NORMAL FLO

RA OF

BODY

DR

YU

GA

ND

AR

WHAT IS NORMAL FLORA ?

mixture of microorganisms regularly

found at on / within the body of a healthy person.

Some of these microorganisms are found in association with humans / animals

only.

Normal Flora• Definition

Normal flora is the mixture

of microorganisms (bacteria

and fungi) that are regularly

found at any anatomical site

of human body

ROLE OF NORMAL FLORA

Immunostimulation

a) They produce antibodies which may contribute to host defenses.b) Some of these antibodies may

cross react with normal tissue components.

May be source of opportunistic infections

e.g : In Patients with impaired defense Mechanisms

Protection from External Invaders a) Because of the normal flora occupy bodys

epithelial surfaces,they are able to prevent other bacteria by blocking receptors

b) competing for essential nutrients c) producing anti-bacteria substances

e.g : Fatty acids, peroxides , Bacteriocins.Nutrition Some of the normal intestinal flora e.g. E. coli & Bacteroids produce Vitamin K in the gut

which is available for use by host.

Production of Carcinogens

Some normal flora may modify, through their enzymes, some chemicals in our diets into carcinogens

e.g. Artificial sweeteners may be enzymatically

modified into bladder carcinogens.

Stimulate development of certain tissues:

Caecum and lymphatic tissues (Peyer’s patches) in GIT

ORIGIN OF NORMAL FLORA

Healthy fetus, in utero, is essentially free of MOs

Infant exposed immediately to MOs when passing

through mothers vaginal tract and then to MOs in

environment

Within few hours, oral and nasopharyngeal flora of

neonate established

Within one day, resident flora of lower intestinal

tract established

NORMAL FLORA

Human body 1013 cells 1014 bacteria

Numbers represent the number of organisms per gram of homogenized tissue or fluid or per square centimeter of skin surface.

NORMAL FLORA OF SKIN

Human adult has 2 square meters of skin

Overall, a hostile environment toward bacteria

Eccrine (simple sweat) glands release Nacl solution : hyperosmotic surface

Apocrine glands : sweat & nutrients,secrete lactic acid pH3-5

Sebaceous glands : a/w hair follicles secrete lipids ,bacteria convert it into unsaturated fatty acids which have antimicrobial activity and

strong odor

MOs that normally live on or in any part of the body

with out causing disease

Two basic types of NF:

Resident NF – normally GROW on/in indicated body site,

presence fixed in body

Transient NF – only TEMPORARILY PRESENT on/in

indicated body site, usually don’t become firmly

attached but simply die within hours

NORMAL FLORA OF SKIN

Transient microbes: in contact with environment

Resident microbes Areas of high moisture content

Axilla, groin, toe webs, perineum, scalp Superficial layers of the epidermis, upper parts of hair follicles, sebaceous glands

Staph epidermidis; Staph aureus; Corynebacteria (Propionibacterium acnes); Streptococci; Clostridium spp. (perineum and upper thighs)

Skin: 3 main micro environments

Axilla, perineum, toe webs

Hands, face and trunk

Upper arms and legs

S. epidermidis

Major inhabitant making up more than 90% of the flora

S. aureus

Nose, perineum, vulva skin

Occurrence in nasal passages varies with age being greatest in

newborns, less in adults

Neither profuse sweating nor washing significantly modifies normal

skin flora

Soap or disinfectant (hexachlorophene) diminish microbial population Soap - 30 second scrubbing

Soap: physically removes organisms Disinfectants: bactericidal

But NF rapidly replenished from sebaceous and sweat glands

Pathogenic organisms eliminated (?)

Re-exposure needed to colonize

Nosocomial infections dramatically diminished or eliminated

Antiseptics applied to the skin, as in the preparation of operation sites

generally remove the transient flora and reduce the resident organisms

Key organisms to learn in skin –

Staphylococcus epidermidis (coagulase-negative

staphylococci);

Propionibacterium sp.,

Corynebacterium sp.

SKIN NORMAL FLORA

Staphylococcus epidermidis

Staphylococcus aureus

C+, clusters

SKIN NORMAL FLORA

Propionibacterium acnes

B+, anaerobic

Diphteroids (club shaped)

Branching bacilli

SKIN NORMAL FLORA

Candida albicans

Yeast, budding

Soil & air

SKIN NORMAL FLORA

Streptococcus species

C+, pairs, chains

SKIN NORMAL FLORA

Corynebacterium

B+, aerobic

Diphtheroids

Numerous bacterial colonies on blood agar plate after swabbing from normal human skin

Staphylococcus aureus, a predominant normal microflora of skin.

