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255018337-cariology-4.ppt
- 1. Cariology IV Microbiology of dental caries Zdenek Broukal, Erika Lencova Institute of Dental Research
- 2. • Ontogenesis of oral microbial ecosystem • Cariogenic microorganisms - Streptococcus mutans - Heterofermentative lactobacilli - Other oral microorganisms • Dental caries as an infectious disease - Concept of the window of infectivity - Mechanisms of oral infection - Contributing factors
- 3. Ontogenesis of oral microbial ecosystem I. • Several hrs after delivery – Transient flora of the birth canal and St. epidermidis from skin surface • Up to 5-6 months – S. viridans, S. oralis, S. sanguinis, lactobacilli, diphteroids • Teeth eruption – 1. Infectious window period for S. mutans, G- cocci (Veilonella, Neisseria), anaerobic streptococci, gramm negative rods • Beginning of physiological dentition exchange – Infectious window period for S. mutans, G+ anaerobic rods (actinomycetes), G+ aerobic rods (Nocardia), G+ Rothia dentocariosa • Adolescence – G- anaerobes (Prevotella), Porphyromonas spp. , Capnocytophaga spp., anaerobic Vibrio spp., spirillae
- 4. Ontogenesis of oral microbial ecosystem II. • In edentulous oral cavity of a newborn/toddler microorganisms adhere partially to mucous membranes, partially form planktonic microflora in the saliva and in salivary biofilm covering mucous membranes • After the teeth eruption 90-95 % of microbial population forms plaque, the rest covers mucous membranes and form planktonic microflora
- 5. Ontogenesis of oral microbial ecosystem III. 0% 20% 40% 60% 80% 100% 0 1 2 3 4 5 7 9 11 13 15 1 2 G+ G+ cocci and rods G- cocci and rods filaments and fusobacteria spirili and vibrio spp. Toothbrushing No oral hygiene Toothbrushing days
- 6. Investigating association of microbial flora with dental caries • 1878 J. Tomes – microorganisms in dentin tubules • 1892 W.D. Miller – chemo-parasite theory of dental caries aetiology • 1922 J. Clark – streptococci in dentin tubules – S. mutans • 1938 G. Soggnaes – experimental dental caries in rodent model – Physiological breed of laboratory rodents – Cariogenic diet – Choosing suitable animal models for experimental dental caries – hamster, rat, (mice, gerbil, …) – Selecting cell lines (rat Sprague-Dawley) • 1963 R.W. Schaedler – gnotobiotic and germ-free animal models in research
- 7. Investigating association of microbial flora with dental caries - evidence • 1960 P.H. Keyes, R.J. Fitzgerald – Caries transmitted experimentally in hamster and rat models with cariogenic diet that were kept together via bedding, excrements etc. – germ-free animals with cariogenic diet had no dental caries – Mono-contamination of germ-free animals with individual oral microbial strains of rodent and human origin – 1964 – “cariogenic“ streptococci, actinomycetes • 1965 J.van Houte, J. Carlsson – “cariogenic“ streptococci = S. mutans • 1967 B. Mejàre and others – levels of S. mutans in saliva correlate with caries experience in humans • 1978 B. Köhler – S. mutans is transmitted from mother to child – window of infectivity
- 8. Taxonomy I. (Bergey´s Manual of Determinative Bacteriology 2nd Edition, 1992) Domain (Bacteria) Kingdom (Procaryotae /Monera/) Phylum (Firmicutes) Class (Bacilli) Order (Lactobacillales) Family (Lactobacillaceae) • Genus (Streptococcus) Species (S. mutans)
- 9. Oral lactobacilli Heterofermentative • main substrate: starch (glucose, maltose, dextrins) • slow metabolism • produce only 50% lactic acid and considerable amounts of ethanol, acetic acid and carbon dioxide • not so low pH – L. casei – L. fermentum – L. salivarius – L. plantarum – L. brevis – L. cellobiosus – L. buchneri Homofermentative • main substrate: glucose, sucrose • rapid metabolism • produce more than 85% lactic acid from glucose • low pH – L. acidophillus – L. acidophillus sensu stricto – L. crispatus – L. gasseri – L. rhamnosus
- 10. S. mutans – ecological requirements • Presence of solid surfaces in the oral cavity – hard dental tissues, dentures, infant obturator in cleft children • Repeated infection • Increased frequency of sucrose intake • Ecological niche – mode of existence that a species has within an ecosystem – smaller proportion of other viridans streptococci (showing alpha haemolysis on blood agar - S. viridans, S. oralis, S. sanguinis)
