTranSfer to the water is signiant andencourages algal productivity ar4 eventualeutrophication of any body of water, but,this does not occur as rapidly as when thewater body is super saturated with CO2from bacterial decay of wastewaterdischtrges or benthie deposits from them.

_r5

r (*.


I'

LAKEZONATIA N

AQUATIC PLANTS

II LITTCM,ZIAL--

LIMNETIC

/

rzo FU NDAL

DO

3

I ik )° ALGAE..SEWAGE

BACTERIADEAD .

DEAD

. 0

0

OXIDATIONBY.

° BACTERIA LIGHT

6

/"----MINERALS

M `g0

MIRA 'MUG.189

;),;:""\

a

4


PITIORAL ZONE, AND,APUIVITIC-PLAHT DEPTH

10 METERS

20 me

,40 m'

70m*

SEED PLANTS

FEROS.

CHARAPHYTES

B Nitrogen arid phosphorus ate essentialfor growth. ,They also are prominently.considered ineutrophication control.Algal cell.ma't4s is about 50% carbon, 415% nitrogehind appoximately 1%phosphorus not considering luxury uptakein. excess of immediate use. Phosphorusks considered as the most controllable*niti.rig nutrient. It's control is corn-

, plicated by the feedback of P frotn.tenthicsediments and surface wash. Phosphorusremoval means solids removal. Goodclarification is essential to obtain goodremoval of P.- This also means improvedremoval of other nutrients- a majoradvantage of the P removal route. Both ,N & P are easily converted from one formto another; most form are water

V SUMMARY

Control dfteutrophication is npt entirelypossible. takes must eventually fill with

* benthic sediments, silt-Lace wast and'vegetation. Natural processes, eventually.causefilling. Increased nutrient dischargesfrorR added activities grossly increase fillingrqte.

a

LAD5PHORA

o

A We produce more nutrients per capita per"'day in the United States than in othernations and much more today than 100years ago. More people in populationcenters accentuate the problem.

B Technology is available to remoZ mostof the nutrients from the water carriagesystem.

C

1 This technology will not be used untesswater is recognized to be in sho'rtsupply.

-. .2 It will not be Used villep.we'PfaCe

realistic dommodity value on the waterand are willing to pay for, cleanup forrense,putposes. 4....

Removal musebe followed by isolation ofacceptable gases to the atmosphereacceptable solids into the spa.....for ienseor storage. Water contact cannot beprevented, but it must be limiie4 oltheenrichment" of the water body is hastened.

4-0190,

6

A

Al ae and Cultural Eutro shicatio4

REFERENCES

1" A collection of articles on e,BiosphereSci Am. 223'':(No.,'3) , jP. 44-208.

-- . ember 1970. 0-, i'l04,

2 Bartch, A,. E. and Ingram,', . M.Stream life and the P-Plluff6n, Environ-ment. Public Works 90:(NO7,) 104--

-July 1959. ,. '',4-'

j.

'A

o.

A

?...k V.Tt,

4'S

1. R

"

'03'; 044 .

ThiS outlin was prepared by F. J. Ludzack,Chemist, National Training Center,

VD, OWPO, USEPA, Cincinnati, Ohio45-

criptors: Algae, Eutrophication

#

.

-

et.

V #

,

9

.0

Nc(. , 44 ,

,

igraI

192

.

,,z

.

4

0

tr

1r

#,

,.9%

-

4 .. JL, !.. It* :4.

t r.

. .4p

5.

6

6

(c

..Is

IA s UV,41.4 -4 4

eis.s °-1.;

°

CALIBRATION AND USE OF,.PLANKTON COUNTING EQUIPMENT

I INTRODUCTIONo

_. '. ''-A With the exception of factory-set ..

instruments,' no two microscopes can .be counted, upon to provide exactly.the

. same magAificat4n witfi ay given com- 9.

bination-of ocuaarl and objectives. For - 4a .0. accurate quantitative studies, it is.there- .

, fore necessary to standardize,or "valibrate"each.instrument against a. known standard:scale. One scale frequently used is aMicroscope slide on which two millimetersare subdivided into tenths, and two addi-,-tional tenths'are subdiviaed into hIndredths."Figlure 3.

. I

Another pracedureis to select,a valueSor each of the variables involved, (inter-pupillary distance,-t oOm, etc.) and,calibrate the scope at that combintion:Then each time the scope is to be used for',quantitative work, re -set each,variableitdthe Value selected. A separ.ate multipli-cation factor must be-calcalated for eachadjustment which changes ,the magnificationof'the insfronent. is.

.. . ..

Since the Whipple Square can be used tomeasure both linear dimensions and ;square areas, both shtild be recorded onan appropriate form. A. suggested format

`,. is ,shown in Figure 6. : ", ,

. ..

(Data written. fn are used asan illustrationand are not intehded to apply to anyPareicular.microicope. An, unused form .is includeckas Figure 6-A: ),.. .

It . i A ''' ,1e

S.i 4. t

,THE CALIBRATION 'PROOEDUR:E ...4

. . , . ... 1,6 , t,

.Installing the Whipple `Square or'Reticuie,.; . o 41 ... .44' To install the-Fetionleip the'dcuti,r.

(Usually-the right one on a binodnWr --:.

microscope)4 carefully unscreVhe'upper .,.* e moun ng a .p e re leg e on

B In orcier to provide an accurate measuring. /.device in thwmic'roscope, aWhipple

'Plankton Counting Square or reticule(Figure 2a) is installed in one ocular(there are many gifferent types of reticules), -0-

N. This square i8 theoretically of such p. siret that with a 10X objective, a 10x ocular,

andla tube length of 160 mm, -the image 4 v.- Uof thesquare covers a square area on the--slide one-mm on a slide: Since this A

4 objectiirg is raceItattained, however, IrnostAwtcPosco, apes must b4 standardized.or 41' do.,

4141e at ea". as described bei,low in orzter, ,

to ascertain the actual size,or the,Whipple ,.;

,

,Square, a seen} throw litte microscopes(herein tet'iefeilre q as'the'tWhipple4eld"r. prTkreseis's,Chematically,-

c. 'reprzlte'd Figt2rfts-5 and'7. If the" Whipple eyepiece is to be used at more 4P

than (Inb. magnification, it 'must be rgcali;' ,orated for each. A basic tYrit of monocular.- microbcope.is shown in Figure 1.

t

. C Microscopes with two-eyepieces (binocular)are a convenience but not essential.' Likemodtrn car3-Ithey are not only great f"performers," but also complicated' toservice or, in this instance, calibrate. ,On some instruments, changing the inter-

---pupalary distance also changes the tubelength, on others it does not. The "zoom"featureon certain scopes is also essentiallya syste'm.for.changiug the tube length.

The resultant is that inddition to calibra.-tion at each combination of eyepiece andobjective, any otherf actor which may.i.

' affect magnification must also betconsidered- In some instances this may mean setting up

a table of calibrations at- a series of micro-:,scope settings.

'4.1 rut ti rid lace th t 1

th$ dircular diaphragm or shelevihichwill ,

be foul* af roxiinatery half way loy.tinside' r(f'igare`*4): eplace the mo,untl.ng &Fickobserve thoCha k* gs on'the reticule. Ifthey are not ih sha p foc s, remove and ".turn the reticule o r

On

., .

reticules with the.markings etched-onone side of a glass disc, the etched sur-face tan tisually,be recognized by shitliqgthe disc at the proper angle in a light.The markings will usually be in the beat

, focus' with the etched surface awn:. If 'themarkings are sandwiched between tvp,g14.as.discs cemented togefher, both aides ateaiSe, and the foCils may not be quite assham d

.B Obdervation of the Stage Microingir

V.

.

Replace the ocular in the- Microscope andtobser4e the stage mitrometer as 's illus- '

' trated schematically in Figure 51* Calibration

,sr.

tqvt61'. miC. id:-6.76 192,

of the Whipple Squar.e. On a suitably ruleddrin`stith as the one illustrated' Figure 6,Calibration Data,' record the°aCtual distance,_in millimeters subtended by the image of

201. c.

q

. ;

alibration and Use of Plankton Cbunting Equipment

AI

the entire WhiPpletfield and also by eachof its subdivisions. This should bedetermined for each significant settling of,the interpupillary distance for a bin,ocularmicroscope, and also for' each combinationof lensesipmp/oyed. Since oculars andobjectives marked with identical magnifi-cation, and since microscope frames toomay differ, the serial or other identifyingnumber of those actually calibrated shouldbe recorded. It is thus apparent that thedeterminations recorded will only be validwhen used With the lenses Vted an, on thatpdrticular microscope.

C Use of the 20X Objective..

Due to the short working distance beneatha 46X (4mm) objective, it is impossibleto focus ,to the bottom. of the,Sedgewick-Rafter plan)cton,counting cell with this'lens.A 10X (16mm) lens on the other hand"wastes" space.between,the front of theleris ..nd the coverglass,, even when focusedon the bettom o` Thee In order to takeIre m-oseteffickenruse possible of this cellthen, an objective of intermediate focal0length is'desirable: ,A lens with a Locallength of approximatelr8 mm, having amaerlification of 20 or 21X will meet thesereqtairements. Subh lenses are available.from American mantfacturers and arerecommended for this type of-work,

. III CHECKING THE CELL

The internal dimensions of a Sedgewick-Rafter%*plankton counting celL8liciuld be 50 mm'longby 20 mm wide by 1 rnm7cleep (Figure 8). . , r

The actualhorizontardimensions of each newcell should be checked With calipers, and thedepth of the ceA checked at several pointsaround the edge using the vertical focuSingscale engraved on the fine adjustment knob of

.most microscopes. OrPe complete rotation ofthe knob usually raises or lowers the ohjeqtive1 mm or I00 microns (andspach single ina.Atequals 1 micron). Thus, approximately tenturns of the fine4adjustment knob shotild raisethe focus from the bottom of the cell to, theunderside 'of a coverglass testing on the rim.Make thesegmeaswicements-on an empty cell.The use of a No. I or-l-r/2,i24 X 60 mmcoverglass is recommended rather than the'heavy coverglass that comes with the S-Rcell, as the thinner glass will somewhat con-,form to any irregularities of the cell rim(hence, also making a tighter seal and reduc-ing evaporation when in actual se). Do notattempt to focus on the upper s' face of the

234

rim of an_empty cell fOr the above depthmeasurements, as the coverglass is supportedby the highest points of the rim only, whichate very difficult to identify. Use the averageof all depth measurements as the "true" depthof the cell. To simplify calculations below, "it, .will be assumed that we are dealing with a _

cell with an average depth of,exactly 1.0 mm.

IV

A

B

PROCEDURE FOR STRIP COUNTS USING'THE SEDGEWICK-RAFTER CELL

Principles

Since the total area of the ce'l is- 1000 mm2,the total volume is 1000 mm or 1 ml. A"strip" the length of the cell thus constitutesa volume (Vi) 50 mm long, 1 mm deep, Aridthe width of -the Whipple field.

The volume of such a strip in riffn2 is:

I

50 X" width of field X depth,

50 X w.X

56 w

In theigexample.given below on the plateentitlffd Calibration Data, at a magnificationof approximately 200X 'with an ,interpupillarysetting of "60", the width of the Whipplefield is recorded as 'approximately 0.55 mm(or 550 microns). In this case, the volumeof the strip is:

V1= 50 w =° 50 >n.55 27.5 (mm3)

Calculation of Multiplier F, actor

In Drder to convert plankton counts perstrip to counts per ml, it is simply,.,necessary to multiply the count obtainedby, a factor (F1) which represents thenumber of times the volume of the stripexamined (V1) would be contained ink 1 ml or1000 mm3. Thus in the example givenabove:

F,1 = volume of cell in mm3vo ume e75.1717-01.rimml-

1000 100027.5 36.36

= approx. 36

If more than one strip is to be counted,the factor for twd, three, etc. , /stripscould be calculated separately using thesame relationship' outlined above, changingonly the measurement-for-the length of

1

a

Ca,libration and Use of Plankton Counting Equipment

G

Fig req.. THE COMPUUA) coarse adjustment; 33) fin adjustment;C) arm or pillar; D) mecha cal stage whichholds slides and is movable n two directiorisby means of the two knobs; ) -pivot or joint.This should not be used or'' rokenll whilecounting plairkton; F) eyepie e (or ocular cf:figure 4); G) draw tube. T k will be foundon monocular microscopes nly (those havingonly one eyepiece). AdjOs ent of this tubeis very helpful in calibradn the microscopefor quantitative cotutt.ing (Se 5. 5. 2. 2. ).H) bddy tube. In some mak s of microscopesthis can be replaced with a body tube havingtwo eyepieces, thus maldrig the 'scope intoa."binocular. " I) revolving nosepiece onwhich the objectives are m unted; J) throughM are objectives,, wily one if which can be .

