MFD 18 (theme 2.1) Flashcards
• To provide an overview of the oral microbiota and the impact of saliva on colonization of the mouth by bacteria. Content • Introduction to the oral microbiota • Microbial habitats within the mouth • Saliva and microorganisms • [Dental plaque and oral soft tissue biofilms are covered in later lectures]
1
Q
What is the difference between Autochthonous microbiota and Allochthonous microbiota?
A
Autochthonous microbiota: micro-organisms characteristically found at a
particular site. These organisms are adapted to survive and grow at that site.
Allochthonous microbiota: micro-organisms transiently present at a site. These organisms do not thrive at the site, but may colonise transiently, or if the site becomes compromised (ie in disease
2
Q
What are the 4 different types of oral microbiota found in the mouth?
A
- Archaea
- Viruses
- Fungi
- Bacteria
3
Q
When is archaea more likely to be isolated from the oral cavity?
A
in periodontal disease state, and not in health
4
Q
How are viruses detected in the oral cavity?
A
through PCR-based methods
5
Q
What viruses do we need to be aware of that are found in the oral cavity?
What is there clinical significance?
A
Herpes simplex type 1 (HSV-1) most common,HIV and hepatitis B
2) HIV and hep B can be carried asymptomatically thus cross infection methods must always be used.
6
Q
What is the most common oral infection of fungus?
2) Give an example of a less common fungus infection ,if occurs usually in immunosuppressed individuals:
A
1) Candida spp.
2) Aspergillus,
7
Q
What oral diseases are bacteria responsible for?
A
caires
periodontitis
abscesses
even infections on oversite of the body e.g. rember atheroscletotic plaques contain high risk periodontal pathogens (possible the cause)
8
Q
What oral bacteria is the most abundant?
A
streptococci
9
Q
Is streptococci , gram + or -, is it alpha-haemolytic or gamma haemolytic
A
gram +
alpha haemolytic
10
Q
What does it mean If bacteria are alpha haemolytic?
A
they don’t contain catalase, but they do produce hydrogen peroxide and release it out ,(this bleaches haemoglobin in kiss plate= yellow) they must have some way to protect themselves from the hydrogen peroxide.
11
Q
What does it mean If bacteria are gamma haemolytic? + e.g
A
no haemolysis occurs e.g staphylococcus
12
Q
Which one is alpha haemolytic and which one is gamma-haemolytic:
1) staphylococcus
2) streptococci
A
1) gamma
2) alpha
13
Q
what is the range of different species of bacteria within a individual in the mouth?
A
100-200
14
Q
How many species of bacteria naturally colonise the mouth? How mnay phyla?
2) How many of these species can we culture?
A
around 700
13 phyla
2) about half
15
Q
How many days does it take to form “mature” dental plaque?
A
1-4 days
16
Q
toothbrushing removed most bacteria from the exposed surfaces ,but dental plaques begins to accumulate with ____1____
A
1) minutes
17
Q
Give an innate defence of soft tissues?
A
slothing
18
Q
Where is the bacteria , and what type of bacteria causes halitosis?
A
anaerobic bacteria , the back of the tongue
19
Q
What bacteria are found in supragingingival plaque of the tooth?
A
streptococcus, actinomyces, haemophilus
20
Q
What bacteria are found in the subgingival plaque?
A
streptococcus, actinomyces, perptostreptococcus, fusobacterium, prophymonase, aggregatibacter
21
Q
What type of streptococcus are found in dental plaque?
A
streptococcus gordonii
streptococcus sanguinis
22
Q
What bacteria are found on the palate?
A
streptococcus and actinomyces
23
Q
What bacteria are found on the tongue?
A
Streptococcus, veillonella, actinomyces, haemophilus, prevotella
24
Q
What cell types is found in the lips, cheeks and palte? what process occurs as part of the innate immune response?
A
Epithelial cells, continually shed (desquamation)
25
Describe the surface of the tongue? What does this mean in terms of whether it habours periodontal bacteria?
Highly papillated
| Reservoir for obligate anaerobes (perio. pathogens) Note that tonsils may also harbour perio pathogens
26
Teeth are non-shedding in humans,
1) Why are there many different microbial populations found on teeth?
2) what are teeth covered with?
