Diagnostic and Typing Methods Flashcards
What bacteria are associated with periodontal disease?
P. gingivalis
A. actinomycetemcomitans
P. intermedia
B. forsythus
What bacteria are associated with dental caries?
S. mutans
What bacteria are associated with root canal infections?
P. endodontalis
F. nucleatum
What are the methods of bacteria detection?
Microbiological culture - culture on agar medium, isolate bacteria and identify by characterisation of enzyme activities and sugar fermentation tests
Molecular biology - DNA probes and PCR
What are the advantages of the microbiological culture method?
Yields isolated bacteria for future testing and study
What are the disadvantages of the microbiological culture method?
Requires viable cells
Insensitive
Only a small number of samples can be analysed at once
Results can be inconclusive
Labour heavy
What are the advantages of PCR?
Highly specific
Sensitive
Can be used to directly detect bacteria in clinical specimens
Doesn’t require viable cells
Less time consuming
What are the disadvantages of PCR?
May detect dead cells
Can only detect pre-selected species
Why is it important to subtype bacteria?
To track routes of transmission during disease outbreaks
To study pathogenicity of specific strains
What are the different molecular (genetic) typing methods?
Restriction enzyme analysis (REA)
Gene probe typing
Ribotyping
16S-23S intervenir spacer region (IGS)
DNA sequencing
What do PCR primers target?
16S rRNA gene
What is REA and what are its problems?
Restriction enzyme analysis
Digests whole genomic DNA with restriction enzymes
Too many DNA fragments are obtained so interpretation is difficult
What is gene probe typing?
Reduces the number of DNA fragments generated by REA using a suitable gene probe
What is ribotyping?
Using E. coli rRNA operon as a DNA probe following REA
rRNA operon is present in multiple copies in bacterial genomes and is generally well conserved
What is IGS?
16S-23S intergenic spacer region
This region is amplified by PCR using primers
PCR product is digested with restriction enzymes to obtain strain-specific fingerprints