MICROSCOPE .

tuKned toward the object being studied. Inthis case J is.a 40X, K is a 010X, L is a 20X,and M is a lOX objective. The productof

. the magnification power of the objective being.used times the magnification power of theeyepiece gives the total magnification of .hemicroscope. Different makes of microscopesemploy objectives of slightly differentpowers,but all are approximately equivalent. N) stageof the microscope; 0) Sedgwick-Rafter cell inplace for observatiori; P) substage condenser;Q) mirror; R) base or staid; note: forinformation on the optibal system, consultreference 31 (Photo by Dbn*Moran. ).

fr

Calibration and Use of Plankton Counting Equipment

Ar

I

a

a

I1

Figure 2

the margin of the large square to facilitate'the estimation of sizes of organisms indifferent parts of the field. (b) Quadrantruling with 8. 0 mm circle, for counting.'bacteria in milk smears for example. (c)Linear scale 5. 0 mm divided into tenths,.

ent of linear diniehsions.(d) Portohr icule for estimating the sizeof pdrticlese.- The sizes of the series of discsis based on the square root of two so that the-areas of successive discs double as:they

. progress in size.

Types ,of -eyepiece micrometev,discs orreticules (reticules, graticules, 'etc.).When dimensions -are mentioned in thefolloWing description, they refer to thpmarkings on the reticule disci and not to.the inpapurements subtended on the micro-scppe Slide. ,The latter must be.determinedby calibration procedures such as thosedescribed elsewhere. (a) Whipple planktoncounting eyepiece. The fine rulings in thesubdivided square are sometimes. extended to

23-4 ,

b

o 0

. t. ) ( (

m11111101

d

Jil

. . rstrip counted, Thus or twostrips in theexample cited above:

0

V2= .100W = 100 X 0.55 = 55 mm31000 1000 18. 2F2= 55V1

.1? .,

It will however be noted that F2

=

I-Likewise a factor F3 for three strips

Flwould equal 7 or approxima ely 12, etc.

Calibration and Use y Plankton Counting Equipment

C\' An-Empirical "Step-Off" Method

A simpler but more empirical procedurefor determining the.factor is to considerthat if'a. strip 20 mm wide were to becounted the length of the cell, t13,9.t theentire 1000 mm3 would lie included 'sincethe cell is 20 mm wide and 1"mm.deep.

Thus in the example cited aboire where atan approximate magnificiation of 200X andwith an interpupillary setting of 60, thewidth of the Whipple field is .55 mm. Then:

.0F1

= 36.36 or approx. 36-5

above)'

If two strips are counted:

. .55

1020 ,and F2 = ITT = 18. 2 = approx. 18, etc.

his same value could be obtained without,withoue use of a stage micrometer by refullyoving the cell sidewise across the field

f vision by the use of a mechanical stage.,Count the number of Whipple fields ih thewidth of the cell. There should be ap)roxi-

ately 36 in the instance cited above.

This 20 mm strip width can be equated to1000 mm3. If a strip (or the total of 2 ormore strips) is less than 20 mm in width,the quotient/of 20 divided by this width willbe a multiplier factor for converting fromcount per strip(s) to count per ml.

SEDGEWICK-RAFTER LEPARATE COUNT USING THE

A Circumstances of Use

The use of concentrated samples, local'established programs, or other circumstantes

Figure 3. STAGE MICROMETERThe type illustrated has two millimeters divided into tenths, plus two additionaltenths subdivided into hundredths.

1mmLI]

Enlargement of Micrometer Scale

,r.

. '23-5

Calibration and Use of Plankton Counting Equipment ,

.

a

*,

0

CALI limn ON OF WHIPPLE SQUARE

as seen with 10X Ocular and 43X Objective(approximately 430X total magnificatiOn)

f'

Wh;PpleSquare asseen through ocular:"

r ("Whipple field")

L

4'

"Small squares" subtendone Ufth of large squares..005r trim or 5 2p

I\"Large square" subtendsone Tenth of entire WhippleSquare: :026 nun or 26p

Apparent lines of sightsubtend .26 mm or 260pon stage micrometerscale

.01,mm(10p)

PORTIOltripf MAGNIFIEDIMAGE OF STAGE MICROMETER SCALE --10.

Min "IP\(1000

, Figure 5

CALIBRATION OF THE WHIPPLE SQUARE .

, The apparent relationship of the Whipple'Square is shown as it is viewed through amicroscope,while looking at -a stage.

11

micron ethr with g magnification ofapproximately "430X (10X ocular .and 43Xobjective).

r,19.0 23-7

ti

Calibration and Use of4PlanktOn Counting Equipment

MICROSCOPE CALIBRATION DATA

Mictoscope No. 6,q3 79

%.

TubeApr "oximate Length, or

.11k41.gnification InterpupillatiirSetting

Linear dimensions of Whipplesquares in millimeters*.

100X, obtained with

Whole Large Small

Factor forConversionto count/ ml

(2 S-8 Stripe)ObjectiveSerial No.

5,710 9,Vaol)

r ,

Sp I. /30 O. 11.3 0, oc226 8.9and OcularSerial No

A:194.74'AI)

, 60 /, / / 5- 0, /I I

0.1/0

Nj5 o.ta,2,

'0. 0;.2,2

9,o

9. /70 I. /DO

. ,

200X, obtained with 6 (2 8-8 Strips)ObjectiveSerial No.

'ilgfV.PI..nd 6-0 0.500 0 05:4 0,0 //a /7. 9and OcularSerial No.

(2967gLGox)

(,0 0.550 0 055 0.0//0 /.19. 2

70 0.5Y5 0 nsg "00/09 /1.3, .

400X, obtained with(Nannoplanlcton).(cell-20 fields )

ObjectiveSerial No.

,

..

2.19/134/49X) i i D ; .005 .and OcularSerial No 0 , . 0..

.005 .,lo..

',1967.9Z00,0 , o 05 AT .

..

4

.

*1 mm z' 1000 microns

11V1icroscope calibration data. The formshown is suggested for the recording ofdata pertaining to a particular microscope.Headings could be modified to suit local

,

Figure

.

situations.' For example, "InterpupillarySetting" could beyeplaced by "Tube Length"or the "2.S-11 Strips could be replaced by"per field" or "per 10 fields."

I

Ir

F

Approximate_Magnification

4

Calibration and Use of Plankton Counting (Equipment .

MICROSCOPE CALIBRATION.DATA

Tubel_iigth, or

InterpupillaitySett ing

100X, obtained With

Linear dimensions of Whipplesquares in millimeters*.

whole if Large I Small

Factof for .

Conversionto count/ ml

(2 S-R Strips)ObjectiveSerial No;

,

' .

..

.

.

.

and OcularSerial No .

.,

.

..

.

. . . .

200X, obtained withObjectiveSerial No.

Wn7ii

Serial No.

(2 S-R Strips)

400X, .obtained with1

(Nannoplankton)(cell- 20' fields )s _

ObjectiveSerial No.,

.

-.

.

..

.

. . .

.,

...:

. ,

and OcularSerial No.

. ,

,

.ice..:

.-

..

. . d

lrnm = 1000 microns

FigUre

AQ, pl. 8. 10. 60,

MICROSCOPE CALIBRATION DATA

Suggetted work sheet for the calibration'of a microscope, Details will need to be adaptedto the particular instrument and situation.

4:-

yr

a

a

(.. .. .Calibratfon and Use of Plankton Counting Equipment

.

S-A COVERGLASS .14Bilir WHIPPLE SQUARE

WATER INS-R CEJL

I -1MM A

PLANKTERS

'/InAnn*,\,,.\s\% A

\; 4-\\

I-THICKNESS S-R SL1DE

01.

. Figure 7

A cube of water as seen through a Whipple square at 100X magnifitation ina Sedgewick-Rafter.cell The figure is draWb as if the micro cope werefocused on the bottom of the tell, making.yisible only th'ose organisms lyingon the bottom of the cell. The little =bug (copepod) halfway up, and thealgae filament at the top wataldbe out of focus. The focus must be rhoved upand down in order to.study,lor count) the entire cube.

0

2 0°1 n O

r

11

Ca14 ration and,Use of Plankton Coun nig..Equipment'

may make it necessary to employ'the mareconventional technique of counting one ormore separate Whipple fields inste of .the strip count method. The basic re tion-ships outliqed above still hold, namel

'volume cell in mm3mF /-\volume examined in n--73

F

B Princ1 ies Involved

The volu e examined in this case willconsist of one or more squares the dimen.?,_sionsof the Whipple field in area and 1 mmin depth (Figure 7). Common practicefor routine work is to examine 10 fields,but exceptionally high or low counts orother circumstances may indicate thatsome other number of fields should beemployed. In this case a "pr field"factor may be determined to be subsequentlydivided by the number of fields examinedas with the strip count. The followingdescription however is based on an assumedcount of 10 fields.

1

,C Calculation of Multiplidr Factor

As stated above, the total volumerepresented in.the fields examined con-Asts of the total area of the Whipple fieldsmultiplied by the depth.

V4= (side of Whipple field)2 X depth-

(1 mm) X no. of fields counted)

f

For example, let us assume an approxi-*mate magnification of 100X (see Figures -6 and 7 and.an interpupillary setting of"50". The observedleneth of one sideof the Whipple field in this case is 1:13mm. -The calculation of V

4 is thus:

side 2 X depth X no, of fieldS

1.13 X 1.13 X 1.5< 10 = 12.8 mm3

V4 =

The multiplier faCtor is obtained asabove (Section IV ,A):

volume cell in mna3 .

4 3vol xaminecl in mm.i0

8(approx.) 78

( one field were counted, the factorwo d be IL, for 100 fields it wouldb e ? . ) . )

0

.`

NANNOPLANKTON COUNTING

For counting nannoplankton using, the highdry power (10X ocular and 43X objective)and the "nannoplankton counting cell"(Figure 9) which is 0.4 mm deep, a minimum

of 20 separate Whipple fields is suggested.The same general relationships presentedabove (Section IV)-can be used to obtain a.multiplier or factor (F5) to convert countsper 20 fields to counts per ml.

To take another example from Figure 4, atan approximate.magnification of 400X and aninthrpuPillary setting of 70 (see also Figure .3)we observe that one side of the Whipple fieldmeasures 0. 260 mm. The volume of thefields examined is thus obtained as follows:

side2 X depth X no. bf fields' 4

. ,

=5

36= 0.26 X 0.26 X 0.4 X 20 = .54 mm

and F 1 0005 = (approx. ) 1 .850 '

It Should be nbted that the volume of thenannoplankton cell, .1 ml, is of-no significancein this particular calculation.

REFERENCES i1 American Public Health Association, et. al.

Standard Methods for the Examination/ of Water, Sewage, and Industrial Wastes.

14th Edition, 'Am. Public Health Asgoc.New York. 1976.

2 Jackson, H. W. and Willis s, L. G.The Calibration and Use of CertainPlankton, Counting Equip. enta Trans.Am. Mic, Soc. LXXXI(1tk:96-103. 1962.

3 Ingram; W. M. and Palmer; C. M. , .

Simplified Procedures for Colle4ing,Examining, and Recording Plankton in

:Water. Jour. Am. Water Works.Assoc. 44(7): 617-724. 1952.

4 Palmer, C. M. Algae in Water Supplies.. U. S. D. H. E. W. Public Health Service ..C`

Pub. No. 657. 1959.

