Species diversity of heterotrophic flagellates in White Sea littoral sites

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European Journal of

PROTISTOLOGY

0932-4739/$ - se

doi:10.1016/j.ej

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(D.V. Tikhonen

European Journal of Protistology 42 (2006) 191–200

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Species diversity of heterotrophic flagellates in White Sea littoral sites

Denis Victorovich Tikhonenkova,�, Yuri Alexandrovich Mazeib,Alexander Petrovich Mylnikova

aInstitute for Biology of Inland Waters, Russian Academy of Sciences, BOROK, Yaroslavskaya obl. 152 742, RussiabDepartment of Zoology and Ecology, Penza V.G. Belinsky State Pedagogical University, Lermontova str. 37,

PENZA 440026, Russia

Received 5 January 2006; accepted 20 May 2006

Abstract

The species diversity and distribution of benthic heterotrophic flagellates in sediment samples from along the salinitygradient in the Chernaya River Estuary and from Velikaya Salma Strait (Kandalaksha Bay, the White Sea) wereinvestigated during August 2004. One hundred and six taxa have been identified by means of phase and interferencecontrast light microscopy and transmission electron microscopy. The majority of observed flagellates werebacterivores. The species diversity of the following groups: choanoflagellates, euglenids, kinetoplastids, bicosoecids,chrysomonads, thaumatomonads and flagellates Incertae sedis was the highest. Ancyromonas sigmoides andPetalomonas pusilla were the most common species. The species richness was lowest in the brackish water estuarinepart with salinity levels between 5% and 8%. The distribution of heterotrophic flagellates conforms to the so-called‘‘rule of critical salinity’’, possessing, apparently, the same universal character for organisms of different size levels.

Heterotrophic flagellate communities in these littoral sites were highly heterogeneous. The curve of ‘‘cumulativespecies number vs. sampling effort’’ is well fitted by equation S ¼ 21.17N0.50 and unsaturated, which indicates thatmore intensive investigations of the heterotrophic flagellates in the White Sea should be expected to reveal more species.r 2006 Elsevier GmbH. All rights reserved.

Keywords: Heterotrophic flagellates; The White Sea; Critical salinity; Species diversity; Flagellate community

Introduction

Heterotrophic flagellates (HF) are an obligatorycomponent of microbial communities in all types ofecosystems containing fluid water. Planktonic HF inmarine and estuarine water areas have been extensivelyinvestigated (e.g. Berninger et al. 1991). In contrast, thespecies composition of benthic HF communities has beenless well studied (Larsen 1987; Larsen and Patterson1990; Lee and Patterson 2000). The species richness of

e front matter r 2006 Elsevier GmbH. All rights reserved.

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sses: [email protected], [email protected]

kov).

benthic HF in the Arctic is currently known for theBarents Sea (Mazei and Tikhonenkov 2006; Mylnikovand Zhgarev 1984), whereas there are no data in theliterature on HF diversity in the White Sea. Thus, the aimof our study was to investigate species diversity of benthicHF in littoral sites of the Chernaya River Estuary and theVelikaya Salma Strait (Kandalaksha Bay, White Sea), inrelation to physico-chemical features, especially salinity.

Materials and methods

This investigation was carried out in August 2004at littoral sites in the Chernaya River Estuary

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ARTICLE IN PRESSD.V. Tikhonenkov et al. / European Journal of Protistology 42 (2006) 191–200192

(Kandalaksha Bay, the White Sea: 3219505000 E,6615108300 N), the Velikaya Salma Strait (northern shoreof the Kindo Peninsula, opposite the Small EremeevskyIsland: 3310800400 E, 6615503500 N) and in Polupresnoe(Kislo-sladkoe) Lake which is separated by a narrowdam from the strait area: 3310800400 E, 6615503300 N(Fig. 1a).

The estuarine samples were collected from fivestations (numbers 1–5), located along the coastline fromthe marine towards the riverine part (Fig. 1b); at eachstation, two samples were taken, one from the interstitial(5 cm depth) and the other from surface regions of thesediments from each of the upper, middle and lowerlevels on the shore. In total, six samples were collected ateach station; all samples from one station were taken onthe same day. Material was taken in the morning at thetime of the ebb. The tidal range is about 2m.

