plaque and biofilm

PRESENTED BY : G.SRINIVAS DATE: 8/7/11 2ND YEAR

POST GRADUATE

CONTENTSIntroduction

Definition -Dental plaque: -Biofilm: Types and Composition of Dental Plaque

Formation of Dental Plaque

Coaggregation Microscopic Structure and Physiologic

Properties of Dental Plaque

Significance of the plaque Environment

Microbial specificity of periodontal diseases

What makes plaque pathogenic?

Microorganisms Associated with Specific Periodontal Diseases:

-Periodontal health -Periodontal diseasesTransmission of periodontal plaque bacteriaReferences

INTRODUCTIONDental plaque is a general term for the diverse

microbial community (predominantly bacteria)

Plaque develops naturally on teeth, and forms part of the defence systems of the host by helping to prevent colonisation of enamel by exogenous (and often pathogenic) microorganisms (colonisation resistance).

Plaque is an example of a biofilm

Plaque is found preferentially at protected and stagnant surfaces, and these are at the greatest risk of disease.

DEFINITION

Dental plaque:

Is defined clinically as a structured, resilient, yellow-grayish substance that adheres tenaciously to the intraoral hard surfaces including removable and fixed restorations.*

*Bowen WH: Nature of plaque. Oral sci Rev 1976;9:3

Because of their unique structure, biofilms are notoriously resistant to

-surfactants -antibiotics -opsonization and -Complement mediated phagocytosis and

killing

TYPES & COMPOSITION OF DENTAL PLAQUETypes of dental plaque:Based on its relationship to the gingival marginDental plaque is broadly classified as

Supragingival plaque -coronal plaque -marginal plaque

Subgingival plaque= 1.Attached plaque a. tooth associated b. tissue associated 2.Unattached plaque

DEFERENCE BETWEEN SUPRA & SUBGINGIVAL PLAQUE

Supragingival

Found at or above the gingival margin.

Contains 50% of matrix.

Bacteria are mostly Gram positive.

Motile bacteria are few.

Aerobic environment.

Predominantly carbohydrate metabolism.

subgingival

Found below the gingival margin.

Contains little or no matrix. Bacteria are mostly Gram

negative.

Motile bacteria are common.

Anaerobic environment. Predominantly protein

metabolism.

SUPRA GINGIVAL PLAQUE:

Tooth surface:Gram positive cocci

Short rods.

Outer surface:Gram negative rods

&filaments Spirochetes.

SUBGINGIVAL PLAQUE:

Tooth associated:Gram +ve rods &cocci

S.mitis,S.sanguis,A.viscosus

A.naeslundii& Eubacterium sps

Tissue assosiated:Gram –ve rods& cocci, filaments flagellated rods, spirochetes

S.oralis,S.intermedius,

Peptostreptococcus,Porphyromonas

Prevotella,Fusobacterium.

Diagram depicting the plaque-bacteria association with tooth surface and periodontal tissues.

Morphologic studies indicate a differentiation of tooth associated and tissue-associated regions of subgingival plaque—Listgarten MA

in certain cases bacteria are found within the host tissues.

-Saglie FR, Carranza FA

COMPOSITION

Dental plaque is composed primarily of microorganisms---70-80% bacteria

One gram of plaque (wet weight) contains approximately 2 x 10 11 bacteria--------Socransky SS

The intercellular matrix, estimated to account for 20% to 30% of the plaque mass

consists of organic and inorganic materials derived from saliva, gingival crevicular fluid, and bacterial products.

Organic constituents = polysaccharides

proteins

glycoproteins, lipid materialGlycoproteins from saliva are an important

component of the pellicle that initially coats a clean tooth surface.

Inorganic components = calcium and phosphorus sodium, potassium,

and fluoride

FORMATION OF DENTAL PLAQUE

The location and rate of plaque formation vary among individuals

determining factors include = oral hygiene as well as

host factors such as diet or

salivary composition and flow

rate . --Manganiello AD,

Socransky SS

Three major phases:

1) Formation of the pellicle

2) Initial adhesion& attachment of bacteria

3) Colonization & plaque maturation

FORMATION OF THE PELLICLE Within nanoseconds after a vigorously polishing the

teeth a thin saliva derived layer called the acquired pellicle covers the tooth surface.

Pellicle consists of glycoproteins proline rich proteins

phospho proteins histidine rich proteins

enzymesFormation of pellicle involves electrostatic,

vanderwaals and hydrophobic forces.

