MICROBIOLOGY OF PERIODONTAL DISEASES

Dr. Ali YaldrumFaculty of Dentistry, SEGi University.

LEARNING OBJECTIVES

• At the end of this session, the student should be able to describe:

What is Periodontium and its roleEcology of Dental Crevice and its roleConditions that affect Periodontal tissueRole of Microorganisms in Periodontal DiseaseComplex relationship between Plaque and periodontal disease

The term ‘periodontal diseases’ embraces a number of conditions in which the supporting tissues of the teeth are attacked.

Supporting tissues are

• Gingivae

• Cementum

• Alveolar Bone

• Periodontal Ligament Fibers (PDL)

ECOLOGY OF DENTAL CREVICE

Ecology of dental crevice is different from other sites in the oral cavity

• more anaerobic

• bathed in gingival crevicular fluid (GCF)

*In diseased state crevice becomes a pocket

• In Periodontal pocket

• Low Eh (oxidation reduction potential)

• increased flow of GCF

➡in gingivitis: 147% increase

➡in periodontitis: 30 fold increase

GINGIVAL CREVICULAR FLUID

GCF contains

• humoral and cellular defense factors

• proteins & glycoproteins

alter the gene expression

alter the competitiveness ofperiodontal pathogens

disturbing the natural balanceof subgingival microflora

growth of proteolyticgram -ve organisms

assachrolytic & proteolytic increased growth & enzyme activity of periodontal pathogen Porphyromonas gingivalis

slight increase during inf lammation

P. gingivalis

enhanced attachment of microbes with epithelium

ENVIRONMENTProteolysis Decreased pH increased temperatureMetabolism

• Flow of GCF can remove planktonic microorganisms

• Prevotella, Porphyromonas, Fusobacterium spp. adheres to Streptococcus & Actinomyces colonizing cementum

• Parvimonas micra adhere crevicular epithelial cells

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destruction directly (and satisfy Koch’s postulates), while in other pockets, different bacteria could fill identical roles.

AETIOLOGY OF PERIODONTAL DISEASE – CONTEMPORARY PERSPECTIVE

The predominant bacteria found in the various types of periodontal disease are different to those that are prevalent in the healthy gingival crevice. One of the most intriguing questions in periodontology, there-fore, concerns the reservoir and source of these potential periodontal pathogens. Some periodonto-pathic bacteria can attach to mucosal surfaces, and a range of putative periodontal pathogens (includ-ing black-pigmented anaerobes, Capnocytophaga spp., spirochaetes and Fusobacterium spp., etc) can be isolated from the dorsum of the tongue and from tonsils. Recently, P. gingivalis, T. forsythia and A. actinomycetemcomitans were detected in 23%, 11% and 30%, respectively, of tongue samples from young children. Most species were detected more frequently from tongue rather than plaque samples, confirming

that the tongue may act as a reservoir for these peri-odontal pathogens. Some pathogens may also persist by invading buccal epithelial cells (Fig. 4.6).

In general, the putative periodontal pathogens are non-competitive with other members of the resident subgingival microflora at healthy sites, and remain at low levels; such levels would not be clinically sig-nificant. If plaque is allowed to accumulate beyond levels that are compatible with health, then the host mounts an inflammatory response. The flow of GCF is increased, and this introduces into the crevice not only components of the host defences but also com-plex host molecules (e.g. transferrin, haemoglobin, etc.) that can be catabolized and used as a nutrient source by the proteolytic Gram negative anaerobes that predominate in advanced periodontal lesions. This metabolism leads to an increase in local pH and a fall in the redox potential (i.e. the pocket becomes slightly alkaline and more anaerobic). These changes in the local environment will up-regulate some of the virulence factors associated with these putative pathogens (e.g. protease activity by P. gingivalis), and favour their growth at the expense of the species asso-ciated with gingival health (i.e. increase the competi-tiveness of the potential pathogens). This can lead

D

D

CC

AA

A

C

Fluid flowTooth

Modulationof host

defences

Immune response –innate and adaptive

Systemic

Cellular

Gingiva

Tissuedestruction

X X

Fig. 6.16 Pathogenic synergy in the aetiology of periodontal diseases. Bacteria capable of causing tissue damage directly (e.g. species X) may be dependent on the presence of other cells (e.g. organisms C and D) for essential nutrients or attachment sites so that they can grow and resist the removal forces provided by the increased flow of GCF. Similarly, both of these groups of bacteria may be reliant for their survival on other organisms (e.g. A and C) to modulate the host defences. Individual bacteria may have more than one role (e.g. organism C) in the aetiology of disease.

