Escherichia coli strain:K-12 J53 Genome sequencing. Escherichia coli strain:K-12 J53

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.Salmonella Typhimurium carrying the multidrug resistance (MDR) plasmid pMG101, was isolated from three burns patients in Boston USA in 1973. pMG101 was transferrable into other Salmonella 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 183.5 Kb 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 we had obtained from two different laboratories. We sequenced the strains to identify whether they were in fact identical. Long-read sequencing (PacBio) of the two strains shows significant differences between them. pMG101-A is a non-transmissible 383 Kb closed-circular plasmid consisting of an IncHI2 plasmid sequence fused to an IncFI/FIIA plasmid. pMG101-B is a mobile closed-circular 168 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 has demonstrated large scale changes in pMG101, loss of resistance and conjugation function, and movement of resistance genes into the chromosome- similar to plasmid variations seen in recently isolated plasmids- suggesting that changes in the plasmids in the laboratory strains are similar to those that occur naturally. This study emphasises the importance of utilising long read sequencing technologies of plasmids and host strains at the earliest opportunity.