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Understanding how multidrug-resistant Enterobacterales (MDRE) are transmitted in low- and middle-income countries (LMICs) is critical for implementing robust policies to curb the increasing burden of antimicrobial resistance (AMR). Here, we analysed samples from surgical site infections (SSIs), hospital surfaces (HSs) and arthropods (summer and winter 2016) to investigate the incidence and transmission of MDRE in a public hospital in Pakistan. We investigated Enterobacterales containing resistance genes (blaCTX-M-15, blaNDM and blaOXA-48-like) for identification, antimicrobial susceptibility testing and whole-genome sequencing. Genotypes, phylogenetic relationships and transmission events for isolates from different sources were investigated using single-nucleotide polymorphism (SNP) analysis with a cut-off of ≤20 SNPs. Escherichia coli (14.3%), Klebsiella pneumoniae (10.9%) and Enterobacter cloacae (16.3%) were the main MDRE species isolated. The carbapenemase gene blaNDM was most commonly detected, with 15.5%, 15.1% and 13.3% of samples positive in SSIs, HSs and arthropods, respectively. SNP (≤20) and spatiotemporal analysis revealed linkages in bacteria between SSIs, HSs and arthropods supporting the One Health approach to underpin infection control policies across LMICs and control AMR. Flies, insects and spiders can serve as vectors of multidrug-resistant Enterobacterales in a public hospital in Pakistan, according to a clinical and molecular epidemiology study.

Understanding how multidrug-resistant Enterobacterales (MDRE) are transmitted in low- and middle-income countries (LMICs) is critical for implementing robust policies to curb the increasing burden of antimicrobial resistance (AMR). Here, we analysed samples from surgical site infections (SSIs), hospital surfaces (HSs) and arthropods (summer and winter 2016) to investigate the incidence and transmission of MDRE in a public hospital in Pakistan. We investigated Enterobacterales containing resistance genes (bla