Microbiology of dental caries Flashcards
What things in the oral environment are relevant to caries?
-Tooth surface
- around 700 species of bacteria
- Saliva
- Gingival crevicular fluid
- Periodic availability of nutrients from host diet
what factors are required for the development of dental caries?
- teeth
-time
-bacteria in plaque
-Fermentable carbohydrate in diet
Are some bacteria more important than others in caries
- Clarke (1924) grew Streptococcus mutans from advanced carious lesions
- 1960s – organisms responsible for “infectious” dental caries in rodent models identified as member of mutans streptococci (MS) group and lactobacilli
- MS in humans – S. mutans and S. sobrinus. Other mammals have their own version, e.g. S. rattus in rats; S. cricetus in hamsters
Evidence implicating
Streptococcus mutans in caries
Streptococcus mutans found in high numbers in carious lesions
Subjects with active caries have higher salivary counts of S. mutans (>106/ml) than subjects without active caries (<103/ml)
What features of S. mutans makes it easier for it to survive?
It’s acidogenic & aciduric- produces high levels of acid & can survive & flourish in low pH conditions
What virulence factors are present in S. mutans that may enable it to contribute to the caries process?
-Produces Antigen I/II so adherence to surfaces
-Glucosyl transferases- produce glucan polymers from sucrose
-Glucan binding proteins- attachment of cells to glucans
Have any vaccines been developed for these virulence factors of s.mutans?
Vaccines have been developed (active and passive) against all three factors but no commercial vaccine yet available
is lactobacillus associated with caries?
Earliest studies associated lactobacilli with caries
Very low levels in individuals without caries
Many different species found in caries and thought to be of equal importance.
What kind of organism is lactobacillus?
It appears to be an opportunist organism that requires a low pH habitat so lactobacilli don’t initiate caries but will colonise existing lesions.
Doesn’t only appear in the mouth but also in the body and fermented food e.g yogurt
slide 8
what kind of bacteria is veillonella
Gram negative anaerobic cocci that is associated with caries.
Describe the importance of Veillonella in caries.
-Requires lactate as substrate for growth
-Lactate is produced by both streptococci and lactobacilli
-May be present in carious lesions because high levels of lactate are present.
-It has been suggested that veillonella may be beneficial to caries by reducing lactate levels but little evidence to support this.
What was the consensus reached about the microbiology of caries from recent culture & DNA-based studies?
-There’s a complex bacterial community in carious lesions
-S. mutans is frequently not present
-Some samples from caries-free sites have high levels of S.
mutans
Other acidogenic species are significantly associated with carious lesions, including Scardovia wiggsiae, lactobacilli etc.
What laboratory experiment did Bradshaw & Marsh carry out which forms the basis of much of our current thinking about the role of bacteria in caries?
-9 oral bacteria species growing together
-Glucose pulsed into them for 10 days.
-Fall in pH seen
-Increase in s.mutans and lactobacillus
-If pH was held at 7, no change in microbiota
Therefore, change in microbiota was in response to change in environment and not its cause
What is the extended ecological plaque hypothesis (Takahashi & Nyved 2008)?
Dynamic stability stage: associated with health; only mild or infrequent acidification
Acidogenic stage: arises if there is frequent/moderate acidification
Aciduric stage: so much carbohydrate is being consumed so regularly that the pH of the whole mouth falls.
look at slide 14 and 15
What was the background information that Takahashi & Nyved used to form their plaque hypothesis?
-S. mutans and lactobacilli are highly aciduric
-Mutans streptococci only a minor component of healthy plaque
look at slide 14 and 15
Describe the dynamic stability stage.
1.Acid produced from sugars by range of bacteria lowers pH of plaque
2.Followed by alkalinisation phase, caused by:
-diffusion of acid
-buffering by plaque constituents and saliva
-production of alkali by bacteria
- Production of alkali:
-ureolysis – ammonia production from urea by urease and reduction of nitrate
-arginine deiminase – dietary arginine catabolised to ammonia
Describe the acidogenic stage.
1.Initiated by:
-Repeated, raised levels of sugar intake
-Reduced salivary flow
-Poor oral hygiene
- Microbiota typically dominated by non-mutans streptococci and Actinomyces
Describe the aciduric stage.
- Occurs after prolonged acidogenic stage:
- the buffering capacity of saliva is lost and bacterial community changes with:
- Selection of aciduric bacteria, particularly S. mutans and lactobacilli and some Bifidobacterium and Propionibacterium species - Thus, the ecological change drives the change in the composition of the microbiota and not vice versa, but the presence of aciduric bacteria compounds the disease state
What is a significant problem since more older people are retaining their teeth?
Caries affecting the exposed roots
Oral hygiene is key because there’s a strong correlation with the amount of plaque on the root surface and root caries
What did microbial analysis find in relation to root caries lesions?
S. mutans and lactobacilli found, together with Actinomyces species & bifidobacteria.
Components of saliva important in plaque and caries
Amylase - 38 (mg/ml)
Lysozyme - 22
IgA - 19
Phosphate - 17
Calcium - 6
What does alpha amylase do?