INITIAL ADHESION &ATTACHMENT OF BACTERIA

This includes 4 phases

Phase 1:Transport to surface

This stage involves the initial transport of bacterium

to the tooth surface.

This occurs through

- brownian motion,

- sedimentation of microorganisms ,

-liquid flow or

- active bacterial movement.

Phase 2:Initial adhesion: This stage results in initial reversible adhesion of the

bacterium to the tooth surface. This occurs through vander waals attractive forces &

electrostative repulsive forces.

Phase 3:Attachment: A firm anchorage between bacterium and surface will be

established by specific interactions(covalent,ionic or hydrogen bonding )

Phase 4:Colonization of the surface and biofilm formation:

3.COLONIZATION & PLAQUE MATURATION

Firmly attached microorganisms start growing and newly formed bacterial clusters remain attached resulting in a biofilm

COAGGREGATION

Cell-to-cell recognition of genetically distinct partner cell types—Kolenbrander PE

Interactions of secondary colonizers with primary colonizers

Primary colonizers: Steptococci,Actinomycetes

Cumulative evidence suggests that such adhesion influences the development of complex multi-species biofilms.

Secondary colonizers: P. intermedia, Capnocytophaga, F. nucleatum, Porphyromonas

Examples of coaggregation: Corncob formation: Steptococci adhere to

filaments of Bacterionema matruchotii or Actinomyces

Testube brush: Gram –ve rods adhere to filamentus bacteria

Experimental gingivitis model (Loe et al. 1965) (a) Human volunteer with clean teeth and clinically healthy gingival tissues at the start of the period of experimental plaque accumulation. (b) Same human volunteer after 21 days of abolished oral hygiene practices leading to plaque deposits covering almost all tooth surfaces and consequently developing a generalized marginal gingival inflammation.

MICROSCOPIC STRUCTURE & PHYSIOLOGIC PROPERTIES OF DENTAL PLAQUESupragingival plaque :Gram-positive cocci and short rods predominate at

the tooth surface,

Whereas gram-negative rods and filaments as well as spirochetes predominate in the outer surface of the mature plaque mass.

Highly specific cell-to-cell interactions are also evident from the "corncob" structures" often observed.

Corncob formations have been observed between rod-shaped bacterial cells (e.g., Bacterionema matruchotii or F. nucleatum)

Subgingival plaque:

The tooth-associated (attached) plaque is characterized by gram-positive rods and cocci, including

Streptococcus mitis,

S. sanguis, A.viscosus, Actinomyces

naeslundii, Eubacterium

spp.

The apical border o f the plaque mass is separated from the junctional epithelium by a layer o f host leukocytes.

Left Diagrammatic representation of the histologic structure of subgingival plaque. Right, Histologic section of subgingival plaque. Arrow with box, Sulcular epithelium. White arrow, Predominantly gram-negative unattached zone. Black arrow, Tooth surface. Asterisk, Predominantly gram-positive attached zone.(Listgarten

The bacteria o f this apical tooth-associated region show an increased concentration of gram-negative rods.

Tissue associated plaque is more loosely organized than the very dense tooth-associated region.

It contains primarily gram-negative rods and cocci, as well as large numbers of filaments, flagellated rods, and spirochetes.

Tooth associated plaque is similar to supragingival plaque.

Tissue associated plaque is covered with flagellated bacteria without a well defined extracellular matrix .

Long-standing supragingival plaque near the gingival margin demonstrates "corncob" arrangement. A central gram negative filamentous core supports the outer coccal cells, which are firmly attached by interbacterial adherence or coaggregation

MICROBIAL COMPLEXESRecent analyses of more than 13,000 plaque

samples of 40 subgingival microorganisms using a DNA-hybridization methodology was used to define "complexes" of periodontal microorganisms.

The composition of the different complexes is based on the frequency with which microorganisms are recovered together.*

*Socransky SS, Haffajee AD, Cugini MA, et al: Microbial complexes in subgingival plaque. J Clin Periodontol 1998; 25:134.

the early colonizers are either independent of defined complexes (A. naeslundii, A. viscosus) or members of the yellow (Streptococcus spp.) or purple complexes (A. odontolyticus).

secondary colonizers fell into the green, orange or red complexes.

The green and orange complexes include species recognized as pathogens in periodontal and nonperiodontal infections.

The red complex is of particular interest because it is associated with bleeding on probing, which is an important clinical parameter of destructive periodontal diseases.*

The existence of complexes of species in plaque is another reflection of bacterial interdependency in the biofilm environment.

