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- 2. Contents Introduction Routes of rootcanal infection Mechanisms of Microbial Pathogenicity and Virulence Factors Biofilm and Community-Based Microbial Pathogenesis Biofilm and Bacterial Interactions Biofilm Community Lifestyle Quorum Sensing— Bacterial Intercommunication Methods for Microbial Identification Diversity of the Endodontic Microbiota Primary Intra radicular Infection Spatial Distribution of the Microbiota Microbial Ecology and the Root Canal Ecosystem Secondary/Persistent Infections and Treatment Failure Extra radicular infection Conclusion
- 3. Introduction Microorganisms causevirtually all pathoses of the pulp and periapical tissues. Once bacterial invasion of pulp tissues has taken place, both non-specific inflammation and specific immunologic response of the host have a profound effect on the progress of the disease. Endodontic infection develops in root canals devoid of host defenses, pulp necrosis (as a sequel to caries, trauma, periodontal disease,or iatrogenic operative procedures) or pulp removal for treatment. Biofilm-induced oral diseases.
- 4. ROUTES OF ROOT CANAL INFECTION •Caries • Trauma-induced fractures • Cracks • Restorative procedures • Scaling and root planing • Attrition • Abrasion • Gaps in the cementoenamel junction at the cervical root surface • Dentinal tubules • Direct pulp exposure • Periodontal disease • Anachoresis Cohen's 11th edition
- 5. Mechanisms of MicrobialPathogenicity and Virulence Factors Pathogenicity : The ability of a microorganism to cause disease. Virulence: Degree of pathogenicity of a microorganism. Some microorganisms routinely cause disease in a given host and are called primary pathogens. Other microorganisms cause disease only when host defenses are impaired and are called opportunistic pathogens by changing the balance of the host–bacteria relationship. v Bacterial strategies that contribute to pathogenicity include the ability to coaggregate and form biofilms. Cohen's 11th edition
- 6. In thepathogenesis of primary apical periodontitis Bacteria in caries lesions form authentic biofilms adhered to dentin. Diffusion of bacterial products through dentinal tubules induces pulpal inflammation After pulp exposure, the exposed pulp tissue is in direct contact with bacteria and their products and responds with severe inflammation. Some tissue invasion by bacteria may also occur. Bacteria in the battlefront have to survive the attack from the host defenses and at the same time acquire nutrients to keep themselves alive. In this bacteria–pulp clash, the latter invariably is “defeated” and becomes necrotic, so bacteria move forward and “occupy the territory”—that is, they colonize the necrotic tissue. Cohen's 11th edition
- 7. These events advancethrough tissue compartments, coalesce, and move toward the apical part of the canal until virtually the entire root canal is necrotic and infected. At this stage, involved bacteria can be regarded as the early root canal colonizers or pioneer species (play an important role in the initiation of the apical periodontitis disease process, modify the environment, making it conducive to the establishment of other bacterial groups)
- 9. Biofilm and Community-BasedMicrobial Pathogenesis Individual microorganisms proliferating in a habitat give rise to Population which interact with one another to form a Community which is a part of a larger system called Ecosystem (a functional self supporting system that includes the microbial community and its environment). • Each population occupies a functional role (niche) within the community • Community profiling studies revealed that bacterial composition of the endodontic microbiota differs consistently between individuals. From the perspective of the single-pathogen concept: ◦ Apical periodontitis can be considered as having no specific microbial etiology. However, based on the community as-a- pathogen concept: ◦ it is possible to infer that some communities are more related to certain forms of the disease Cohen's 11th edition
- 10. Biofilm: asessile multicellular microbial community characterized by cells that are firmly attached to a surface and enmeshed in a self-produced matrix of extracellular polymeric substance (EPS), usually polysaccharide. Biofilm infections account for an estimated 65% to 80% of bacterial infections that affect humans in the developed world. Community members form, o distinct populations or microcolonies separated by open water channels that traverse the biofilm matrix and create primitive circulatory systems. o vital nutrients and communication molecules can diffuse, and wastes can be washed out through these channels. During the early stages of biofilm formation, bacteria bind to many host proteins and coaggregate with other bacteria. changes in growth rate, gene expression, and protein production. Biofilm and Bacterial Interactions Cohen's 11th edition
- 11. Development of biofilm First stage of biofilm involves the adsorption of macromolecules in the planktonic phase to surface- a conditioning film forms- (transport of microbe to the substrate surface) Second stage – adhesion and co-adhesion of microbes and attachment strengthened by polymer production and unfolding of cell surface structures- (initial non-specific microbial-substrate adherence phase) Third stage involves the multiplication and metabolism of attached microorganisms -(bacterial growth and biofilm expansion) Fourth stage involves detachment of biofilm.
