Introduction Terminologies Oral ecosystem Normal microbial oral flora and its development Factors determining bacterial oral flora Dental implications of oral microbiology:

1.periodontal diseases

2. dental caries Fungi and parasite in oral cavity Role of oral flora in systemic diseases. Preventive approach to oral infection Conclusion References

Microbes are the part of our lives in more ways than most understand. They have shaped the present environment and their activities will greatly influence our future.

The oral cavity is a border area between the environment and the human organism, a gateway for food, an outpost of the immune system and above all a multilayered habitat inhabited by countless microorganisms.

The mouth is continually exposed to organisms

from the external environment ,beginning with

the passage through the birth canal.

In time a ecological balance is reached that

serves to establish a resident microbial flora

that remains fairly stable throughout life.

It has been estimated that the human body is made up of over 1014 cells of which only around 10% are mammalian. The remainder are the micro-organisms that comprise the resident micro flora of the host.

(Sanders & Sanders, 1984)

Microorganisms in mouth were first described by Anton von Leeuwenhoek in 1683.

Habitat: the site where a microorganism grows

Ecology: Study of relationship between organisms and their environment.

Ecological niche: The functional position of an organism in its environment comprising the living space, periods of time during which it is active there and resources it obtains there.

Aerobes: These organisms require oxygen for aerobic cellular respiration to obtain energy.

Obligate aerobe: is an organism that can survive and grow only in an oxygenated environment

Facultative anaerobes: can use oxygen, but also have anaerobic methods of energy production.

Capnophiles: are which thrive in the presence of high concentrations of carbon dioxide.

An oligotroph: is an organism that can live in an environment that offers very low levels of nutrients.

Microaerophiles: are organisms that may use oxygen, but only at low concentrations

Indigenous Flora(Resident): It comprises those indigenous species

that are almost always present in high numbers, greater than 1 percent of the total viable count.

Supplemental Flora: The supplemental flora are those bacterial

species that are nearly always present, but in low numbers, less than 1 percent of total viable count

Transient Flora:

Transient flora comprise organisms "just passing through" a host. At any given time a particular species may or may not be

represented in the flora.

Pathogens Micro organisms that have the potential to

cause disease are termed pathogens. Opportunistic Pathogens Micro-organisms that cause disease only

under exceptional circumstances . True Pathogens

Micro-organisms that are consistently

associated with a particular disease .

Four major ecosystems are present in the oral cavity

-Buccal epithelium

-Dorsum of the tongue

-Supragingival tooth surface

-Subgingival tooth and crevicular epithelial surfaces

Tooth surface• mitis group• mutans group• anginosus group

Saliva• S. salivarius• S. mitis• S. oralis• mitis group• anginosus group

Tongue• S. salivarius• S. mitis

Pharynx• S. mitis• S. oralis• mitis group• anginosus group

Nasopharynx• S. pneumoniae

Vestibular mucosa• S. vestibularis

Buccal mucosa• S. mitis• mitis group• anginosus group

Tonsils• S. sanguis• S. mitis• S. mutans• mitis group• anginosus group

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The buccal epithelium has more of gram positive streptococci in contrast to tongue which has more of gram positive filaments.

The subgingival region is essentially anaerobic as compared to the supragingival region.

Mutans streptococci (S. mutans, S. sorbinus, S. cricetus, S. rattus) and S. Sanguis are found in large number on teeth, whereas S. Salivarius is isolated mainly from the tongue.

S. mutans and S. sanguis appear only after eruption of teeth.

In the buccal mucosa the predominant bacterium is Streptococcus mitior with Streptococcus sanguis and salivarius.

Hard palate flora also resembles that of buccal mucosa, with predominance of streptococci. The soft palate harbours respiratory tract bacteria Corynebacterium, Neisseria, Haemophilus.

Tongue is an ideal site for retention of microorganisms owing to its keratinized dorsal surface.

Streptococcus salivarius is the predominant flora accounting for upto 50% of the total.

Streptococcus mitior is also common here.

Gingival crevice has the most numerous bacterial population among all sites in mouth.

As many as 1010-1011 organisms are recovered per gram wet weight of gingival debris.

It is considered to be due to absence of dislodging forces and crevicular fluid acting as a rich nutrient medium.

