title: Real-time whole-genome sequencing for routine typing, surveillance, and outbreak detection of verotoxigenic Escherichia coli.
abstract: Fast and accurate identification and typing of pathogens are essential for effective surveillance and outbreak detection. The current routine procedure is based on a variety of techniques, making the procedure laborious, time-consuming, and expensive. With whole-genome sequencing (WGS) becoming cheaper, it has huge potential in both diagnostics and routine surveillance. The aim of this study was to perform a real-time evaluation of WGS for routine typing and surveillance of verocytotoxin-producing Escherichia coli (VTEC). In Denmark, the Statens Serum Institut (SSI) routinely receives all suspected VTEC isolates. During a 7-week period in the fall of 2012, all incoming isolates were concurrently subjected to WGS using IonTorrent PGM. Real-time bioinformatics analysis was performed using web-tools (www.genomicepidemiology.org) for species determination, multilocus sequence type (MLST) typing, and determination of phylogenetic relationship, and a specific VirulenceFinder for detection of E. coli virulence genes was developed as part of this study. In total, 46 suspected VTEC isolates were characterized in parallel during the study. VirulenceFinder proved successful in detecting virulence genes included in routine typing, explicitly verocytotoxin 1 (vtx1), verocytotoxin 2 (vtx2), and intimin (eae), and also detected additional virulence genes. VirulenceFinder is also a robust method for assigning verocytotoxin (vtx) subtypes. A real-time clustering of isolates in agreement with the epidemiology was established from WGS, enabling discrimination between sporadic and outbreak isolates. Overall, WGS typing produced results faster and at a lower cost than the current routine. Therefore, WGS typing is a superior alternative to conventional typing strategies. This approach may also be applied to typing and surveillance of other pathogens.
authors.affiliation: National Food Institute, Division for Epidemiology and Microbial Genomics, Technical University of Denmark, Kongens Lyngby, Denmark.
authors.forename: Katrine Grimstrup
authors.initials: KG
authors.lastname: Joensen
authors.forename: Flemming
authors.initials: F
authors.lastname: Scheutz
authors.forename: Ole
authors.initials: O
authors.lastname: Lund
authors.forename: Henrik
authors.initials: H
authors.lastname: Hasman
authors.forename: Rolf S
authors.initials: RS
authors.lastname: Kaas
authors.forename: Eva M
authors.initials: EM
authors.lastname: Nielsen
authors.forename: Frank M
authors.initials: FM
authors.lastname: Aarestrup
mesh_terms: D018829:Adhesins, Bacterial / Q000235:genetics; D015373:Bacterial Typing Techniques / Q000379:methods; D003718:Denmark; D004196:Disease Outbreaks; D004927:Escherichia coli Infections / Q000175:diagnosis* / Q000382:microbiology; D029968:Escherichia coli Proteins / Q000235:genetics; D016680:Genome, Bacterial / Q000235:genetics; D055106:Genome-Wide Association Study / Q000379:methods; D010802:Phylogeny; D017422:Sequence Analysis, DNA / Q000379:methods; D022622:Shiga Toxin 1 / Q000235:genetics; D022641:Shiga Toxin 2 / Q000235:genetics; D054323:Shiga-Toxigenic Escherichia coli / Q000235:genetics*; D014774:Virulence / Q000235:genetics
chemical_list: D018829:Adhesins, Bacterial; D029968:Escherichia coli Proteins; D022622:Shiga Toxin 1; D022641:Shiga Toxin 2; C430449:verocytotoxin 2; C073819:eaeA protein, E coli