IS257-mediated amplification of tet(L) variant as a novel mechanism of enhanced tigecycline resistance in Staphylococcus cohnii

The mechanism of enhanced tigecycline MIC in Staphylococcus cohnii after in vitro tigecycline exposure was investigated. S. cohnii 11-B-312 was exposed to incremental concentrations of tigecycline (2–32 mg/L) and the mutants growing at 8, 16 and 32 mg/L were determined by AST and WGS. Copy number and relative transcription level of the tet(L) gene were determined by quantitative PCR. The fitness cost was evaluated by growth kinetics and competition assays. The results revealed that enhanced tigecycline MIC was identified in S. cohnii mutants. Copy number and relative transcription level of tet(L) in the mutants increased 8-, 20-, and 23-fold and 20-, 34-, and 39-fold in the presence of 8, 16, and 32 mg/L tigecycline, respectively. The read-mapping depth ratio analysis indicated that a multidrug resistance region carrying the tet(L) variant has a gradually increased copy number, correlating with the tigecycline selection pressure. S. cohnii strain 11-B-312_32 had a fitness cost, and enhanced tigecycline MIC can revert to the initial level in the absence of tigecycline. In summary, enhanced tigecycline MIC develops with extensive amplification of an IS257-flanked tet(L)-carrying segment in S. cohnii. IS257 seems to play a vital role in the gain and loss of the amplification product.