Pathogenic synergy in the aetiology of periodontal diseases. Bacteria capable of causing tissue damage directly (e.g. species X) may be dependent on the presence of other cells (e.g. organisms C and D) for essential nutrients or attachment sites so that they can grow and resist the removal forces provided by the increased flow of GCF. Similarly, both of these groups of bacteria may be reliant for their survival on other organisms (e.g. A and C) to modulate the host defences. Individual bacteria may have more than one role (e.g. organism C) in the aetiology of disease.

• The main aetiological agent of periodontal disease is microflora inhabiting subgingival plaque.

• Host tissues and its specific and non- specific host defense mechanisms play crucial modulating roles.

HOST TISSUE

• Dentogingival junction is most vulnerable site for microbial attack but can be maintained healthy with good oral hygiene

• Plaque accumulates close to the gingival margin, the host defenses are overcome and gingival inflammation (gingivitis) and subsequent periodontal inflammation with loss of attachment ensues (periodontitis).

Specific Host Defense Factors Non-specific Host Defense Factors

B and T LymphocytesPolymorphsMacrophages

Antibodies: IgA, IgG, IgM

Complement systemProteasesLyzozymeLactoferrin

IgG, Immunoglobulin G.

Dental Plaque is an essential aetiological agent in the development and progression of periodontal diseases and is shown by the following

� Epidemiological data� Clinical Studies� Topical application of antimicrobial agents� Initiation of disease in gnotobiotic animals by peridontopathogenic bacteria

PLAQUE SPECIFIC HYPOTHESIS

� In certain disease states such as necrotizing ulcerative gingivitis the key aetiologic agents are fusobacteria and spirochaetes. Direct involvement of Aggregatibacter actinomycetemcomitans in aggressive (juvenile) periodontitis

� Disease can be resolved by appropriate antibiotics active against anaerobes (metronidazole) and tetracycline.

NON-SPECIFIC PLAQUE HYPOTHESIS

� Collective groups or consortia of different bacteria have the total complement of virulence factors required for periodontal tissue destruction

� some bacteria can substitute for others absent from the pathogenic consortium.

� This hypothesis implies that plaque will cause disease irrespective of its composition, and it is supported by the clinical findings of numerous bacterial species in diseased periodontal pockets.

ECOLOGICAL PLAQUE HYPOTHESIS

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to a shift in the proportions of the resident subgin-gival microflora (Fig. 6.1). This is analogous to the increases in mutans streptococci and Lactobacillus spp. seen prior to caries development following the repeated ingestion of dietary carbohydrates.

Evidence for these bacterial population shifts has come from laboratory studies. The growth of subgin-gival plaque on human serum (used to mimic GCF) led to the selection of species associated with peri-odontal destruction, such as black-pigmented anaer-obes, anaerobic streptococci, Fusobacterium spp. and spirochaetes; most of these species could not be detected in the original samples. Likewise, in the laboratory, a rise in pH from 7.0 to 7.5 (as can occur during inflammation) allowed the proportions of P. gingivalis to rise from <1% to >99% of a microbial community of black-pigmented anaerobes. If similar events occur in a pocket, then periodontal diseases can be regarded as endogenous infections, caused by an imbalance in the composition of the resident microflora at a site, due to an alteration in the ecol-ogy of the local habitat. This view is formulated in the ‘ecological plaque hypothesis’, which describes the dynamic relationship between the resident micro-flora and the host in health and disease in ecological terms (Fig. 6.17). A consequence of this hypothesis is that disease can be prevented not only by targeting the putative pathogens, but also by interfering with the environmental factors that drive the changes in the balance in the microflora, e.g. such as by reduc-ing the severity of the inflammatory response, or by altering the redox potential of the pocket to prevent the growth of the obligate anaerobes. Other relevant changes in the local environment that could perturb

the host-microbe balance could come from trauma, an alteration in the immune status of the host (e.g. during systemic disease or after drug therapy), or from tobacco smoking.

Transmission of P. gingivalis and A. actinomycetem-comitans can occur among family members, with par-ents and children sharing identical strains. Likewise, some married couples have the same clonal types of P. gingivalis and A. actinomycetemcomitans indicat-ing that these species can be transmitted between spouses (Ch. 4). Therefore, treatment might also require elimination or suppression of putative peri-odontal pathogens from their primary oral reser-voirs. However, even if a pathogen is acquired from another individual, it will be present initially at only low cell levels. As described earlier, a major ecologi-cal disturbance will have to occur at a site if these putative pathogens are going to be able to out-com-pete the other members of the plaque microflora, and achieve numerical dominance and clinical sig-nificance (Fig. 6.11).