The presence of nutrients, epithelial debris, and secretions makes the mouth a favorable habitat for a great variety of bacteria.

Oral bacteria include streptococci, lactobacilli, staphylococci and corynebacteria, with a great number of anaerobes, especially bacteroides.

The mouth presents a succession of different ecological situations with age, and this corresponds with changes in the composition of the normal flora

The process begins with the colonization of habitat by pioneer microbial populations.

In oral cavity of newborns, streptococci are the pioneer organisms.

They fill the niche of the new environment and modify the habitat and new population develops.

When no additional niche is available for new population, a stable assemblage of bacterial population is achieved called as climax community.

At birth:The mouth of full term foetus is usually sterile, transient flora from the birth canal may be acquired.Mouth then rapidly acquires organisms from mother and from the environment.It consists of several streptococcal and staphylococcal species with Lactobacilli, Bacillus, Neisseria and Yeasts.Streptococcus salivarius is the most common and forms the pioneer community with Staphylococcus albus.

Infancy & Early ChildhoodThe infant comes into contact with an ever-increasing range of microorganisms and some become established as part of commensal flora.The eruption of deciduous teeth provides a new attachment surface and turns Streptococcus sanguis and mutans as regular inhabitants of oral cavity.Anaerobes are few in number due to absence of deep gingival crevice.Actinomyces , Lactobacilli are found regularly.

Adolescence The greatest number of organisms in mouth occur when permanent teeth erupt.These teeth have deep fissures, larger inter proximal spaces and deeper gingival crevice, allowing a great increase in anaerobes. AdulthoodIts chief characteristic is its complexity of oral flora.There is an increase in Bacteroides and Spirochetes with maturity of dental plaque.

As the teeth are lost the available sites for microbial colonisation decreases and several species diminish disproportionately in numbers.

Edentulous persons harbour few Spirochetes or Bacteroides but carriage of Yeast increases.

S.sanguis & mutans disappear.

HUMAN ORAL FLORA

Gram-positive facultative cocci

Gram-negative facultative rods

Staphylococcus epidermidis

Staph. aureus

Streptococcus mutans

Strep. sanguis

Strep. Mitis

Strep. Salivarius

Strep. Faecalis

Beta-hemolytic streptococci

Enterobacteriaceae

Hemophilus influenzae

Eikenella corrodens

Actinobacillus

Actinomycetemcomitans

Gram-positive anaerobic cocci

Gram-positive anaerobic rods

Peptostreptococcus sp Actinomyces israelii

A. odonotolyticus

A. Viscosus

Lactobacillus

Gram-negative anaerobic cocci

Gram-negative aerobic or facultative cocci

Diphtheroids

Corynebacterium

Eubacterium

Neisseria sicca

N. Flavescens

Spirochetes Yeasts

Treponema denticola

T. Microdentium

Candida albicans

Geotrichum sp.

Protozoa Mycoplasma

Entamoeba gingivalis

Tirchomonas tenax

Mycoplasma orale

M. pneumoniae

Physicochemical factors

- Temperature

- Oxygen tension

- Hydrogen ion concentration Host factors

Nutrient sources

Temperature: Although the average temperature in the oral

cavity is approximately 37' C, temperatures can vary considerably, especially on the mucosal surfaces and on the clinical crowns of teeth.

Periodontal pockets with active disease have a higher temperature – up to 39 C compared with healthy sites (mean value 36.80 C).

Such changes in temperature affect gene expression in periodontal pathogens, such as Porphyromonas gingivalis.

Oxygen tension : The oxygen concentrations at different

locations in the oral cavity vary widely. As may be expected, the dorsum of the

tongue and the buccal and palatal mucosa are in an essentially aerobic environment.

The oxygen tension inside a periodontal pocket is very low, with the species having a tendency to become reduced rather than oxidised, explaining the survival of obligate anaerobe.

Therefore obligate aerobic organisms (which require oxygen) cannot survive, whereas obligate anaerobic organisms (which cannot tolerate the presence of oxygen) are able to thrive.

Hydrogen ion concentration: The term pH refers to the negative logarithm of

hydrogen ion concentration; thus, the lower the pH value, the higher is the hydrogen ion concentration.

