Molecular Epidemiology of mcr-1, bla KPC-2, and bla NDM-1 Harboring Clinically Isolated Escherichia coli from Pakistan.

The multiple-drug resistant Escherichia coli are among the deadliest pathogens causing life-threatening infections. This study was planned to determine the molecular epidemiology of mcr-1, bla KPC-2, and bla NDM-1 harboring clinically isolated E. coli from Pakistan. In total, 545 strains of E. coli from clinical samples were collected from June 2018 to September 2019. All the isolates were screened for colistin-resistance, extended-spectrum-β-lactamases (ESBL), and carbapenemases through the micro-dilution method, Double-Disk-Synergy-Test (DDST), and Modified-Hodge-Test (MHT). The detection, sequence-typing, conjugal transfer, S1-PFGE, plasmid-replicon-typing, and southern-blotting for mcr, ESBL, and carbapenemase-encoding genes were performed. A total of four (0.73%) colistin-resistant strains carrying alongside mcr-1 and bla CTX-M-15 genes, three of these strains also had the bla TEM-1 gene. The presence of ESBL genes was detected in 139 (25.5%) isolates harboring bla CTXM-15 (74.82%), bla TEM (34.53%), bla SHV (28.06%) and bla OXA-1 (28.78%). In 129 carbapenemase-producers, 35.83% possessed bla NDM-1, 26.67% bla KPC-2, 8.3% bla OXA-48, 25% bla VIM-1, and 20.83% bla IMP-1 genes. The sequence typing revealed that mcr-1 harboring isolates belonged to ST405, ST117, and ST156. Fifty percent of bla KPC-2 and 48.83% of bla NDM-1 were found on ST131 and ST1196, respectively. Two rare types of STs, ST7584, and ST8671 were also identified in this study. The mcr-1 gene was located on Incl2 (60-kb) plasmid. The bla KPC-2 was present on (140-kb) IncH12, (100-kb) IncN, (90-kb) Incl1, while bla NDM-1 was located on (70-kb) IncFIIK, (140-kb) IncH12, (100-kb) IncN, (60-kb) IncA/C, and (45-kb) IncFII plasmids, which were successfully trans-conjugated. Among the plasmid types, the Incl1 carrying bla KPC-2, IncH12 harboring bla KPC-2 and bla NDM-1, and IncFIIK carrying bla NDM-1 were for the first time detected in Pakistan. The mcr-1, bla KPC-2, and bla NDM-1 genes finding in various clonal and plasmids types indicate that a substantial selection of the resistance genes had occurred in our clinical strains.