Stefan Monecke (1), Ralf Ehricht (2), Peter Slickers (2), Hui-Leen Tan (3), Geoffrey Coombs (3) (1) Institute for Medical Microbiology and Hygiene, Technical University of Dresden, Germany, (2) CLONDIAG GmbH, Jena, Germany, (3) Department of Microbiology and Infectious Diseases, PathWest Laboratory Medicine Western Australia -Royal Perth Hospital, Australia Between 2003 and 2008, seventy-six clinical isolates of a Panton-Valentine leucocidin (PVL)-positive ST8-MRSA-IV strain were isolated from seventy-two non-hospitalised patients living in Western Australia. All isolates had a USA300-like pulsed field gel electrophoresis pulsotype. Approximately 95% of isolates were recovered from skin and soft tissue infections.Genotyping was performed on all isolates using a diagnostic DNA-array which included species markers, resistance factors, virulence-associated determinants, and MSCRAMM genes. Strains varied in their carriage of the blaZ/I/R, ermC, msrA+mpbBM, aadD+mupR, aphA3+sat, tetK, qacC, merA/B/R/T resistance genes, the beta-haemolysin-converting phages, and the enterotoxin (sek+seq) genes. The argininecatabolic mobile element (ACME) was absent in 15.8% of isolates. The mercury resistance (mer) operon, which is usually associated with SCCmec type III elements was found in several ACME-negative isolates. Hybridisation profiles identified sixteen variants or subtypes. 47 (61.8%) strains were identified as “Variant A” which was identical to the sequenced USA300-TC1516 strain. The earliest isolate of USA300detected in Western Australia (July 2003) belonged to this subtype suggesting most other variants may have evolved from variant A as a result of a limited number of gene losses or acquisitions. Investigations on family outbreaks showed that the analysis of such intrastrain variability might be helpful for tracing transmission.
Introduction: Several distinguishable PVL-positive CA-MRSA strainsfrom very diverse clonal groups with different geographic distributions have evolved. A clonal group 8, spa type t008 MRSA strain carrying the mecA-containing Staphylococcal Chromosomal Cassette (SCCmec) type IV element has recently emerged as the dominant MRSA strain in several regions of North America in both the community and hospital setting. Colloquially known as USA300 sporadic reports ofthis strain have been reported from several countries including Australia. Material & Methods: USA300 isolates were identified based on PFGE profile (O'Brien, 2006) and positive PVL-PCR (Fey, 2003). All isolates were examined using DNA-microarrays covering 334 targets (approximately 185 distinct genes and their alleles). Related protocols and procedures have been described previously (Monecke 2007,2008). In short, for every target, one specific primer was developed, and a mixture of these primers was used for a multiplex primer elongation reaction incorporating Biotin-16-dUTP into the amplicons. The labelled sample was hybridised against the DNA-array and horseradish peroxidase coupled streptavidin was added. The ArrayTube was then placed into a reading device (ATR 01 or ArrayMate,CLONDIAG), and Seramun Green was applied for precipitation staining. Scans were recorded and analysed.
Table 1: Variants of USA300/WA MRSA12 based on the variability of 31 out of 334 targets. Sequenced strains USA300-FPR3757 and USA300TCH1516 are shown for comparison.
Results: Between 2003 and February 2008, 76 WA MRSA-12/USA300 were isolated from 72 patients living in the Perth metropolitan area ofWestern Australia. Skin and soft tissue infections were reported in 68 (94.4%) patients. The remaining 4 patients' isolates were from contact screening specimens. Cases of necrotising pneumonia were not observed. The first case of an infection with WA MRSA12/USA300 was reported in 2003. Since this time, an increase in the number of people infected with this strain has occurred, particularly...