The buffering capacity of plaque and saliva maintain the pH in oral cavity at about 7.

pH can vary due to:

1) exogenous materials placed in the mouth2) production of hydrogen ion by bacteria as a

result of carbohydrate fermentation3) buffering system in saliva (bicarbonates)

The second reason is responsible for dramatic drop in pH leading to Dental Caries.

Most oral bacteria grow best at a pH of about 7 (essentially neutral). Most of the time the pH in the oral cavity is maintained at about 7 by the buffering systems in plaque and saliva.

1) Saliva 2) Crevicular fluid 1. Saliva : Various salivary components interact with oral

flora in ways that can either enhance or inhibit ability of these organisms to survive.

IgG is seen in gingival inflammation contributed by GCF.

These make it more difficult for bacteria to bind to oral epithelium or hard tissue surfaces.

Non-specific factors: These are lysozyme, lactoferrin and

lactoperoxidase. Lysozyme degrades bacterial peptidoglycan i.e

the cell wall, rendering bacteria susceptible to osmotic disruption and death.

Lactoferrin binds to iron which is a growth limiting substance in bacteria. Making iron unavailable to bacteria lactoferrin limits bacterial growth.

Lactoperoxidase catalyses the formation of hypothiocyanate ion ,inactivating bacterial enzymes and finally death.

2. Crevicular fluid : It is an inflammatory exudate derived from

plasma with large amount of antibody and complement proteins.

The predominant immunoglobulin is IgG, derived from plasma cells located in periodontal tissues as well as from circulating plasma.

These antibodies keep the subgingival flora in check by inhibiting colonization or activating complement system.

1.Periodontaldiseases 2. Dental caries

According to WHO : Plaque is a highly specific and selective but structurally variable clinical entity characterized by sequential colonization of microorganisms on the surface of teeth, restoration and other parts of the oral cavity.

It is made up of mucins, desquamative epithelial cells and microorganisms embedded in an extracellular matrix.

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Subgingival Tissue associated

St. oralis, St. intermedius Peptostreptcoccus micros

P. gingivalis, P. intermedia

T. Forsythis, F. Nucleatum

Pellicle formation Attachment of single bacterial cell (0-

4h) Growth of attached bacteria leading to

formation of distinct micro colonies. (4-24h) Microbial succession and co-

aggregation .(1-14 days) Climax community plaque.(2 weeks or

older)

Microrganisms don’t colonize on the mineralised tooth surface.

The teeth are always covered by an acellular proteinaceous film ,the pellicle that forms on the naked tooth surface within mins to hours.

The bacteria colonize the tooth surface only when pellicle is in place for hours.

Function of pellicle- Protect enamel. Reduces friction. Provide matrix for re-mineralization.

Pellicle contains-lysozyme,albumin,IgA,IgG.

Early colonisation : Plaque builds up first in small defects or pits on the

enamel surface and then spreads over the tooth surface.

Early succession evolves adhesion between pellicle and pioneer organism.

S.sanguis ,A.viscosus,A.naeslundii and peptostreptoccous attaches within 1 hr.

The first organism to attach include Streptococcus sanguis, other Streptococci and gram-negative cocci (Neisseria and Branhamella). After developing for about 24 hours, plaque consists largely of Streptococci and Veillonella, Corynebacterium, Actinomyces, Lactobacillus and Rothia.

Veillonella is first anaerobe to appear followed by facultative Actinomycetes and anaerobic Actinomyces israelli.

As plaque ages, number of anaerobes increases and after 7 days Fusobacteria and Bacterioides can be detected.

Responsible of causing specific disease.

Early stage lack pathogenicity-aerobic, lack sufficient production of damaging metabolites.

Change of aerobic to anaerobic environment.

Secondary colonizers are the microorganisms that do not initially colonize clean tooth surfaces, including P. intermedia, P. loescheii, Capnocytophaga spp., F.nucleatum, and P. gingivalis.

These microorganisms adhere to cells of bacteria already in the plaque mass.

Extensive laboratory studies have documented the ability of different species and genera of plaque microorganisms to adhere to one another, a process known as coaggregation.

Coaggregation is based on the specific interaction of a proteinaceous adhesion procedure by one bacterium and a respective carbohydrate or protein receptor found on the surface of another bacterium.

