NORMAL FLORA

Oral microorganisms comprises of:

Bacteria Fungi Virus Protozoa Mycoplasma

- smallest known cell

(~0.1µm in diameter)

- were considered as

viruses for years

- currently is categorise

due to lack of a cell

wall

Introduction to Oral Ecosystem…

• Of the various sites of the body, the oral cavity is one of

the most densely populated.

Bacterial Cell Morphology

- Shape & Size -

Bacteria are classified by shape into 3 basic

groups: (i) Cocci (spherical)

(ii) Bacilli (rod & bacillus shape)

(iii) Spirochaetes (helical)

Some bacteria with variable shapes, appearing

both coccal & bacillary forms are called

PLEOMORPHIC

The size of bacteria ranges from about 2 m - 5m

Bacterial Cell Morphology…..

- Arrangement-

Which ever shape the

bacterial may be, the

cells arranged

themselves as :

Single

Pairs

Chains

Clusters

Cocci Arrangement

Single

Pairs

Chains

Clusters

Structure of a Typical Bacterium….

Bacteria have a rigid cell wall outside the cytoplasmic

membrane.

Bacterial cell wall is porous & permeable to substances of

low molecular weight.

A typical

bacterial Cell

The cell wall of both Gram –ve & Gram +ve bacteria is

made up of peptidoglycan.

Peptidoglycan layer in Gram +ve is thicker than Gram –ve.

For Gram –ve bacteria, the peptidoglycan is covered by

an outer membrane.

outer

membrane

cytoplasmic membrane

peptidoglycan cell

wall cytoplasmic membrane

peptidoglycan

outer

membrane

cytoplasmic membrane

peptidoglycan cell

wall

Lipop

oly

saccha

ride

Outer membrane is composed of:

lipopolysaccharide (LPS)

lipoprotein

phospholipid

These forms ‘porins’,

through which

hydrophilic molecules

are transported in & out

of the organism

Bacterial Cell Morphology

- Gram Staining Characteristics-

Bacteria can be classified according to the staining

characteristics of their cell walls.

(i) Gram Positive

- will be stained BLUE with gram staining reagents

(ii) Gram Negative

- will be stained RED with gram staining reagents

Gram staining property of bacteria is useful for:

(i) bacterial identification

(ii) therapy of bacterial infections (Gram +ve are more susceptible to penicillins than Gram –ve)

Gram Staining Principle

(1) Crystal violet will penetrate the cell wall & cell membrane of both Gram +ve and

Gram –ve bacteria.

(2) The addition of iodine allows it to complex with crystal violet. Iodine is a trapping

agent that prevents the removal of the crystal violet dye and therefore, color the cell.

(3) A decoloriser, acetone will interact with the lipids of the cell membrane causes:

(i) Gram +ve bacterial cell walls to dehydrate, causing the closure of the pores

of the cell wall (peptidoglycan), thus resulting in retention of the crystal violet

(ii) Gram –ve bacterial will lose its outer lipopolysaccharide membrane, and the

inner peptidoglycan layer is left exposed.

(4) Finally when safranin is added:

(i) the cell wall of Gram +ve bacteria cannot absorb the safranin, therefore

maintaining the blue colour stain.

(ii) The peptidoglycan of Gram –ve bacteria will absorb/stained red by the

stain.

Bacterial Physiology….

Bacteria requires nutrients for metabolic purposes & cell division

Oxygen & hydrogen - obtained from water

Carbon - obtained by bacteria via 2 ways :

autotrophs - obtain carbon from CO2

heterotrophs - obtain carbon by breaking down other carbon

molecules in complex organic substances eg. sugars

In-organic ions

nitrogen, sulphur, phosphate, magnesium, potassium & trace elements

(iron, copper, molybdenum, & zinc) are required for bacterial growth

Organic nutrients

CHO (energy source), amino acids (crucial for growth), vitamins (B1, B2,

B6), purines & pyrimidines

Bacterial Physiology….

Bacterial Growth Cycle:

4 phases :

Lag phase

Log phase

Stationary phase

Death phase

Bacterial Growth Cycle

Growth

Phases

Description of activities

Lag phase

Adaptation period

• Bacteria do not divide immediately

• Undergoes vigorous metabolic activity

• May last for a few minutes or for many hours

Log phase

Rapid cell division occurs exponentially

• is influenced by the environmental conditions

Stationary

phase

Is reached when nutrient depletion or toxic products

cause growth to slow

• the number of new cells produced balances the number of

cells that die

Death phase

A decline in the number of live bacteria

Bacterial Physiology….

Aerobic & anaerobic growth:

Bacteria can be classified according to their ability to

live in the presence of O2 or O2-free environment.

Bacterial Physiology….

• Sodium thioglycolate medium

can be used to demonstrate the

degree of oxygenation for

bacteria as the medium forms an

oxygen gradient during growth.

1) Obligate aerobe organisms grow at the top

2) Obligate anaerobes organisms grow at the bottom

3) Facultative anaerobes grow throughout the medium

4) Microaerophilic requires low level of O2

5) Capnophilic grow in the presence of CO2

Bacterial Genera O2 tolerence Gram

Streptococcus Anaerobe (Facultative) G/+

Actinomyces Anaerobe (Facultative) G/+

Lactobacillus Anaerobe (Facultative) G/+

Actinobacillus Anaerobe (Facultative) G/-

Porphyromonas Anaerobe (Obligate) G/-

Prevotella Anaerobe (Obligate) G/-

Fusobacterium Anaerobe (Obligate) G/-

The MAJOR (predominant) bacterial

genera in the oral cavity

ORAL MICROORGANISMS

• The mouth has natural microorganisms.