In the Polupresnoe Lake and the Velikaya SalmaStrait, two samples were taken from the surface of thesediments at each place on the same day.

The sediment samples, including water, were placed in50-ml flasks, and maintained at approximately 3 1Cduring transportation to the laboratory. Two replicates,each of 5ml, were analyzed in Petri dishes from everysample in order to study the species composition.

Light microscopical observations were made with theaid of Biolam-I microscope (Russia) equipped withphase contrast and water immersion objectives giving atotal magnification 770� , as well as a Reichert(Austria) microscope with Nomarski interference con-

Fig. 1. (a), (b) Location of studied sites: (a) western part of the

Kandalakscha Bay: 1–2, sampling areas; (b) Chernaya River

estuary: 1–5, sampling sites.

trast and glycerin immersion objectives (1000� ). Themicroscopes were equipped with an analog video cameraAVT HORN MC—1009/S, which was connected to aPanasonic NV-HS 850 video recorder. Image acquisi-tion in VHS and S-VHS modes, followed by digitizationof images and preservation of fragments of video film asAVI files, was carried out in order to facilitate moreprecise identification of HF. HF were identified bymeans of observations on living cells with the exceptionof scale-bearing species. Drops of suspended scale-bearing cells were placed on copper grids coated withFormvar film and prepared as whole mounts by themethod described by Moestrup and Thomsen (1980).Grids were shadowed with tungsten oxide, and wereobserved with a JEM-100C transmission electronmicroscope.

The authors have used the systems of classifyingeukaryotes and protists presented in papers by Adl et al.(2005) and Lee and Patterson (2000).

The levels of salinity, pH and Eh in samples weremeasured in situ, simultaneously with the collection ofeach sample, using a Kruss S-10 salinometer, a Hannapocket-sized pH meter with automatic temperaturecompensation and a Redox meter.

Results and discussion

The results of measuring environmental variables(Table 1) show a considerable range of hydrochemicalparameters between the biotopes. The salinity of waterfrom estuarine samples varied between 24% in themarine part and 0.2% in the riverine part. The criticallevel of 5–8% (see below) and maximum variability ofsalinity were recorded in the estuarine station 4 and inPolupresnoe Lake. The biggest within-station variationwas at station 4 (from 0.5% to 14%), where the lowestsalinity was recorded from the surface samples takenfrom the lower level on the shore, and the highestsalinity from the interstitial samples taken from theupper level on the shore. The salinity of the VelikayaSalma Strait was 25% in all samples, and at thePolupresnoe Lake sediment surface (30 cm depth)stations, it varied between 4% and 13% in differentparts of the lake with a mean of 8.5% (from twomeasurements on the same date). The pH fluctuatedmore within any station than the means differedbetween stations, whereas reduction–oxidation (redox)properties show a gradation (anaerobic conditions areencountered more frequently at the marine stations thanat the lower salinity stations). Detailed hydrochemicalcharacteristics of the biotopes studied have beenpresented by Burkovsky and Mazei (2001), Mazeiet al. (2002) and Schaporenko (2003).

One hundred and six species and forms of HF wereidentified in the investigated biotopes. Five forms of

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Table 1. Basic hydrochemical characteristics of investigated biotopes

Factors Stations

Chernaya River Estuary

1 2 3 4 5

Salinity (%)

Mean 23.7 18.2 11.6 6.6 1.8

Standard deviation 0.6 5.6 4.8 6.4 2.8

Range 23.0–24.0 12.0–24.0 9.0–20.0 0.5–14.0 0.2–5.0

pH

Mean 7.3 7.6 7.7 7.3 7.3


Range 6.7–7.9 7.1–8.1 7.4–8.9 6.6–8.4 6.3–7.8

Eh (mV)

Mean �238 �119 �80 �76 �37


Range �100y�270 �80y�180 �20y�170 �10y�150 �80y70

D.V. Tikhonenkov et al. / European Journal of Protistology 42 (2006) 191–200 193

bikont HF (numbered flagellate sp. 1–5) could not beidentified (Table 2).

