The aim of this study was to investigate the prevalence of the polymyxin resistance gene mcr-1 in Enterobacteriaceae from environmental water sources in Hangzhou, China. Colistin-resistant bacteria were isolated from environmental water samples using an enrichment broth culture method, were screened for mcr-1, and then were analyzed for the location and transferability of mcr-1 Isolates positive for mcr-1 were further examined to determine their susceptibility profiles and were screened for the presence of additional resistance genes. Twenty-three mcr-1-positive isolates (16 Escherichia coli, two Citrobacter freundii, two Klebsiella oxytoca, two Citrobacter braakii, and one Enterobacter cloacae) were isolated from 7/9 sampling locations; of those, eight mcr-1-positive isolates also contained β-lactamase-resistance genes, eight contained qnrS, and 10 contained oqx No mcr-2-positive isolates were identified. The majority of isolates demonstrated a low to moderate level of colistin resistance. Transconjugation was successfully conducted from 14 of the 23 mcr-1-positive isolates, and mcr-1 was identified on plasmids ranging from 60 to 220 kb in these isolates. Conjugation and hybridization experiments revealed that mcr-1 was chromosome-borne in only three isolates. Pulsed-field gel electrophoresis showed that the majority of E. coli isolates belonged to different clonal lineages. Multilocus sequence typing analysis revealed that sequence type 10 (ST10) was the most prevalent, followed by ST181 and ST206. This study demonstrates the utility of enrichment broth culture for identifying environmental mcr-1-positive isolates. Furthermore, it highlights the importance of responsible agriculture and clinical use of polymyxins to prevent further widespread dissemination of polymyxin-resistant pathogens.

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