5 Palmer, C. PI,. and Maloney; T. E. ANew Counting Slide for Nannoplankton.American Soc. Limnol and Oceanog.Special Publications No; 21. pp. 1-6.1954.

-

20°

A -

23-11 *,*

rs

-,

. ---Calibration and Use of Plankton Counting Equipment

O

AreaUncounted

I litrips,,Counted

1,--,\ Figure 8 . ,. \. ,

Sedgewick-Rafter counting cell showing bottom scored across fdr ease in countingstrips. The "strips" as shown in the illustration simply represent the area counted,and are not marked On the slide. The conventignaPdi,mensions are 50 X 20 )< 1 mm, bufthese should b'e checked for accurate work.

,

_

/

:Jr, .t rrrrrr-7.1en r.

f:(Ii

Figure 9t

at /Nannoplanktpn cell.* Dimensions of the circular part of the cellare 17.9 mm diameterX 0.4 mm depth. When covered with a coverglass, the volume contained is 0. I ml.The channels for the introduction of sample,and the release of air are 2-mm wide andapproximately 5 vim long.. This slide is designed to be used with the 4 mm or 43XThigh dry) objective.

'6 Welch, PaulBlakiston1948.

7' Whipple, G.Whipple,

.0.23..12

4

S. Limnological Methods,Company. Phila. T,oronto..

49±7

C., Fair, G. M., andM, C. The Micrqscopy of

.$

Dyinking Water. John Wiley and Sons.New York. 1948.

,, This Aline was prepared by HI W. Jackson,forrrier Chief Biologist, National Training

9 Center, and revised by R. M. *Sinclair,4quatic tiologist, National Training CenterOTD, OWP01.USEPA, Cincinnati, Ohio 45268

.

. 4

Descriptors: Plankton,

:.203

Microscopy

. s``

,

a

LABORATORY: PROPORTIONAL COUNTING OF MIRED LIQUOR

. .. ,

I OBJECTIVE

To learn the techniques of propoftional8ounting of mixed liquor, samples andbecome familiar with cornmon typesof microorganisms.

II MATERIALS

A Mixed liquor samples, each con-. taining a number of microscopic

forms.

'B Glass slides, cover slips, and droppingpipets. ,

III PROCEDURES

A Make a wet mount of the sample(s)proviaed./Do not alloy,' the sliltlemountto evaporate. Add a dropto the Side as necessary.

13 First, scan the slide. On the sheetmarked, "Conimon types of Protozoa

. and I\ietazoa ", make a check next toeach type you find. If you find,,atype not illustrated'ipake a simple-sketch on the .reverse of the above ,sheet. The objective here is tobecoine familiar with alla of the .

Common types of, microorganisms4,) found in the sample. .

7..

,C Proportional Counting

1 Fifty- count0

A

a Moving the slide at random counteach type, until a total of 50organisms have been "jo.9kted:

q b Tally the results and compute thepercentage Of each type.

IV RESULTS

A Record your results on the board. 4-

B Discuss the methods and the use ofthe proportional count results.

This outline ,wfts prepared by Ralph. Slnclair,,National Training and Operational TechnologyCbnter, OWPO, 14SEPA, Cincinnati,'Ohio 4526 .

\,'Descriptors: Analytical TechniqUes,Activated SludgeS

44

o .4

0

.4

X laboratory: propor,i6nal CoUnting of Mixed Liquor

o .

ESTIMATING THE SIZE OF 'A PROTOZOAN

o

10x OBJECTIVE10x EYEPIECE= 100 <x TOTAL

TRACHELOPHYLLUM PUSILLUM

v

24-2

e.

.

40x OBJECTIVE:10x EYEPIECE5,--400. TOTAL.

4

.

O

Laboratory: Proportional Counting of Mixed Liquor

COMMON TYPESOF,PROTOZOA AND METAZOA

ROTIFER - For size comparison with other

Ki514microorganisMe

Aspidisca111

I.'.

17,1

,E5Crawl' ng

STALKED CILIATES

Epistylis I.1 9

PER ITRICHSLateral

2ta 11z

NAKEDCC.

0Lucc

AMOEBAE FLAGELLATES

4:10-t OTHER PROTOZOA

rvrPosterior

12

13

c.) 1

't0

80

C

telotrochstage

NEMATODE

14Chironomus

WORM-LIKE MULTICELLULAR ANIMALS .

I

4-3

7

Laboratory: Proportional Counting pf Mixed Liquor

PROPORTIONAL COUNTS --PROTOZOA AND METAZOANS

Count Organisms in random swoop across slideDo not search for #articular typosIgnore rare organisms ``"

. Stop at total count of oither 50 or 100 organisms.

Tally each organism twice, once ina total taffy box, nd once in theappropriate tally box below

Paramecium

Trachelophyllum --ry6

5

DateAnalystPlant

TOTAL 50-0. TOTAL 100

1

r.

r?-Aspidisca

1.

Epistylis

Imp

.

."T

WI*

10

Vaginicola

tel otrochs

11

-96.

15

Pleuromonas

Amoebae

4

207

7-% Aelosoin a719

ROTIFER6

;


PROPORTIONAL COUNTS -

PROTOZOA AND METAZOANS°

Count organisms in a random sweep across slideDo not'search for particular typesIgnore rare organismsStop at a total count of either 50 or 100 organisms

Tallt each organa total tally box,appropriate tally

dun twice, once inand once in thebox below

DateAnalystPlant

TOTAL 50

9 1TOTAL 100 Law-

r.

1

2

Trachelophyllum

-14

6

Aspidisca

1N

a.

10,

Vaginicola

telotrochs

-36

15

Pleuromonas

Ionotus-

/Stentor

-34

MMIMMINO

1=IMMMI

IMMIAMM

Epistylis

IMMIMMIM

2033

Im10..0

19.

ROTIFER

M.m.=

IN= =ION

Im=

-34=IP

11imi

1s

A

%

it ).

Laboratory: Proportional Counting of Mixed Li. , .

1

A

PRO,PORTIONAL COUNTS -PROTOZOA AND METAZOANS

t..t

Count organisms in a random sliver" across slideDO not search for particular typesIgnore tars organismr=-Stop at a total count of either 50 or 100 organisms

Tally each grganIsm twice. once ina tole, tally box, and once in theappropriate tally box below

DataAnalystPlant

TOTAL SO

1

C

TOTAL 100 --aw

IParamecium

2

n*.

Trachelophyllum

l

)

Litonotus

Stentor

6ti

Aspidisca

0.

telotrochs'

11

15-

Pleuromonas

Epistylis =44

.209.

ROTIFER

,111011.111110

1

0 1

. ,

Laboratory: PropOrtional Counting of Mixed Liquor:

, I

<55

PliaiPORTIONAL COUNTS -PROTOZOA AND METAZOANS

Count organisms in a random sweep across slideDo not search for particular typesIgnore rare organismsStop at a total count of either 50 or 100 organisms

7'Tally each organism twice, once in

total tally box, and once in theappropriate tally box below !")

DateAnal*Plant

TOTAL 50

Paramecium

"Iv

TOTAL 100 --ow

6;NA

-

Trachelophyllumtelotrochs

-16

Pleuromonas

1611

1

/

Amoebae

4

Stentor,"_ Epistylis

210

Aelosoma


r

PROPORTIONAL COUNTS.-PROTOZOA AND METAZOANS

Count organisms in a random swoop across slideDo not search for particular typesIgnore-ram organismsStop at a total count of *idler 60 or 100 organisms

Tally each organism twice. once ina total tally box, and once in thiappropriate tally box below

DateAnalystPlant

TOTAL 60

Peranema

trachelophyllum

3.

Pleuromonas

4Litonotus Amoebae

%.

1.1

.4.1

°soma

Epistylia'

5.

4

3

o:

ReITIFER.

A

*:*

Laboratory: Proportional.Counting of Mixed Liquor

r

PROPORTIONAL COUNTS 7..

PROTOZOA AND METAZOANS

Count organisms in a random sweep across slideDo not.march for particular typesIgnore' ram organismsStop at a total count of either 50 or 100 organisms

Tally each organa total tally box,appropriate tally

7'4ism twice, once inand once in thebox below .)

Plant

TOTAL 50

1

Paramecium

2

TrachelophyIlum

3

1

/

TOTAL 100 --am-

6

%Aspidisca

4

Litonotus

Stentor

1

F.pistylis

1...,

11

telotrochs

Th

'*Pleuromonas

Amoebae

212"

19

ROTIFER

1001114

.11

s, KEY TO SELECTED GROUPS OF FRESHWATER ANIMALS

The following key- is intended to prOvidean introduction to same of the morecommon freshwater animals Technicallanguage is kept to a minimum.

In using this key, start with the firstcouplet (la, lb), and select the alternativethat seems most reasonable. If you -

selected,"la" you have identified the

BI.4 21b. 6. 76

O

el

animal as a mernbe-r sof the group PhylumPROTOZOA. If you selected "lb' s, proceedto the couplet indicated Continue-thisprocess,until the selected statement isterminated with the name.).4 a group,.

If you wish more information About thegroup, consult references. (See referencelist. )

213

.S

N1.

251-1

0

Key to Selected Grou s of Freshwater nimals:"

la The body of the organ' n comprisinga single microscopic iependentcell, or many similar indepen-dently functioning cells associatedin a colony with little or no differ-ence between the cells i. e. , with-out forming tissues; or body co.-n-prised of rnaSses of nultinucleateprotoplasm. Mostly microscopic,single celled animals. -

Phylum PROTOZOA

lb The body of the organism com-prised of many cells of differentkinds, i.e..vforming tissues. .

May be microscopic or macro-scopic.

O

15b Body soft and/or wormlike,. 'Nr 'Skin may range from soft to

parchment-like.

6a Three or'more pairs o ellformed jointed legs pr ent.

Phylum ./0Tf113.0PODA (Fig. 4)

5a Skeleton or shell present. Skel- 15eton may be external or internal..,

t-6b Legs or appendages, if present,

limited to pairs of bumps or hpoks.2 ,-"lobes or tenacles; if present,

soft and fleshy, not jointed.

Body strongly depresited or 8flattened in cross section.

7b Body oval, round, or shaped like -10

anfinverted."U" in Cross section.

8a Parasitic inside bodice of higheranimals, Extremely long and flat,

-divided into _Sections like a-Roman. , girdle.' Life historymay involve:

4 an intermediate host. Tape wormClass' CESTODA 51. 4

2a _Body or colony usually for'm'ing 3

irregular masses or layers sometimes 'cylindrical, goblet shaped,vase shaped, or tree like. Sizerange from barely visible to

2b Body or colony shows some typeof definite symmetry.

'3a Colony surface rough or bristlyin appearante under microscope

,or hand lens. Grey, green, orbrown. Sponges.

Phylum PORIFERA (Fig. 1)

3b CoyiriSr surface relatively smooth.'General texture of mass gelatinous,transparent. Clumps of minuteindividual organisms variouslydrstributed. Moss animals,bryozoans.

Phylum BRYOZOA (Fig. 2)

4a Microscopic: Action of twociliated (fringed) lobes at an-terior (front) end in life oftengives appearance of wheels.,Body often segmented, accordian-like. Free swimming or attached.Rotifers or wheel 4nimalcaes.

Phylum TROCHELMIIITHES(Rotifers) (Fig. 3)"

19

7

4b Larger, wormlike, or having 5

strong skeleton b shell.

25 -2

8b "Body a single unit. Mouth anddigestive systempresent, but noamts. }

9a External or internM parasite ofhigher animals. Sucking discspresent for attachment. -Life his-tory may involve two or more in-termediate hosts or stages. Flukes.

Clpss TREMATODA,

9b Fr e living.. Entire body coveredwith locomotive cilia. Eye areasin head often appear "crossed".Free living fllitworms.'

ClassjURBELLARIA (Fig. 6)

10a Long, slender; with snake -like- motion in life. Covered with glis-

tening'cutiele. Par-asitic or free-living. Microscopic tosix feet inlength. Round worms.

