Epidemiological links Flashcards
list the order of microbial taxonomy
- domain- bacteria
- kingdom
- phylum- proteobacteria
- class- gamma proteobacteria
- order- enterobacteriales
- family- enterobacteriacea
- genus- escherichia
- species- escherichia coli
what is meant by microbial strain in a taxonomic sense
a strain is made up of the descendants of a single isolation in pure culture and is usually made up of a succession of cultures ultimately derived from an initial single colony
what is the definition in nature of microbial strain
a strain is an isolate or group of isolates that can be distinguished from other isolates of the same genus and species by phenotypic characteristics or genotypic characteristics or both
how can we differentiate microorganisms below species level
- phenotypic typing methods
- antibiotic resistance typing
- phage typing (staphylococcus aureus)
- serotyping (salmonella) - genetic typing methods- genotyping
- electrophoretic methods (PCR fingerprinting)
- sequence base typing (single gene sequencing)
- other methods (SNPs and microsatillite analysis )
what are phenotypic typing methods based on
based on analysis of phenotypic characteristics
what are genetic typing methods based on
based on analysis of genetic characteristics
what are the reagents needed in PCR
- DNA template extracted from culture or sample
- DNA polymerase
- nucleotide bases (dNTPs)
- primers flanking the region of interest
- water
- polymerase buffer
what is PCR
exponential amplification of nucleic acids by a series of alternating heating and cooling steps
what does PCR stand for
polymerase chain reaction
describe the process of amplification of nucleic acids
- analysis of amplified DNA fragments typically involves an electrophoresis in agarose or polyacrylamide gel
- fragments are separated according to their sizes
- gels are visualised under UV light
give examples of application of genotyping methods
- outbreak of health care associated Burkholderia bacteremia and infection attributed to contaminated sterile gel used for central line insertion
- whole genome analysis of Exserohilum rostrum from an outbreak of fungal meningitis
describe the outbreak of BCC
- a group of gram - aerobic bacilli
- patients with BCC are typically unwell, with multiple comorbidities, including being immunocompromised
- common sources of infection are usually contaminated IV fluids, contaminated ventilation equipment or contaminated fomites
- this outbreak is attributed to contaminated ultrasound gel
- molecular typing of the isolates used several techniques including a MLST method
what does MLST stand for
multi locus sequence typing
what is MLST
an unambiguous procedure for characterising microbial isolates, using the sequences of internal fragments of usually 7 house keeping genes
- approx 450-500bp internal fragments of each gene are used, as these can be accurately sequenced on both strands using an automated DNA sequencer
what occurs for each housekeeping gene in MLST
For each housekeeping gene, the different sequences present within a microbial species are assigned as distinct alleles
what occurs for each microbial isolate in MLST
For each microbial isolate, the alleles at the end of each loci define the allelic profile or sequence type
what is each isolate characterised by in MLST
characterised by a series of 7 numbers which correspond to the alleles at the 7 housekeeping loci
what is the prevelance of the allelic profiles in MLST
- most bacterial species have sufficient variation within housekeeping genes to provide many alleles per locus
- allowing billions of distinct allelic profiles to be distinguished using 7 housekeeping loci - the allelic profiles of isolates can easily be compared to those in a large central database via the internet
explain the link between MLST and the B.cenocepacia outbreak
- MLST typing demonstrated that B.cenocepacia isolates from patients linked to the use of the ultrasound gel and an isolate grown from the gel were highly genetically related to each other
- most isolates related to the infection shared the same MLST sequence type
- the ultrasound gel was identified as the most likely source of the infection outbreak
- within 24 hours of being notified, the therapeutic goods administration contacted the distributor and manufacturer, issued a notice of recall and classified the event as life threatening
describe the outbreak of fungal meningitis
- more than 13000 people were exposed to contaminated methylprednisolone and 750 developed fungal infections
- vast majority of infections were caused by E.rostratum , a saprobic mould found in soil
- WGST was used to investigate if the fungal strains identified from patients and lots of methylprednisolone were the same or different
- 2 next generation sequencing platforms were used
what does WGST stand for
whole genome sequence typing
what did the whole genome analysis find in the outbreak of fungal meningitis
- showed that most outbreak isolates had identical genomes
- no differences found between strains isolated from patients and lots of methylprednisolone
- data demonstrated that the outbreak strains were clonal and suggested a single common source
- isolates from the 2 different methylprednisolone lots were indistinguishable suggesting that the source of contamination was persistent
