5 oral flora


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5 oral flora

  2. 2. Introduction: In a healthy animal, the internal tissues, e.g. blood, brain, muscle, etc., are normally free ofmicroorganisms. surface tissues oral mucous membrane skin 2
  3. 3.  The mixture of organisms regularly found at anyanatomical site is referred to as the normal flora. Researchers - "indigenous microbiota". 3
  4. 4. Normal flora:• Normal flora refers to the populations of microorganismsthat inhabit the skin and the mucous membranes of normalhuman body. Resident flora Indigenous flora Supplemental flora Transient flora 4
  5. 5. Oral flora Oral flora comprises a diverse array of organisms andincludes eubacteria, fungi, protozoa and possibly a viralflora which persist from time to time. Bacteria – 350 cultivable species Oral bacteria classification a. Gram-positive b. Gram-negative 5
  6. 6.  Depending upon the effect of oxygen divided as: Obligate aerobe Micro aerophilic / Micro aerophilic strict anaerobe Facultative anearobes Strict obligate anaerobe Capnophilic 6
  7. 7. Bacteria commonly found on the surfaces of the human bodyBacterium Skin Nose Pharnyx MouthStaphylococcu ++ ++ ++ ++s epidermidisStaphylococcu + + + +s aureusStreptococcus ++ ++salivariusStreptococcus + ++mutansEnterococcus +/- +faecalisStreptococcus +/- + +pneumoniaeStreptococcus +/- + +pyogenes 7
  8. 8. Bacterium Skin Pharnyx MouthNeisseria sp. ++ +Neisseria ++meningitidis +Enterobacteriaceae +/- + ++(Escherichia coli)Pseudomonas +aeruginosaHaemophilus +/- +influenzae 8
  9. 9.  Association between Humans and the Normal flora Not much known dynamic interactions mutualistic Some normal flora parasitic pathogenic 9
  10. 10. Tissue specificity:Most members of the normal bacterial flora prefer tocolonize certain tissues and not others.This “tissue specificity” is usually due to properties of boththe host and the bacterium. 1. Tissue tropism essential nutrients growth factors, suitable oxygen, pH 10
  11. 11. Lactobacillus acidophilus, informally known as "Doderleins bacillus"colonizes the vagina because glycogen is produced which provides thebacteria with a source of sugar that they ferment to lactic acid 11
  12. 12. 2. Specific adherence 12
  13. 13. Bacterium Bacterial adhesion Attachment siteStreptococcus Cell-bound protein (M- Pharyngeal epitheliumpyogenes protein)Streptococcus Cell- bound protein Pellicle of toothmutans (Glycosyl transferase)Streptococcus Buccal epithelium of Lipoteichoic acidsalivarius tongueStreptococcus Cell-bound protein (choline- Mucosal epitheliumpneumoniae binding protein)Staphylococcus Cell-bound protein Mucosal epitheliumaureusNeisseria N-methylphenyl- alanine Urethral/cervicalgonorrhoeae pili epitheliumEnterotoxigenic Type-1 fimbriae Intestinal epitheliumE. coli 13
  14. 14. 3. Biofilm formation:Some of the indigenous bacteria are able to constructbiofilms on a tissue surface, or they are able to colonize abiofilm built by another bacterial species. Many biofilms are a mixture of microbes, although onemember is responsible for maintaining the biofilm and maypredominate. 14
  15. 15. Cartoon depicting biofilm formation. 15
  16. 16.  The classic biofilm - oral cavity dental plaque on the teeth. Plaque is a naturally-constructed biofilm, thickness of 300-500 cells - teeth. 16
  17. 17.  These accumulations subject the teeth and gingivaltissues to high concentrations of bacterial metabolites,which result in dental diseases like caries. 17
  18. 18. The Composition of Normal floraThe makeup of the normal flora influenced genetics age sex stress nutrition and diet of the individual. 18