HUMAN BENEFITS FROM NORMAL FLORA

Prevention of colonization by more pathogenic species

(colonization resistance).

Competition for receptors (tropism)

Competition for nutrients (interference)

Direct antagonism through bacteriocins, toxic chemical byproducts

Example of protective effect

Normal mouse - requires ~106 Salmonella to become infected

Germ free mouse - infectious dose is ~10 Salmonella

Competitive Protective Role of Normal Microflora

Kloos and Schleifer scheme, Baird–Parker system for

classifying staphylococci and micrococci

Kloos and Schleifer : Staph species are able to produce

acid aerobically from glycerol in the presence of

erythromycin

they are sensitive to lysostaphin and nitrofuran at defined

concentrations, whereas Micrococcus species are not.

NORMAL FLORA - CONJUNCTIVA

Variety of bacteria: low numbers present

High moisture

Blinking mechanically removes bacteria

Lachrymal secretions include lysozyme

Staph epidermidis and Propoinibacterium acnes dominate

Staph aureus, Haemophilus sp. and Neisseria sp.

occasionally found

NORMAL FLORA - RESPIRATORY TRACT

Nares (nostrils)

20% of human population are carriers for Staphylococcus aureus Staphylococcus epidermidis and corynebacteria

Nasopharynx (above soft palate)

Small numbers of Streptococci, Neisseria spp., Haemophilus

influenzae Strep pneumoniae, Strep pyogenes, Haemophilus influenzae, Neisseria meningitidis, Mycoplasma spp.,

pneumococci, Bordetella pertussis, C.diphtheriae

Oropharynx: between soft palate and upper edge of epiglottis

Large numbers of Staphylococcus aureus and S. epidermidis

Alpha-hemolytic streptococci, diphtheroids, Neisseria sp

NORMAL FLORA OF LOWER RT

Usually sterile

Ciliated epithelium

Mucus blanket: entrapment

Removed by coughing, sneezing

Contains lysozyme

Alveolar macrophages If breached: opportunistic infections, H. influenzae,

Streptococcus pneumoniae,

Ecology : Normal flora of Oral Cavity

Birth: sterile mouth

Neonate Within 4-12 hours (lactobacilli, streptococci)

Colonized from environment (especially first feeding)

Streptococcus salivarius, staphylococci, Neisseriae, Moraxella

catarrhalis

Normal flora of Oral Cavity

NORMAL FLORA OF ORAL CAVITY

Teeth appear Tooth is nondesquamating surface

Streptococcus mutans, Streptococcus parasanguis Gingival crevice area (supporting structures of teeth)

Anaerobic species, yeasts

Puberty Bacteroides, spirochetes

108 bacteria/mL of saliva; potentially >700 species

ORAL & OP NORMAL FLORA

Resist mechanical removal by adhering to various

surfaces such as gums and teeth

MOs that can’t resist mechanical flushing of oral cavity,

swallowed, destroyed by HCl in stomach

Comfortable environment for MOs due to availability of

water and nutrients

Key organisms to learn:

Mouth - Strep viridans (e.g. S. sanguis, salivarius, mutans, mitis)

Nose - Staphylococcus aureus

NORMAL FLORA - GASTROINTESTINAL TRACT

Ecology Birth: sterile

Breast-fed - Bifidobacteria species

Switch to cow’s milk

Bifidobacteria species joined Enteric, bacteroides, enterococci, lactobacilli and clostridia

Switch to solid food

Microflora similar to parents

GI ecology : Varies along the tract (longitudinal levels)

Varies along horizontal level: lumen vs. epithelium

Esophagus

Microbes associated with saliva and food

Stomach

Low pH limits population numbers (10 bacteria/ml) , Helicobacter

pylori, acid-tolerant lactobacilli

Reduction of acidity (increase alkalinity)

Increases population of resident flora

Small intestine Proximal small intestine (duodenum and jejunum)

Sparse (<103 bacteria/ml fluid) due to acid from stomach,

bile and pancreatic secretions

Lactobacilli, Enterococcus faecalis Distal small intestine (ileum)

