Optisch-Molekulare Diagnostik und Systemtechnologie

Background: A variety of rapid molecular PCR tests has been developed and commercialised that interrogate the junction site between the staphylococcal core genome, and the mobile genetic element (SCCmec) which harbours the gene responsible for methicillin-/beta-lactam-resistance. Aim: The purpose of the present study was to investigate why a clinical MRSA isolate yielded false negative test results with widely used, commercial orfX/SCCmec junction site PCR tests. Methods: A collection of isolates that belonged to the same epidemic strain as the index isolate were investigated with commercial MRSA assays and all isolates were sequenced in order to explain this observation. Results: It was found that isolates of the epidemic European CC1-MRSA-IV clone, which likely originated in South-Eastern Europe and subsequently spread to Western Europe, generally exhibit this behaviour. The failure of the assays was attributable to a characteristic large insertion in the orfX/SCCmec integration site presumably targeted by such tests. In contrast to MW2 (GenBank BA000033.2, a CC1 USA400 strain which also harbours SCCmec IVa), the European CC1 clone harbours an insertion of ca. 5,350 nucleotides adjacent to orfX. This sequence starts with a novel SCC terminal sequence alternate to dcs and encodes six different hypothetical proteins (E7MHX1, ydiL2, C5QAP8, A8YYX4, npd-SCC, H4AYD7; nucleotide positions 280,690-286,024 of GenBank RBVO000005.1). An SCCmec element with the same insertion was previously found in a Staphylococcus epidermidis isolate ( GenBank MH188467.1) suggesting transfer between staphylococcal species. Conclusion: In order to ensure the reliability of molecular MRSA tests, it is vital to monitor the emergence of new SCCmec junction sites, not only in Staphylococcus aureus but also in coagulase-negative staphylococci.