Laboratory Stock Variants of the Archetype Silver Resistance Plasmid pMG101 Demonstrate Plasmid Fusion, Loss of Transmissibility, and Transposition of Tn7/pco/sil Into the Host Chromosome

Abstract

Salmonella Typhimurium carrying the multidrug resistance (MDR) plasmid pMG101 was isolated from three burns patients in Boston United States in 1973. pMG101 was transferrable into other Salmonella spp. and Escherichia coli hosts and carried what was a novel and unusual combination of AMR genes and silver resistance. Previously published short-read DNA sequence of pMG101 showed that it was a 183.5Kb IncHI plasmid, where a Tn7-mediated transposition of pco/sil resistance genes into the chromosome of the E. coli K-12 J53 host strain had occurred. We noticed differences in streptomycin resistance and plasmid size between two stocks of E. coli K-12 J53 pMG101 we possessed, which had been obtained from two different laboratories (pMG101-A and pMG101-B). Long-read sequencing (PacBio) of the two strains unexpectedly revealed plasmid and chromosomal rearrangements in both. pMG101-A is a non-transmissible 383Kb closed-circular plasmid consisting of an IncHI2 plasmid sequence fused to an IncFI/FIIA plasmid. pMG101-B is a mobile closed-circular 154 Kb IncFI/FIIA plasmid. Sequence identity of pMG101-B with the fused IncFI/IncFIIA region of pMG101-A was >99%. Assembled host sequence reads of pMG101-B showed Tn7-mediated transposition of pco/sil into the E. coli J53 chromosome between yhiM and yhiN. Long read sequence data in combination with laboratory experiments have demonstrated large scale changes in pMG101. Loss of conjugation function and movement of resistance genes into the chromosome suggest that even under long-term laboratory storage, mobile genetic elements such as transposons and insertion sequences can drive the evolution of plasmids and host. This study emphasises the importance of utilising long read sequencing technologies of plasmids and host strains at the earliest opportunity.

Global genomic epidemiology, resistome, virulome and plasmidome of the Extraintestinal Pathogenic Escherichia coli (ExPEC) ST38 lineage

“…Interestingly, all pKPC-CAV1321-positive ST38 genomes co-harbored the IncHI2 plasmid. The IncHI2 replicon has been associated with human and animal disease, representing a threat in the context of One Health (Hooton et al, 2021) and, although abundant among Enterobacteriaceae (Wong et al, 2016; Matamoros et al, 2017; Zhao et al, 2018), the IncHI2 had never been reported in ST38 and is rare among other ExPEC strains (Shaik et al, 2017).…”

Laboratory strains of Escherichia coli K-12: things are seldom what they seem

“…Heterogeneity in bacterial laboratory strains and plasmids has been observed many times and we are at a stage when even the same E. coli K-12 stock strains can produce different outcomes, calling reproducibility into question [27,48,[56][57][58][59]. It is clear that there are significant major differences between K-12 and other commensal E. coli strains, and these differences became fixed in the ancestors of the very widely used MG1655 and W3110 strains.…”

Laboratory strains of Escherichia coli K-12: not such perfect role models after all

“…Heterogeneity in bacterial lab strains and plasmids has been observed many times and we are at a stage when even the same E. coli K-12 stock strains can produce different outcomes, calling reproducibility into question (27,48,(56)(57)(58)(59). It is clear that there are significant major differences between K-12 and other commensal E. coli strains, and these differences became fixed in the ancestors of the very widely used MG1655 and W3110…”