| Summary: | Tetracycline resistance (Tet<sup>R</sup>) has been evidenced as a good phenotypic marker for detection of livestock-associated methicillin-resistant <i>Staphylococcus aureus</i> (LA-MRSA) isolates of the clonal complex CC398. The aim of this study was to characterise a collection of 95 Tet<sup>R</sup>-MRSA isolates, not belonging to the lineage CC398, that were obtained in a previous multicentre study, to detect other MRSA clonal complexes that could be associated with this phenotypic Tet<sup>R</sup> marker. The Tet<sup>R</sup>-MRSA isolates were recovered from 20 Spanish hospitals during 2016 and they were characterised to determine their antimicrobial resistance and virulence phenotypes/genotypes as well as the presence of the immune evasion cluster (IEC). A high proportion of isolates belonging to the CC1 lineage (46%) were observed, as well as to the CC5, CC8 and CC45 lineages (11% each one). Thirty-two different <i>spa</i>-types were identified, being predominantly CC1-t127 (40%) and CC45-t1081 (11%). The IEC system (with the gene <i>scn</i> as marker) was present in 73% of isolates and 16% produced the Panton Valentine leucocidin (PVL). A high proportion of MRSA-CC1 isolates were <i>scn</i>-negative (38.6%) and 52.9% of them were <i>blaZ</i>-negative. A multidrug resistance (MDR) phenotype was identified in 86% of MRSA isolates. The knowledge of other Tet<sup>R</sup>-MRSA genetic lineages, in addition to CC398, is highly relevant, since most of them were MDR and some of them presented important virulence factors. Strains potentially associated with livestock (as the subpopulation CC1-t127-<i>scn</i>-negative) or with humans (as the CC45 lineage or the subpopulation CC1-<i>scn</i>-positive) have been found in this study. The use of tetracycline-resistance for detection, not only of CC398 but also of other LA-MRSA lineages should be tracked in the future.
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