Flagellates from the following groups: choanoflagel-lates, euglenids, kinetoplastids, bicosoecids, chrysomo-nads, thaumatomonads and a group of Incertae sedis

species, were dominant in terms of species richness. It isnotable that choanoflagellates and bicosoecids were thedominating taxa in the White Sea microbenthic com-munities, although they are often considered to bemainly planktonic or periphytonic forms (Zhukov andKarpov 1985; Zhukov 1993). This finding confirms theview that a great number of water column HF areubiquitous species (Ekebom et al. 1995/1996), which,furthermore, may be explained in terms of the similarityof microconditions between the water column and thesediments (Bamforth, 1963). Only euglenids, thaumato-monads and some Incertae sedis species among thenoted dominants are generally regarded as typicalbenthic inhabitants.

Cercomonads are typical protists in benthic condi-tions rich in organic matter (Hanel 1979; Zhukov andMylnikov 1983). However, species from the genusCercomonas were rare in samples from the White Sea.Most likely this is connected with physiological pre-ferences of these organisms, which are extremelysensitive to increasing salinity (Arndt et al. 2000;Mylnikov 1983a). The low species richness of cercomo-nads in the lowest salinity estuarine station 5 does notlook strange, since large salinity fluctuations, exceedingthe physiological tolerance of these organisms, occur atthis site (Mazei and Burkovsky 2002).

The highest numbers of species were from the generaSalpingoeca (9 species), Petalomonas (7), Paraphysomo-

nas (6) and Cafeteria (5). The most common species(observed in more than 50% of samples) were Ancyr-

omonas sigmoides (83%), Petalomonas pusilla (83%),Bodo designis (79%), Cafeteria roenbergensis (69%),flagellate sp.1 (69%), Paraphysomonas imperforata

(59%), Paraphysomonas vestita (55%) and Metopion

fluens (52%). However, 31 species (29% of the totalspecies richness) were detected in samples only once,which is evidence of great heterogeneity of the commu-nity.

Nine species (Ancyromonas sigmoides, Bodo designis,B. saliens, Cafeteria minuta, C. roenbergensis, Paraphy-

somonas vestita, Petalomonas pusilla, Phyllomitus gran-

ulatus, Rhynchomonas nasuta) were found at all stationsinvestigated, which is evidence of their wide ecologicaldistribution.

The majority of flagellates observed were bacterivor-ous (according to Sanders (1991); Larsen and Patterson(1990); Tikhonenkov pers. comm.). Among flagellatesencountered, 16 species of possible predators wereidentified: Allantion tachyploon, Amphidinium incolora-

tum, Amphidinium sp., Colpodella sp., Kathablepharis

remigera, Kathablepharis sp., Katodinium asymmetricum,Katodinium sp., Metopion fluens, Metromonas grandis,M. simplex, Multicilia marina, Phyllomitus apiculatus,Ph. granulatus, Rhynchobodo simius, Stephanopogon

colpoda. The greatest number of carnivorous species(12) was observed at depths of 5 cm at the interstitialsites, whereas at the surface of the sediments this speciesrichness was only half this level. The number ofomnivorous species was not high, but increased inboth marine (stations 1 and 2) and riverine habitats(station 5). The community trophic structure variedslightly from station to station (Fig. 2). Bacterio-detritophage collector feeders and filter feeders, whichplay important roles in the control of abundance,production and structure of bacteriocoenoses (Berninger

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Table 2. Species composition and % frequency of occurrence of species of heterotrophic flagellates at the stations studied

