Genomic characterisation of multi-drug resistant Escherichia coli and Klebsiella pneumoniae co-harbouring mcr-1 and mcr-3 genes on a single plasmid from paediatric clinical cases

Abstract

Emergence of plasmid-born mobile colistin resistance (mcr) gene, a growing concern in healthcare. Therefore, this study aims to genomically characterize multidrug-resistant Escherichia coli and Klebsiella pneumoniae co-harboring the mcr-1 and mcr-3 genes in young children. E. coli (n=3) and K. pneumoniae (n=2) were collected from abdominal secretion and blood, respectively. The isolates were screened using tryptone soya broth (TSB) with 4µl/ml polymyxin-B. Growing bacteria were identified using the VITEK-2 system, MALDI-TOF and 16s RNA sequencing, followed by antibiotic susceptibility testing, as well as metallo-β-lactamase (MBL) and extended spectrum β-lactamase (ESBL) production was detected. Afterwards, strains were subjected to molecular screening targeting mcr variants, and ESBLs/MBL encoding genes. Conjugation, PFGE, southern hybridisation, MLST, and Phylogenic group detection were performed. Plasmid-genome sequencing and bioinformatic analysis were performed. E. coli named EC-19-322, 323, and 331 and K. pneumoniae isolates named KP-19-225 and 226 harboured both mcr-1 and mcr-3 genes. These strains were also found to be resistant to more than three classes of antibiotics. The conjugation experiment revealed the presence of mcr-1 and mcr-3 on the single plasmid and the transmission frequency was 10-2 to 10-3. Both strains were found to be able to produce ESBLs and MBL. E. coli EC-19-322 and 323 were identified as an ST131(O25a:H41), SP-19-331as an ST1577 (O16:H30) and K. pneumoniae as an ST231 (K2). All E. coli strains belonged to phylogenetic group B2, and the results of PFGE supported the MLST findings. This study, reported the co-occurrence of mcr-1 and mcr-3 genes on single plasmid in pathogenic ESBLs/MBL-producing E. coli and K. pneumoniae isolated from young children.