Phylum NEIViAttIELMINTHES(Fig. 7)

10b. Divided into ,sections pr seginents, 11

2 1 .4. .

,

4'

O

it

I'

Key to Selected Groups of Frethwater Animals

10c Vnsegmented, head blunt one 18,

or two retractile 'tentacles.Flat pointed, tail.

Ila Head a more or less well-fornied,hard, capsule with jaws, eyes,and antennae..

Class INSECTA order DIPTERA(Figs. 8A, 8C)

1lb Head structure pOft: exceptjaws (if present). 8E.).

sucking parasites on higher animals,often found unattached to host.

. Leaches.Ma's HIRUDINEA (Fig. 98)

15a Skeleton internal, of true bone.(Vertebrates)

15b Body covered with an external 16skeleton or shell.

12 (Figs. 10, 13 17, 18,- 24,

40

12a Head conical gr rounded, lateral 13appendages not conspicuous ornumerous.

12b Head somewhat broad and blunt, 14

Retractile jaws usually present...00*L Soft fleshy lobes or tentacles,

often somewhat flattened, may bepresent in the-head region. Tailusually narrow. Lateral lobesor fleshy appendages on eachsegment unless there is a large.sucker disci at .rear end.

Phylum ANNELIDAN ig. 9)/

r

.13a Minute dark colored retractile?jaws present, body taperingsomewhat at both ends, pairs orrings of bumps or "legs" oftenpresent, even, near tail.

Class INSECTA Order DIPTERA(Fig. 8)'

13b No jaws, sides of body generallyparallel except at ends, Thicken-ed area or ring usually presentif not all the way back on body.Clumps of minute bristles on mostsegments. Earthwo-rms, sludge-worms.

Order OLIGOCHAETA

14

14a Segments with brittles and/or fleshy' 'lobes-orlother extensions. Tubebuilders, borers, or burrow6rs..

: Often reddish or greenish incolor. Brackish or fresh water.Nereid, worms.

Order POLYCHAETA (Fig. DA)

14b Sucker disc at each end, the'largeone posteriOr. x rnal.blood-

25, 28) 0

16a External skeleton jointed, shellcovers legs. and other appendageS,often leathery.in nature..

phylum ARTHROPODA

1-6b External shell entire, not jointed,unless composed of two ri-

-7-like halves.(Figs. 10, 11, 12)

'17a. Half inch or less in length. Twoleathery, clam-like shells. Soft,parts inVittle include delicate,.jointed appendages. Phyllopodsor branchiopods.

Class CRUSTACEA, Sub,classesBRANCHIOPODA (Fig.. 12)

and OSTRACODA (Fig. 11)

19

17

17b /Soft parts covered with thin 18skin, mulcous:produced; no: jointed legs.

phylum.MOLLUSCA

18a Shell single, may be a spiral cone.Snails.

ClasS.GASTROPODA (Fig. 13)4

18b Shell double, two halves, hingedat one point. Mussels, clams.

Class BIVALVIA (Pig. 10)

194 Three pairs of regular walkinglegs, ortheir rudiments. Wingspresent in all ad'Ults and rudimentsiz*some larvae,

Class INSECTA (Figs. 22;. 225, 26, 2ft:'29)

19b More than three pairs'of legsapparently` present.

e20a Body elongated, head broad and flat.

._ 25-3

0

29

20

t

rKey to Selected'Groups Frvshwater Animals

with strong jaws. Appendages follow-ing first, three pairs Of legs are rclund-

tppering filaments. Up to 3inches long. 5obson fly and fish flylarvae.

Class liNISCTA. Order'MEGALOPTERN (Fig. 14)

20b FOU1' or more pairs of legs.

21a Four pairs of legs. Body rounded,bufbous, head minute. Oftenbrown or red. Water mitOs.

Phylum ARTIIROPODA, ClassARACIINII)A, Order ACARI(Fig. 15)

21b Five or more pairs of walliingor swimming legs: gills, twopairs of antennae. Crustaceans.

Phylum ARTIIROI'ODA,Crass CRHSTACE

22a Ten ormore pairs of fleaflike \swimming andappendages. Mhny spedconstantly in /life; some swimupside doWn, Fairy shrimpspityllopods. o'rbranchipods.

Subclass BRANCBIOPODA(Fig. 16)

22b .1,ess than ten pairs of swimmingor respiratory appendages.

3. 23a Body arid legs inclosedvalved (2 halves) shell wliich Nayor may not completely hide them.

3b Body'and legs mht enclosed inbivalve shell. May he large 9rminute.

(Figs. 17, 18, 19)

d,atorywim

24a One pair.of branched antennaeenlarged for locomotion, extendoutside of shell (carapace).Single eye usually visible."Water: fleas"

Subclass CLADQCERA (Fig. 12).24b Locomotion abcornplished by

body legs, not by antennae.

N

25-4

21

J

25a Appendages leaflike, flattened,more than' ten pairs.

Subclass BRANCHIOPODA(See 22 a)

1

25bAiiiimal less than 3 mm, in length.Append4ges more or less slenderand jointed, often used for walking.S'hells opaque. Ostracods,.

(Fig. 11) Subclass OSTRACODA

2C) Body q series of six or Moresimilar segments, differing.mainly imsize.

261) Front part of body enlarged intoa somewhat separate body unit

cqcophalothora0 often covered22 with a single-piece of shell (cara-

pace). Back part (abdomen) may berelatively small, even foldedunderneath front part. (Fig. 19b)

23

24

26

27a Body coropressdgaterally,organism is tall and thin. Seuds.atnphipods.

Subclass AMPIIIPODA (Fig. 17)

27b Body' compressed dorsoventrally,i.e., organism low and britad.

A Flat gills contained in chainberirteneath tail, Sowbugs.

Subclass ISOPODA (I+ ig. 18)

28a Abdomen extending straight outhohind, Gilding in two small pro-jections. One or two lai-ge masses' ofeggs are often attached to female.Locomotion by means of two enlarged,unbranched antennae, the only largeappendages on the body. 'Copepods.

Subclass COPEPODA (Fig: 19)

28b Abdomen extending rut behind endingin,an expanded "flipper" or swim-ming paddle. Crayfish or craw fish.Eyes on movable stalks. Size rangeusually from one to six inches.

4 Subdlass DECAPODA

29a Two pairs of functional wings,one pair may be more or less har-

25 dened as protection for the otherpair. Adult insects which normallylive on or in the water. 25, 28)

21G 0

39

4

a

. es

lir

"b. .

Key to'_Selected Groups of Freshwater Animals

Mb No functional wings, thoughpads in which wings are develop-ing may be visible. Some may

. resemble adult insects veryclosely, others pay differ ex-tremely from adults.

30

30a External pads or cases in which 35 .

wings develop clearly v24,26,27)

$0b More or less wormlike, or at 31

least no external evidence of -

wing development.

31a No jointed legs present.. Other.,structures such as hooks, suckerdiscs, breathing tubes may bepresent. Larvae of flies,midges, etc.

Order DIPTERA ('Fig. 8)

31b Three pair,s of jointed thoracic 32legs, head capsule well formed.

.32a Minute (2-4mm) living on the

water surface film. Tail a .

strong organ that can be hookedinto a "catch" beneath thethorax. 'When rele-ased animaljumps into the air. No wingsare ever grown. Adulttails.

Order COLLEMBOLA

32b Larger (usually over 5 mm)wormlike, living beneath thesurface.

33a Live in cases or webs in water.Cases or webs have a silkfoundation to which tiny sticks.stones, and/or bib of 46brisare attached. Abdominal segmentsoften with tiiinute gill filaments. ..Gen,erally cylindric in shape.Caddisy larVae:

be Order TRICHOPTERA (Fig. 21)

33b' Free living, build-no cases.

34a Somewhat flattened in crosssection apd massive in appear-ances Each abdominal segmentwith rrather stout, tapering, lateralfilaments about as long as body

is wide. Alderflies, .fishflies, sanddobsonflies.

Order MEGALOPTERA (Fig. 22, 14)

(34b Gene ally rounded in cross section..Late al filaments if present tendto b long and thin. A few formsextremely flattened, like a suctioncup. Beetle larvae.

Order C014tOPTERA (Fig. 23)

35a Two or three filaments or other.structures extending out fromend Of abdomen.

35b Abdomen ending abruptly, unlessterminal s ment itself is extendedas single rticture.(kgs. 24A, 24C)

37

36a Mo th parts adopted for chewing.*# FrOnt o cc civered by extensible

folded mouthparts often called a.; "mask". Head broad, eyes widely

spaced. Nymphs of dragonfliesor dar'n,ing needles.

Order ODONATA (Figs.24A, 24C, 24E)

361) Mouthparts for piercing and sucking.Legs often adapted for water lo-comotion. Body forms various.Water bugs, water scorpions, waterboatmen, backswimmers electriclightbugs., water stridems watermeasurers, etc.

33 Order IIF-MIPTERA (Fig. 25)

37a extensions (caudal' filaments)two. --Stonefly larvae.

Order pLEcoRTErifi (Fig. 26)

371) Tail extensions three, at timesgreatly reduced in size. d.

1

38a Tail extensions long and Blender.;Rows of hairs may give. extensionsa feather-like appearance.Mayfly larvae.

Order EPtAMEROPTRA(Fig. 27)

38b Tail extensions flat, elongatedplates. Head broad With widelyspaced eyes, abdomen relativelylong and slender. Damselflynymths:

Order ODONATA.(1ìf. 24D'

25.-5

38

5'

4.

Ku to Selected Groups. of 'Freshwater Animals

"""39a External wings or wing coversform a hard protective domeover theinner wings foldedbeneath, and over the abdomen.

a(las.-Order COLEOPTERA

(1}g. 28)

39b E'xterinal wings leathery at base.-Membranaceous at tip. Wingssbmetime;;°yer6r short. Mouth-parts fciVpiering and suckingBody form Various. True bugs.

Order FIEMIPTEFIA (Fig. 25)

40a Appendage present in,pais.(fins, legsv-wings)

40b.. N,o paired appendages. Mouth 41a round suction disc.

42

41a Body- long and slender Severaltyples along side of headLampreys."'

Sub Phylum VERTETillATAClass 'N'('7.0STOMATA

41h 13odv plump, oval Tail extendingout abruptly Larvae of frogs andtoads, Legs appear one 'at a,til-neduring metamorphosis to adultform. Tadpoles.

Class 41MPHIBIA

25 -6

A

515

42a Paired rippendages arelegs

42b Paired appendages are fins.gills covered by a flap(operculum).----T rue'Tishes

Class PISCES

43

43a' Digits with claws, nails, or hoofs 44

43b Skin naked. No claws or digits.Frogs, toads.\ and salamanders.

ClaSS A MPHIBIA

44a, Warip blboded; :\',N "f''\

. 44h Cold blooded Bo y coveredwith horny scales r plates

'Class REPTIL A --""

45a Body Covered with fathersBirds

Class A VES

45b Body entered with hair"Mammals

Class MAMMALIA

5 5

a

a

4'5

4

II

I ,

4Key to Selected Groups of.Ereshwater Animals,

c

REFERENCES - Invertebrates

1 Eddyi,,Samuel and Hodson, A. C.litionornic Keys to the CommonAnimals of the North Central States.Burgess Pub. Company, Minneapolis.162 p. 1961.

