An epidemic CC1 MRSA IV clone yields false negative test results in molecular MRSA identification assays: a note of caution

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 (SCC mec ) 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 /SCC mec 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 /SCC mec integration site presumably targeted by such tests. In contrast to MW2 (GenBank [BA000033.2][1], a CC1 “USA400” strain which also harbours SCC mec 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, ydiL 2, C5QAP8, A8YYX4, npd- SCC, H4AYD7; nucleotide positions 280,690–286,024 of GenBank RBVO000005.1). An SCC mec element with the same insertion was previously found in a Staphylococcus epidermidis isolate (GenBank [MH188467.1][2]) 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 SCC mec junction sites, not only in Staphylococcus aureus but also in coagulase-negative staphylococci. ### Competing Interest Statement The authors have declared no competing interest. ### Funding Statement There was no external funding for this study. ### Author Declarations All relevant ethical guidelines have been followed; any necessary IRB and/or ethics committee approvals have been obtained and details of the IRB/oversight body are included in the manuscript. Yes All necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines and uploaded the relevant EQUATOR Network research reporting checklist(s) and other pertinent material as supplementary files, if applicable. Yes Relevant data are either covered by the manuscript and the supplemental file, or (in case of one sequence) are suvbmitted to GenBank; accession no. [MT380478][3], currently still being processed) [1]: /lookup/external-ref?link_type=GEN&access_num=BA000033.2&atom=%2Fmedrxiv%2Fearly%2F2020%2F05%2F05%2F2020.04.28.20083048.atom [2]: /lookup/external-ref?link_type=GEN&access_num=MH188467.1&atom=%2Fmedrxiv%2Fearly%2F2020%2F05%2F05%2F2020.04.28.20083048.atom [3]: /lookup/external-ref?link_type=GEN&access_num=MT380478&atom=%2Fmedrxiv%2Fearly%2F2020%2F05%2F05%2F2020.04.28.20083048.atom