Increases (108 bacteria/ml) due to pH change

Lactobacilli, Enterococcus faecalis, coliforms,

Bacteroides, bifidobacteria

Large intestine 109-1011/ml

>350 species

E. coli = 0.1% of total population Primarily anaerobic

Facultative aerobes deplete oxygen Adult excretes 3x1013 bacteria/day

25%-35% of fecal mass = bacteria

NORMAL FLORA - GASTROINTESTINAL TRACT

Location (adult) Bacteria/gram

contents

duodenum 103-106

jejunum and ileum 105-108

cecum and transverse colon 108-1010

sigmoid colon and rectum 1011

In the normal adult colon, 96–99% of the resident

bacterial flora consists of anaerobes:

bacteroides species, especially B fragilis;

fusobacterium species;

anaerobic lactobacilli, e.g., bifidobacteria; clostridia

(C perfringens); and

anaerobic gram-positive cocci (Peptostreptococcus

species).

Site

skin

large intestine

vagina

Ratio of Anaerobes:

Aerobes

~ 1:1

1000:1

10:1

ANAEROBIC NORMAL FLORA

BACTERIAL PROCESSES OCCURRING IN GUT

Process Examples

Vitamin Synthesis Thiamine, riboflavin, pyridoxine, B12, K

Gas Production CO2, CH4, H2

Odor production H2S, NH3, amines, indole, butyric

Organic Acid production Acetic, propionic, butyric

Polysaccharide metabolism Extraction of calories from otherwise indigestible polysaccharides

Steroid metabolism Esterification, dehydroxylation, oxidation, reduction, inversion

Colonocyte nutrition - provides protection against cancer and ulcerative colitis. May affect obesity

Butyric Acid production, esp by Firmicutes (GP organisms).

Key colonizing organisms to learn:

Stomach - Helicobacter pylori

Colon -Facultative anaerobes - Escherichia coli, Enterococcus sp.

Obligate anaerobes - Bacteroides fragilis

Normal Flora of the Gastrointestinal Tract (GIT)

• In breast-fed Bifidobacteria account for more than 90% of the total intestinal bacteria.

Normal Flora of the Gastrointestinal Tract (GIT)

• In bottle-fed infants• Bifidobacteria are not

predominant. When breast-fed infants are switched to a diet of cow's milk or solid food, bifidobacteria are progressively joined by:

1. Enterics2. Bacteroides3. Enterococci4. Lactobacilli5. Clostridia

Normal Flora of the Gastrointestinal Tract

(GIT)• The proximal small intestine

1.Lactobacilli2.Enterococcus

faecalis3.Coliforms4.Bacteroides

Normal Flora of the Gastrointestinal Tract (GIT)

In the upper GIT of adult humans

• mainly acid-tolerant lactobacilli

e.g. Helicobacter pylori

Recent studies have also shown that the composition of

your gut flora can also affect how much you weight.

Persons with more Firmacutes (primarily gram positive

organisms) are more obese than those with more

Bacteroides (primarily gram negative).

NORMAL FLORA OF UROGENITAL TRACT

Upper urinary tract (kidneys, ureters, bladder) usually sterile

Male anterior urethra

Low numbers (102-104/ml)

Staphylococcus epidermidis, Enterococcus faecalis,

Escherichia coli, Proteus Male anterior urethra Same as Skin + enteric + enterococcus

NORMAL FLORA OF UROGENITAL TRACT

Vagina: complex microbiota

At birth: glycogen,pH5(Lactobacillus spp.)Same as mother (PH 5)

Neonate (one month) to puberty - (pH neutral),Same as skin + enteric

+ strept (PH 7)

At puberty

L. acidophilus, corynebacteria, pepto streptococci, staphylococci,

streptococci, and Bacteroides L. acidophilus converts glycogen to lactic acid Lowers to pH 5

At menopause : return to prepuberty flora

Normal flora of the Urogenital Tract

a) The anterior urethra1. Staphylococcus

epidermidis2. Enterococcus faecalis3. Alpha-hemolytic

streptococci. 4. Some enteric bacteria

(e.g. E. coli, Proteus sp.)5. Corynebacteria sp.6. Acinetobacter sp.7. Mycoplasma sp.8. Candida sp.9. Mycobacterium

smegmatis

Normal flora of the Urogenital Tract

b) The vagina 1. Corynebacterium sp.2. Staphylococci3. Nonpyogenic

streptococci4. Escherichia coli5. Lactobacillus

acidophilus6. Flavobacterium sp.7. Clostridium sp.8. Viridans streptococci9. Other Enterobacteria

Key organism to learn in

GUT - Lactobacillus sp

LACTOBACILLUS SPP. FOUND IN UROGENITAL SYSTEM

HAIR FOLLICLES

inhabited by anaerobes (Propionibacterium sp)

in their deeper parts and nearer the surface aerobic cocci,Malassezia species of yeasts.