› F. nucleatum with S. sanguis,

› P. loescheii with A. viscosus,

Tooth surface is stable and covered with pellicle , so it is an ideal surface for the attachment of many oral streptococci .

Tooth habitats favorable for harboring pathogenic plaque includes :

Pits and fissures- Community is dominated by S.sanguis and other streptococci.

Smooth surface -proximal areas of very young patient are less favorable habitat for MS.

Root surface Gingival recession favors the formation of plaque in

this area.

Sub gingival areas

Initial occupants of sulcus are merely extension of immediate tooth surface

Plaque community changes from masses of cocci to a community dominated by filamentous bacteria and spirochetes in sub gingival area .

B. melaninogenicous can explore this habitat, because protein and heme is available.

Characteristic Early Mature

Gram reaction + +/-

Morphotypes Cocci, branching rods cocci, rod,

spirochetes

Energy metabolism Facultative F-

anaerobic

Tolerated by host Well Can cause caries

and gingivitis

Gram positive: Actinomyces viscosus Actinomyces naeslundii Streptococcus sanguis Streptococcus mitis Peptosreptococcus microsGram negative: Fusobactecterium nucleatum P. intermedia Veillonella parvula Wolinella Haemophilus species

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Fusobacterium (fusobacterium nucleatum)

Oral spirochaetes (treponema species)

Fusospirochaetal complexOthers – provetella

intermedia,veillonella and spirochaetes

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CHRONIC PERIODONTITIS:P. gingivalisP. intermediaA. actinomycetemcomitans

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gram negative ,non motile pleomorphic short rods

gram negative obligate anaerobe. P. gingivalis is a member of "black-pigmented

Bacteroides" group Size – 0.5 * 1 microns Doesn’t ferment carbohydrate. Major site of colonization – gingival sulcus

of human oral cavity aggressive periodontal pathogen.

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Microbiota is predominantly composed of gram –ve anaerobic, capnophilic rods.

Microbiological studies indicate- All disease sites harbor A.comitans which may compose as much as 90 % of total cultivable microbiota.

Other organisms found in significant levels include P. gingivalis, C.rectus & F. nucleatum.

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This is a small, non-motile, Gram-negative saccharolytic, capnophilic, round-ended rod that forms small, convex colonies with a "star-shaped" center when grown on blood agar plates.

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May be isolated on non selective blood agar incubated anaerobically.

Its growth is stimulated by addition of carbon dioxide

Ferments glucose and fructose Site – subgingival sites in oral cavity Virulence factors – LPS, leukotoxin,

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PRE-PUBERTAL PERIODONTITIS P. IntermediaA. ActinomycetemcomitansFusobacterium species P. Gingivalis

JUVENILE PERIODONTITIS

A. actinomycetemcomitans

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REFRACTORY PERIODONTITISA. actinomycetecomitans

P. gingivalisP.intermediaB.forsythus

PAPILLON-LEFEVRE SYNDROMEA. actinomycetemcomitans

P. intermediaP.gingivalisF.nucleatumE.corredens

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The micro flora contains mainly gram –ve anaerobic rods

There is high prevalence of putative pathogens F.nucleatum , P. intermedia , P. gingivalis & T. forsythia.

MOST COMMONLY BY Staphylococcal aureus or enterobacteria

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Dental caries is a specific and treatable bacterial infection due to mutans streptococcus (MS) and in the later stages to  lactobacillus.” (Oral Sci Rev. 1976).

“ Caries is a dynamic process of demineralization of  the dental hard tissues by the products of bacterial metabolism, alternating with periods of remineralization.”Larsen and Bruun (Clinical Cariology, 1994)

In 1976 Loesch (Oral Sci Rev, 1976) postulated that dental caries is a specific and treatable bacterial infection due primarily to MS and in the later stages to lactobacillus

T here is evidence that some bacteria – S. mutans, Lactobacilli and actinomyces- are more important than others.

S. MUTANS: The feature that supports its role as

cariogenic organisms are its: Rapid generation time Acidogenic nature Production of extra cellular polysaccharides

from sucrose which aids on adherence and acts as a nutrient

Isolated in high numbers in caries active mouth in incipient lesions

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Capacity to attain critical pH for enamel demineralization more rapidly than other plaque bacteria.

