High resolution melt analysis of the spa region demonstrates diversity within ST93-MRSA from northern Australia

  • Steven Tong, Menzies School of Health Research, Charles Darwin University, Australia
  • Rachael Lilliebridge, Menzies School of Health Research, Charles Darwin University, Australia
  • Dr Deborah Holt, Menzies School of Health Research, Charles Darwin University, Australia
  • Dr Malcolm McDonald, Menzies School of Health Research, Charles Darwin University, Australia
  • Dr Bart Currie, Menzies School of Health Research, Charles Darwin University, Australia
  • Dr Philip Giffard, Menzies School of Health Research, Charles Darwin University, Australia

    • Objectives
    ST93 is a singleton Staphylococcus aureus clone unique to Australia. ST93-MRSA is one of the two major clones of PVL+ community-associated MRSA extant in Australia, and it is now apparent that it is also present in northern Australia. The objective of this study was to obtain insight into the diversity of this clone, and its MSSA equivalent, and to infer recent transmission patterns.

    Methods
    Isolates were defined as belonging to ST93 using a set of seven SNPs that discriminate the major S. aureus clones. Two additional confirmatory SNPs were used. We assessed diversity within the clone by high resolution melt analysis of the spa locus for initial high throughput screening, and by sequence determination when appropriate. Parallel experiments were carried out using isolates assigned to the ST30-MRSA clone, as the recent history and diversity of this clone are quite well understood and it provides a valuable comparison point. We performed HRM analysis on 30 isolates for each group: ST93-MRSA, ST93-MSSA and ST30-MRSA.

    Results
    There was greater diversity within ST93-MRSA, with at least four distinct spa types discriminated compared to only one within ST30-MRSA. HRM analysis of a glpF SNP demonstrated the ability to consistently discriminate a one base pair difference between isolates.

    Conclusions
    Diversity of spa type within ST93-MRSA compared to ST30-MRSA suggests that ST93 may represent a clone that has been resident in Australia for a long time, as distinct from the recently imported ST30-MRSA. This is consistent with the absence of ST93 or close relatives from other parts of the world. Diversity may have emerged by either the diversification of a single ST93-MRSA progenitor or multiple independent acquisitions of mecA by ST93. The study also showed that HRM analysis is a robust, efficient and rapid means for determining diversity within the spa locus.