Propionibacteria are anaerobic, Grampositive, rod-shaped organisms,

P. acnes, and P. granulosum

widespread, follicles that have large sebaceous glands

over the face and upper trunk.

both been associated with acne lesions

The third species, P. avidum, is found in moist sites,

particularly the axillae and groins. Its pathogenic

potential unclear

Propionibacterium species require the increased skin

lipid levels of puberty before becoming established

Present at very low levels before the onset of puberty.

Propionibacterium acnes & Gram-positive cocci are

capable of hydrolysing lipids of sebum to produce

free fatty acids

The acidic milieu which results inhibits the growth of

other organisms such as Streptococcus pyogenes

NORMAL FLORA OF EAR

External auditory meatus

In addition to coagulase-negative staphylococci and coryneforms

Proteus species, Escherichia coli, Neisseria,Psuedomonas

NORMAL FLORA OF EAR

Pseudomonas aeruginosa frequently isolated from ear

NORMAL FLORA OF AXILLA

supports a very high level of bacterial colonization

mostly staphylococci, micrococci and coryneforms

Propionibacterium acnes is usually present and

P. avidum is often found

NORMAL FLORA OF TOE CLEFTS

In shoe wearers, 4 toe cleft is often hyperhydrated and

the skin macerated.

large number of bacteria present

The toe web is an important site for Brevibacterium

species, and not surprisingly Acinetobacter species are often

isolated, as are Alkaligenes species.

In the elderly and in tropical Climates coliforms and other

organisms of the intestinal flora may be present

NORMAL FLORA OF UMBILICUS

The umbilicus of the newborn is frequently colonized by

Staphylococcus aureus shortly after birth

The umbilicus of the newborn may also be colonized by

Streptococcus pyogenes, and look normal

The organisms can readily spread from infant to infant in a

hospital nursery.

Searches for the source of infection in a maternity unit

should always include umbilical swabs from the babies.

Importance of The Normal Flora (Disadvantages)

1. They can cause disease in the following:a) When individuals become immunocompromised or debilitated.b) When they change their usual anatomic location.

Importance of The Normal Flora

2. The oral flora of humans may harm their host since some of these bacteria are pathogens

NORMAL FLORA - RISKS AND OPPORTUNISTIC

Clinical conditions that may be caused by members of the normal flora

“viable microbial food supplement that beneficially influences the

health of the host”

Oral administration of living organisms to promote health

Mechanism speculative: competition with other bacteria;

stimulation of nonspecific immunity

Examples:

Lactobacillus augments protective gut barrier function against

diarrheagenic E. coli

Lactobacillus stimulate IgA secretion, providing protection

against pathogens such as rotavirus

.

PROBIOTICS

PREBIOTICS Non- digestible food ,Non essential food components used to affect microbial populations

Increases bifidobacteria and lactobacillus bacteria

Typically a carbohydrate: soluble fiber

(e.g. non digestible oligosaccharides)

Result in formation of short chain fatty acids that affect lymphocyte proliferation, cytokine production etc

ORGANISMS PER GRAM OF HOMOGENIZED TISSUE OR FLUID OR PER SQUARE CENTIMETER OF SKIN SURFACE

Noncolonized areas: Blood, body fluids, tissues,

internal organs Lower respiratory tract Upper urinary tract

Sterile tissuesIn a healthy human,

the internal tissues such as:

• blood• brain• muscle• cerbrospinal fluid

(csf.) are normally free of

microorganisms.

Normal Flora

GNOTOBIOLOGY

Gnotobiotic animals : “germfree”

Fetus is sterile Caesarean sections to obtain fetus

Fetus growing in sterile isolator

Not anatomically or physiologically normal

Poorly developed lymphoid system, thin intestinal wall, enlarged

caecum, low antibody titers

Die of intestinal atonia,Require vitamin K and B comp

No dental caries or plaque More susceptible to pathogens

WHY ARE ARMPITS SMELLY?