S. mutans can produce glucan by using glucosyl transferase enzyme.It is this glucans which help in the attachment of the bacteria to the tooth.

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Gram positive cocci forming chains Non motile, facultative anaerobes. Size – 0.5-0.75 microns in diameter Usually alpha hemolytic Selectively cultured in Mitis salivarius

(20% sucrose + 0.2% unit/ml of bacitracin)

Cultureblood agar– grey to whitesucrose-containing media –produce

extracellular polysachharides67

Collection of seven different species1. S. mutans2. S. sobrinus3. S. cricetus4. S. ferus5. S. rattus6. S. macacae7. S. downei

Primary habitat – human oral cavity to colonize smooth surface of teeth. Doesn’t appear prior to eruption of teeth and disappear after eruption

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Gram negative ,non spore forming rods Grow under micro-aerophillic conditions They are both acidogenic and aciduric They are secondary invaders Lactobacilli for many years was believed to

be the causative agent of dental caries as High numbers were obtained in most enamel

caries Able to synthesis extra cellular and intra

cellular polysaccharides form glucose.69

They produces lactic acid at pH < 5 Although these properties seemed valuable to

a cariogenic organism, it was also seen that their affinity for the tooth surface and their number prior to development of caries was low.

They were, in fact, absent from incipient lesions while present is significant numbers in developed caries.

Thus, they were categorized as secondary invaders which caused progression of caries due to their acidogenic and aciduric properties.

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Gram positive non acid fast non motile non spore.

Strict anaerobic or facultative anaerobes. Main species –• [A. naesulundi + A. viscosus] : facultative

anaerobes• [A.israelli +A.odontolyticus] : strict

anaerobes

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Found in increase numbers in plaque overlying root surface and sound root surface

Actinomyces species: A.viscous, some other Gram +ve bacilli are involved in the initiation of lesions on root surface.

Role played by a large number of other bacteria isolated from caries such as Arachnia, Eubacterium, Rothia etc are not yet clearly known.

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Antibacterial effects of fluoride are attributable to the weak acid nature of hydrofluoric acid allowing its permeation at low pH into bacterial cells.

The unionised hydrofluoric acid here dissociates due to alkaline intracellular pH, which positions fluoride to affect a variety of vital enzymatic cell functions.

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The significant ones are inhibition of enolase, potassium and inorganic phosphate transport.

The mutans streptococci are considerably more sensitive to fluoride inhibition than A. viscosus or Lactobacilli.

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Most commonly found :- candida species (C.albicans, C. tropicalis, C. stellatoidea, C. parapsilosis, C. guilliermondi)

Other rhodotorula & torulopsis ( denture wearer)

Conditions – thrush ,erythematous candidiasis,hyperplastic candiasis,angular chelitis

Oral sampling – imprint culture Medium used – sabouraand’s agar(peptone-

glucose) Indentification – psuedohyphae, septate hyphae

and germ tubes

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Entamoeba gingivalis Trichomonas tenax

E. gingivalis – found in soft calculus,periodontal pockets and

infection of tonsils Can become opportunistic pathogen T.tenax – only parasitic flagellate in oral cavity --number increases in periodontitis

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Recently it has been recognized that plaque related oral diseases, especially periodontitis, may alter the course and pathogenesis of a number of systemic diseases.

These includes: Cardiovascular diseases:• — Infective endocarditis• Coronary heart disease (atherosclerosis, MI)• stroke Bacterial pneumonia Diabetic mellitus AIDS

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DIABETES MELLITUS: capnocytophaga species P. intermedia A. actinomycetemcomitans P. gingivalisAIDS fusobacterium species A. actinomycetemcomitas P. micros P. intermedia

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Oral health is an integral element of general health and well-being.

Oral diseases are highly prevalent and their impact on both individual and society is significant.

There are many ways to prevent or reduce the risk of infection or diseases. Brushing and Flossing are most basic ways to reduce these risks. These two help to get rid of bacteria which stick to surfaces in our mouth to form plaque.