Stations

Species and forms 1 2 3 4 5 P and V

Amoebozoa (Luhe 1913) emend. Cavalier-Smith 1998

Mastigamoeba sp. 0.0 0.0 16.7 0.0 0.0

Multicilia marina Cienkowski 1881 50.0 0.0 33.3 0.0 0.0 P+V

Phalansterium sp. 0.0 0.0 0.0 0.0 0.0 V

Choanomonada Kent 1880

Codonosiga botrytis Kent 1880 16.7 66.7 16.7 0.0 0.0 P

Diploeca angulosa de Saedeleer 1927 0.0 0.0 16.7 0.0 0.0

Monosiga ovata Kent 1880 33.3 33.3 16.7 0.0 0.0 P

Salpingoeca amphoridium Clark 1868 0.0 0.0 0.0 25.0 0.0 P

S. globulosa Zhukov 1978 0.0 0.0 16.7 0.0 0.0

S. infusionum Kent 1880 33.3 50.0 50.0 0.0 0.0 P+V

S. marina James-Clark 1867 50.0 33.3 0.0 25.0 0.0 P

S. massarti de Saedeleer 1927 0.0 16.7 0.0 0.0 0.0 P

S. megachelia Ellis 1929 0.0 0.0 0.0 0.0 0.0 P

S. pelagica Laval 1971 0.0 0.0 0.0 25.0 0.0

Salpingoeca sp. 0.0 0.0 0.0 0.0 0.0 P

S. vaginicola Stein 1878 0.0 16.7 0.0 0.0 0.0

Stephanoeca diplocostata Ellis 1930 16.7 0.0 0.0 0.0 0.0 V

Cercozoa Cavalier-Smith 1998, emend. Adl et al. 2005

Cercomonadidae Kent 1880, emend. Mylnikov and Karpov 2004

Cercomonas aff. agilis (Moroff 1904) Mylnikov and

Karpov 2004

0.0 0.0 0.0 0.0 33.3

C. angustus (Skuja 1948) Mylnikov and Karpov 2004 16.7 0.0 0.0 0.0 0.0

C. granulatus Lee et Patterson 2000 16.7 16.7 0.0 0.0 0.0

Cercomonas sp. 0.0 0.0 0.0 25.0 0.0

Heteromitidae Kent 1880, emend. Mylnikov 1990, emend. Mylnikov and Karpov 2004 Incertae sedis

Allantion tachyploon Sandon 1924 16.7 0.0 0.0 0.0 66.7 V

Protaspis gemmifera Larsen et Patterson 1990 16.7 0.0 0.0 0.0 0.0

P. simplex Vørs 1992 16.7 16.7 0.0 0.0 0.0

P. verrucosa Larsen and Patterson 1990 16.7 0.0 16.7 0.0 0.0

Protaspis sp. 50.0 16.7 0.0 0.0 0.0 V

Thaumatomonadida Shirkina 1987

Thaumatomonas/Thaumatomastix sp. 83.3 33.3 16.7 0.0 0.0

Thaumatomonas seravini Mylnikov et Karpov 1993 0.0 0.0 0.0 0.0 33.3

Thaumatomonas sp. 16.7 0.0 0.0 0.0 0.0 V

Incertae sedis Cercozoa

Massisteria marina Larsen et Patterson 1990 16.7 0.0 16.7 0.0 0.0 P+V

Metopion fluens Larsen et Patterson 1990 50.0 100.0 0.0 50.0 33.3 P+V

Cryptophyceae Pascher 1913, emend. Schoenichen 1925

Goniomonas amphinema Larsen et Patterson 1990 0.0 16.7 0.0 0.0 0.0

G. pacifica Larsen et Patterson 1990 0.0 0.0 0.0 0.0 33.3

Goniomonas sp. 0.0 33.3 0.0 0.0 33.3

Bicosoecida Grasse 1926, emend. Karpov 1998

Caecitellus parvulus (Griessmann 1913)

Patterson et al. 1993

16.7 0.0 16.7 0.0 0.0

Cafeteria ligulifera Larsen et Patterson 1990 16.7 16.7 16.7 0.0 0.0

C. marsupialis Larsen et Patterson 1990 50.0 0.0 33.3 25.0 33.3

C. minuta (Ruinen 1938) Larsen et Patterson 1990 16.7 33.3 33.3 25.0 33.3

C. roenbergensis Fenchel et Patterson 1988 100.0 50.0 66.7 50.0 66.7 P+V

Cafeteria sp. 33.3 0.0 0.0 0.0 0.0

Discocelis saleuta Vørs 1992 83.3 66.7 33.3 25.0 0.0

D.V. Tikhonenkov et al. / European Journal of Protistology 42 (2006) 191–200194

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Table 2. (continued )