1) many different surfaces
| 2) acquired enamel pellicle
27
What are kiss plates made of ?
| 2) What do they allow you to identify? +observations
1) haemoglobin
| 2) turns yellow if alpha-haemolytic bacteria are resent, remains the same colour if gamma-haemolytic bacteria present
28
What type of plate is this (put IMAGE)?
blood agar, of micrococcus luteus
29
What type of plate is this (put IMAGE)?
blood agar, of micrococcus luteus
30
How do you estimate the numbers of different species of bacteria in clinical samples?
by culturing and counting colonies
31
Why do we culture bacteria?
to get an isolated bacteria
32
once we have cultured bacteria, what tests performed on the isolated bacteria allow us find out:
o understand basic physiology/biochemistry
o link organism to disease (Koch-Henle postulates)
o identify pathogenesis mechanisms
o test antibiotics.
33
What type of anomaly occurs when you perform a plate count?
| 2) Why does this happen?
Counting the number of bacteria on an agar plate greatly underestimates the number present.
2) • Some bacteria are dormant and not easily reactivated.
• Some species are fastidious – do not grow on nutrient agar.
• Estimates suggest that 50% of oral bacteria have never been isolated.
34
What is a more accurate way of finding the number of bacterial cells in a population than performing a plate count?
using a microscope
35
After extracting total community DNA, which approach tells you :
1) the single-gene phylogenetic tree
2) the total gene pool of the community
1) community sampling approach
| 2) environmental genomics approach
36
What is the outcome of the community sampling approach?
a single-gene phylogenetic tree
1. phylogenetic snapshot of most members of the community
2. identification of novel phylotypes (fancy way of saying new species)
37
What is the outcome of the environmental genomics approach?
total gene pool of the community in the form of partial complete genomes
1. identification of all gene categories
2. discovery of new genes
3. linking of genes to phylotypes
38
What is the method community sampling approach?
amplify a single gene (using PCR) e.g.gene encoding 12s rRNA
2) sequence
3) generate phylogenetic tree
39
What is the methods of environmental geneomics approach?
```
A)METHOD 1
1) restriction digest total DNA
2) shotgun the sequence
METHOD 2
1) sequence directly (without cloning) using a high-throughput DNA sequencer
B) assemble and annotate genes??
```
40
What are the main differences between the community sampling approach and the environmental genomics approach?
Community sampling approach focuses on 1 gene, cheap and tells you which microrganisms are there, while environmental genemoics aproach, sequences everything which is more costly and more complex analysis, lets you find the genes as well as the species that are present.
41
community sampling approach and the environmental genomics approach do require isolation of an organism
TRUE or FALSE
FALSE- it doesn't require isolation (but can aid isolation and identificaiton)
therefore no need to culture after sampling
42
What gene is targeted in "microbiome" analysis when using "next generation sequencing"?
2) What is the term used when full sequence is looked at using this technique?
identifies all DNA present, but when targerting a particulat gene it is usually 16sRNA
2) metagenomics
43
1) What does shotgun sequencing allow you to know?
| 2) How does it work (don't need to know this, but so you understand)?
lets you read longer DNA sequences
2) shotgun sequencing,[1][2] DNA is broken up randomly into numerous small segments, which are sequenced using the chain termination method to obtain reads. Multiple overlapping reads for the target DNA are obtained by performing several rounds of this fragmentation and sequencing. Computer programs then use the overlapping ends of different reads to assemble them into a continuous sequence.
44
What is a holoboint?
us and our resident microbes that have coevolved and coexisted with us in a mostly harmonious symbiotic relationship.
45
What is a phylotype?
a type of bacterium defined by its placement in a phylogenetic tree on the basis of it 16sRNA gene sequence
46
Define coevolution?
the parallel evolution of interacting species
47
What does next-generation sequencing mean?
an umbrella term for the number of different modern high through put sequencing technologies
48
Define symbiosis
2 or more species living closely together in a long-term relationship
49
Bacteria are removed from oral surfaces by:
* sloughing of epithelial cells
* mechanical debridement
* active release (possibly). E.g. your chewing as saliva from parotid “dilutes” normal saliva and changes location where your sampling bacteria from.
50
Saliva is ___1___ when secreted, but rapidly
accumulates ___2___ per ml
Approx. ___3___ times ___4 __bacteria are attached to epithelial cells than are ‘free’ in saliva.