  19. 19.  Three developmental changes in humans ,weaning,the eruption of the teeth, and the onset and cessation ofthe ovarian functions, invariably affect the compositionof the normal flora in the oral cavity. The Bacterial Flora of Humans © 2007 Kenneth Todar University of Wisconsin-Madison Department of Bacteriology 19
  20. 20. Table . Predominant bacteria at various anatomical locations in adults.Anatomical Location Predominant bacteriaSkin staphylococci and corynebacteriaConjunctiva sparse, Gram-positive cocci and Gram-negative rodsOral cavity Teeth Streptococci, lactobacilli mucous membranes Streptococci and lactic acid bacteriaUpper respiratory tract nares (nasal membranes) staphylococci and corynebacteria pharynx (throat) streptococci, neisseria, Gram-negative rods and cocci 20
  21. 21. Normal Flora of the oral Cavity: Various streptococci in a biofilm in the oral cavity. 21
  22. 22. Flora of the oral cavity: GRAM POSITIVE COCCI:Genus streptococcus: Streptococcus mutans. Gram stain 22
  23. 23. The mutans group:Main species Strep mutans, strep sorbinus, strep cricetusCultural characteristics Mitis Salivarius Agar (MSA)Intra oral sites Teeth andinfections Dental caries 23
  24. 24. The salivarius group:Main species Strep salivarius, Strep vestibularisCultural characteristics Mitis Salivarius Agar (MSA)Intra oral sites dorsum of the tongue saliva. andinfections does not cause major oral pathogenesis 24
  25. 25. The anginosus group:Main species Strep constellatus, Strep intermedius, Strep anginosusCultural characteristics Mitis Salivarius Agar (MSA).Intra oral sites Gingival crevice and Dentoalveolar and endodonticinfections infections. 25
  26. 26. The mitis groupMain species Strep mitis; Strep sanguis Strep gordonii Strep oralisCultural characteristics Mitis Salivarius Agar (MSA).Intra oral sites dental plaque biofilms , tongue and cheek. andinfections dental caries. 26
  27. 27. Anaerobic streptococciMain species Micromonas micros Finegoldia magnusCultural characteristicsIntra oral sites teeth, especially the carious dentine. andinfections periodontal and dentoalveolar abscesses 27
  28. 28. Genus stomatococcusMain species StomatococcusCultural characteristics facultative anaerobesIntra oral sites tongue and the gingival crevice. andinfections major opportunistic pathogen 28
  29. 29. Genus stomatococcus 29
  30. 30. Genus StaphylococcusMain species Staph aureus, staph epidermidis, Staph saprophyticusCultural characteristics blood agarIntra oral sites Present on the buccal mucosa andinfections Angular chelitis 30
  31. 31. Genus Staphylococcus 31
  32. 32. GRAM POSITIVE RODS AND FILAMENTS: These organisms are very commonly isolatedfrom the biofilms of dental plaque.This group consists of the following organisms – Actinomycetes Lactobacilli Eubacteria Propionibacteria. 32
  33. 33. Genus actinomycetesMain species Actino israeli Actino gerensceriae Actino odontolyticus Actino naeslundiiCultural characteristics facultative anaerobes.Intra oral sites andinfections 33
  34. 34. Actinomycetes 34
  35. 35. Main intraoral sites and infections: They are seen on the gingiva, on the mucosal and alsoon the teeth surfaces. Actinomyces odontolyticus is related to the earlieststages of enamel demineralization and the progression ofsmall caries lesions. Actinomyces naeslundii has been related to rootsurface caries and gingivitis. 35
  36. 36. Actinomyces israeli is an opportunistic pathogencausing cervicofacial and ileocecal actinomycosis.Actinomyces gerensceriae and Actinomyces georgiaeare considered to be the minor components of healthygingival flora. 36
  37. 37. Genus Lactobacillus -Main species Lacto casei Lacto fermentum Lacto acidophilus Lacto salivarius Lacto rhamnosus.Cultural characteristics Rogosa agar.Intra oral sites dental plaque biofilm, advancing front of dental caries andinfections 37
