Colistin Resistance and Molecular Characterization of the Genomes of -Positive Clinical Isolates.

Research on resistance against polymyxins induced by the gene is gaining interest. In this study, using agar dilution method, polymerase chain reaction, and comparative genomic analysis, we investigated the colistin resistance mechanism of clinical isolates. The minimum inhibitory concentration (MIC) analysis results revealed that of the 515 isolates tested, bacteria with significantly increased MIC levels against colistin were isolated in 2019. Approximately one-fifth (17.14% to 19.65%) of the isolates showed MIC values ≥1 mg/L against colistin in 2015, 2016, and 2017. However, in 2019, up to three-quarters (74.11%, 146/197) of the isolates showed MIC values ≥1 mg/L against colistin indicating an increase in colistin resistance. Six isolates (EC7518, EC4968, EC3769, EC16, EC117, EC195, 1.13%, 6/515) were found to carry the gene and a novel variant with Met2Ile mutation was identified in EC3769. All six strains showed higher MIC levels (MIC=4 mg/L) than any negative strains (MIC ≤ 2 mg/L). Whole-genome sequencing of the six -positive isolates revealed that EC195 carried the highest number of resistance genes (n = 28), nearly a half more than those of the following EC117 (n = 19). Thus, EC195 showed a wider resistance spectrum and higher MIC levels against the antimicrobials tested than the other five isolates. Multi-locus sequence typing demonstrated that these -positive strains belonged to six different sequence types. The six genes were located in three different incompatibility group plasmids (IncI2, IncHI2 and IncX4). The genetic context of was related to a sequence derived from Tn (IS---IS). Investigations into the colistin resistance mechanism and characterization of the molecular background of the genes may help trace the development and spread of colistin resistance in clinical settings.