Phylogenetic diversity and mutational analysis of New Delhi Metallo-β-lactamase (NDM) producing E. coli strains from pediatric patients in Pakistan

The evolution of NDM genes (bla(NDM)) in E. coli is accounted for expansive multidrug resistance (MDR), causing severe infections and morbidities in the pediatric population. This study aimed to analyze the phylogeny and mutations in NDM variants of E. coli recovered from the pediatric population. Carbapenem-resistant clinical strains of E. coli were identified using microbiological phenotypic techniques. PCR technique used to amplify the bla(NDM) genes, identified on agarose gel, and analyzed by DNA sequencing. The amino acid substitutions were examined for mutations after aligning with wild types. Mutational and phylogenetic analysis was performed using Lasergene, NCBI blastn, Clustal Omega, and MEGA software, whereas PHYRE2 software was used for the protein structure predictions. PCR amplification of the bla(NDM) genes detected 113 clinical strains of E. coli with the contribution of bla(NDM-1) (46%), bla(NDM-4) (3.5%), and bla(NDM-5) (50%) variants. DNA sequencing of bla(NDM) variants showed homology to the previously described bla(NDM-1), bla(NDM-4,) and bla(NDM-5) genes available at GenBank and NCBI database. In addition, the mutational analysis revealed in frame substitutions of Pro60Ala and Pro59Ala in bla(NDM-4) and bla(NDM-5,) respectively. The bla(NDM-1) was ortholog with related sequences of E. coli available at GenBank. The phylogenetic analysis indicated that the NDM gene variants resemble other microbes reported globally with some new mutational sites.