  38. 38. Lactobacillus 38
  39. 39. Genus EubacteriumMain species Eubact brachy Eubact nodatum Eubact saphenum Eubact yurii.Cultural characteristics Obligate anaerobesIntra oral sites Dental plaque biofilm, calculus and Periodontal diseaseinfections Eubacterium yurii is involved in the “corn-cob” formation in the dental plaque 39
  40. 40. Rod shaped eubacterium 40
  41. 41. Genus propionibacteriumMain species Propionibacterium acnesCultural characteristics Strict anaerobesIntra oral sites gingival pockets and plaque biofilms andinfections root surface caries 41
  42. 42. OTHER NOTABLE GRAM-POSITIVE ORGANISMS:• Rothia dentocariosa - Gram-positive branching filament -strict aerobeFound in plaque & isolated from patients with infectiveendocarditis.• Bifidobacterium dentium - Gram-positive, strict anaerobe,Regularly isolated from the biofilms of plaque, and its role indisease is unclear. 42
  44. 44. Genus NisseriaMain species Neisseria subflava Neisseria mucosa Neisseria siccaCultural characteristics facultative anaerobesIntra oral sites tongue, saliva, oral mucosa and early plaque. andinfections rarely associated with any oral disease. 44
  45. 45. Neisseria 45
  46. 46. Genus VeillonellaMain species Veillonella parvula Veillonella dispar Veillonella atypicalCultural characteristics Rogosa vancomycin agarIntra oral sites tongue, saliva, plaque biofilm. and not associated with any oral disease.infections 46
  48. 48. Genus Haemophilus Gram-negative coccobacilliMain species Haemophilusparainfluenzae Haemophilus segnis Haemophilus aphrophilus Haemophilus haemolyticusCultural characteristics Facultative anaerobesIntra oral sites tongue, saliva, plaque biofilm. and dentoalveolar infections, acuteinfections sialadenitis, and infective endocarditis 48
  49. 49. Genus ActinobacillusMain species Actinobacillus actinomycetemcomitansCultural characteristicsIntra oral sites andinfections 49
  50. 50. Culture characteristics:The freshly isolated strains contain fimbriae that are loston subculture.Actinobacillus produces many virulence factors like –leukotoxin; epitheliotoxin; collagenase; protease thatcleaves immunoglobulin G (IgG). 50
  51. 51. microscopic picture of Actinobacillus 51
  52. 52. Main intraoral sites and infections:Mostly seen in the periodontal pockets. They are implicated in aggressive forms of periodontaldisease (e.g. localized and generalized forms of aggressiveperiodontitis) They are often isolated as co-pathogens from thecervicofacial Actinomyces infections. 52
  53. 53. Genus Eikenella Gram negative coccobacilliMain species Eikenella corrodensCultural characteristics blood agarIntra oral sites plaque biofilms andinfections chronic periodontitis dentoalveolar abscesses 53
  54. 54. Genus Capnocytophaga Gram-negative fusiform rodsMain species Capnocytophaga gingivalis C. Sputigena C. Ochracea C. Granulose C. HaemolyticaCultural characteristics Capnophilic organismsIntra oral sites Plaque, mucosal surfaces, saliva. andinfections Periodontal disease 54
  55. 55. GRAM-NEGATIVE RODS – OBLIGATE ANAEROBICGENERA:Form large portion of the plaque biofilms 55
  56. 56. Genus PorphyromonasMain species Porphyromonas gingivalis P. Endodontalis P. CatoniaeCultural characteristics Strict anaerobesIntra oral sites Gingival crevice and the subgingival plaque andinfections chronic periodontitis and dentoalveolar abscess. 56
  57. 57. Porphyromonas 57
  58. 58. Genus Fusobacterium Gram-negative rodsMain species Fusobacterium nucleatum F. Alocis F. Sulci F. periodonticum.Cultural characteristics Strict anaerobesIntra oral sites Normal gingival crevice, tonsils andinfections acute ulcerative gingivitis, halitosis 58
  59. 59. SEM picture of Fusobacterium 59
