Development and Validation of a New Diagnostic PCR Assay for vanA Gene Encoding Vancomycin Resistance in Staphylococcus aureus
Year: 2014
Type: article
Abstract: Staphylococcus aureus is associated with several human diseases.Following the introduction of meticillin for the treatment of infections associated with penicillin-resistant Staphylococcus aureus, meticillin resistant S. aureus (MRSA) emerged.Recent reports indicate that some MRSA strains have acquired vanA gene encoding staphylococcal vancomycin resistance, thus constituting further pressures on clinical antibiotic use and infection control.To prevent the spread of such clones, rapid diagnostic assays are needed.We used all the vanA nucleotide sequences available at the National Centre for Biotechnology Information to perform multiple alignments in the clustalW suite at the European Bioinformatics Institute from which we designed and developed a new vanA PCR assay.The new vanA PCR assay generated 235bp amplicon through 40-cycles in 55 min with negative predictive value of 100% and positive predictive value of 100% and sensitivity of one colony-forming unit.Following primary isolation, the turn-around time was 2.5 h against ≥ 24 h by bacteriological methods.The new assay was validated on reference vancomycin-susceptible and vancomycinresistant staphylococcal strains (N=345) obtained from local, national and international standard supplies.PCR products were confirmed by nucleotide sequencing.Staphylococcal vancomycin resistance testing methods recently recommended by the CLSI were used to confirm staphylococcal vancomycin resistant phenotype.Use of multiple sequence data for oligo-design will help stay ahead of the bugs.The new vanA PCR assay can easily be integrated into the routine microbiology workflow especially for laboratories with slim budgets.Richer laboratories could easily convert the new vanA PCR assay from end-point platform to real-time PCR using fluorescent probes.
Language: English
Subfield: Infectious Diseases
Field: Medicine
Domain: Health Sciences
Open Access status: green
OpenAlex ID: W2183233247