The New Delhi Metallo-β-lactamase (NDM) is among the most threatening forms of carbapenemases produced by K. pneumoniae, well-known to cause severe worldwide infections. The molecular epidemiology of bla_NDM-1-harboring K. pneumoniae is not well elucidated in Pakistan. Herein, we aim to determine the antibiotics-resistance profile, genes type, molecular type, and plasmid analysis of 125 clinically isolated K. pneumoniae strains from urine samples during July 2018 to January 2019 in Pakistan. A total of 34 (27.2%) K. pneumoniae isolates were carbapenemases producers, and 23 (18.4%) harbored the bla_NDM-1 gene. The other carbapenemases encoding genes, i.e., bla_IMP-1 (7.2%), bla_VIM-1 (3.2%), and bla_OXA-48 (2.4%) were also detected. The Multi Locus Sequence Typing (MLST) results revealed that all bla_NDM-1-harboring isolates were ST11. The other sequence types detected were ST1, ST37, and ST105. The cluster analysis of Xbal Pulsed Field Gel Electrophoresis (PFGE) revealed variation amongst the clusters of the identical sequence type isolates. The bla_NDM-1 gene in all of the isolates was located on a 45-kb IncX3 plasmid, successfully transconjugated. For the first time, bla_NDM-1-bearing IncX3 plasmids were identified from Pakistan, and this might be a new primary vehicle for disseminating bla_NDM-1 in Enterobacteriaceae as it has a high rate of transferability.

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