Stations


Pseudobodo tremulans Griessmann 1913 0.0 16.7 16.7 25.0 0.0 V

Pseudodendromonas vlkii (Vlk 1938) Bourrelly 1953 0.0 0.0 0.0 0.0 33.3

Chrysophyceae Pascher 1914

Paraphysomonas bandaiensis Takahashi 1976 0.0 0.0 0.0 50.0 0.0

P. curcumvallata ssp. curcumvallata Thomsen 1981 0.0 0.0 0.0 25.0 0.0

P. imperforata Lucas 1967 100.0 33.3 66.7 0.0 33.3 P+V

P. vestita (Stokes 1885) De Saedeleer 1929 100.0 33.3 83.3 50.0 33.3

P. vestita (Stokes 1885) De Saedeleer 1929 spp. vestita 0.0 0.0 0.0 50.0 33.3

Paraphysomonas sp. 0.0 83.3 16.7 0.0 66.7

Spumella sp. 0.0 0.0 0.0 0.0 66.7 P

Pedinellales Zimmermann, Møestrup and Hallfors 1984

Actinomonas mirabilis Kent 1880 0.0 33.3 0.0 0.0 0.0 V

Ciliophrys infusionum Cienkowsky 1876 33.3 33.3 16.7 25.0 0.0 V

Pteridomonas danica Patterson et Fenchel 1985 0.0 33.3 33.3 25.0 33.3 P

Dinoflagellata Butschli 1885, emend. Fensome, Taylor, Sarjeant, Norris, Wharton, and Williams 1993, emend. Adl et al. 2005

Amphidinium incoloratum Campbell 1973 0.0 0.0 0.0 0.0 0.0 P

Amphidinium sp. 0.0 0.0 0.0 0.0 0.0 V

Katodinium asymmetricum (Massart 1920) Loeblich 1965 16.7 0.0 16.7 25.0 0.0 V

Katodinium sp. 16.7 0.0 16.7 0.0 0.0

Colpodellida Cavalier-Smith 1993, emend. Adl et al. 2005

Colpodella sp. 0.0 0.0 0.0 0.0 33.3 P

Heterolobosea Page and Blanton 1985

Percolomonas cosmopolitus (Ruinen 1938)

