Rapid detection of the pandemic methicillin-resistant Staphylococcus aureus clone ST 239, a dominant strain in Asian hospitals

Abstract

We describe and validate a novel PCR assay to detect the pandemic hospital-acquired methicillin-resistant Staphylococcus aureus (HA-MRSA) lineage ST 239. Results based on previously uncharacterized isolates from a hospital in northeast Thailand support the view that at least 90% of HA-MRSA isolates in mainland Asia correspond to ST 239 or close relatives.

FIG. 1.

Primer design. Two pairs of primers were designed based on SA0317 and SA2003 loci (gene codes from the N315 genome and annotated as hypothetical protein and hysA, respectively). These genes were identified as being the most variable of those sequenced previously by Robinson and Enright (21), which facilitates the design of primers specific for ST 30- and ST 8-like sequences. The ∼550-kb ST 30-like replacement spans the origin of replication and is shown as the hatched block. The positions of SA0317 and SA2003 are identified by arrows. SA0317 falls within the ST 30-like replacement, while SA2003 falls outside of it. The primer sequences are shown in boldface type, with the positions given in vertical numbers above the sites. Primer direction is indicated by arrows. SA2003 forward and reverse primers are specific for the ST 239/ST 8-like sequence, while the SA0317 forward primer is specific for the ST 239/ST 30-like sequence (note the 3-base indel in the corresponding ST 8 sequence in this case).

FIG. 2.

(a) Agarose gel electrophoresis of amplicons using initial control strains. Lane 1, EMRSA-4 (ST 239); lane 2, C507 (ST 30); lane 3, C125 (ST 8); lane 4, negative control (−ve) (no DNA). Both amplicons are present for ST 239, only the larger one is present for ST 30, and the smaller one is present for ST 8. The sizes of the amplicons are as predicted. (b) Agarose gel electrophoresis of five Thai isolates characterized by MLST. Lane 1, MSSA (ST 88) (only the small band amplifies); lanes 2 to 4, MRSA (ST 239) (both bands amplify); lane 5, MRSA (ST 779) (only the small band amplifies).