Phenotypic and genotypic characterisation of linezolid-resistant coagulase-negative staphylococci possessing cfr-carrying plasmid

Abstract

Linezolid-dependent growth has contributed to wide dissemination of Staphylococcus epidermidis in hospitals. This study aimed to characterise linezolid-resistant coagulase-negative staphylococci (CoNS) isolates and the possibility of linezolid dependence in China. Phenotypic and genotypic resistance of 13 CoNS isolates were investigated by antimicrobial susceptibility testing and PCR. Similarity of isolates was determined by pulsed-field gel electrophoresis (PFGE). Characterisation of the cfr-carrying plasmid was performed by S1 nuclease PFGE, Southern blotting and whole-genome sequencing (WGS). A maximum likelihood phylogenetic tree was constructed for phylogenetic analysis. Growth curve analysis was performed with and without linezolid to determinate the possible contribution of linezolid dependence to linezolid-resistant CoNS dissemination. Thirteen CoNS isolates showed linezolid MICs of 8 mg/L to >256 mg/L, typed into three PFGE profiles. Southern blotting and WGS indicated that the cfr gene was located on a 39.5-kb plasmid with 99% identity to cfr-harbouring plasmids pSR01, pLRSA417 and pH29-46. The cfr gene was flanked by two copies of an IS256-like element ISEnfa4 family transposase, indicating the transferability of linezolid resistance conferred by the cfr gene. Comparative phylogenetic analysis revealed that Staphylococcus capitis XZ03 shared high similarity with linezolid-resistant S. capitis isolates in Huashan Hospital, Shanghai. Thirteen CoNS isolates did not exhibit linezolid dependence on exposure from 8-32 mg/L. The endemic CoNS clone carrying the cfr gene in our hospital showed high-level linezolid resistance, threatening linezolid use. Linezolid-dependent growth under linezolid selective pressure was not observed, indicating that it may not yet be a common phenotype in Staphylococcus spp.