- 12. Biofilm Community Lifestyle ABroader Habitat Range for Growth The metabolism of early colonizers alters the local environment, setting the stage for attachment and growth of latecomers (including more fastidious species) Increased Metabolic Diversity and Efficiency Take part in a number of nutritional interrelationships, and food webs. Products of the metabolism of one species may become the main source of nutrients for other species. Byproducts of the degradation of complex nutrients are trapped in the biofilm matrix and shared with other community members Cohen's 11th edition
- 13. Protection From CompetingMicroorganisms, Host Defenses, Antimicrobial Agents, and Environmental Stress Beta-lactamases, catalase, and proteinases Retained in the biofilm matrix protect other bacteria against antimicrobials and host defenses. Genetic Exchanges Horizontal gene transfer in the community. Conjugation, transformation, and transduction. Dissemination of virulence and antibiotic- resistance genes. Cohen's 11th edition
- 14. A diverse rangeof virulence traits are required for these particular stages of the disease process, Require the concerted action of bacteria in a community Bacterial species that individually have low virulence and are unable to cause disease can do so when in association with others as part of a mixed consortium (pathogenic synergism) Enhanced Pathogenicity Cohen's 11th edition
- 15. Resistance to AntimicrobialAgents Biofilm Structure May Restrict Penetration of Antimicrobial Agents The antibiotic concentration required to kill bacteria in the biofilm About 100 to 1000 times higher than that needed to kill the same species in planktonic state Altered Growth Rate of Biofilm Bacteria : Bacteria grow slowly under conditions of low availability of nutrients in an established biofilm much less susceptible than faster-dividing cells. Most antibiotics require at least some degree of cellular activity to be effective. Cohen's 11th edition
- 16. Presence of“Persister” Bacteria Increased tolerance of some biofilms to antibiotics may be largely due to the presence of a subpopulation of specialized survivor cells known as persisters. Quorum Sensing –Bacterial Intercommunication Cell-cell communication that regulate gene expression in a cell density–dependent manner Quorum sensing Production, release, and subsequent detection of diffusible signaling molecules Autoinducers
- 17. Two predominant typesof autoinducers: ◦ N-acyl-l-homoserine lactones (AHLs) ◦ Post translationally modified peptides Used by gram negative and gram-positive bacteria, respectively. Bacteria can perform specific functions only when living in groups: ◦ Regulate virulence ◦ Competence for DNA uptake ◦ Entry into stationary phase, ◦ Biofilm formation Entry into stationary phase dramatically alters patterns of gene expression Allow extended cell survival in the absence of nutrients.
- 18. METHODS FOR MICROBIALIDENTIFICATION Endodontic samples are collected and transported to the laboratory ◦ in a viability-preserving, non-supportive, anaerobic medium. Dispersed by sonication or vortex mixing Diluted, distributed onto various types of agar media cultivated under aerobic or anaerobic conditions After a suitable period of incubation, Individual colonies are sub cultivated and Identified on the basis of multiple phenotype- based aspects colony and cellular morphology, gram-staining pattern, oxygen tolerance, comprehensive biochemical characterization, metabolic end-product analysis by gas-liquid chromatography Microbiologic culture method Cohen's 11th edition
- 19. The outer cellularmembrane protein profile as examined by gel electrophoresis, fluorescence under ultraviolet light Susceptibility tests to selected antibiotics can be needed for identification of some species. Marketed packaged kits that test for preformed enzymes have also been used for rapid identification of several species. Not all microorganisms can be cultivated under artificial conditions Nutritional and physiologic needs of most microorganisms are still unknown. Cohen's 11th edition
- 20. To sidestepthe Limitations of culture Molecular biology substantially improved to achieve a more realistic description of the microbial world without the need for cultivation. Molecular approaches for microbial identification rely on certain genes that contain revealing information about the microbial identity. 16S rRNA gene (or 16S rDNA) has been the most widely used because it is universally distributed among bacteria, Cohen's 11th edition
- 21. There are anestimated 10 billion bacterial cells in the oral cavity. Over 50% to 60% of the oral microbiota still remains to be cultivated and fully characterized. More than 400 different microbial species/phylotypes have been found in infected root canals. Endodontic infections develop in a previously sterile place that does not contain a normal microbiota. Culture and molecular studies reveal only prevalence of species. DIVERSITY OF THE ENDODONTIC MICROBIOTA Cohen's 11th edition