Destructive disease is the outcome of a complex interplay of interactions between the host and the altered microbial challenge (Fig. 6.18). This inter-play can be considered to occur between several discrete, interactive compartments: (a) the micro-bial challenge, (b) the immune and inflammatory response of the host, and (c) connective tissue and bone metabolism. These interactions are influenced by disease modifiers, which may be genetic (e.g. neutrophil defects) or environmental (e.g. tobacco smoking) factors. The clinical signs reflect the sum of these interactions, and the severity of the disease can feed back to influence the microbial challenge,

Fig. 6.17 A schematic representation of the ‘ecological plaque hypothesis’ in relation to periodontal disease. Plaque accumulation produces an inflammatory host response; this causes changes in the local environmental conditions which favour the growth of proteolytic and anaerobic Gram negative bacteria. Disease could be prevented by not only targeting the putative pathogens, but also by interfering with the factors driving their selection.

Plaquereduction

Plaqueaccumulation

Gin

giva

lhe

alth

Gin

givi

tis

Reducedinflammation

Low GCF flowHigher Eh

PredominantlyGram positive microflora

Facultativeanaerobes

Increasedinflammation

PredominantlyGram negative microflora

Obligateanaerobes

High GCF flowLower Eh

Inflammatoryresponse

Environmentalchange

Ecologicalshift

Ecology of Healthy Gingival Crevice

Ecology in Gingivitis

Ecology in Periodontitis

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and Treponema denticola, and their presence was often preceded by members of the orange complex, which was also often found in deeper pockets, but was more diverse in membership. In contrast, species of the yellow, green and purple complexes, together with A. naeslundii, were considered to be ‘host com-patible’, and were generally associated with healthy sites. Aggregatibacter (formerly Actinobacillus) actin-omycetemcomitans serotype b did not fall within a complex, and is associated more with aggressive periodontitis (see later). Thus, chronic periodon-titis appears to result from the activity of mixtures of interacting bacteria, and therefore has a polymi-crobial aetiology. There is a progressive change in the composition of the microflora from health and gingivitis to periodontitis. This change involves not only the emergence of apparently previously unde-tected species, but also modifications to the num-bers, or proportions, of a variety of species already present.

Two theories have been proposed to explain the emergence of previously undetected species. It may be due to the selective growth (enrichment) of a microorganism that is present in health in only very low numbers, due to a change in the environment during disease. Alternatively, it might be due to the exogenous acquisition of periodon-tal pathogens from other diseased sites or sub-jects. The recent application of sensitive molecular

approaches has identified low levels of many of the putative pathogens at healthy sites, while evi-dence of transmission of organisms such as P. gingivalis and A. actinomycetemcomitans between spouses has been obtained. However, in either sit-uation, a major change to the ecology of the habi-tat has to occur in order to enable low levels of an organism to outcompete the existing members of the resident microflora and reach clinically signifi-cant proportions within the subgingival biofilm. The most likely environmental changes capable of causing such a shift in the microflora are asso-ciated with the host inflammatory response (Fig. 6.11). The increased flow of gingival crevicular fluid (GCF) not only introduces components of the host defences but also a range of novel nutri-ents that selects for the growth of asaccharolytic, anaerobic bacteria. This proteolytic metabolism leads to a rise in local pH, which also favours the growth of many of the bacteria that predominate in periodontitis (Fig. 2.4).

The application of molecular methods to char-acterize the subgingival microflora of sites with chronic periodontitis has further emphasized the diversity of bacteria found in these sites (Table 6.7). A large proportion of clones identified by 16S rRNA gene sequencing belong to novel phylo-types, some of which have no cultivable represen-tatives. Some studies have detected unculturable

V. parvulaA. odontolyticus

S. oralisS. mitisE. corrodens

Streptococcus spp.S. gordoniiS. intermedius

E. corrodensC. gingivalisC. sputigenaC. ochraceaeC. concisusA. a serotype a

Aggregatibacterium (Actinobacillus)actinomycetemcomitans b.