  60. 60. Genus TreponemaMain species Treponema denticola Treponema macrodentium Treponema skoliodontium Treponema sokranskii Treponema maltophilumCultural characteristics Strict anaerobesIntra oral sites Gingival crevice and acute ulcerative gingivitis,infections destructive periodontal disease 60
  61. 61. ORAL PROTOZOA: Genus EntamoebaMain species Entamoeba gingivalisCultural characteristics Strict anaerobesIntra oral sites and Periodontal diseaseinfections 61
  62. 62. Genus TrichomonasMain species Trichomonas tenaxCultural characteristics Strict anaerobesIntra oral sites Gingival crevice andinfections Unclear 62
  63. 63. Beneficial effects of the normal flora:1.Can synthesize and excrete vitamins2.Prevent colonization by pathogens3.May antagonize other flora4.Stimulate the production of natural antibodies 63
  65. 65. MYCOPLASMASMain species Mycoplasma pneumoniae Mycoplasma buccale Mycoplasma oral Mycoplasma salivarium; Mycoplasma fauciumCultural characteristics Special agar and broth mediaIntra oral sites oro-pharyngeal region, saliva, dental plaque andinfections Mucocutaneous lesions 65
  66. 66. FUNGI YeastsMain species Cryptococcus RhodotorulaIntra oral sites Mucosa andinfections Mucosal ulcers in immunocompramised patients 66
  67. 67. Yeast-like fungigenus CandidaMain species Candida albicans Candida tropicalis Candida krusei Candida glabrata Candida guillermondii Candida parapsilosis Candida kefyrIntra oral sites Mucosa andinfections Mucosal ulcers in immunocompramised patients 67
  68. 68. Virulence:The degree of pathogenicity of a microorganism as indicatedby the severity of disease produced and the ability to invadethe tissue of the host ; by extension, the competence of anyinfectious agent to produce pathologic effects. ( Dorland’s Medical Dictionary) 68
  69. 69. The main etiological agent of periodontal disease is microflora– dental plaqueDental plaque:It is defined as a structured, resilient, yellow-grayish substancethat adheres to the intraoral hard surfaces, including removableand fixed restorations. 69
  70. 70. Composition Bacteria (matrix of salivary glycoproteins & extracellular poysaccharides)1 gm = 10 11 bacteria> 500 species30% uncultivable species 70
  71. 71. Dental plaque Supra gingival plaque: Demonstrates a multi-layered accumulation ofbacterial morphotypes gram positive cocci + rods - tooth surface gram negative rods, filaments - outer surface Sub-gingival bacteria anerobic bacteria 71
  72. 72. Plaque formation of at the ultra-structural levelSaliva derived layer – acquired pellicle - glycoproteins, proline-rich proteinsStreptococcus & Actinomycetes bind to specificsalivary proteinsVeillonellae, Capnocytophagae, Prevotella – bind toStreptococci, Actinomycetes Primary colonizers Secondary colonizers 72
  73. 73. Microorganisms associated with periodontal disease:Bacteria associated – gram +ve Streptococcus( s.sanguis, s.mitis) Actinomycetes (A.viscosus, A.naeslendi) gram –ve capnocytophage neisseria 73
  74. 74. The currently recognized key Gram negativeperiodontopathogens include: Porphyromonas gingivalis (P.g), Prevotella intermedia (P.i), Bacteroides forsythus (B.f), Aggregatibacter actinomycetemcomitans (A.a), Fusobacteriumnucleatum (F.n), Capnocytophaga species(C.sp), Campylobacter rectus (C.r) 74
  75. 75. Porphyromonas gingivalispreviously known as Bacteroidesstrictly anaerobic, Gram negativeVirulence mechanism: carbohydrate capsule – prevents opsonizationVirulence factors: fimbriae - adhesion proteases – degrade collagen fibers, cytotoxins hemolysin 75