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Caries:Frequent mechanical removal of plaqueUse of fluoridesDiet modificationsStimulation of saliva after mealsCaries vaccine

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  Periodontal diseases:Mechanical removal of plaqueMouthwash and AntibioticsAnti-inflammatory agents to reduce flow of GCFRefractory - Antibiotics

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Current dental therapy is focussed on removing whole dental plaque. There are new approaches towards selective inhibition of pathogens and modulations of microbial composition of dental plaque to control community-based oral microbial pathogenesis.

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Brief outline of ORALMICROFLORA IN DISEASE

INFECTIONS OF THE MOUTHINFECTIONS OF THE MOUTH

InfectionInfection OrganismOrganism

Dental cariesDental caries Streptococcus mutansStreptococcus mutans

Periodontal diseasesPeriodontal diseases Bacteroides, ActinomycesBacteroides, Actinomyces

Surgical infectionSurgical infection

a)a) Dry socketDry socket

b)b) Dental abscessDental abscess

c)c) OsteomyelitisOsteomyelitis

d)d) Ludwig’s anginaLudwig’s angina

e)e) PericoronitisPericoronitis

ActinomycesActinomyces

Oral streptococciOral streptococci

Staphylococcus aureusStaphylococcus aureus

ββ - -haemolytic streptococcihaemolytic streptococci

BacteroidesBacteroides

INFECTIONS OF THE MOUTHINFECTIONS OF THE MOUTHInfectionInfection OrganismOrganism

Soft tissue infectionsSoft tissue infectionsa)a) DiphtheriaDiphtheriab)b) ANUGANUGc)c) Cancrum orisCancrum orisd)d) TuberculosisTuberculosise)e) LeprosyLeprosy

C. DiphtheriaeC. DiphtheriaeFuso-spirochaetesFuso-spirochaetesFuso-spirochaetesFuso-spirochaetesM. TuberculosisM. TuberculosisM. LepraeM. Leprae

Viral infectionsViral infectionsa)a) Herpetic stomatitisHerpetic stomatitisb)b) Herpes ZosterHerpes Zosterc)c) MumpsMumpsd)d) MeaslesMeasles

Herpes simplexHerpes simplexVaricella-zosterVaricella-zosterMumps virusMumps virusMeasles virusMeasles virus

INFECTIONS OF THE MOUTHINFECTIONS OF THE MOUTH

InfectionInfection OrganismOrganism

Fungal infectionsFungal infections

a)a) CandidosisCandidosis

b)b) HistoplasmosisHistoplasmosis

Candida albicansCandida albicans

H. CapsulatumH. Capsulatum

MiscellaneousMiscellaneous

a)a) Erythema multiformeErythema multiforme

b)b) StevensJohnson StevensJohnson syndromesyndrome

As a public health dentist it is very important to in detail about the oral microflora, as we commonly come across these micro organisms while treating the diseases like dental caries ,periodontitis etc.

We also should know about the various types of microflora in oral cavity, the mechanism by which they cause various diseases so that we can effectively prevent the disease.

Especially at the camp sites we come across many diseases like lichen planus ,herpes , candidiasis etc. some of which can be potentially infectious. Thus a deeper understanding about the microflora can be beneficial for us.

Also, there are microoraganisms which are beneficial for the body thus treatment planning of any disease should not interfere with these microorganisms

Conclusion  There are wide variety of organisms present,

each with a distinctive property. This determines the ways in which they will react with their hosts therefore contribute to the characteristics of the disease they cause.The normal flora play a very important role in protection against these established pathogenic microbes.

Identification of the microorganisms found during the different phases of the disease process is technically challenging. The interpretation of microbiologic data is greatly influenced by the clinical classification of disease status, that promise to provide much information about the basic mechanisms involved in the disease process.

Technologic advances in molecular microbiology have improved the ability to detect specific bacteria and their products, which may serve as markers of ongoing disease or predictors of future disease.

References 1. Textbook of Microbiology- Anantnarayan

2. Carranza’s clinical periodontology. 9th edition3. Dental caries –disease and its clinical managenent –

Ole Fejerskov and Edwina A.M Kidd

4. Textbook of cariology– Ernest Newburn

5. An introduction to risk prediction and preventive dentistry – Per Axelsson and Karlstad

6. Prevention of oral diseases – John J. Murray,June H Nunn and James G. Steele