Abstract
The aim of this study was to use single-molecule, nanopore sequencing to explore the genomic environment of the resistance determinants in a multidrug-resistant (MDR) strain of enteroaggregative Escherichia coli serotype O51 : H30, sequence type (ST) 38. Sequencing was performed on the MinION Flow cell MIN-106 R9.4. Nanopore raw FAST5 reads were base-called using Albacore v1.2.1, converted to FASTA and FASTQ formats using Poretools v0.6.0, and assembled using Unicycler v0.4.2, combining the long-read sequencing data with short-read data produced by Illumina sequencing. The genome was interrogated against an antimicrobial resistance (AMR) gene reference database using blast. The majority of the 12 AMR determinants identified were clustered together on the chromosome at three separate locations flanked by integrases and/or insertion elements [region 1 –catA, bla OXA-1, aac(6′)-Ib-cr, tetA and bla CTX-M-15; region 2 – dfrA1 and aadA1; region 3 – catA, bla TEM-1, tetA and sul2]. AMR determinants located outside these three regions were a chromosomally encoded bla CMY-16, mutations in gyrA and parC, and two plasmid-encoded AMR determinants, bla OXA-181 and qnrS1 located on the same IncX3 plasmid. Long-read analysis of whole genome sequencing data identified mobile genetic elements on which AMR determinants were located and revealed the combination of different AMR determinants co-located on the same mobile element. These data contribute to a better understanding of the transmission of co-located AMR determinants in MDR E. coli causing gastrointestinal and extra-intestinal infections.
Highlights
Enteroaggregative Escherichia coli (EAEC) cause a range of gastrointestinal symptoms including acute or persistent, watery or mucoid diarrhoea, often accompanied by severe abdominal pain [1]
In 2017, we identified a strain of EAEC serotype O51 : H30 ST38, isolated from a patient who developed persistent diarrhoea shortly before returning to the UK from Pakistan, that had 12 antimicrobial resistance (AMR) genetic determinants including blaOXA-181, blaCMY-16, blaCTX-M-15, blaTEM-1, blaOXA-1, aadA1b, aac(6¢)-Ib-cr, gyrA [83 : S-L];parC[80 : S-I;84 : E-V], qnrS1, dfrA1, tetA, sul2 and catA [2]
Long-read analysis of whole genome sequencing (WGS) data enabled us to determine the mobile genetic elements on which the key AMR determinants are located, and to characterize the combination of different AMR determinants co-located on the same mobile element
Summary
Enteroaggregative Escherichia coli (EAEC) cause a range of gastrointestinal symptoms including acute or persistent, watery or mucoid diarrhoea, often accompanied by severe abdominal pain [1]. The implementation of whole genome sequencing (WGS) at Public Health England (PHE) has improved surveillance of antimicrobial resistance (AMR) of gastrointestinal pathogens [4]. Analysis of these surveillance data indicates that the incidence of AMR in strains of E. coli belonging to the EAEC pathotype is high in comparison with other diarrhoeagenic E. coli pathotypes [2]. In 2017, we identified a strain of EAEC serotype O51 : H30 ST38, isolated from a patient who developed persistent diarrhoea shortly before returning to the UK from Pakistan, that had 12 AMR genetic determinants including blaOXA-181, blaCMY-16, blaCTX-M-15, blaTEM-1, blaOXA-1, aadA1b, aac(6¢)-Ib-cr, gyrA [83 : S-L];parC[80 : S-I;84 : E-V], qnrS1, dfrA1, tetA, sul and catA [2]. The isolate was confirmed phenotypically by antimicrobial susceptibility testing to be resistant to the third-generation cephalosporins, ciprofloxacin, streptomycin, trimethoprim, tetracycline, sulphonamide and chloramphenicol [2]
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