- 22. TYPES OF ENDODONTICINFECTIONS Endodontic infections can be classified according to the anatomic location as: Intra radicular infection is caused by microorganisms colonizing the root canal system and can be subdivided into three categories : primary infection: caused by microorganisms that initially invade and colonize the necrotic pulp tissue. secondary infection: caused by microorganisms not present in the primary infection but introduced in the root canal at some time after professional intervention. persistent infection: caused by microorganisms that were members of a primary or secondary infection and in some way resisted intracanal antimicrobial procedures and were able to endure periods of nutrient deprivation in treated canals. Extra radicular infection in turn is characterized by microbial invasion of the inflamed peri radicular tissues and is a sequel to the intra radicular infection. Extra radicular infections can be dependent on or independent of the intra radicular infection. Cohen's 11th edition
- 23. PRIMARY INTRARADICULAR INFECTION Mixed community conspicuously dominated by anaerobic bacteria. The number of bacterial cells may vary from 10 3 – 10 8 per root Canal. Molecular studies have disclosed a mean of 10 to 20 species/phylotypes per infected canal. Canals of teeth with sinus tracts exhibit a mean number of 17 species. The size of the apical periodontitis lesion has been shown to be proportional to the number of bacterial species and cells in the root canal. The larger the lesion, the higher the bacterial diversity and density in the canal. Geographic Influence patients residing in distinct geographic locations and suggested that significant differences in the prevalence of some important species can actually exist. Cohen's 11th edition
- 25. SPATIAL DISTRIBUTION OFTHE MICROBIOTA Bacterial cells from endodontic biofilms are very often seen penetrating the dentinal tubules. Dentinal tubule infection can occur in about 70% to 80% of the teeth with apical periodontitis. Bacteria present as planktonic cells in the main root canal may be easily accessed and eliminated by instruments and substances used during treatment. Those organized in biofilms: Attached to the canal walls Located into isthmuses, lateral canals, Dentinal tubules More difficult to reach and may require special therapeutic strategies to be eradicated. Cohen's 11th edition
- 26. Whenever dentinis exposed, Pulp is put at risk of infection Permeability of normal dentin dictated by its tubular structure largest diameter located near the pulp (mean, 2.5 μm) Smallest diameter in the periphery, near the enamel or cementum. (mean, 0.9 μm) The smallest tubule diameter is entirely compatible with the cell diameter of most oral bacterial species: Which usually ranges from 0.2 to 0.7 μm Bacterial invasion of dentinal tubules occurs more rapidly in nonvital teeth than in vital. Presence of tubular contents (In Vital teeth) the functional or physiologic diameter of the tubules is only 5% to 10% of the anatomic diameter. Cohen's 11th edition
- 27. Most of thebacteria in the carious process are non-motile Invade dentin by repeated cell division which pushes cells into tubules Bacterial cells may also be forced into tubules by hydrostatic pressures Developed on dentin during mastication. Cohen's 11th edition
- 28. The rootcanal infection is a dynamic process, and different bacterial species apparently dominate at different stages. In the very initial phases of the pulpal infectious process facultative bacteria predominate. After a few days or weeks, oxygen is depleted loss of blood circulation in the necrotic pulp. Growth of obligate anaerobic bacteria. Microbial Ecology and the Root Canal Ecosystem Cohen's 11th edition
- 29. The main sourcesof nutrients for bacteria colonizing the root canal system include: The necrotic pulp tissue. Proteins and glycoproteins from tissue fluids and exudate that seep into the root canal system via apical and lateral foramens. Components of saliva that may coronally penetrate into the root canal. Products of the metabolism of other bacteria. Cohen's 11th edition
- 30. Interbacterial nutritionalinteractions are important ecologic determinants that result in higher metabolic efficiency of the whole community. Nutritional interactions are mainly represented by food webs, including utilization of metabolic end products from one species by another and bacterial cooperation for the breakdown of complex host-derived substrates. Cohen's 11th edition
- 31. Other Microorganisms inEndodontic Infections: Fungi: Candida species Archaea Viruses o Noninflamed vital pulps of Patients infected with the human immunodeficiency virus. o Human cytomegalovirus (HCMV) and Epstein-Barr virus (EBV) have been detected in apical periodontitis lesions. Cohen's 11th edition
- 32. PERSISTENT/SECONDARY ENDODONTIC INFECTIONSand Treatment Failure Persistent or secondary intraradicular infections are the major causes of endodontic treatment failure. Involved microorganisms are remnants of a primary or secondary infection. Microorganisms that at some time entered the root canal system secondary to professional intervention. Bacteria at the Root Canal–Filling Stage: Diligent antimicrobial treatment may still fail to completely eliminate bacteria from the infected root canal System. Persisting bacteria are either resistant or inaccessible to treatment procedures. When bacteria resist treatment procedures, gram- positive bacteria are more frequently present. Cohen's 11th edition