*Socransky SS, Haffajee AD, Cugini MA, et al: Microbial complexes in subgingival plaque. J Clin Periodontol 1998; 25:134.

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MICROBIAL SPECIFICITY OF PERIODONTAL DISEASESWalter Loesche proposed two hypothesis in 1976.1.Non specific hypothesis2.Specific hypothesis

Nonspecific Plaque Hypothesis (NPH):

The NPH maintains that periodontal disease results from the "elaboration of noxious products by the entire plaque flora*.

*Loesche WJ: Importance of nutrition in gingival crevice microbial ecology. Periodontics 1968; 6:245.

According to this thinking, when only small amounts of plaque are present, the noxious products are neutralized by the host.

NPH is the concept that control of periodontal disease depends on control of the amount of plaque accumulation.

Specific Plaque Hypothesis:(SP)The SP states that only certain plaque is pathogenic, and its

pathogenicity depends on the presence of or increase in specific microorganisms *.

This concept predicts that plaque harboring specific bacterial pathogens results in periodontal disease.

*Loesche WJ: Importance of nutrition in gingival crevice microbial ecology. Periodontics 1968; 6:245.

The resistance of bacteria to antimicrobial agents is significantly increased in the biofilm environment.

----Allison DG, Costerton JW, Helmerhorst EJ

The resistance to antimicrobials inherent to biofilm bacteria may relate to

1.limited diffusion of substances into the biofilm matrix,

2.the slow rate of cell growth in the biofilm environment, possibly to altered properties of bacteria in response to growth on a surface ---Costerton JW

WHAT MAKES PLAQUE PATHOGENIC ?

The following are the possible pathogenic mechanisms by which the plaque microorganisms can cause periodontal disease.

a. physical nature of plaque b. invasion of tissues by bacteria c. release of toxic and inflammatory

substances d. role of bacterial specificity

MICROORGANISMS ASSOCIATED WITH SPECIPHIC

PERIODONTAL DISEASES

The microbiota associated with periodontal health and

disease has been studied with a wide variety of

techniques for sampling, cultivation, and/or detection of bacteria

by DNA hybridization--- Colombo AP, Haffajee AD, Dzink

JL, Socransky SS, Loesche WJ.

PERIODONTAL HEALTH

Actinomyces (viscosus and naeslundii)

Streptococcus (S. mitis and S. sangius)

Veillonella parvula

Small amounts of Gram-negative species are also found.

MICROBIAL SHIFT DURING DISEASE

Comparing the microbiota in health, gingivitis , and periodontitis , the following microbial shifts can be identified:

-From G+ve to G-ve -From cocci to rods (and at a later stage to

spirochetes). -From non motile to motile organisms. -From facultative anaerobes to obligate

anaerobes. -From fermentating to proteolytic species.

CHRONIC GINGIVITIS

Gram-positive (56%), Gram-negative (44%) organisms are found. Predominant gram-positive species include, S.sangius, S.mitis, S.oralis, A.viscosus, A.naeslundii, Peptostreptococcus micros.

Gram-negative organisms are: Fusobacterium nucleatum Prevotella intermedia Veillonella parvula as well as Hemophilus,

Capnocytophaga and Campylobacter species

Pregnancy-associated gingivitis

-Prevotella intermedia

Acute necrotizing ulcerative gingivitis

-Spirochetes -Prevotella intermedia

ADULT PERIODONTITISPorphyromonas gingivalisBacteroides forsythus Prevotella intermediaCampylobacter rectusEikenella corrodensFusobacterium nucleatumActinobacillus actinomycetemcomitansPeptostreptococcus microsTreponema, andEubacterium species.

Viruses such as:EBV-1 (Ebstein-Barr virus)HCMV (Human cytomegalovirus)

Localized juvenile periodontitis Actinobacillus actinomycetemcomitansPorphyromonas gingivalisEikenella corrodensCampylobacter rectusFusobacterium nucleatumBacteroides Eubacterium CapnocytophagaHerpes virus.