- 11. S. mutans – topography • Pits and fissures • Interdental spaces • Carious defects • SM is not necessarily present on all teeth • SM can be transmitted iatrogenically • Rare on mucous membranes • Proportion in salivary planktonic bacteria reflects the proportion in plaque – saliva as material for microbial examination • Treatment of carious defects does not decrease the proportion of SM in plaque
- 12. S. mutans – colonies morphology Colonies of S. mutans on the spoon contaminated with saliva and dipped for 36 hrs in BHI broth + 5% sucrose Colonies of S. mutans on MSA broth, incubated for 24 hrs, GasPak
- 13. Lactobacilli - ecological requirements • Presence of solid surfaces in the oral cavity – hard dental tissues, dentures, infant obturator in cleft children • Increased proportion of aciduric flora • Increased frequency of sucrose intake • Carious defects • Increased amount of plaque
- 14. Lactobacilli – topography • Carious defects • Proportion in salivary planktonic bacteria partially reflects the proportion in plaque – saliva as material for microbial examination • Treatment of carious defects does decrease the proportion of SM in plaque
- 15. Actinomycetes - ecological requirements • Presence of solid surfaces in the oral cavity – hard dental tissues, dentures, infant obturator in cleft children • Wide range of plaque flora • Increased frequency of sucrose intake • Carious defects • Increased amount of plaque
- 16. Actinomycetes – topography • Interdental spaces • Subgingival plaque • Carious defects • Proportion in salivary planktonic bacteria does not reflect the proportion in plaque • Treatment of carious defects does not decrease the proportion of A. in plaque • Abundant in the human mouth and induce root surface caries in hamsters and gnotobiotic animals
- 17. Virulence factors – cariogenicity I. • S. mutans – Energy utilisation - α,(1-2) glycosidic bonds in sucrose – Intensive glycolysis under wide range of pH – Production of EPS – intermicrobial matrix in plaque – Production of IPS – intracellular glycogen (ADPglucose pyrophosphorylase (GlgC) and glycogen synthase (GlgA) – Specific adherence to acquired pellicle – Tolerance towards high sucrose concentration - up to 40 % – Microaerofilic • Heterofermentative lactobacilli – Optimum pH for metabolism 3.5 – 5.5 – Enzymatic set for intensive glycolysis – Amylase
- 18. Virulence factors – cariogenicity II. • A. odontolyticus, A. naeslundii – Glycolysis under wide range of pH – Production of IPS – intracellular glycogen – Specific adherence to acquired pellicle – Wide range of proteases – Microaerophilic
- 19. Relationship of other microorganisms to dental caries • Mixed flora of dental plaque – Microaerophilic conditions for cariogenic microorganisms – Aciduricity (tolerance of, and growth at low pH) – Co-aggregation – increased cohesion of plaque – proteases – destruction of protein matrix of enamel/dentin – amylase – hydrolysis of starch to oligosaccharides
- 20. Koch’s postulates • Suspected pathogenic microorganism should be present in all cases of the disease and absent from healthy animals • The microorganism should be grown in pure culture • Cells from pure culture should cause disease in a healthy animal • The microorganism should be re-isolated and shown to be the same as the original
- 21. Metabolic activity of cariogenic microorganisms – pH of the environment hetero- fermenta- tive lactobacilli S. mutans 0 20 40 60 80 100 7 6 5 4 3 2 pH %
- 22. Production of polysaccharides by SM – Extracellular polysaccharides (α 1-3, 1-4, 1-6 glucane) – adhesion factor • glucosyltranspherase (GTF) – Intracellular polysaccharides (intracellular glycogen-like) – energy source • ADPglucose pyrophosphorylase (GlgC • glycogen synthase (GlgA)
- 23. Extracellular polysaccharides Microorganism Extracellular polysaccharides S. mutans α1-4, α1-3, glucane α1-6, glucane mutane dextrane lactobacilli β2-6, fructane levane actinomycetes β2-6, fructane α1-6, glucane levane dextrane
- 25. Chemo-parasite theory W.D.Miller 1892 Dental caries occurs as a result of hard dental tissues demineralization with organic acids – end products of sucrose metabolism of oral microorganisms.