Fenchel et Patterson 1986

50.0 66.7 16.7 0.0 0.0 V

P. denhami Tong 1997 33.3 0.0 0.0 0.0 0.0

P. membranifera Larsen et Patterson 1990 16.7 16.7 0.0 0.0 0.0

Euglenida Butschli 1884, emend. Simpson 1997

Anisonema graciale Larsen et Patterson 1990 0.0 16.7 0.0 0.0 0.0

A. trepidum Larsen 1987 0.0 0.0 0.0 0.0 33.3

Heteronema exaratum Larsen et Patterson 1990 16.7 0.0 0.0 0.0 0.0 V

H. ovale Kahl 1928 16.7 0.0 0.0 0.0 0.0 V

Petalomonas abscissa (Dujardin 1841) Stein 1859 0.0 0.0 0.0 0.0 0.0 P

P. labrum Lee et Patterson 2000 33.3 0.0 0.0 0.0 0.0 V

P. minor Larsen et Patterson 1990 50.0 33.3 33.3 25.0 0.0 P+V

P. minuta Hollande 1942 0.0 16.7 16.7 25.0 33.3 P

P. pusilla Skuja 1948 100.0 100.0 83.3 25.0 66.7 P+V

Petalomonas sp. 1 66.7 16.7 33.3 0.0 0.0

Petalomonas sp. 2 0.0 16.7 33.3 0.0 0.0

Ploeotia corrugata Larsen et Patterson 1990 16.7 0.0 16.7 0.0 0.0 P

Urceolus pascheri Skvortzov 1924 16.7 0.0 0.0 0.0 0.0

Kinetoplastea Honigberg 1963

Bodo curvifilis Griessmann 1913 0.0 0.0 0.0 25.0 0.0

B. designis Skuja 1948 66.7 100.0 83.3 75.0 100.0 P

B. saliens Larsen et Patterson 1990 33.3 50.0 50.0 25.0 33.3 P+V

B. saltans Ehrenberg 1832 16.7 0.0 0.0 0.0 100.0

Cryptaulax marina Throndsen 1969 0.0 0.0 0.0 0.0 0.0 V

Klosteria bodomorphis Mylnikov et Nikolaev 2003 16.7 50.0 33.3 50.0 0.0 P

Parabodo nitrophilus Skuja 1948 0.0 0.0 0.0 0.0 33.3

Procryptobia sorokini (Zhukov 1975) Frolov,

Karpov et Mylnikov 2001

33.3 0.0 50.0 0.0 33.3

Rhynchobodo simius Patterson et Simpson 1996 0.0 0.0 0.0 0.0 66.7

Rhynchomonas nasuta (Stokes 1888) Klebs 1892 16.7 16.7 33.3 25.0 100.0 P


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Table 2. (continued )

Stations


Incertae sedis Kinetoplastea

Bordnamonas tropicana Larsen et Patterson 1990 100.0 50.0 33.3 0.0 0.0

Incertae sedis Eukaryota

Amastigomonas caudata Zhukov 1975 33.3 0.0 0.0 0.0 33.3

A. debruynei De Saedeleer 1931 83.3 0.0 0.0 0.0 0.0 P+V

A. griebenis Mylnikov 1999 33.3 0.0 0.0 0.0 0.0

A. mutabilis (Griessmann 1913) Molina et Nerad 1991 66.7 16.7 50.0 0.0 0.0

Ancyromonas contorta (Klebs 1893) Lemmermann 1910 16.7 0.0 16.7 0.0 0.0

A. sigmoides Kent 1880 100.0 100.0 83.3 50.0 33.3 P+V

Clautriavia cavus Lee et Patterson 2000 16.7 0.0 0.0 0.0 0.0 V

Flagellate sp.1 83.3 100.0 83.3 75.0 0.0 V

Flagellate sp.2 33.3 16.7 16.7 0.0 0.0

Flagellate sp.3 33.3 33.3 0.0 50.0 0.0

Flagellate sp.4 33.3 16.7 16.7 0.0 0.0

Flagellate sp.5 16.7 0.0 0.0 0.0 0.0

Glissandra innuerende Patterson et Simpson 1996 16.7 0.0 0.0 0.0 0.0

Kathablepharis remigera (Vørs 1992)

Clay et Kugrens 1999

16.7 0.0 0.0 0.0 0.0 V

Kathablepharis sp. 0.0 16.7 0.0 0.0 0.0 P

Kiitoksia ystava Vørs 1992 16.7 16.7 0.0 0.0 0.0 P

Metromonas grandis Larsen et Patterson 1990 33.3 0.0 16.7 25.0 33.3

M. simplex (Griessmann 1913) Larsen et Patterson 1990 33.3 16.7 0.0 50.0 33.3 P+V

Phyllomitus apiculatus Skuja 1948 0.0 0.0 0.0 0.0 33.3

P. granulatus Larsen et Patterson 1990 50.0 66.7 50.0 50.0 33.3 P

Phyllomitus sp. 0.0 0.0 0.0 0.0 33.3

Stephanopogon colpoda Entz 1884 16.7 0.0 0.0 0.0 0.0

1–5, Chernaya River Estuary stations from highest to lowest salinity as in Table 1. P and V, present at other sites; P, present in Polupresnoe Lake

samples; V, present in Velikaja Salma Strait samples.


et al. 1991, 1993; Sanders et al. 1992; Weisse 1991), wereprevalent at all sites. The prevalence of bacterivorousflagellates in microbial communities is known for waterbodies of many types (Kosolapova 2005).

