title: Comparative Evaluation of Genomic and Laboratory Approaches for Determination of Shiga Toxin Subtypes in Escherichia coli.
abstract: The determination of Shiga toxin (ST) subtypes can be an important element in the risk characterization of foodborne ST-producing Escherichia coli (STEC) isolates for making risk management decisions. ST subtyping methods include PCR techniques based on electrophoretic or pyrosequencing analysis of amplicons and in silico techniques based on whole genome sequence analysis using algorithms that can be readily incorporated into bioinformatics analysis pipelines for characterization of isolates by their genetic composition. The choice of technique will depend on the performance characteristics of the method and an individual laboratory's access to specialized equipment or personnel. We developed two whole genome sequence-based ST subtyping tools: (i) an in silico PCR algorithm requiring genome assembly to replicate a reference PCR-based method developed by the Statens Serum Institut (SSI) and (ii) an assembly-independent routine in which raw sequencing results are mapped to a database of known ST subtype sequence variants (V-Typer). These tools were evaluated alongside the SSI reference PCR method and a recently described PCR-based pyrosequencing technique. The V-Typer method results corresponded closely with the reference method in the analysis of 67 STEC cultures obtained from a World Health Organization National Reference Laboratory. In contrast, the in silico PCR method failed to detect ST subtypes in several cases, a result which we attribute to assembly-induced errors typically encountered with repetitive gene sequences. The V-Typer can be readily integrated into bioinformatics protocols used in the identification and characterization of foodborne STEC isolates.
authors.affiliation: Food Microbiology Research Team, Canadian Food Inspection Agency, 960 Carling Avenue, Building 22, Central Experimental Farm, Ottawa, Ontario, Canada K1A 0C6.
authors.forename: Catherine D
authors.initials: CD
authors.lastname: Carrillo
authors.affiliation: Food Microbiology Research Team, Canadian Food Inspection Agency, 960 Carling Avenue, Building 22, Central Experimental Farm, Ottawa, Ontario, Canada K1A 0C6.
authors.forename: Adam G
authors.initials: AG
authors.lastname: Koziol
authors.affiliation: Food Microbiology Research Team, Canadian Food Inspection Agency, 960 Carling Avenue, Building 22, Central Experimental Farm, Ottawa, Ontario, Canada K1A 0C6.
authors.forename: Amit
authors.initials: A
authors.lastname: Mathews
authors.affiliation: Food Microbiology Research Team, Canadian Food Inspection Agency, 960 Carling Avenue, Building 22, Central Experimental Farm, Ottawa, Ontario, Canada K1A 0C6.
authors.forename: Noriko
authors.initials: N
authors.lastname: Goji
authors.affiliation: Food Microbiology Research Team, Canadian Food Inspection Agency, 960 Carling Avenue, Building 22, Central Experimental Farm, Ottawa, Ontario, Canada K1A 0C6.
authors.forename: Dominic
authors.initials: D
authors.lastname: Lambert
authors.affiliation: Food Microbiology Research Team, Canadian Food Inspection Agency, 960 Carling Avenue, Building 22, Central Experimental Farm, Ottawa, Ontario, Canada K1A 0C6.
authors.forename: George
authors.initials: G
authors.lastname: Huszczynski
authors.affiliation: Food Microbiology Research Team, Canadian Food Inspection Agency, 960 Carling Avenue, Building 22, Central Experimental Farm, Ottawa, Ontario, Canada K1A 0C6.
authors.forename: Martine
authors.initials: M
authors.lastname: Gauthier
authors.affiliation: Food Microbiology Research Team, Canadian Food Inspection Agency, 960 Carling Avenue, Building 22, Central Experimental Farm, Ottawa, Ontario, Canada K1A 0C6.
authors.forename: Kingsley
authors.initials: K
authors.lastname: Amoako
authors.affiliation: Food Microbiology Research Team, Canadian Food Inspection Agency, 960 Carling Avenue, Building 22, Central Experimental Farm, Ottawa, Ontario, Canada K1A 0C6.
authors.forename: Burton W
authors.initials: BW
authors.lastname: Blais
mesh_terms: D004927:Escherichia coli Infections / Q000175:diagnosis*; D029968:Escherichia coli Proteins / Q000235:genetics; D023281:Genomics; D006801:Humans; D022621:Shiga Toxin / Q000235:genetics*; D022622:Shiga Toxin 1 / Q000235:genetics; D022641:Shiga Toxin 2 / Q000235:genetics; D054323:Shiga-Toxigenic Escherichia coli / Q000302:isolation & purification
chemical_list: D029968:Escherichia coli Proteins; D022622:Shiga Toxin 1; D022641:Shiga Toxin 2; D022621:Shiga Toxin
keywords: Genomics tool; Shiga toxin subtype; Shiga toxin–producing Escherichia coli