- 26. Ecologic factors in dental caries S. Socransky 1992 S. mutans S. sanguis Selective ecological pressure (nutrition) lactobacilli actinomycetes Shortage of sucrose neutral pH Enough of sucrose acidic pH Healthy tooth caries
- 27. Microbial agent Defending mechanisms Genotype hypothesis of SM cariogenicity M. W. Russell 1994 Defending mechanisms Microbial agent Defending mechanisms specific pathogen or virulent genotype
- 28. Hypotheses integration Non-cariogenic spectrum of plaque Selective ecological pressure aciduric flora (S. mutans) Selective physical- chemical pressure acidophilic flora (lactobacilli) Selective genetic pressure Cariogenic spectrum of plaque virulent SM and lactobacilli genotypes
- 29. Window of infectivity for cariogenic microorganisms • Conditions suitable for transmission of microorganisms to planktonic flora • Conditions suitable for adhesion of microorganisms to dental surface/obturator/dentures • Conditions suitable for repeated infection • Contributing factors – Frequent presence of sucrose in the oral cavity – Decreased salivary secretion (during sleep) – Quantity of infection
- 30. 0 6 12 18 24 30 36 5 6 7 age months years Beginning of the eruption of permanent teeth Window of infectivity Beginning of the eruption of temporary teeth Succedaneous window of infectivity? Window of infectivity C.W. Caulfield 1993 Repeated infection Repeated infection
- 31. Mechanism of oral infection Infection source Transmission vector Infection entry window of infectivity Contributing factors Persistent infection Repeated transmission Transmission way
- 32. Contributing factors of oral infection • Risk behaviour of mother (siblings, father, grandparents, carers) • Untreated carious dentition of mother/others • High levels of cariogenic microorganisms in saliva • High frequency of sucrose intake especially before sleeping (decreased salivary flow) • Later start of tooth brushing • Administration of sweetened medications • Prolonged nursing/nursing as tranquilizing practice
- 33. Evidence of SM transpher from mother to child Li and Caufield - 1995: 34 pairs mother - child; 70.6 % children had SM genotype identical with mother De Soet et al. - 1998, 21 pairs mother – child (with lip or palate cleft); 38 % had SM genotype identical with mother Kozai et al. - 1999: 20 families, 51 % had SM genotype identical with mother, 31 % with father, 19 % other type of SM genotype Dušková a Broukal – 1997: 40 families, 60 % had SM genotype identical with mother, 8 % with father, 32 % other type of SM genotype
- 34. Proportions of SM and lactobacillli in fissure plaque Caries affected % z TVC (total viable count) Tooth Caries experience S. mutans S. sanguinis lactobacilli With caries low 2.0 5 1 With caries high 25 8 4 Caries free intact dentition < 1 13 0 Caries free low 2.0 21 < 1 Caries free high 8.0 6 < 1
- 35. Hereditary fructose intolerance • autosomal recessive disorder of fructose metabolism first described in 1956 • incidence 1:22 000 of live births • deficiency of fructose-1-phosphate aldolase (EC 2.1.2.13) activity • accumulated fructose-1-phosphate inhibits glycogen breakdown and glucose synthesis, thereby causing severe hypoglycaemia following ingestion of fructose • patients develop a strong distaste for sweet food • almost all affected persons have intact dentition and do not have S. mutans present in the mouth