• What is Normal Oral Microbiota?

- Microorganisms which are normally found in the oral cavity. - Exist in harmony with the host.

- Is also known as commensal microorganims.

Other important terminologies !!!

• Transient microbes :

- Microorganisms that survive in the mouth only

for a short period of time.

• Opportunistic microbes :

- Microorganisms that take advantage of certain

opportunities to cause disease.

• Pathogenic microbes :

- Microorganism that can cause infection in

individuals with normal host defenses.

- do not always comprise the normal microflora

ACQUISITION OF THE COMMENSAL ORAL FLORA

STERILE

New born A few hours after birth

A few months old

Encountered microbes:

(i) Passive contamination from mother, food, milk and water.

(ii) Saliva of individuals in close proximity to the baby.

(iii) From the birth canal itself may be of only limited

significance.

* Pioneer species : Streptococcus sp.

Few hours later....

- The mouth is highly selective for microorganisms.

- Pioneer species colonizing the mouth will collectively

make up the pioneer community.

- During the development of the pioneer community one

genus or species is usually predominant. (.. eg.

Streptococcus sp.)

- The pioneer species continue to grow & colonize.

With time, the metabolic activity of the pioneer

community modifies the environment.

This provides conditions suitable for colonization by a

succession of other populations – causes the number of

microbial groups to increase.

Reached a high species diversity : Climax community

• The next evolutionary change in this community occurs

during & after tooth eruption as 2 further niches are

provided for bacterial colonisation (enamel & gingival

crevice)

• Oral flora on the child’s 1st b’day usually consist of

streptococci, staphylococci, neisseriae & lactobacilli,

together with some anaerobes such as Veillonella&

Fusobacteria. Less frequently isolated are Lactobacillus,

Actinomyces, Prevotella & Fusobacterium

• Introduction of a prosthetic appliance changes the

microbial composition once again, fovouring the growth of

Candida sp.

* As one grows older and

becomes edentulous, oral flora

that colonize the mouth are

similar to a child before tooth

eruption.

ORAL HABITATS

(1) Buccal mucosa

- the cheek mucosa is sparsely colonized due to frequent

shedding of the mucosal surface.

(2) Dorsum of tongue

- highly papillated surface & has a large surface area.

- suppports a higher bacterial density than other oral

mucosal surfaces.

(3) Teeth

- large bacteria & their products accumulate on tooth

surfaces to produce dental plaque.

- as teeth are non-shedding surfaces, the highest

concentrations of microorganisms are found in stagnant

sites which afford protection from removal forces.

….eg. SMOOTH SURFACES are

colonized by a smaller number of

species than PIT and FISSURES

(4) Crevicular epithelium & gingival crevice

- is a minor region of bacterial colonization.

However, bacteria that colonize the crevicular area play a

critical role in the initiation &development of gingival &

periodontal disease.

(5) Prothodontic & Orthodontic Appliances

- dental appliances may act as inanimate reservoirs of

bacteria & yeasts – if not kept meticulously clean.

FACTORS WHICH INFLUENCE THE GROWTH OF MICROORGANISMS IN

THE ORAL CAVITY

(1) Saliva

1) salivary pellicle facilitates bacterial adhesion and thus,

contribute to plaque formation.

2) has a good buffering capacity, thus pH is maintained.

3) acts as a primary source of food.

4) facilitate bacterial clearance from the mouth.

5) contribute in host defense by having :

• non-specific defense factors (eg. lysozyme, lactoferrin, histatins).

• specific defense factors (eg. immunoglobulins).

(3) Gingival crevicular fluid (GCF)

- The GCF composition is similar to serum, thus, the crevice is protected by the specific & non-specific defense factors.

- GCF flow increases during inflammation.

- same as no 2-5 (saliva)

(2) Anatomical factors

1) the structure & shape of the teeth (eg. fissures),

2) malalignment of teeth,

3) poor quality of restorations (eg. fillings, bridges)

Mechanisms that accomplish include :

1) Competition for receptors for adhesion

at colonizing sites.

2) Production of toxins that kill other

bacteria of the same/different sp.

3) Production of metabolic end products

4) Use of metabolic end products of

other bacteria for nutritional purposes.

5) Able to co-aggregate with the same

or different bacterial species.

Oral

microbes

Oral

microbes

Oral

microbes

Can interact

both in

promoting &

suppressing the

neighbouring

bacteria

(4) Microbial factors

(5) Environmental pH

• The acidity of most oral surfaces is regulated by saliva

(saliva has mean pH 6.7)

• Many microbes require a neutral pH for growth.

• However, the environment pH depends on the

frequency of dietary carbohydrate intake.

(6) Oxidation-reduction potential

(redox potential / Eh)

Eh potential varies in different locations of the mouth.

eg.

Supragingival plaque bacteria are living at sites

which are exposed to O2 (highly oxidised – thus having

higher Eh).

• Thus, gives rise to the growth of facultative bacteria.

Subgingival plaque bacteria occupies periodontal

pockets which are protected from O2 (reduced Eh).

• Thus, giving advantage for the growth of obligately

anaerobic bacteria.

*Certain antibiotics & antiseptics affect the oral flora.

eg. (i) tetracycline – can wipe out most of the endogenous microbes

(ii) mouth rinses with antimicrobial agents

- chlorhexidine, hexetidine

(7) Antimicrobial therapy

(8) Diet & Nutrition

(i) Exogenus nutrient source : FOOD

(ii) Endogenus nutrient source : HOST, MICROBES

(a) HOST : saliva (amino acids, vitamins, glycoprotein)

crevicular fluid (proteins, glycoprotein)

(b) MICROBES : intracellular polysaccharides (glycogen)

extracellular microbial products