  76. 76. Actinobacillus actinomycetemcomitansGram –veVirulence factors: leukotoxin forms pores – death – osmotic pressure collagenase destruction of connective tissue lipopolysaccharides macrophages – IL 1, IL2 – bone resorptionMicrobial etiology of periodontal disease – a mini reviewMedicine and Biology Vol.15, No 1, 2008 - Ljiljana Kesic 76
  77. 77. Fusobacterium nucleatumVirulence factors: induce – apoptotic cell death – PMN release – cytokinesAs fusobacterium co-aggregates with most oralmicroorganisms – binding organismsMicrobial etiology of periodontal disease – a mini reviewMedicine and Biology Vol.15, No 1, 2008 - Ljiljana Kesic 77
  78. 78. Capnocytophaga spGram –veVrulence: lipopolysaccharides – activity on alveolar bone proteolytic enzymes 78
  79. 79. Spirochetes:Motile, flagella Treponema denticola Treponema vincentii• a lipopolysaccharide, and unusual metabolic endproducts,like indole, hydrogen sulphide, ammonia, which arepotentially toxic to host cells.• T.d – proteolytic enzyme – destroy (IgA, IgM, Ig G) 79
  80. 80. Dental caries Multifactorial 80
  81. 81. Virulence factors: Specific adherence to tooth surface using antigenI/II adhesin and GTF (glycosyl transferase ) Production of extracellular polysaccharides(dextran) allows the cariogenic bacteria to stick ontothe teeth and form a biofilm. acid-tolerance (aciduricity) Able to maintain microbial growth and continueacid production at low pH values. 81
  82. 82.  Rapid metabolism of sugars to lactic and otherorganic acids Lower the pH to below 5.5, the critical pH. Drivesthe dissolution of calcium phosphate (hydroxyapatite)of the tooth enamel (acidogenicity) Accumulation of intracellular polysaccharides(carbon/energy reserve) 82
  83. 83. Virulence propertiesstreptococus mutansAdhesion sucrose – independent adhesion sucrose – dependent adhesionAcidogenicityAcid-tolerance 83
  84. 84. AdhesionSucrose – independent adhesion• influenced by antigen I/II• proteins of this family share structural similarity• role of antigen I/II adhesion of s. mutans to saliva – coatedhydroxyapetiteOhat et al : characterization of a cell – surface proteinantigen of hydrophilic streptococcus mutans strain.J Gen Microbiology 135, 981-988 84
  85. 85. Gram stain - streptococcus 85
  86. 86. Interaction between antigen I/II & salivary components alanine- rich & proline-rich domainsProved - mutant lacking P1 ( antigen I/II) - did not bindBowen et al : Role of a cell surface-associated protein inadherence and dental caries . Infect immunology 59,4606-4609 86
  87. 87. Sucrose-Dependent Adhesion• The action of glucosyltransferases (GTFs) in the synthesisof glucans is the major mechanism behind sucrose -dependent adhesion.• GTFs – sucrase activity• sucrose glucose + fructose added growing polymer of glucan 87
  88. 88. s.mutans - 3 GTFs - encoded by gftb, gftc, gftdSo , GTFs – synthesizeWater- soluble glucan water- insoluble glucan(dextran) (mutan) 88
  89. 89. • The ability of glucan to facilitate adhesion of S.mutansmay be due to hydrogen bonding of the glucan polymers toboth the salivary pellicle and the bacteria.• This glucan could be synthesized by extracellular GTFsthat bound the salivary pellicle, S. mutans that hadpreviously adhered via sucrose-independent means, orperhaps by other oral streptococci. 89
  90. 90. • It is not known why S. mutans requires multiple GTFs,but there is evidence that the different GTFs havediffering affinities for the bacterial surface or salivarypellicle , and that a particular ratio of each is necessaryfor optimal sucrose-dependent adhesion.Ooshima et al: contbutions of three glucosyltransferasesto sucrose-dependent adherence of streptococcusmutans. J dental res :80 , 1672-1677 90
  91. 91. • Another possibility - WapA ( antigen A) its yet uncertain• Carbohydrate Metabolism: additional putative factors Gene product Hypothesized function Ftf Catalize the synthesize of fructans FruA Breakdown fructans for energy DexA Glucan synthesis 91
