Rapid discrimination of Staphylococcus epidermidis genotypes in a routine clinical microbiological laboratory using single nucleotide polymorphisms in housekeeping genes.
PURPOSE
Staphylococcus epidermidis colonies often display several morphologies and antimicrobial susceptibility patterns when cultured from device-related infections, and may represent one or multiple genotypes. Genotyping may be helpful in the clinical interpretation, but is time consuming and expensive. We wanted to establish a method for rapid discrimination of S. epidermidis genotypes for use in a routine microbiology laboratory.


METHODOLOGY
A real-time PCR targeting eight discriminatory class I or II single-nucleotide polymorphisms (SNPs) in six of the seven housekeeping genes was constructed. Post PCR, high-resolution melt (HRM) analysis using EvaGreen as fluorophore discriminated amplicons based on their percentage GC content.


RESULTS
In silico, 42 representative sequence types (STs), including all major MLST group and subgroup founders, were separated into 23 different cluster profiles with a Simpson's index of diversity of 0.97. By HRM-PCR, 11 commonly encountered hospital and outbreak STs were separated into eight HRM patterns.


CONCLUSION
This method can rapidly establish whether S. epidermidis strains belong to different genotypes. It can be used in patients with S. epidermidis infections, as an aid in outbreak investigations and to select strains for investigation with more discriminatory methods, saving workload and costs. Results may be obtained the same day as culture results. Its strength lies mainly in indicating differences, as some STs may have the same melt profile. Changes in S. epidermidis epidemiology may warrant alterations in the inclusion of SNPs. We believe this method can reduce the threshold for performing genotyping analysis on an increasingly important nosocomial pathogen.
['Larssen|Kjersti Wik|KW|', 'Nor|Anne|A|', 'Bergh|Kåre|K|']
D015373:Bacterial Typing Techniques D003428:Cross Infection D004269:DNA, Bacterial D020043:Genes, Essential D005838:Genotype D006801:Humans D058885:Multilocus Sequence Typing D020641:Polymorphism, Single Nucleotide D060888:Real-Time Polymerase Chain Reaction D013203:Staphylococcal Infections D013212:Staphylococcus epidermidis
single nucleotide polymorphisms S. epidermidis sequence types HRM-PCR clinical routine rapid genotyping