Generalized juvenile periodontitisActinobacillus actinomycetemcomitansPorphyromonas gingivalisPrevotella intermedia CapnocytophagaEikenella corrodens Neisseria

Refractory periodontitisActinobacillus actinomycetemcomitansBacteroides ForsythusPorphyromonas gingivalisPrevotella intermediaWolinella recta

Abscesses of the periodontium

Fusobacterium nucleatum

Prevotella intermedia

Peptostreptococcus micros

Bacteroides forsythus

Porphyromonas gingivalis

TRANSMITION OF PERIODONTAL PLAQUE BACTERIASeveral transmission studies have demonstrated that

the same clonal types of P gingivalis, Aa, P. intermedia, Porphyrornonas endodontalis and Streptococcus mutans are present in family members

---Petit MD, van Steenbergen TJ, Scholte LM

But different clonal types are found in unrelated individuals

This suggests that transmission most likely occurs through intimate contact

It would be interesting to determine what clonal type was present in spouses before they were married.

Assessment of plaque can be done by using 1.plaque index

2. PHP index

Plaque control can be done by 1.mechanical methods 2.chemical methods

BIOFILM

Introduction:Commonly encountered as a layer on slime clogging

drainpipesbiofilms appear in our everyday life in more than one

form.Bacteria suspended in water or aqueous cultures are

"Planktonic“.In such aqueous environments when bacteria adhere

to surfaces and become "Sessile", secreting a slimy-glue like substance for anchorage, they form a biofilm*

* Costerton JW, Lewandowski Z, Caldwell DE, Korber DR, Lappin -Scott HM. Microbial biofilms. Ann Rev Microbiol 1995; 49:711-45.

Definition:

“Matrix ---enclosed bacterial populations adherent to each other and or/to surface or interfaces—Costerton.

The term biofilm describes the relatively undefinable microbial community associated with a tooth surface or any other hard, non-shedding material (Wilderer & Charaklis 1989)

A single bacterial species can form a biofilm,

But in natural environment often biofilms are formed from various species of bacteria, fungi, algae, protazoa, debris along with corrosion products Adhesion to surfaces.

provides considerable advantage for the biofilm forming bacteria, such as protection from anti-microbial agents, exchange of nutrients, metabolites or genetic material from close proximity to other micro organisms.

FORMATION OF BIOFILM

Pioneering studies by Cholodny, Henrici and Zo Bell commenced more than 50-60 years ago.

Usually the methodology involved was unmersion of glass slides into natural environments and observing the biofilms developed under microscope.

Biofilm formation usually commences with the colonization of a surface by bacteria.

The adhesion and attraction of the bacteria to the surface may be brought about by different mechanisms.

-surface charge, -gravity, -Brownian motion

and - chemo attraction,After attraction, attachment of the bacteria to the

surface occurs by a two-step process comprised of reversible binding*

1. The reversible binding is usually brought about by weak Vander Waal forces

2. stronger attachment can form by a combination of both physical or chemical forces.

Jenkinson HF, Lappin-Scott HM. Biofilms adhere to stay. Trends Microbiol. 2001; 9(1):9-10.

Factors influencing bacterial attachment to surfaces :

-Nutrient availability at the surface, -nutrient concentration, -pH available, - temperature, - electrolyte concentration, -the flux of materials and surface types such as : a) High surface energy materials: These are

negatively charged hydrophilic materials such as glass, metal or minerals.

b) Low surface energy materials : These are either low positively or low negatively charged hydrophobic materials such as plastics made up of organic polymers.

Limiting of a biofilm is brought about by the removal of sessile cells or sections of the biofilm, as a result of erosion, sloughing or abrasion processes.

l. Erosion leads to a continuous loss of cells from

biofilm. Thickness of biofilm, fluid shear stress and fluid velocity all affect the rate of erosion.

2. Sloughing is observed in cases of bulky biofilms

and involves a large and rapid loss of material from the biofilm.

3. Abrasion occurs when some object repeatedly

collides with the biofilm.

BIOFILMS IN MEDICAL SYSTEMS

Orthopedic implants (Screws, pins, plates, and other prosthetic devices) :

Ocular lenses, heart values, vascular grafts, intra uterine devices,

temporary indwelling catheters, intra-venous catheters ports/caths and reverse osmosis membrane filters

REFERENCE

Carranza ; clinical periodontology-9th & 10th edition

Jan Lindhe ; clinical periodontology& Implantology

Max A ,Listgarten; The structure of dental plaque. Periodontal 2000;5:1994

RICHARD P . DARVEAAUN, NET ANNER& ROYC . PAGE; the microbial challenge in periodontitis Periodontology 2000, Vol. 14, 1997, 12-32

Sigmund S, Socransky, Anne D. Haffajee. Dental biofilms: difficult therapeutic targets; periodontal 2000;28:2002

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