Abstract
Detailed laboratory characterization of Escherichia coli O157 is essential to inform epidemiological investigations. This study assessed the utility of whole-genome sequencing (WGS) for outbreak detection and epidemiological surveillance of E. coli O157, and the data were used to identify discernible associations between genotypes and clinical outcomes. One hundred five E. coli O157 strains isolated over a 5-year period from human fecal samples in Lothian, Scotland, were sequenced with the Ion Torrent Personal Genome Machine. A total of 8,721 variable sites in the core genome were identified among the 105 isolates; 47% of the single nucleotide polymorphisms (SNPs) were attributable to six “atypical” E. coli O157 strains and included recombinant regions. Phylogenetic analyses showed that WGS correlated well with the epidemiological data. Epidemiological links existed between cases whose isolates differed by three or fewer SNPs. WGS also correlated well with multilocus variable-number tandem repeat analysis (MLVA) typing data, with only three discordant results observed, all among isolates from cases not known to be epidemiologically related. WGS produced a better-supported, higher-resolution phylogeny than MLVA, confirming that the method is more suitable for epidemiological surveillance of E. coli O157. A combination of in silico analyses (VirulenceFinder, ResFinder, and local BLAST searches) were used to determine stx subtypes, multilocus sequence types (15 loci), and the presence of virulence and acquired antimicrobial resistance genes. There was a high level of correlation between the WGS data and our routine typing methods, although some discordant results were observed, mostly related to the limitation of short sequence read assembly. The data were used to identify sublineages and clades of E. coli O157, and when they were correlated with the clinical outcome data, they showed that one clade, Ic3, was significantly associated with severe disease. Together, the results show that WGS data can provide higher resolution of the relationships between E. coli O157 isolates than that provided by MLVA. The method has the potential to streamline the laboratory workflow and provide detailed information for the clinical management of patients and public health interventions.
Highlights
Scottish E. coli O157/VTEC Reference Laboratory (SERL), Royal Infirmary of Edinburgh, Edinburgh, Scotlanda; Wellcome Trust Clinical Research Facility (WTCRF), University of Edinburgh, Western General Hospital, Edinburgh, Scotlandb; Centre for Immunity, Infection and Evolution, School of Biological Sciences, University of Edinburgh, Edinburgh, Scotlandc; Gastrointestinal Bacterial Reference Unit, Microbial Services Division, Public Health England, London, United Kingdomd
Sanger sequencing of 792 bp of the mutL gene, which lies within the putative recombinant region, confirmed that the single nucleotide polymorphisms (SNPs) detected by whole-genome sequencing (WGS) were genuine, and a BLAST analysis of this gene region showed that it was identical to the mutL gene of E. coli K-12
This study showed that the identification of core genome SNPs following WGS provided a highly discriminatory method for subtyping of E. coli O157, giving results concordant with the epidemiological data
Summary
Scottish E. coli O157/VTEC Reference Laboratory (SERL), Royal Infirmary of Edinburgh, Edinburgh, Scotlanda; Wellcome Trust Clinical Research Facility (WTCRF), University of Edinburgh, Western General Hospital, Edinburgh, Scotlandb; Centre for Immunity, Infection and Evolution, School of Biological Sciences, University of Edinburgh, Edinburgh, Scotlandc; Gastrointestinal Bacterial Reference Unit, Microbial Services Division, Public Health England, London, United Kingdomd. This study assessed the utility of whole-genome sequencing (WGS) for outbreak detection and epidemiological surveillance of E. coli O157, and the data were used to identify discernible associations between genotypes and clinical outcomes. In Scotland, the epidemiology and clinical outcome of E. coli O157 infection in humans has been closely monitored by Health Protection Scotland (HPS) since the introduction of enhanced surveillance in 1999, following a rapid increase in the number of microbiologically confirmed cases of STEC infection during the mid-1990s. The Scottish Escherichia coli O157/Verotoxigenic E. coli Reference Laboratory (SERL) works closely with Scottish National Health Service (NHS) boards and with HPS and plays a pivotal role in case confirmation and outbreak detection by providing identification and typing services for E. coli O157 and other STEC isolates.
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