The species richness of communities fluctuated from 7to 38 species per sample. The average number of speciesin a sample was 17.84; however, this quantity changedappreciably from station to station (Fig. 3). Samplesfrom the marine part of the estuary were the richest,whereas the community in the brackish water part of theestuary (station 4) was characterized by the lowestspecies richness (12). Such a pattern of species richnessdistribution may be explained in terms of ‘‘the brackishwaters paradox’’ (Remane 1934): at salinities between5% and 8% marine and freshwater fauna are not mixed(Khlebovich 1974). Many of the species observed(Cercomonas aff. agilis, Colpodella sp., Parabodo nitro-

philus, Phyllomitus apiculatus, Phyllomitus sp., Pseudo-

dendromonas vlkii, Rhynchobodo simius, Spumella sp.,Thaumatomonas seravini) are known from fresh waters(Mazei et al. 2005; Mylnikov and Kosolapova 2004;

Skuja 1956; Zhukov 1993), and were found during ourinvestigation in biotopes with a salinity of 5% or less.However, the halopathy curve obtained for HF showsless expressed depression, than is displayed for otherorganisms (Mordukhai-Boltovskoi 1953, 1960). This isprobably a consequence of the large number of euryha-line species in the HF community (compared with largesize organisms) such as Bodo saltans, Bodo designis,Goniomonas pacifica, Percolomonas cosmopolitus, Amas-

tigomonas caudata, Rhynchomonas nasuta, Cafeteria

roenbergensis, Petalomonas minor, Ploeotia corrugata,whose halotolerance has been shown in experimentaland natural conditions (Arndt et al. 2000; Gorjachevaet al. 1978; Mylnikov 1983a; Patterson and Simpson1996; Zhukov 1970).

Morphological features of HF species identified fromboth marine and low-salinity parts of the estuary aresimilar to and correspond with those reported elsewhere(Al-Quassab et al. 2002; Larsen and Patterson 1990,2000; Mylnikov and Kosolapova 2004; Patterson andSimpson 1996; Skuja 1956; Vørs 1992; Zhukov 1971,

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Fig. 2. Trophic structure of heterotrophic flagellate communities at the seven sites, based on published information on the diet of

the constituent species. 1–5, Chernaya River estuarine stations; 6, Polupresnoe Lake; 7, Velikaja Salma Strait littoral.

Fig. 3. Species richness in communities at each station. Error bars represent the standard error of the arithmetic mean.


1993), with the exception of contractile vacuoles, which,as a rule, are absent in organisms dwelling at higher-salinity (more than 10%) estuarine parts. It is possiblethat in this case we observe different ‘‘physiologicalspecies’’, for which morphological criteria cannot beapplied to distinguish them taxonomically. According tothe recent report by Koch and Ekelund (2005),individuals of the same ‘‘morphospecies’’, Bodo designis,isolated from different marine, freshwater and soilhabitats, showed a different degree of tolerance tosalinity changes and were characterized by high geneticheterogeneity, so it can be proposed that the clonesinvestigated belong to several different biologicalspecies.

Even in the investigated biotopes with lowest Eh-level(redox level), HF without mitochondria, such asdiplomonads, retortomonads and trichomonads, werenot observed, while they are generally the most commonflagellates in anoxic habitats (Brugerolle and Muller2000; Lackey 1932, 1938; Mylnikov 1983b, 1985, 1991).The absence of these flagellates can be connected to thehigh salinity levels in marine parts of the estuary andsignificant seasonal fluctuations in the freshwater part,since it has been shown experimentally that anaerobicflagellates are very sensitive to increase of salinity

(Gorjacheva et al. 1978; Mylnikov 1983a), but areobserved in freshwater and brackish coastal sediments(Bernard et al. 2000; Fenchel et al. 1995).

The species richness of the communities formed onthe surface of the sediments and in interstitial water is atthe same level (80 and 75 species, respectively). This iscaused by large concentrations of microaerobes, facul-tative anaerobes and anaerotolerant species in thesecommunities (Bernard and Fenchel 1996), which arerepresented by euglenids, dinoflagellates, stramenopilesand choanoflagellates (Fenchel et al. 1995) that occurvery frequently in the investigated samples.