Hydrolyses starch to glucose & maltose, thus making it available for fermentation
It also binds with high affinity to Streptococci and retains activity when bound,
so it’s involved in bacterial adherence to the pellicle
In what ways can sugar become available to oral bacteria for fermentation?
Can be present in diet & directly obtained from that route
Can be broken down from starch by amylase
Can be derived from salivary glycoproteins by bacteria themselves
How are the sugars then transported to the interior of the bacterial cell?
PTS (phosphotransferase system) & other systems
What happens once the sugars have entered the bacterial cell?
They enter glycolysis at different stages of the pathway, but all sugars eventually give rise to pyruvate which is a key metabolic intermediate
Where does the ATP for the PTS system in the bacterial cell come from?
The conversion of phosphoenolpyruvate to pyruvate.
look at slide 24 and 25
What are the major pathways of glucose fermentation?
pyruvate can be converted into lactate or acetate.
What does the bacteria in plaque do to sucrose?
Metabolises it to synthesise extracellular polysaccharides (EPS) which constitute the plaque matrix
Some bacteria e.g. S. mutans also form intracellular polysaccharides
What are EPS (extracellular polysaccharides) and what do they contribute to?
“Sticky” polymers & they contribute to the adherence of bacteria to the pellicle and plaque
They also make bacteria more resistant to being washed away by dietary food & drink & salivary flow
It can also be used as a source of sugars for metabolism & growth when nutrients are limited
What are the enzymes that form EPS called?
Glucosyl transferases
How many glucosyl transferases does S. mutans produce?
3: GTFB, GTFC & GTFD (B,C,D form)
GTFB & GTFC (B and C) make water-insoluble glucans, rich in alpha-1,3-glucosidic linkages
GTFD makes water-soluble glucans rich in alpha-1,6-glucosidic linkages
Why are insoluble glucans more problematic?
Because they, in particular, give plaque bulk & form a sticky mass which aids adhesion
What do glucan binding proteins (GBPs) mediate
The specific attachment of bacterial cells to glucan
Describe the process of glucan & fructan synthesis
Glucan synthesis:
Sucrose + (G)n primer –>
Glucan (G)n+1 + fructose
Enzyme - glucosyl transferase (GTF; “dextran sucrase”)
Fructan synthesis:
Sucrose + (F)n primer –>
Fructan (F)n+1 + glucose
Enzyme - fructosyl transferase (FTF; “levan sucrase”)
Describe the process of glucan & fructan hydrolysis.
Glucan hydrolysis:
Glucan (G)n + water –> Glucan (G)n-1 + glucose
Enzyme - glucanase (“dextranase”)
Fructan hydrolysis:
Fructan (F)n + water –> Fructan (F)n-1 + fructose
Enzyme – fructanase (“levanase”)
What is enolase?
A key regulatory enzyme of the glycolytic pathway
mediating the conversion of 2-phosphoglycerate to phosphoenolpyruvate
slide 32
What inhibits the action of enolase?
Fluoride
What are the effects of fluoride inhibition of enolase?
Reduced sugar transport because PEP is a key component of PTS system
Reduced acid production because pyruvate production is reduced
What is xylitol?
Sweetening agent
Describe the study carried out with xylitol and the findings.
In 2-year study (Turku), when sucrose was replaced in diet by xylitol, the mean increment of decayed, missing and filled teeth was 0.0, compared to 7.2 in sucrose control group
Partly due to lack of sucrose but also few oral bacteria produce acid from xylitol
Xylitol inhibits growth of S. mutans in vitro
What is the primary mode of action of xylitol thought to be?
Passive; in that if the diet is thought to be rich in xylitol, it’s likely to be lower in sugar
What direct effect does xylitol have on S. mutans?
It’s transported into S. mutans cells by fructose-PTS system but enters “futile cycle” of phosphorylation, dephosphorylation & export from cell
The futile cycle uses energy but produces none for the cell
Is xylitol of high cariogenicity?
No, it’s of low cariogenicity & inhibits glycosyl transferase in cariogenic bacteria
how could Xylitiol be delivered to caries?
Xylitol frequently delivered in chewing gum
Reduced caries for xylitol gum compared to sugared gum
can Probiotics be used for oral diseases?
-Number of probiotic preparations now available for oral diseases
-Aim is to inhibit disease-associated oral bacteria
-Mainly re-badged “gut” probiotics
-Potential concern that many are lactobacilli – caries associated
-Evidence for efficacy against caries is lacking
What is the aim of replacement therapy for caries?
Aim is to replace S. mutans with other strains / species
Attempted with other Streptococcus species that are less acidogenic and aciduric but were found to compete poorly with S. mutans
Alternative is to modify S. mutans strains so that they produce less acid. Strategy then is to use antimicrobials to reduce plaque levels and then re- seed with avirulent strains. Large scale clinical trials awaited
what are the Microbiological tests for caries prediction?
-Commercial test kits are available
-Test for numbers of S. mutans, and some also for lactobacilli
-Effective for populations, less so for individuals
-Caries is multifactorial disease – individual bacteria are not primary cause
-Not widely used except for research