2 Edmondson, W. T. (ed.) and Ward andWhipple's Freshwater Biology. JohnWiley & Soni, New-york. pp. 1-1248.1959.

3 Jahn, T. L. nd Jahn, F. F. How to Knowthe Pr to Wm. C. Brown Company,aubuque, Iowa. pp. 1-234. 1949.

4 Klotsr-Elei-e"B. The New Field Book'ofFreshwater Life. G.P. Putnam's ns.398 pp. .1966

5 Kudo, R. Protozoology. Charles C.Thomas, Publisher, Springfield, Illinois.pp. 1-778. 1950. or.

6 Palmer, E. Lawrence. Fieldbook ofN,atural History. Whittlesey House,McGraw-Hill Book Company, Inc.New York. 1949.

7 Pennak, R. W. . Freshwater Invertebratesof the United States. The Ronald Press,Company, New York. pp. 1-769, 1953

-

8 Piinentel, Richard A. InvertebrateIdentification Manual. ReinholdPublishing Corp. 151 pp. 1967.

Pratt, HW.- A Manual of the CommonI Invertebrate Animals Exclusive of

Insects. The Blaikston Company,Philadelphia, Pa. pp. 1-854. 1951

REFERENCES - Fishes

1 American Fisheries Society. A List ofCommon and Scientific Names of FishesJ from the United States and Canada.-Special Publication No. 2, Am. Fishsoc. Executive Secretary AFS.,Washington Bld. Suite 1040, 15th &New York Aventie, N.W. Washington,DC 20005. (Price $4.00 paper,$7. 00 cloth). 1970

2 Bailey, Reeve M. A ReVised List ofthe Fishes of Iowa with Keys forIdentification, IN: Iowa Fish andFishing. State of Iowa, Super. ofPrinting. 1956 (Excellent ,colorpictures).

3 Eddy, Samuel. How to Know theFreshwater Fishes. Wm. C. BrownCompany, Dubuque, Iowa. 1957.

.,.4 Hubbs, C. L. and Laglesi, K. F. Fishesof,the Great Lakes Region. Bull.Cranbrook Inst. Science, BloomfieldHills, Michigan. 1949.

5- Lagler, K. F. Freshwater FisheryBiology. Wm. C. Brown 'Company,Dubuque, Iowa. 1952.

6 Trautman, M. B. 'The- Fishes of Ohio.Ohio State University Press; Columbus,.1957. (An outstanding example of aState st zcly)%

Descriptors: Aquatic Life, Systematics.

Key to Selectsid Groups of Freshwatei. Ahimap

...wlA,.

tiy

.\t cei

1. SeaTilla spieulesUp to .2 mm.lpag.

'IA. Rotifer, FolyarthraUp to .3 mm.'

3C. Rotifer, PhilodinaUp to .4 mm.

3B. Rotifer, KeratellaUp to .3 mm.

4A. Jointed legCaddisfly

25-8x:

2B. Bryozoal mass. Up toseveral feet diam.

214. Bryozoa, Plumatella. Individuals- upto 2 mm. Intertwined masses may hevery extensive. 1-

4B. Jointed legCtayfish

aria, MesostdmaJ IA? a %ATI.

4C. Jointed legOstracbd

5. Tapeworm head,Taenia . Up to25 yds. long

6B. Turbellaria, Dugesia Nematodes. Free livingUp to 1:6 cm. forms commonly up to--I mm.. occasionally

more.

2210

-

I

I

I

I

I

I

I

I

I

I

I

I

(

.-

,.

Key to Selected Groups of Fregliwater AnimalsA

813. Diptera, Mosquitopupa. Up to 5 mm.

13A. Dipt, rd, :Mosquito larvaeUp to 15 mm. long.

8C. Diptera, chironomid 8E. Diptera, crane fly. pupa. I:p to 2. 5 cm.

larvae.' Up to 2 cm. ,

9A. Annelid,segmentedworm, up to1/2 meter

9D. Diptera,. Rattailed maggotUp to 25 mm. without tube.

1013. AlasiniOnta, end view.-

ri.

010A, Pelecyopod, Alasmidonta

- Side view,. up' to 18 Cm.long.

9B. Annelid, leech up to 20 cm.

1

4 11A. Ostracod, CypericusSide view,: up to 7 mm.-

.

12A. BratichioPod,,Daphnia. Upto 4mm.

c

:12B. Branchiopod,Bosmina. Up -

____.., to 2mm.

1113. Cyper us, end view. 25 -9

4

$

ii.ey to grjeted-Orotts-of Festmate Animals

a

13. Gastropod, CaMpelomaUp to 3 inches.

14. Megaloptera, Sialis

ALderfly larvaeUp to 25 min,

16. Fairy Shrimp, EubranchipusUp to 5 cm.

J10111111 Ilk,4Wert Nt,

/1.7C\I

17. Amphipod, pcmaiaporefaUp to'25 mm.

9.

.11

15. Wkter mite,up to 3 mm.

18. Isopod, AsellusUp-to 25 mm.

O

20. Collembola, PoduraUp to 2 mm.

25,10

iffillite. 41Io

19A. Calanoid copepod,

, Female 19B. yclopoid copepodUp to 3 am. Female

Up to 25 mm.

I.

2° 904#0*.e

21A.

3

21B. 21C.

21D. 21E.

Trichoptvra, 1 aryal cases,mostly 1-2 cm.

Key to Selected Groups of Freshwater Animals

22.. Megaloptera,--alderflyUp to 2 cm.

23A. Beetle larvae, 23B. Beetleklarvae, 24A. Odonata, dragonflyDytisidael Hydrophilidae nymph up to 3,orUsually about 2 cm. Usually about 4cm

cm. 4

Odonata, tailof damselflynymph(side view)

SuborderZygoptera(24B, D)

A

24E, Odonata, front viewof dragonfly-nymphshowing "mask"partially extended

SuborderAnisoptera

(24A, E, C). Odonata, damselflynymph (top view)

223

24C. Odonata, tail ofdragonfly nymph(top view)

25-11

e

Key to Selected Groups of Freshwater Animals

25 -12

25A. Hemiptera,Water BoatmanAbout 1 cm.

27 .Epheme ropte ra.Mayfly nymphUp to 3cm.

,

258. Hemiptera,Water ScorpionAbout 4 cm,

28A. Coleoptera,Water scavengerbeetle. Up to 4 cm.

29A. Diptera, Cranefly. Up to 23 cm.

26. Plecopte ra,Stonefly nymphUp to 5cm.

28Bs. Coleoptera.Dytiacid beetleUsually up to 4 CM.

29B. 4Diptera. MosquitoUp to 20 mm.

,

1

1

A.

A .Kay FOR THE INITIAL SEPARATION OF SOME COMMONPLANKTON ORGANISMS

1. No chlorophyll present, 'unless 'through ingestion 8

1. At least some chlorophyll present0

2. Pigments not in plastids Oyanophyta2. Pigments in one or more plast s 3

3. Cell wall .of over-lapping halves and distinctly sculptured Bacillariophyta'3. Cell wall not of over-lappingchalves, , or if so, then not sculptured 4

4. Pyrenoids present; color usually. bright green Chlorophyta4. Pyrenoids absent; color green, yellow-green yellow-brown 5

5. Bright green, motile, usually withone 'anterior flagellum Euglenophyta6

J .st.1.5.!

Yellowish to brownish, motile or not

6,, With a ,distinct lateral groove, motile Pyrrophyta6. '"Without a lateral. groove 7

I ,-

'7. Seldom' motile; unicellular, colonial or filamentous , . . . Xanthophyta7. Motile, unicellular or colonial r . . . Chrysophyta

8. Unicellular; naked or enclosed in a smooth or sculptured shell 9

8. Multicellular; body usually with a distinct exoskeleton . . ... . . . . . 11

9. Amoeboid; sometimes with shell, no cilia or flagella Ameoboid Protozoa9. Actively motile; never with shell; cilia or flagella obvious 10

10. Body more or less covered by short cilia;movement "darting" . Ciliate Protozoa

10.. Body with one or more flexible whip-like falgella;movement "continous" . ;Flagellate Protozoa

11. Shell bivalved (clam-like) r .1211. Shell not compoOd of two halves 13

12. With distinct head anterior to valves12. No head anterior to valves

CladoceraOstracoda

-.

.13. Usually microscopic; body extended into a tail or foot' with oneor more toes Rotiferat .

\ 13. Usually macroscopic if rilatur ; ' A14

.... .,- Copepoda14. Appendages bilateral; heed not prominent .. .

14. Appendages unilateral; head prominent Phyllopoda.

o o

..

CHLOROPHYTA

A Key,,, to' Sorrre of the Comnion Filamehtous Genera

Alf1.1.

. .

,Filaments unbranched _Filaments branched (sometimes parenchymatous)

2. Chloroplast single, parietal band2. Chloroplast one or more, If parietal not a band'

3. Chloroplast encircling more than half the cell (Nap -ring like)-3. Chloroplast .encircling less than hdlf the cell . . . .

4. Filaments of indefinite length; cells with square ends4. Filaments usually short, of 3.8 cells, with ends round

5. Cell wall of Ilk pieces; pyienoids Iaaing <

5. Cell wall not of H pieces

6. Some cells with apical caps6. Cells without apical taps 1

7. Chloroplast (s) parietalChloroplast (s) axial .. ti t

8; Chloroplast one or more, spiral bands..8. Chloroplast several, longitudinal bands

9. Cell walls- without a median constriction,9. Cell walls with a median constriction

I.

10. Chlor plast stellate ;10. Chlor Oast an axial band

11. cylindrical11.

.FilamentsFilaments triangular, twisted

2

12

3

5

Ulothrix4

HormidivmStichococcus

Microspora

Oedogonium4 . 7

8

9

SpirogyraSirogonium

10. 11

ZemaMouygngeotia

HyalothacaDesmidium

12. Coenocytic dichotomously, branched, with constrictions Dichotomosiphon12. Filaments with regular cross walls 13

I o# 4*

13. Parenchymattius, discoid, epiphytic . a . . Coleochaete13.- Not parenchymatous 01°4

- - °14. Main axis cells much broader than branch cells . ... . _._. . -Draparnaltia-14. Main axis and branch _cells 'approximately- thesanie breadth

..15, Maitraxi arilateral branches attenuated into long multicellualr hairs ,.16

15. Axis and branches not attenuated into long multicellular hairs d' .. 17

16'

16. Sparsely or loosely branched Stigeoclonium.16. ;1Densely and compactlybrarich d Chaetophora

t..

e .0. 0.

17; Cells bearing swollen or bulbous-bd.sed setae 18

17. Cells without setae 19

18, Swollen-based ,setae on dorsal surface of cells; rostrate1 \ .

epiphytes; little or not al all branched Apbanochaete`18. Bulbous-based setae terminal..onbranches;,itot S . .

. , Bitlbochaeteprostrate epiphytes _a , -, -

19. With .terminal and/or intercalary akinites ° Pithophora19. . Without ,distinctive akinites 20

;r1

20. Cells of erect filaments becoming shbrter and broader towardfilament apex;-UsuallY growing on back of turtles; branching only .

from base ' . , Ba.sicladia. . 20; Thallus not as above . 21

21. Filaments irregularly branched; branches short 1 - or few celled . . zoclonium21. Filaments repeatedly°branched; branches narrowed toward tips C dophora

J.

0

e 0

s,,,

°

0.

,0 0o n

o .°

O

*s

0

et

. .

i°

0

CHLOROPHYTA

A Key to Some Common Non Filamentous Genera4

47'

I.

1.

3.'.3.

5.5.

o

7.7.

9.9.

UnicellularColonial

.. ,

2. Motile2. Non

CellsCells

4. 'Cell4. Cell.

CellsCells.

6. Cells6". Cells

Cellscells

8. \ength8,

CellsCells

. . . - 1

-in vegetative -state,, flagella 2 -4

- motile in *vegetative state ... : .. .

-with 2 flagellawith .4 flagella

enclosed by bicovex shell . . .. .rnot enclosed by a'Shell . . ",'. :

with median constriction (often slight), or of chloroplast only ..without a median constriction

..

..lunate :not lunate in any degree

i

cylindrical, noticeably 'longer than broad . ti'''almost .never cylindrical; flattened or triangular in apical view

2-3 times the breadth, contriction nearly lacking . . . .ength much greater than breadth, nodulose at constriction

___.

,.

triangular in end view 1

not triangular in end vie)w z :-

2

27I

3.5

4. . .4

Carteria

:

PhacotusChlamydomonas

. 615

palClosteri# 7

89

. CylindrocystisPleurotaenium

Staurastrum'10..

