The emergence and biological characteristics of linezolid-resistant <i>Clostridioides difficile</i> isolates in the Asia-Pacific region

Linezolid-resistant <i>Clostridioides difficile,</i> conferred by the acquisition of <i>cfr</i>-like genes, has been reported in Europe and America. However, the emergence of linezolid resistance in <i>C. difficile</i> in the Asia-Pacific region and its impacts on <i>C. difficile</i> pathogenicity remain unclear. In this study, 881 <i>C. difficile</i> isolates from the Asia-Pacific region were screened for <i>cfr</i>-like genes. Whole genome sequencing was performed on 16 <i>cfr</i>-like gene-positive isolates from four countries. Thirteen isolates possessed <i>cfr</i>(B), which was located within Tn<i>6218</i>, while three isolates possessed <i>cfr</i>(C), which was located within the integrative and conjugative elements (ICE) F548 and DA275. Fourteen (87.5%, 14/16) of the <i>cfr</i>-like gene-positive isolates were resistant to linezolid. In comparison to the two isolates susceptible to linezolid, these 14 isolates exhibited significantly higher mRNA expression levels of <i>cfr</i>(B) and <i>cfr</i>(C), along with significantly higher bacterial density at 12 h. Conversely, they demonstrated reduced abilities for sporulation and biofilm formation. After the <i>cfr</i>(B) gene was knocked down by the CRISPR interference, the <i>C. difficile</i> strain presented lower bacterial density at 12 h, higher toxin production and stronger sporulation and biofilm formation abilities. Our findings reveal the emergence of <i>cfr</i>-like genes <i>C. difficile</i> isolates in the Asia-Pacific region, highlighting that <i>cfr</i>-like genes not only mediate linezolid resistance but also contribute to regulating pathogenic potential. Linezolid resistance in CDI should be closely monitored in specific patients.