Limited Genetic Diversity of blaCMY-2-Containing IncI1-pST12 Plasmids from Enterobacteriaceae of Human and Broiler Chicken Origin in The Netherlands

Abstract

Distinguishing epidemiologically related and unrelated plasmids is essential to confirm plasmid transmission. We compared IncI1–pST12 plasmids from both human and livestock origin and explored the degree of sequence similarity between plasmids from Enterobacteriaceae with different epidemiological links. Short-read sequence data of Enterobacteriaceae cultured from humans and broilers were screened for the presence of both a blaCMY-2 gene and an IncI1–pST12 replicon. Isolates were long-read sequenced on a MinION sequencer (OxfordNanopore Technologies). After plasmid reconstruction using hybrid assembly, pairwise single nucleotide polymorphisms (SNPs) were determined. The plasmids were annotated, and a pan-genome was constructed to compare genes variably present between the different plasmids. Nine Escherichia coli sequences of broiler origin, four Escherichia coli sequences, and one Salmonella enterica sequence of human origin were selected for the current analysis. A circular contig with the IncI1–pST12 replicon and blaCMY-2 gene was extracted from the assembly graph of all fourteen isolates. Analysis of the IncI1–pST12 plasmids revealed a low number of SNP differences (range of 0–9 SNPs). The range of SNP differences overlapped in isolates with different epidemiological links. One-hundred and twelve from a total of 113 genes of the pan-genome were present in all plasmid constructs. Next generation sequencing analysis of blaCMY-2-containing IncI1–pST12 plasmids isolated from Enterobacteriaceae with different epidemiological links show a high degree of sequence similarity in terms of SNP differences and the number of shared genes. Therefore, statements on the horizontal transfer of these plasmids based on genetic identity should be made with caution.

Global transmission of extended-spectrum cephalosporin resistance in Escherichia coli driven by epidemic plasmids

“…Previous studies have implicated highly similar IncI1 pST3 plasmids as the main mechanism of spread for bla CTX-M-1 in isolates from turkeys 21 and major production animals from Canada, 8 veal calves and humans from France, 26 , 71 food isolates from Germany, 12 wild animals from Portugal, 71 and amongst human, animal and food products in Denmark, 29 Reunion Island 30 and Japan. 28 Similarly, pST12 and pST2 bla CMY-2 subtypes were previously reported in isolates from humans, dogs and poultry in Denmark, 11 , 32 humans and chickens in the Netherlands, 27 humans, poultry and food isolates from Germany, 9 and in dogs in France. 72 , 73 Here we also found these two bla CMY-2 epidemic plasmids in multiple sources and countries.…”

“… 45 , 46 Previous studies using only short-read data reported ESC-R plasmid subtypes, 9 , 14 , 20 and recent studies using long- and short-read data have investigated the degree to which these plasmid subtypes were conserved. 8 , 21 , 27 , 31 However, the majority of these previous studies were limited to a single ESC-R gene or were specific to a particular country and/or source. 8 , 21 , 27 , 28 , 29 , 30 , 31 , 32 , 33 , 34 , 35 Therefore, here we investigated the main ESC-R plasmid types and subtypes at high resolution and at a larger scale, using ESC-R complete genomes from diverse sources and multiple countries.…”

“… 8 , 21 , 27 , 31 However, the majority of these previous studies were limited to a single ESC-R gene or were specific to a particular country and/or source. 8 , 21 , 27 , 28 , 29 , 30 , 31 , 32 , 33 , 34 , 35 Therefore, here we investigated the main ESC-R plasmid types and subtypes at high resolution and at a larger scale, using ESC-R complete genomes from diverse sources and multiple countries. Our approach allowed identification of five dominant and promiscuous IncI1 and IncF plasmid subtypes.…”

“…Despite the recent numbers of genomic studies reporting plasmid sequences harbouring ESC-R genes, 12 , 14 , 26 most are based on short-read data 12 , 14 , 26 and thus, there is often ambiguity as to the genomic location of the ESC-R genes -- plasmid or chromosome -- and complete plasmid identification. The majority of studies using long-read sequences generally focus on particular geographical regions, 21 , 27 , 28 , 29 , 30 , 31 certain ESC-R genes 8 , 32 and/or sources. 33 , 34 , 35 A recent global food products study identified a potential epidemic plasmid, but this was limited to a small number of genomes (n = 7), 36 and a recent review exploring IncI1 ESC-R plasmids in a global context was based on the five genes of the pMLST scheme.…”

Genetic characterization of extended-β-lactamase (ESBL) plasmids captured from dairy manures

“…Although ESBL-producing E. coli carrying only the bla CMY-2 gene was reported previously, there was not a clear explanation for this phenomenon (3). IncI1 plasmids carrying the bla CMY-2 gene were shown to widely spread among E. coli (35), and they shared a high degree of sequence similarity when isolated from Enterobacteriaceae with different epidemiological links (36). Unlike other plasmids carrying ESBL genes in this study, insertion sequences were not located proximal to the bla CMY-2 gene.…”