1) sterile
2) around 10^9 bacterial cells
3) 3
4) more
51
Saliva is ___1___ when secreted, but rapidly
accumulates ___2___ per ml
Approx. ___3___ times ___4 __bacteria are attached to epithelial cells than are ‘free’ in saliva.
1) sterile
2) around 10^9 bacterial cells
3) 3
4) more
52
What do you sample to get an overall reflection of the oral microbiome?
saliva
53
What are the limitations when using saliva as overall reflection of the oral microbiome?
* proportions of microbial species are different from plaque/soft tissue biofilms
* Care is required when sampling to ‘standardise’
54
1) In health the saliva microbiome has a core, present in most individuals, this includes how many phyla?
2) And a peripheral microbiome, which includes what?
3) Microbiomes of different individuals form what?
3) Clusters are associated with differences in...
1) around 13 phyla
2) species present in some and not others
3) clusters
3) metabolome
55
Revise saliva components from 1st year
just do it!
56
What is the role of acquired enamel pellicle in oral disease?
it protects the enamel
57
What is the role of salivary compounds in oral disease/
a) salivary compounds can help control plaque accumulation through:
o Aggregating bacteria (which are then swallowed)
o Antimicrobial effects.
b) key nutrient for bacteria
58
What salivary molecules can bind to bacteria due to their sticky glycoproteins?
2) What else can they bind to?
3) what effect do these salivary molecules have on bacteria? (2)
4) What do these salivary molecules exhibit, like anything else to do with binding to a receptor/antigen>?
MG2 (muc7) saliva, agglytinin (gp340), PRPs and statherin
2) teeth
3) • Agglutinate/aggregate or inhibit bacteria
• May promote or inhibit microbial colonisation
4) specificity
59
Aggregation/ agglutination
1) What phase does this occur in?
2) what does it result in?
3) How does this benefit the immune response?
1) in fluid phase
2) large clumps
3) • Large clumps adhere poorly and are removed (swallowing).
60
Adhesion
1) What phase does this occur in?
2) what does it result in?
3) How does this benefit the immune response?
1) on surface of teeth (remember some proteinsexhibit different bacteria binding properties when in solution than when on a surface)
2) adhesion of bacterial cells to teeth
3) trick is doesn't
61
Give examples of salivary proteins that exhibit different bacteria binding properties when in solution than when on a surface:
e.g. statherin, PRPs, gp340
| These doe not allow bacteria to adhere to teeth but do cause them to aggregate in fluid phase
62
What happens to concentration of secreoty IgA, an immunoglobin, in saliva when comparing resting flro to stimulated flow?
* 33mg/100mL resting flow
| * 6mg/100mL stimulated flow
63
What do immunoglobins do?
agglutinate bacteria (removed by swallowing)
64
What immuniglobins are present in gingival fluid?
| 2) What do they activated?
1) IgG, IgM
| 2) Activate complement/opsonization
65
What is this the structure of? and where is ti found?
| attach image
an immhunoglobin , IgA is found in saliva
66
what is the role of MG2?
2) what is the name of the gene encoding it?
3) what secretees it?
4) what size is it?
5) is it monomeric ? What does this word mean?
6) Describe its viscoeleastic properties? what does this mean?
1) a muccin which binds to bacteria,
2) muc7
3) Secreted by serous acini of submandibular & sublingual and minor glands
4) Relatively small (125kDa),
5) yes it is monomeric, a molecule that can undergoe polymerisation
6) it has low viscoelastic properties , property of materials that exhibit both viscous and elastic characteristics when undergoing deformation
67
What type of biological molecule is cf MG1?
2) what percentage is it carbohydrate?
3) What is the chemical name?
4) What bacteria does it bind to?
1) glycoprotein
2) around 70%
3) Sialyated di-tri-saccharides
4) • Streptococcus sanguinis, S. mitis, S. gordonii, Aggregatibacter actinomycetemcomitans, Pseudomonas aeruginosa, E. coli
68
cf MG1
1) What antigens is it lacking?
2) What property does Non-glycosylated peptide domains give it?
3) What can • Histidine rich N-term domain do?
4) What is surface found on teeth is it not found?
1) ABO blood group determinants
2) bacteriocydal
3) disturb cell membranes
4) pellicle
69
What is the name given to human salivary agglutinin?
gp340
70
2) what is the name of the gene encoding GP340?
3) what secretes it?