1. Anaerobic bacteria in the pit use sebum

from sweat glands to produce short

chain fatty acids

2. The sweat and sebum produced by the

pit is naturally aromatic

3. Use of deodorants with antibacterial

substances, inhibit growth of G(+), may

lead to growth of G(-) MOs and infection

WHAT DO YOU THINK HAPPENS TO THE FLORA ON YOUR SKIN WHEN IT IS COVERED WITH AN OCCLUSIVE BANDAGE FOR A FEW DAYS?

A waterproof plaster applied to a cut on the forearm

for one or two days causes an increase in the

resident population - increase in gram negative

bacilli

Removal of particlesincluding microorganismsby rapid passage of airover cilia in nasopharynx

Skin is a physical barrier,produces antimicrobial fattyacids, and its normal florainhibit pathogencolonization

Stomach acidity(pH 2) inhibitsmicrobial growth

Normal floracompete withpathogens

Flushing of urinarytract preventscolonization

Lysozyme in tears andother secretions dissolvescell walls

Mucus, cilia lining tracheasuspend and move micro-organisms out of the body

Mucus and phagocytesin lungs prevent colonization

Blood and lymphproteins inhibitmicrobial growth

Rapid pH changeinhibits microbialgrowth

Normal flora compete with pathogens in the gut

.

Methods of sampling :

Simple qualitative studies by swabbing

The number of organisms is increased by the duration of

rubbing, pressure exerted and moistening the swab

The best quantitative estimates of the total bacterial flora are

determined by applying an open-ended cylinder of known

cross sectional area to the skin, introducing a small,

known volume of suitable liquid vehicle and scrubbing

the surface of the skin to free the organisms

Full-thickness skin-biopsy material should be the best sample

Disposal of organisms can be studied quantitatively by air sampling techniques using either settle plates or an impaction

sampler

PCR

ISOLATION MEDIA

use of ordinary blood

serum agar for aerobic organisms

solid Brewer’s thioglycolate medium without indicator

1% Tween 80 indicator for Propionibacterium

acnes

VARIABLES AFFECTING NORMAL MICROFLORA - GENETICS

Relatedness of colonic bacterial flora in identical twins (even twins living apart) is higher

than that of unrelated persons, fraternal twins or married persons

VARIABLES AFFECTING NORMAL MICROFLORA - RACES

Racial differences have been demonstrated in the nasal carriage of S. aureus in children

white individuals being more likely carriers than black individuals

VARIABLES AFFECTING NORMAL MICROFLORA - AGE

Bacterial flora develops in orderly succession from birth

to stable adult flora : Example : mouth microflora

Infant - mouth sterile at birth

Rapidly develops flora, primarily from mother

Strep salivarius (on tongue); lactobacilli

Changes with eruption of teeth(Strep sanguis; mutans)

Ecologic system develops- micro environments of anaerobic

and aerobic flora

Neonates are covered with vernix caseosa contain the antimicrobial peptide cathelicidin

and also lysozyme,Skin is sterile

VARIABLES AFFECTING NORMAL MICROFLORA - GENDER AND STRESS

Gender - Examples

Presence of different mucosal surfaces (esp.

genitourinary)

Hormonal effects on flora in females - e.g. change in

vaginal flora with onset of puberty

Stress - Examples

Alterations in bowel flora recognized with severe stress

There is evidence that males carry higher numbers of bacteria, aerobic organisms than

females

Men are more likely to be disseminators of Staphylococcus aureus than females

VARIABLES AFFECTING NORMAL MICROFLORA - NUTRITION AND DIET OF HOST

Examples : Infant feeding

Breast-fed infants - GI flora is predominantly

Bifidobacterium sp. (anaerobic gram positive bacillus)

Bottle-fed infants - GI flora more closely resembles that of

adults (enteric gram negative bacilli, mixed anaerobes,

staph) Adult diets

Predominant species found in colonic flora differs among

vegetarians, fish eaters, and omnivores

Limburger cheese is made from the same bacteria (Brevibacterium sp.) that grow between your toes

NORMAL FLORA TRIVIA

NORMAL FLORAAcquired rapidly during & after birth

Reflects sex of person

Reflects environment of person

90% is S. epidermidis; S. aureus, may be in moist areas

Reflects genetics of person, nutrition of person, age of person

Changes continuously through out life

Protection against other bacteria

THANK YOU