- 33. Gram-positive facultativesor anaerobes often detected in these samples include: Streptococci, P. micra, Actinomyces species, Propionibacterium species, P. alactolyticus, lactobacilli, E. faecalis, Olsenella uli Gram positive bacteria can be more resistant to antimicrobial treatment measures and have the ability to adapt to the harsh environmental conditions in instrumented and medicated root canals. Bacteria persisting in the root canal after chemomechanical procedures or intracanal medication will not always maintain an infectious process Cohen's 11th edition
- 34. Some apical periodontitislesions healed even after bacteria were found in the canal at the filling stage: Residual bacteria may die after filling because of toxic effects of the filling material, access denied to nutrients, or disruption of bacterial ecology. Residual bacteria may be present in quantities and virulence subcritical to sustaining periradicular inflammation. Residual bacteria remain in locations where access to periradicular tissues is denied. Host resistance to infection is also an important and probably decisive counteracting factor. Cohen's 11th edition
- 35. Microbiota in EndodonticallyTreated Teeth Several culture and molecular biology studies have revealed that E. faecalis is the most frequent species in root canal– treated teeth, with prevalence values reaching up to 90% of cases commonly recovered from teeth treated at multiple visits or left open for drainage Canals apparently well treated harbor one to five species, but the number of species in canals with inadequate treatment can reach up to 10 to 20 species, which is similar to untreated canals. Cohen's 11th edition
- 36. The abilityof E. faecalis : to penetrate dentinal tubules, sometimes to a deep extent enable it to escape the action of endodontic instruments and irrigants It is resistant to calcium hydroxide acidify the cytoplasm Cohen's 11th edition
- 37. E. faecaliscan enter a so-called viable but noncultivable (VBNC) state In the VBNC state, bacteria lose the ability to grow in culture media maintain viability and pathogenicity can resume division when optimal environmental conditions are restored. Cohen's 11th edition
- 38. E. faecalis asthe main causative agent of endodontic failures: ♦ Despite being easily cultivated, E. faecalis is not detected in all studies evaluating the microbiota of root canal–treated teeth with posttreatment disease. ♦ Even when present, E. faecalis is rarely one of the most dominant species in retreatment cases. ♦ E. faecalis has been found not to be more prevalent in root canal–treated teeth with lesions when compared with treated teeth with no lesions Cohen's 11th edition
- 39. EXTRARADICULAR INFECTIONS Microorganismscan overcome the defense barrier and establish an extraradicular infection. Eg- acute apical abscess. Some conditions entail the establishment of microorganisms in the periradicular tissues either by adherence to the apical external root surface in the form of extraradicular biofilm structures or by formation of cohesive actinomycotic colonies within the body of the inflammatory lesion. Extraradicular infection can develop as follows: It can be a result of direct advance of some bacterial species that overcome host defenses concentrated near or beyond the apical foramen, an extension of the intraradicular infectious process, or bacterial penetration into the lumen of pocket (bay) cysts, which are in direct communication with the apical foramen. It can result from bacterial persistence in the apical periodontitis lesion after remission of an acute apical abscess. It can be a sequel to apical extrusion of debris during root canal instrumentation (particularly after overinstrumentation). Cohen's 11th edition
- 40. The extraradicularinfection can be dependent on or independent of the intraradicular infection; Independent extraradicular infections would be those that are no longer fostered by the intraradicular infection and can persist even after successful eradication of the latter. The main bacterial species implicated in independent extraradicular infections are Actinomyces species. These bacteria form cohesive colonies that may be collectively resistant to phagocytosis Those extra radicular infections which are caused by intra radicular bacteria are treated easily by RCT. ▪Example; Sinus Tract If the extra radicular infection which are caused by outer side are treated only by Endodontic Surgery. ▪Example; Actinomycosis Cohen's 11th edition
- 41. CONCLUSION Microbes seeking toestablish in the root canal must leave the nutritionally rich and diverse environment of the oral cavity. Breach enamel, invade dentine. overwhelm the immune response of the pulp and settle in the remaining necrotic tissue within the root canal. During that time they have to compete in a limited space with other microbes for the available nutrition. The microbiota of root canal– treated teeth with apical periodontitis is more complex than previously anticipated by culture studies. The bacterial community profiles in treated cases vary from individual to individual, indicating that distinct bacterial combinations can play a role in treatment failure.
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