10. Semicells with lateral incisions, appearing lobed /11

10. Semicells without lgeral incisions 12

11. Lateral incisions few, shallow, lobes rounded . . .... . Euastrum11.- Lateral incisions many, deep, lobes angular Micrasterias

12. Semicells with radiating arms , . . StaurastrumSemicells without arms; spines, granules, or teeth may be present 13

13. Semicells without spinesOt404. 13. Semicells with spines

14. Spines few, usually at the apical corners14, Spines numerous, scattered

Cosmarium14

15. Cells elongate, sometimes needle-like . . . .

'ArihrodesmusXanthidium

16,, 15. Cells 'spherical,- ovid, angular; not needle-like 18

--., 46. Ceiis with terminal setae16. Cells without terminal setae

Schroederia

...17: Cells acicular, withOut a row of pyrenoids. .. Ankistrodesmus17. Cells acicular, very long, with a row, of 10-12 pyrenoids Closteropsis

..e

18. Cells wittioitt spines br processes 19

18. Cells with spines or processes 23

.19. Cells embedded- in a gelatinous matrix

2201.19.1 Cells without a. gelatinous matrix.

20; Gelatin obvious, lamellate; dhloroplast Cup-shaped . .Asterococcus

. . . . .-Gloeocystis20. Gelatin sometimes obvious, chloroplast, star-like

4

21. Cells spherical , -.±. . ., 2221..: Cells angular Tetraedron

.. , . .

22. Cell wall smooth . Chlorella. 22. -Cell wall sculptured' . . Trochiscia

. ' 7

23. Cells angular 24 s... .

23. ps spherical or. oval, with spines .. .- . . . . 23-'.,.: . .

. , -24. Angles with furcated _processes .,..

, -, . Tetraedron.-...-2-4. Angles with spines ... IN

12 Polredriopsis ...

25. Cells spherical, spines delicatec,

. .. . ._ ... ". .. , G lenkinia.....

25: Cells oval, spines evident . .... . . . 26 ..... .14

.,1/4

26. Spines localized at ends of cell . .. La.geheimii -26. Spines distribbted over cell . . Franceia

27. .Motile,. each cell with 2 equal-length flagella .t? 2827. Non-motile invegetative state 33

28. Colony. a "flat" plate 2928. Colony spherical or ovid 30

29. Colony "horse-shoe" shaped Platydorina29. Colony quadriangular or circular ,Gonium

30. Cells 8-16, crowded-pyrifOrm 1 . . Pandorina30. Cells more than 16, not crowded, spherical .or nearly so 31

.

31. Celld more than 300 in number.31. Cells less than 300 in nitalier

32. Cells (16) - 32k nur per Eudorina-32. .Cells 64-128 - (256), in number . Pleodcirina

. _

1.Volvox

3?

33. , Cells of colony lying in one plane 3433. Cells not in a conspicuous single plane , 37

. . .I .

34'. Colony, ciroillar (rarely cnuciate) . . .'Pediastrum34. Colony not circular I

..4

40,

. 1 2 2

38

26.5

-

,

35. Colony, a flat strip; cells side by side , Scenedesmus35, Colony quadriangular , 36

Colony usually large, cells in 4ts, no *pines Crucigenia36. Colony of 4 cells, leach with 1 or 2 marginal spines . . . . . . Tetrastrum

37. Cells acicular (needle-like) Ankistrodesmus37. Cells not acicular

38. Colony without a gelatinous envelope 3938. Colony with a more or less conspicuous gelatinous envelope 47

39. 2 - 8 oval enclosed by a distinct sheath39. ICells hot enclosed by a sheath

40. Cells with long Spities or setae40. Cells without spines or setae 42

38

. Oocystis40

41

41. Colony pyramidate, cell spherical,.

41. Colony quadrate, - or a tetrahedron,go - .

42. Cells linear, radiating from a42. Cells no linear

long spines Errerella.. . Micractiniumi .long setae . .

\common center A ctinastrum .

43,.. . . .. . ,.

43. Cells .strongly lunate' often "back-to-back" Selenastrum43. Cells not. lunate - 44

-44..` Cells arranged in a hallow sphere 4544. Cells not arranged in a hallow sphere 46

...45. cells spherics1, sometimes joined by processes '. 1,..- . .

45:' e.Cells not spherical, oute* angles extended into stout, blunt.,teeth or spines .7.-;. -.:. ... ...

. ,

46. Cells uniform, spherical, in groups of 4 :- 846. -Cells notluniformi ellipsoid, (oblong) or reniform Dim

. .,4 7 , Cells curved tlo strongly lunate 1 . . 48

Cells, not curved or innate"- .,A ... . .. . . 49&& , . && ..

...d.

8. Cells innate., loosely arranged in colony Kirchneriella .

48. Cells curved or reniform, colony compact, distinct Neptirocytium. .

rlo

. Coelastraim

orr. i

' 49 Cells connected by' branching central strands . Dictyosphaerium'49. Cells not connected by stands 50

or

\

\ 5150. Cells cyli -drical or fusiform50. Cells

..sp rical or s ghtly ovoid 52

..'. v-, .-. 41

4,,,...**N,... . -

..4,..,.51: Celli in parallel "bun s f 2 - 8 ' . r Quadrigula.

. 51. Cells longitudinally arranged, not ,grouped laterally77

Ela\katothrix. 4

.rIr

230

J

52. Cells ellipsoid or ovoid, envelope lamellatedft.

Glosocystis -

52. Cells spherical, or nearly so , . . . 53

53. Chloroplast axial, star - shaped53. Chloroplast parietal, not star- shaped

Asterococcua`54

54, Cells enclosed by lamellated sheaths . . Gldeocystis54.. Cells in homogenous envelopes Sphaerocystis

o^

SI%

ti

(

O

23.t

O

26-7

Ao.

-.

a

1.

1.

..

.

3..3.

5.

5.

7.

7.

ETIGLENOP HYTA

..)

-

,

i

IIII

A

4111ie.r

Colacitnn

4.

i.

Vegetative cells sessileVegetatiye cells- motile

'2. Cells with _chlorophyll, -a

2. Cells without chlorophyll.

Protoplast° withina lorica or teatProtoplast naked, no lorica

,..4. Body strongly metabolic4. Body -rigid .

With two laminate, longitudinal chloroplastsWith numerous chloroplasts .

.

6. Body conspicuously flattened, sometimes twisted,

6. Body not compressed, radially syMmetrical

Body broadly ellipsoid to ovoidBody elongate, narrow

2

3

.. 8

Trachelomonas

4

... . Euglena, 5

Chryptoglena

6

_,PhacuS

6.3,%

Lepocinclis

Euglena

..

e. Cells with one flagellum .. iistisia8. Cells with two flagella Peranema

.

1, t

9

II

°

Is,

,o

.

WI

.............

le--?.....)--:

P

* ',Si

.

I

1

1

I

1

I

I

I

1

1

1

1

1

i

KEY TO SOME COMMON DIATOM GENERA (BACILLARIOPHYTA) OFMICHIGAN

. Diatains in Valve View

1. 4Valves without a dividing line or cleft; markings of' valve radiate about a central4point. (.centric diatom genera) 2

2. Frustules usually in filaments or zig-zag chains; rectangular in girdleviewi, valve round, oblong, triangular or elongate 3

a. Frustules cylindi:ical; inarkings4rominent on girdle; sucuspreseht; seldom seen in valve view Melosira

3. Frustule not cylindrical 4

4. Valve ovate to oblong; two horns or processes present on valve face,scatter small spines often present Biddulphia

.

4. Valve not ovate to oblong, horns or processes absent

5., Valve face appearing as two; three sided pieCes giving theappearance of six processes Hydrosira

..5. Valve face three to several times longei° than broads; marginsundulate.: "costae" evident Terpsinoe

,5

2. Frustules usually solitary (sometimes forsning short chains) 6

6.. Frustules usually elongated; many intercalary bands; frustulescylindrical 7

7. Each valve with a single long spine Rhizosolenia

7. Each valve with two long,spines.

6. Valves discoid; cylindrical; or spherical; somtimesprocesses . . . . ....

9,

8. Ornamentation of valves in two concentric parts of unlikepattei71

with spines ora. 8

CyclotelIa

8. 'Ornamentation of valves radiate; continuous from center tomargi9 of valves

.9

9.. Ornamentation of valves distinctly radiate; rows of punditaesingle at center becoming multtheriate at the margin; marginof valve with recurved spines Stephanodiscus

9. Ornamentation of .valves not distinctly radiate or,becomingmultiseriafe at margin 10

233

10. Isolated large punctum evident at the margin *pinesnot evident . . . . r Thalassiosira

10. Isolated punctum not evident at the margin,' shortspines present,

1." Valve_ with a- dividing line ,o cleft; marking of wall bilaterallydisposed to an axial or exce tric line (pennate diatom genera) 11

11. Both valves with a'pseudoraphe 12

12. Valves asymetrical to['one axis 13

13. Valve asmetrical to longitudinal axis, striae present . . Ceratoneis

Cascinodiscus

'

. 13. Valve apymetrical to transverse axis 14

14. Valvep clavate 15

14; :Valves not claate; striae present; valves wittr-unequalcapitate ends; often forming star like colonies . . Asterionella

15. . "Costae" present, frustules may be joined faceto face to form fan like filaments. Meridion

15. "Costae: present, frustules single (appeawrias asymetrical Fragilaria) Opephora

12. Valves symetricalto both axes 16

16. Frustules septate or "costate"; often appearing inzig- zag chains 17

17. Septae present; usually many partially septateintercalary bands; valves triundulate Tabellaria

17.4. Septae absent; prominant "costae" on valve . . . Diatoma

16. Frustules occuring Ire or attached in filaments;sometimes forming scicles ..... . 18

18. Frustulea typically forming long filaments; usuallynot more than 5 or 6 times longer than broads;often appearing costate . . . Fragilaria

18. Frustules usually solitary or forming. fascicles;usually many times longer than broad Synedra(note: the above two genera are actually seperatedonly of the basis of growth habit)

11. At least one valve with a pseudoraphe 19

19. One valve with a true raphe the other valve with a pseudoraphe '20

20. Valve asymetrical to the 'transverse axis; partial terminal septae;bent about the transverse axis Rhoicosphenia .

20. Valves symetrical to both axes . . 21.

21. Valve elliptical; valves with- a marginal and/or submarginalhyaline ring; often loctiliferous; bent about the longitudinalaxis .. .

Cocconeis

21. Valves usually lanceolate or linera lanceolate"; bent- 'around the transverse- axis Acfmanthes

(Those with a sigmoid raphe` are sometimes put inAc4nanthidium or Eucocconeis)-

19. Both valves with a 'raphe 22

.22. Raphe median or nearly so, never completely marginal;not in a canal 23

ft23. Valkres sigmoid'in outline 24.

24. Raphe sigmoid; punctae in two series; one tran verseand one longitudinal row forming a 900 angle . . . Gyrosigma

24. Raphe sigmoid; punctae in three series forming,angles of other than 90° Pleurosigma

23. Valve not sigmoid in outline 25

25. Valves symetrieal to-,both axes s . . .26'

26. Frustules with sept'ate intercalary bands 27

27. Intercalary bands with marginal loculi, piInctaedistinct Mastagloia

.27. Intercalary _bands with two large faramen along

apical axis, punctae indistinct . ., . .. ...... Diatomella.

2,6. Frustules without septate intercalary bands 28

28. Valve face with a sigmoid saggital keel; ,

"hourglass" shape outline in girdle. view . . . Amphiprora. .

28. Valve face without a sigmoid saggital keel . . . - . . . 29M1'

' 1 ,, ..,.29. Valve with undulate or zig-zag irregular logitudinel lines or

. .blank spaces . . Anomoeneis

29. Valve without undulate or zig-zag irregular logitudinal' lines or,blank spaces 30

.r

T

2 3 5

26-11

a

t

7

1N.