S. constellatus

C. gracillisC. rectus

E. nodatum

P. gingivalisT. forsythiaT. denticola

P. intermediaP. nigrescensP. microsF. nucleatum

Fig. 6.10 The grouping of bacteria into complexes to reflect their relationship with the host in health and periodontal disease. The ‘red complex’ is found most frequently in deep periodontal pockets, and their presence was usually preceded by members of the ‘orange complex’. Members of the ‘yellow’, ‘green’ and ‘purple’ complexes were generally associated with healthy sites.Purple, Yellow, Green: Healthy Gingival sulcus Red, Orange: Periodontal pockets

Plaque-associated gingivit is has been separated into three stages

• Stage 1: The init ial lesion

• Stage 2: The ear ly lesion

• Stage 3: Establ ished lesion

STAGE 1:THE INITIAL LESION

• develops within 4 days of plaque accumulation.

• Micro-f lora consists mostly of Gram-posit ive cocci (Streptococcus spp.) .

• Histological ly, there is an acute inf lammator y reaction.

• The lesion is character ized by increased f low of GCF, migration of PMN leukocytes into the gingival sulcus from the local vasculature .

• Adjacent to the junctional and sulcular epithel ia, the inf lammator y infi ltrate occupies approximately 5-10% of the gingival connective tissue .

• This init ial lesion is not vis ible cl inical ly.

STAGE 2: THE EARLY LESION

• Appear s after approximately 7 days of plaque accumulation, detectable cl inical ly as gingivit is .

• lower oxygen tension and the plaque f lora shifts to more Actinomyces spp. , spirochaetes and capnophil ic organisms.

• Histological ly, the gingival infi ltrate in the ear ly lesion is dominated by lymphocytes (75%) and macro-phages,few plasma cel ls

• The infi ltrated area occupies approximately 15% of the marginal gingival connective t issue , with some local destruction of col lagen.

• Migration of polymorphonuclear leukocytes into the gingival sulcus and crevicular f luid peaks at 6 to 12 days fol lowing the onset of cl inical ly

STAGE 3: ESTABLISHED LESION

• After a var iable per iod of t ime the subgingival microflora develops into an environment that can suppor t the growth of obligate anaerobes such as Porphyromonas gingival is and Prevotel la intermedia.

• Histological ly, there is a fur ther increase in the size of the inf lammator y lesion within the affected gingiva, with a shift to a predominance of plasma cel ls and B-lymphocytes.

• The junctional and pocket epithel ia are heavi ly infi ltrated with neutrophils . Plasma cel ls are found at the per ipher y of die lesion, while macrophages and lymphocytes are present in the lamina propr ia of the pocket wall .

• Establ ished lesions may per sist for months or year s without progression to per iodontit is .

TYPES OF GINGIVITIS

• Chronic Marginal gingivitis

• Acute Necrotizing Ulcerative gingivitis

• Medication influenced gingivitis

• Gingivitis associated with systemic diseases

• Acute Herpetic gingivostomatitis

PERIODONTITIS

Per iodontit is may be defined cl inical ly as inf lammation of the suppor ting t issues of the teeth.

• Maintains al l the features of the establ ished lesion of gingivit is

• Migration of the junctional epithel ium down the root surface , alveolar bone resorption and subsequent pocket formation

• Character ized by progressively destructive changes destroying alveolar bone and per iodontal l igament, with an attachment loss of more than 3 mm.

• Histological ly, the conver sion of the establ ished lesion of gingivit is into per iodontit is is character ized by destruction of the connective tissue attachment to the root surface and by alveolar bone loss.

FORMATION OF DENTAL POCKET

• Creates highly anaerobic environment

• pH shifts from 6.9 to approximately 7.4 to 7.8 and the pocket is continual ly bathed by the protein-r ich GCF, which encourages growth of proteolytic bacter ia.

• Subgingival plaque have a dense zone of mostly Gram-posit ive bacter ia attached to the tooth surface and a less densely packed zone of mainly Gram-negative organisms next to the gingival surface .

REFERENCES

Philip D. Marsh, Michael V Martin, “Plaque mediated diseases- Dental Caries and Periodontal diseases” in Oral Microbiology, 5th Edition, Churchil Livingstone, 2009, pp 117-137

J. Bagg, T. W. Macfarlane, I. R. Poxton and A. J. Smith, “Periodontal Diseases” in Essentials of Microbiology for Dental Students, 2nd Edition, Oxford University Press, 2006 pp 249-259

Lakshman Samaranayake, “Microbiology of Periodontal diseases” in Essential Microbiology for Dentistry, 3rd Edition, Churchil Livingstone, Elsevier, pp 275 - 287.