As in any study of this type, the total number ofspecies recorded (S) increased with an increase ofsampling effort (N) (Fig. 4). The rate of increasegradually reduced, although the curve did not reach aplateau, which indicates that further sampling shouldincrease the number of species recorded from the WhiteSea. The curve representing cumulative species numbervs. sampling effort, combining data from all Chernayaestuary stations, is described by the equation S ¼

21:17N0:50 (R2 ¼ 0:98). It is interesting that the powerparameter (reflecting ‘‘rate of increase’’ of speciesrichness with an increase of sampling effort) appearedrather similar to that obtained for sublittoral

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Table 3. Equations of the ‘‘cumulative species number vs.

sampling effort’’ curves for HF and ciliates in the Chernaya

River Estuary based on six samples at each station

Stations Heterotrophic

flagellates

Ciliates

1 S ¼ 25:87N0:54;

R2 ¼ 0:99

S ¼ 47:89N0:19;

R2 ¼ 0:982 S ¼ 18:77N0:53;

R2 ¼ 0:99

S ¼ 33:09N0:29;

R2 ¼ 0:983 S ¼ 15:84N0:60;

R2 ¼ 0:99

S ¼ 33:89N0:27;

R2 ¼ 0:994 S ¼ 11:73N0:73;

R2 ¼ 0:99

S ¼ 19:42N0:44;

R2 ¼ 0:995 S ¼ 15:38N0:74;

R2 ¼ 0:99

S ¼ 25:32N0:32;

R2 ¼ 0:99

1–5, Chernaya River Estuary stations from highest to lowest salinity as

in Table 1.

Fig. 4. Relationship of the number of species recorded with

the ‘‘sampling effort’’.


(S ¼ 5:16N0:64) and littoral (S ¼ 8:23N0:57) Barents SeaHF communities (Mazei and Tikhonenkov 2006). Thisfact could indicate similar patterns of specific richness indifferent communities. The obtained equations for thetwo seas differ considerably with respect to the firstparameter of the equation, reflecting the level ofcommunity alpha-diversity (number of species persample). Littoral White Sea communities appeared tobe approximately 2.5� as rich in species whencompared with the littoral communities of the BarentsSea. This probably reflects features of estuarine bio-topes, which are characterized by a significant amountof accumulated organic matter (Safjanov 1987).

Ciliate community distribution had been investigatedat the same stations in the Chernaya River estuary inAugust 2000 by the same sampling methodology andanalysis (Mazei and Burkovsky 2006). Tendencies ofchanges of parameters of the equation ‘‘cumulativespecies number vs. sampling effort’’ within stations aresimilar for HF and ciliates (Table 3). The maximalsaturation of the curve is observed in marine parts ofthe estuary and minimal in the brackish water part. Thealpha species diversity gradually decreases from themarine part to brackish water, reaching a minimal valueat station 4 and increases a little at the lowest salinitystation 5.

The cumulative curve for ciliates from 30 samples wassaturated (S ¼ 39:39N0:28; R2 ¼ 0:94), unlike that forHF. The ciliate alpha-diversity is considerably higher(about 40 species observed per sample on the average)than HF (21 species), and the first parameter of theequation is considerably higher. Ciliate cumulativecurves are saturated in all stations (power parametersof equations are less than 0.45), whereas HF curves are

not saturated at any station; consequently, furthersampling should increase the number of HF speciesrecorded from all the investigated stations (throughoutthe salinity range). This can be interpreted as meaningthat the heterogeneity of microhabitats is greater for HFthan for ciliates, or the success of analyzing samples ishigher for ciliates than for flagellates.

Conclusion

Heterotrophic flagellate (HF) littoral communities ofthe White Sea are characterized by both rich speciesdiversity and heterogeneity. The HF distribution con-forms to the so-called ‘‘rule of critical salinity’’, whichobviously has universal character and may be seen evenat the level of nano-organisms. The nonsaturatedcharacter of the ‘‘cumulative species number vs.sampling effort’’ curve reveals the amount of samplingrequired to show the rich HF species diversity in theinvestigated region.

Acknowledgments

This study was supported by the Russian Foundationfor Basic Research (Grant nos. 04-04-48338 and 05-04-48180) and a grant of the President of the RussianFederation (Project no. MK-7388.2006.4).

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Documents

Species diversity of heterotrophic flagellates in White Sea littoral sites