30. Valve with thickened, non-punctate central' area; (stauros)pseudoseptae sometimes present, longitudinal lines and blankspaces lacking ........... . - . Stauronels

30. Valve with or without stauros, septal absent 31.1/4

31. Valve with longitudinal lines or blank spaces . . . 32

32. Proximal ends of raphe usually curved in oppositedirections; "defaut regulariei." toward valve apkces;.longitudinal blank spaces . Neidium

j. 32. Proximal ends of raphe straight; valves with finestriae that appear as costae; longitudinal 'linenear margin. Caloneis

31. .Valves without longitudinal lines or blank spaces 33

-33. Valves with siliceous ribs along each side of the raphe 34

34. Raphe bisects siliceous ribs on valve; central tarea small and orbicular; striae and punctaevery distinct Diploneis,

r,34. Raphe short; less than 1/2 length of valve; central

area long and narrow; terminal nodules evident,elongate 3 5

F-

35. Raphe short; 1/4 or less the length of the valve;.striae not evident Amphipleura

35. Raphe longr; usually about 1/3 the length of.the valve; striae. usually fine, but evident . .

33. Valve without siliceous ribs 36

36. Valves with smooth transverse costae; rapheoften ribbon like. Pinnularia

36. Valves with transverse-striae 37

37. Raphe sigrnoid Scoliopleura

37. Raphe straight

'38. Raphe in straight and raised keel Tropodoneis

38. Raphe 'straight and not in a keel

`38

39. Striae daubly punctate, central area long and narrow . . Brebessonia

39. Striae single to lineate, central area variable . . . . Navicula

39

1..,

25; Valves symetical to one axis only I 40......... . 4

40. Valves symetrical to the longitudinal axis .. 41...,

41. Punctae in one series; longitudinal line absent Gomyhbnema

41. Punctae in two series; longitudinal line present' Gomphoneis.. \ . t .

40. Valves symetrical to transverse axis . . .. t 42

.42. Raphe short; vestigial terminal; with evident terminal nodules,central nodule lacking

A43

43. Colonial; forming tree like coldriies; valves usually withevident spines Desinogonium

43. Usually nbt in colonies or if colonial, forming onlyshort chains or stellate clumps 44

44. Cells shaped like. the femur of a chicken. . . .Actinella

44. Valves various shaped; lunate to nearly straight;valve often with undulate dorsal and/or ventralmargin; raphe prominant in gir le view 45

45. Dorsal margin convex, ve4tral margin slightlycOncave, both margins sinvate-dentste . . . Amphicampa

45_. Dorsal margin convex, Ventrd(l margin straightto concave, one, both or neither margin wavy,pseudoraphe often present on ventral margin . . .Eunotib.

42. Raphe not vestigial; usually as nearly as 'long as the valve;.valves usually cymbiform 46

- .'46. Valves convex; central nodule usually lies very close to the

ventral margin; both. raphe visible In girdle view . . . . Amphort.

46. Valves flat or nearly so; raphe a smooth curve with the`.samecurvature, asthe axial field; raphe not visible in girdleview Cymbella

22. Raphe marginal and in a canal .. . . .

47. Valves 'with a.single canal that is usually marginal but may appearto be somewhat central 48

c:

s

48. Valves symetrical to, both axes . . . . 49'

49. Tranaver,se internal septae that appear as "costae"; canalnearly Median . Denticula:

49. Transverse "costae" lacking; carnial dots present . . . Nitzschia

r

0'

48. Valves asymetrical to longitudinal axis r-50

50. "Costae" quite evident 51

51. Axial field forming an acute angle at the central nodule;raphe along the ventral margin of valve Ephithemia

51. Axial field forming a less acute angle at the centralnodule; raphe along the dorsal margin, of the valve . . Rhopalodia

50. "Costae" not evident; carnal dots along the raphe 52

52. Raphe of, One valvethe othet valve

52. Raphe.of one valveother valve

diagonally opposite the raphe onNitzschia

directly Opposite the raphe on theHantzschia

47. Valves with a canal next to both lateral margins 53

53. Valve face transversely undulate; band of short costae aloqg each .

lateral margin' appearing' like a row of beads Cymatopleura

53. VOve faceortot transversely undtilate 54

54. Valve shaped like a Sadd Camyylodiscus

54. Valve face flat; either isopolar or heteropolar; sometimesthe frustule -is slightly spiral in shape ... . , . . . Surirella

t

2mvv

r.

4

CHRYSOPHYTA

A Key to Some More or Less Common Genera

1. Filamentous, branchedNot filamentous

Phaeothamnion2

2. (Unicellular 3

2. Colonial.,

3. Protoplast enclosed by a lorica 4

3. Protoplast not enclosed by a lorica . 6

4. Epiphytic or epizooic \ 5...,

4. Motile; cells with siliceous scales many of vh ich have long,siliceous spines

5. Epiphytic; lorica flask-shaped

5.. Epiphytic or epizooic; lorica cylindrical

Mallomonas

Lagynion

Epipyxis(= Hyalobryon)

Ochromonas (art associ)6. Motile; protoplast naked.,

6. Non-motile; protoplast with long delicate, pseudopodia . - Rhizochrysis

7. Sessile; each cell In a long, cylindrical lorica Epipyxis (Hyalobryon)

7. Motile 8

8. Each cell within a companulate; basally pointed lorica Dinobryon

8. Lorica absent 9

9. Colony bracelet-shaped

9. Colony spherical -

Cyclonexis

10

10. Colony bristling with long siliceous rods Chrysosphaerella

10. Colony without siliceous rods from each cell .: 11

11. Colonies not enclosed by a gelatinous sheath. / Synura

11. Colonies enclosed by a distinct gelatinous sheath . 12

12. Shorter flagellum more than 1/2 length of longer flagellum Uroglenopsis

12. Shorter flagellum less than 1/2 length. of longer. flagellwn Uroglena

239 26-15

.r"

CYANOPHYTA

A Key to Some of the Common Genera

. .

1.1.

Cells not in trichomes; unicellular or colonialCells in trihomei

. 210

2. Colony with some regular arrangement of cells 3/ 2. Colony amorphilius, no definite form.

6

.3. C-olony a flat 'plate 4

3. Colony spherical, cells peripheral . . 5'

4. Cells regularly arranged Merismopedfa

5.

4. Cells irregularly arranged

Colony with a central branching system

Holop,edium,

Gomphosphaeria

5. Colony withOut a central branching system Coelosphaerium

. many celled6. 'Colony mostly few celled

Cells elongateCells spherical

8. Cells close together8. Cells more than 2-'3 diameters apart

9. Cells usually hemispherical, with of without, definite gelatinous9. Cells sphNgal or olod, 'gelatinoys sheaths very distinct

9it4

Aphanothece8

MicroczstisAphanoc4iisa

sheaths . . ChroococcusGloe ocapsa

1 0. Trichomes without sheath (not filamentous)10. Trichomes in a sheath (filamentous)

11. "Heterocysts absent ._

,. ., 12

,/ 14

11. -Heterocysts present '

c,12. Trichomes straight . . . . . Osciliatoria12. Trichomes' regula y spiralled 13

13. Cross walls dist13. Cross walls lacking .

14, Heterocysts terminaleterocysts intercala'ry

15. - Trichrnes cylindrical . . . CylindrospremurbTrichomes ,attenuate .

. 16

ArthrospiraSpirulina

, 15

.1 s- g, -18. ,

Trichomes\golitary14. Trichomes in masses

26.'46

f

t .

Calottirix.. . . 17

O

I

17. Trichomes without akinetes Rivularia17. Trichome's with akinetes 1 Gloeotrichia

18. Trichomes straight, parallel in bundles Apha.nizomenonTrichomes solitary, or :if in masses, not parallel 19

11.

19. Trichomes solitary, or if numerous, then not in a firm gelatinouses, matrix . . . _. , . .- . Anabaena

19.' Trichomes entangled in a firm gelatinous matrix Nostoc-;.' ',

. 20. Many parallel trichemes in a sheath . . . . . ., t' Microcoleus20. A single trichome or row of irichomet in a sheath 21

21. Filaments not branched . . . . 22

21. Filaments. branched Cl 23

22. Sheath obvious firm .22.. Sheath indistinct; delicate

.4 LyngbyaPhormidium

23. Filaments with false branching 24

23. /Filaments with true branching 26

24. , Without heterocysts Plectonema24, With heterocysts. 25

25. False branches arising single Tolypothrix25. F*alse- branches in 'pairs- .. --- . Scytonema

. . --,26. Trichomes always uniseriate Haplosiphon26.' Trichomes wholly or in part nultiseriate Stigonema

;

Filamentous . .

1. Not filamentous

XANTHOPHYCEAE

A Key to' Som Common Genera

N

Z. Filaments branched, siphopceaous -Vaucheria3

24

2. Filaments not branched

3. Cell wall of stout H pieces, cells spmetimes barrel-shaped Tribonenv.3. Cell wall of delicate H'pieces, cells short and cylinaileal*. . , .

4. Cells embedded in a gelatirious matrix . . ... - . . ,,. 4 5

4. Cells not embedded in a gelatinous matrix . . Ifs 7

5. Colonial gelatinous envelope dichotomously branched . . °Mischococcus5. Colonial gelatinous enveloi4 not, branched . .. 1, . . .6

. ,. te,6. Colonial envelope, 'tough Hheav-P cartilaginous Botryococcus6. Colciniatl envelope' iAratery;:coloniis-small, 'free floating Chlorobotrys

7. Cells epiphytiak. 0 , ,

7. Cells not epiphytic,. . .' ., . . ° .. ' . . . . ..-.;....... ...... 11 .,.. , ..

8. Stipe long,.t

seta-like, :, -- Stipitococcus8. Stipe short; dr cell-sessile .... ... . . .°: .. ,,,' Peroniella

9. Cell vase-like, apex flattened °. . . .. ' i .. l',. - Stipitococcus,. , . ,9. Cell oval or spherical . ... .. °.. ' ... . ./: . 4.4'. . . :' . . , . . .. Peroniella

. Ct ,-..

, ,:, .. 10. CelTs ' cylindricai, ends broadly rounded . . ..°'.. ' . , .. .; . . . . ,. Ophiocytium ,10. Cells not ,cylindrical 4 . Characiopsis .

t . ,- .. . t

11. Cells cylin rical straight or contorted . . -.: . .. .. . - . OPhiocytium.

11. 'Cells glob , with rhyzoids, on soil : - . .. . - Botrydiumt

8.

,

242

4

4,ii4,

Acicular: needle-like in shape.*.(Ankistrodesmus).A*.

Aerial: algal habitat on moist soil, rocks, trees, e ; involving a, thin film of water;sometimes only somewhat aerial.

Akinete; A type of spore formed by the transformation of a vegetative cell into a thick-wallet resting cell, containing a concentration of food material. (Pithophora)

Amoeboid: like an amoeba; creeping by extensions of highly plastic proto lasm(pseudopodia). (Chrysamoeba)

Amorphous: without definite shape; without regular form.

Anastomose: to sepa and come together again; a meshwork.

Antapical: the posterior o rear pole or -region of,.an orgaririm, or of a colony or cells.

7

Anterior: the forward end; toward the top...,Antheridium: a single cell or a series of cells in which male gametes are produced; a

multicellular, globular male organ in the Characeae; more properly calleda globule (a complicated and specialized branch in which antheridial cellsare produced. cr. a

Antherozoid: male sex cell; sperm.

Apex: the summit, the terminus, end of a projection, of an incision, or of a filament, ,zt of cells.

,Apical:. Forward tip.

APlanospore4 nonmotile, thick-welled spore formed_ many within an tinspecializedvegetative cell; a small resting ftp or e.

A ibuscular: branched or growing like a tree or bush.

Armored: See theca.s- .

AtAktuate: narrowing to a point or becoming reduced in diame er. (Gloeotrichia)

f

ik .

Autospores: spore-like bodies cut out- of the contents ore cell which arefirsmall replicaiof the parent cell and which only enlarge to become mature plants. (Coelastrum)

. -

Axial: along a median line bisecting an object either transversely or longitttilinally(especially the later,. e.g.. an axial chloroplast).

Bacilliformi rod- shaped.

Bilobed; with two lobes or extensions.

243

Bipapillate: with. two`` small protrusion's; nipples.-Biscuit- shaped: a thickened pad; Pillow-shaped.

,Bivalve (wall): wall of cell which is in two sections, one usually slightly largerthan the others.