  92. 92. Acidogenicity: S.mutans lactate, formate, acetate, ethanol ( fermentation end-products)Glucose 92
  93. 93. •The velocity with which S. mutans produces acid whentested at a pH in the range from 7.0 to 5.0 exceeds that ofother oral streptococci in most instances.Soet, J.J., B. Nyvad, & M. Kilian: Strain-relatedacid production by oral streptococci. Caries Res 34, 486-490, (2000)• The acidogenicity of s.mutans---- ecological changes in the plaque flora that includes an increase inproportion of s.mutans 93
  94. 94. Acid – tolerance• Growth-inhibitory pH• distinguish s.mutans• this is largely mediated by - F1F0 – ATPase ATR - gene and protein expression• acid – tolerance may be aided by the synthesis ofwater-insoluble glucan and the formation of biofilm 94
  95. 95. • s.mutans with in biofilm – outside ATR , physical characteristics of the biofilm 95
  96. 96. The role of lactobaccillli: Absent – incipient lesions Pioneer organisms in the advancing front of cariousprocessVirulence : synthesize sucrose their ability to grow low- pH  lactic acidThe exact role – not well defined 96
  97. 97. The results of this study suggest that Lactobacilli colonizessites in the oral cavity (including the tongue and saliva)other than the tooth surface in rats.The effect of Lactobacilli in plaque toward reduction of S.mutans-induced dental caries in ratsOral Ecology and Virulence of Lactobacillus casei and Streptococcusmutans in Gnotobiotic Rats : SUZANNE M. MICHALEK el alDepartment ofMicrobiology and Institute of Dental Research, TheUniversity ofAlabama in Birmingham, Birmingham, Alabama 3529 97
  98. 98. Candidiasis:The physiological state host – etiologyVirulence factors: hyphal formation, surface recognition molecules, extracellular hydrolytic enzyme production 98
  99. 99. Hydrolytic enzymes: production – proteinases CATALIZE hydrolysis of peptide bonds in proteins secreated aspartyl proteinase (SAP genes) 99
  100. 100. Correlation between Sap Production In Vitro and CandidaVirulenceMain focus points.(i) The virulence of C. albicans species appears to correlatewith the level of Sap activity in vitro and may correlate withthe number of SAP genes.(ii) Infected patients (oral or vaginal) harbor C. albicansstrains that are significantly more proteolytic than areisolates from asymptomatic carriers.. 100
  101. 101. (iii) HIV infection appears to lead to the selection of C.albicans strains with heightened virulence attributes suchas proteinase production 101
  102. 102. Association of Sap Production with Other VirulenceProcesses of C. albicansMain focus points. (i) Sap proteins facilitate C. albicans adherence tomany host tissues and cell types.(ii) Hypha formation and SAP4 to SAP6 expression arecoordinately regulated, but the signaling pathwaysremain to be elucidated. 102
  103. 103. (iii) SAP1 appears to be regulated by phenotypicswitching, but the contribution of switching to C. albicansvirulence in vivo is not yet clear. 103
  104. 104. How do Sap proteins contribute to adherence? Not clear Two hypothesis i. C. albicans proteinases could act as ligands tosurface moieties on host cells, which does not necessarilyrequire activity of the enzymes. ii. C. albicans utilizes Sap proteins as activeenzymes to modify target proteins or ligands on the fungalsurface or on host cells (i.e., epithelial cells), which mayalter surface hydrophobicity or lead to conformationalchanges, thus allowing better adhesion of the fungus 104
  105. 105. Candida albicans Secreted Aspartyl Proteinases inVirulence and PathogenesisJulian R. Naglik et al; Microbiology and Molecularbiology 105
  106. 106. THANK YOU 106