4) what size is it?
5) is it monomeric ?
6) What is it ? what is its role?
2) DMBT1
3) all salivary glands
4) 340kDa
5) yes
6) it was the orginal strep.mutan salivary aggregator, S.salivarius and S.sanguinis agglutination.
71
What bacteria cna gp340 bind to?
2) Where is it found in the mouth?
3) What is a molecule identical gp340 used for clinically?
4) How does it work
1) S. mutans, S. gordonii, S. salivarius and S. sanguinis
2) pellicle
3) it is a glycoprotein in lung washings
4) it binds to surfactant protein-D, enhances phagocytosis and killing of microorganism by neutrophils and macrophages
72
Which one of these if found in the pellicle?
| the mucin, MG2 OR salivary glycoprotein, gp340
gp340
73
How many mucins are present in saliva? What are there propterties and roles?
MG1 (produces a viscous saliva) and MG2 (relatively small, not a typical viscous mucin, binds to bacteria so swallowed)
74
Some bacteria will only adhere to gp340, some will only aggregate with gp340, some do both.
TRUE OR FALSE
Why?
True,
| we don't know why these are just observations
75
What is the name of the C terminus in proline-rich proteins (PRPs) which bacteria have evolved to bind to in solid state?
cryptitope
76
What are the domains of PRPs?
N-terminus
| and C-terminus
77
What does PRP stand for?
proline-rich proteins
78
In proline-rich proteins , what binds to
1) N-terminus
2) C-terminus
1) hydroxyapatite
2) • Actinomyces spp.
• Streptococcus mutans
• Streptococcus sanguinis
79
What is the main function of PRPs?
2) Where is it found in high concentrations?
3) What types are there?
1) stabilise calcium phosphate
2) parotid and submandibular saliva (70% of total protein)
3) acidic, basic and glyosylated
80
What secretes statherin?
parotid and submandibular
| gland
81
What is the main function of statherin?
calcium phosphate stabilisation (with PRPs)
82
What does it bind to?
• Binds to hydroxyapatite
• Binds Porphyromonas gingivalis and
Actinomyces spp.
83
Do statherins binding properties change when as a precipitate and when in soluble state? if so how?
Does not bind bacteria when in soluble state
84
What is the molecular weight of statherin?
• Low molecular weight (43 aas)
85
Give some of the many antibacterials in saliva?
* Lysozyme
* Lactoperoxidase/thiocyanate/hydrogen peroxide system
* Lactoferrin
86
Where are lysozymes found?
2) is it equally effective against all bacteria?
3) Which bacteria are the most tolerant to it?
4) How does it effect oral bacteria?
5) How?
tears, saliva and sweat
2) no
3) no
3) oral bacteria
4) reduces viability
5) enzymatically: it can cleave bacterial cell wall peptidoglycan
non-enzymatically, causes cell degradtion
87
1) What does lactoperoxidase do?
2) How does it cause damdge to the bacteria?3) What are the reactants and where do they come from?
4) what produces lactoperoxidase
it converts hydrogen peroxide, into a more toxic agent (Reaction product is
hypothiocyanite plus some
cyanosulphurous acid and cyanosulphuric acid)
2) which can cross membranes, as uncharged, and caus damdge
3) thiocynate (which is present in saliva as KSCN)
And
Hydrogenperoxide (which is a metabolic by-product of many oral streptococci)
4) host and bacteria
88
When will the presence of lactoperoxidase product, OSCN (hypothiocynite) be most toxic to bacteria?
2) Why?
the more acidic the conditions are ,
2) The pK for HOSCN/OSCN- is 5.3
• More acid favours HOSCN
• Due to uncharged nature, HOSCN penetrates bacterial cell envelope better
89
What does lactoferrin do?
bind to iron , which stops bacteria from having a source of iron
90
Why do bacteria need iron?
it is an enzyme co-factor
91
What do some bacteria produce to compete for host iron against lactoferrin?
• Some bacteria produce iron-binding proteins called siderophores to compete for host iron
92
Do we use antibacterial enzymes in dentistry?
2) What are they used for and who to treat?
3) how does Biotène: work?
4) How does BioXtra: work?
yes ,
2) a) bought by patients with dry mouth,
b) for pets and children.
3) Dual enzyme system
4) Contains lysozyme, lactoperoxidase, and lactoferrin