.-Blepharoplast: a/granular body in a swimming organism from which a. flagellum arises.

.,

Bristle: a stiff hair; a nee -like spine. (Mallomonas)

CapPlate: with an enlargement or a head at one end. (as in some species of Oscillatoria)

r

Qarotene: Orange-yellow plant pigment of which there are four kinds in algae; ahydrocarbon, C, H.

Chlorophyll;.- a green pigment of which there 11 are five kinds in the algae chlorophyll-a, occurring in all of the algal divisions.

Chloroplast: a body of various shapes within the cell containing the pigments of whichchlorophyll is the predominating one.

Chromatophore: body 'within a cell contining the pigments of which some other tanchlorophyll is the predo inating one;" may be red, yellow, yellow-greenor brown. Go, t

Chrysolaminarin: See leucosin.

Coenobium: a colony with 2N number of cells. (Scenedesmus)'

Coe ocytia! with thultinucleate cells, or -cell-like units; a multinucleatp-,...non-cellulatplant. (Vaucheria)

Collar: A thickened ring or neck surrounding the opening in a shell or lorica throughwhich a flagellum projects from the 4nclosed organism. (Trachelomonas)

Colony: a group or closely associated cluster of. cells, adjoined or merely inclosed py a-a common investing mucilage or sheath; cells not arranged in a linear series toform a filament;,4either aggregate or coenobium.

Constricted: cut in or ipOseds usually form two opposite points on a cell so that anisthmus is formed between two parts or cell halves; indented as at thejoints between cells of a filament. (Cosmarium)

9Contractile vacuole: a small vacuole (cavity) which, is bounded by a membran that

pillsates, expanding and contracting.

Crenulate; wavy with small scallops; with small crenations,,

4 Crescent: an arc of a circle; a curved figure tapering to horn-like points from a wider,cylindrical midiegion,

-

- .Cross wall: a cyoss petition.

L 24

1,

Cup- shaped:

Cylindrical:

Daughter

a mere or less complete plate (as a chloroplast) which lies just within -

the cell w41, open at one side to forlif a cup.

a figure, round in cross section). elongate. with parallel lateral marginswhen seen 'from the side, the ends square or truncate. (H7alotheca)

cells: cells produced directly from the division of 'a primary or parent cell;cells produced from the same mother cell.

Dichotomous: dividing or branched by repeated forkings, usually into two equal portionsOD segments.

Pisc; Disc-shaped: a flat (usually circular) figure: a circular plate.

Distal: the forwar or anterior end or region as op osed to the basal end.

Ellipsoid:

Epiphyte:

an ellipse, a plane figure with curved margins, the poles more sharplyrounded than the lateral margins of an elongate figure.

living upon a plant,' sometimes living 'internally' also.

Euplankton: true or openwater plankton (floating) organisms.

Eye spot: a granular or complex of granules (red or browil) sensitive t6 ligh) andrelated to responses to light by swimming organisms of spores. (Pandorina)

False Branch: a, branch formed by lateral growth of one or'both ends of a brokenfilament a branch not formed by lateral division of cells in an unbrokenfilament. (Tolypothrix)

Filament: a thread of cells; one or more rows of cells; in the ikuegreen ,algae thethread of cells together with a sheath that inay.be present, the thread of .cells alone referred `to as a trichome.

*Flagellum (flagglla): a relatively coarse, whip-like organ of locomotion, arising from

a special, granule, the *blephIroplast, within a cell. (Eugl4na)

Fucoxanthin: a broWn pigment predominant in the Phaeophyta and Chrysophyta. (Synura)

Fusiform: a figure broadest in the iijuidegion and gradually tapering to both poles Whichmay be acute or bluntly rounded; shaped like a spindle. (Closferium)

Gamete: a sex cell; cells which unite to produce a fertilized egg or zygospdre.

Gas vacuole: See pseudovacuole.

/Gelatin (gelatinous) a mucilage-like substance.

Glycogen: a starch -like Storage product questionably identified in food granules of theCyanopjyta. (Chroodocpcus)" j.

Gregarious: an association; groupings of individuals not necessarily joined together butclosely associated.

.245 26-21

I.

C

'

, .

."

Gullet: a canal leading frdrn the opening of flagellated cells into the reservoir in theanterior end. (Euglena)

Gypsum: granules of calcium sulphate which occur in the vacuoles of somedesmids. (Closterium)

Haernatoohrome: a red orbrange pigment, . especially in some Chlorophyta rand,Euglenophyta, which masks the green chlorophyll,

Heterocyst: an enlarged cell in some of the filamentous" blue- green algae, usuallyempty and different in shape from the vegetative cells, (Anabeana) . e,

Hold-fast cell: a basal cell'of a filament or thallus, differentiated to form anan attaching organ: (Oectgonium)

H- pieces: wall of overlapping H- shaped structures. T rib onema)

Intercalary: arranged in the same series, as spores' or heterocysts occur inseries with the vegetative cells rather than 'being terminal or lateraliN.,(heterocyst of Anabeana) .

Laminate: plate-like; layered...

Literal groove: a groove in Dinoflagellates encircling the cell. (CeratiumY

Leucosin: a whitish food reserve characteristic of many 'of,the Chrysophytam especiallythe Heterokontae; gives S. metallic lustre to cell'CoMents. (Difibbryor,1),

Ili

Lorica: a shell-like structure of varying shapes which houses ai4 organism, .has anopening through which organs of locomotion are.' extended. (11.nobryon)

4

. Lunate: crescent-shaped; as of the new moon shape. (Selenastrum)

Median construction: See constricted.

Metabolic: changing shape in

Micron:

S.

tion-es in many Euglena.

a unit of microscopical measurement:._ 1 if a millimeterby using a micrometer' in the eyepiece of thecalibrated with a standard stage micrometer; expressed the symbol..

microsco . as been

Moniliform:"

Mother.

MUltinucleate:

Multiteriate:

Oblong:

I et

arranged like a string of beads; beadlike; lemon-shaped. '( Anabeana)

the -cell which by mitosis or by internal cleavage gives rise toogler' cells (usually spores).

with many nuclei.o

.

cells arranged in more tligr one row; a filament two or more cells indeter. (Stigeonema)

-motion cau sed by cilia or flagella. (Volvox)

.a curved figtxice, elongate With the ends broadly rounded but more"than the lateral margins.

. . ..,

, ,.

. %., 2.46

6'

sharply curved.

Obovate: an ovate figure, broader at the anterior end than at the posterior..

Oogonium: a female .sex organ, usually an enlarged cell; an egg se.

Oval: an elongate, curved figure with convex margins and with ends broadly andsymmetrically curved but more sharply. so than the, lateral margin.

Ovoid: shaped like an egg; a carved figure broader at one end than at the other.

Paramylum: a solid, starch-like storage product in the Euglenophyta. (Euglena)

Parietal: along the wall; arre.neekl at the circumference; marginal as opposed tocentral or azial in location.' .

Pellicle: a thin membrane. ( Euglena)

Peridinin: a brown pigment characteristic of the Dinoflagellatg. (Ceratium)

Perip1ast: bounding membrane of cells in Euglenoids and Chrysophytes.

P,hycocyanin: 'blue pigment lound in the Cyanophyta, and in some Rhodophyta.It

Phycoerythrin: a, red pigment found in the Rhodophta, and in some Cyanophyta.0%,

Plankton: organisms srffting in the ivater, or if swimming, not able to move4against currents.

ti

Plastid: a body or organelle of the cell, either containing pigments or in someinstances colorless.

Plate: sections, p6lygonal in shape, composing the cell wall of some Dinoflagellata(the thecate or armored dinoflagellates).

Poster-ior: toward the rear; the end oppoSite the forward (anterior) end of a cellor of an organism.' ° .

-Protoplast: the living part of the cell; the cell membrane and its contents usually

.1

enclosed by a'ce1.1 wall of dead material.

Pseudocilia: meaning false cilia; flagella-like structures not 'used for locomotion asin Apiobystis and Tetraspora.

Psgrdopa'renchymatious: a false cushion; ,a pillow like mound of cells (usually attached)which. actually is a` compact series of short, often branchedfilaments. (Coledchaete)

Pseudovacuole: . Meaning a f se vacuole; a pocket in the cytoplasm of many blue-greenalgae Which co ains gas ors mucilage; is light refractive. (Microcystis)

Pyrenoid; .a 4otein ,body around which starch or 'paramylum collects in a cell, usually,

1

_buried in a chloroplast but sometimes free within the cytoplasm. (Oedogonium)

Pyiform: pear- shaped. (Pandorina)

t

26-,23

,

Reniform: kidney- shaped; bean- shaped. (Dimorphocacus)

Replicate:: infolded; folded back as in the cross walls of sonic specks of Spirogyra; nota plane or straight wall.

Reticulate: ntt4ii, arranged fo form a network; with ormings.

Scale: siliceous or inorganic material covering the cell. (Mallomonas)

Semicell: a ,eell-half, as in the Plaeoderm desmids in which the cell has two partsthat Pare mirror images of one another, the two parts often connected by a *.

narrow isthmus.' (Stain estrum)

Septum: a cross-partition, cross wall or a membrane complete or incomplete throughthe short diameter of, a cell, sometimes parallel with the long axis.

Setae: a hair, usually arising from within a cell wall; or a hair-like extensionibrined,py tapering of a filament of cells to a fine point.

Sheath: a covering, usually of mucilage,: soft or firm; the covering of a colonyof cells, or an_envelopd about one or more filaments of cells.

,Siphonous: a tube; a thallus without cross partitiOns (Vaucheria)

Solitary: unicellular; solitary. (Chlamydomonas).,

Spine: a sharply-pointed projection from the eel], wall. (Mallomonas)

Sproangium: a cell4 (-sometimes an unspecialized vegetative cell) which gives rise to. spores; the case which forms aboi t the zygospores in the Zygnemalales.

Star- shape d: See stellate

,Stellate: with radiating projections from a common center; star-like. (Zygnema)

'Stigma: see eyespot.

- Sutiire:, a groove between plates, as in some Dinalagellata; a cleft-like crack or line;in some zygospores of the Zygnematacete: (Ceratium) . - '

,Thallus: a plant body which' is not differentiated into root, stem and leal organs; a, frodd; the algal plant. ;

,

'1

Theca; Thecate: a firm outer Agfall;'a shell, sometimes..with plgtes, as in theJ '. .Dinof ia gellata, (Peridinium) .'

. 0 .iTest: a shell Or covering external to the yell it lf. See Lorica.

'

Transverse furrow (groove): a-groove extending ground the cell as in theDinoflagellata. ( Ceratium)

Trichrome: in blue-greent% a series of cells joined end to end. (Oadilltoria)

26.44

I

24 .

-c*

True Branch:

Tychoplankton:

a branch formed by means of lateral division of cells in a main filament.Includes all branched algae except those blue -green algae with falsebranching.

the plankton of waters near shore; organisms floating andamong weeds and in algal mats, not in the open water ofor stream.

Undulate: regularly wavy.

Unicellular:

Vegetative:

Xanthophyll:

Zoospore:

Seeviolitary..

entangleda lake

referring to a non-reproductive stage, activity, or.cell as opposed toactivities and stages involved in reproduction, especially sexual reproduction.

,

a yellow pigment of several kinds associated With chlorphyll, C46H5602

an nimal -like spore equipped with flagella and usually .with an eye-spot.

This key was prepared by Dr. Matthew H. Hohn,Professor of Biology, Central MiohiganUniversity, Mt. Pleasant, Michigan.

24 9

Descriptors: Plankton, Identification Keys

26-25s

PAGES 1-198"FIELD KEY TO SOME GENERA OF ALGAE" AND"CILIATED PROTOZOA" REMOVED DUE TO COPYRIGHT RESTRICTIONS.

Documents

DOCUMENT RESUME L - ERIC · POP DE. O. Jun 80, NOTE 250p.; Contains ... U.S DEPARTMENT OF EDUCATION NATIONAL INSTITUTE OF EDVCATION ... Laboratory: Proportional Counting. of Mixed