Computational protocol: Whole Genome Sequencing for Public Health Surveillance of Shiga Toxin Producing Escherichia coli Other than Serogroup O157

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Protocol publication

[…] All 167 strains of non-O157 STEC isolated during 2014 were phenotypically serotyped by the agglutination of antibodies raised in rabbits to the lipopolysaccharide O antigen and to the flagella H antigen (Gross and Rowe, ). Real-time PCR targeting stx1 and stx2 and the stx subtyping PCR was performed as previously described (Persson et al., ; Jenkins et al., ).Genomic DNA extracted from 140 of the 167 strains of non-O157 STEC was fragmented and tagged for multiplexing with Nextera XT DNA Sample Preparation Kits (Illumina) and sequenced using the Illumina HiSeq 2500. A reference database, SerotypeFinder, containing the gene sequences encoding the 180 O antigen groups (wzx, wzy, wzm, and wzt) and the 53 H antigens (fliC, flkA, fllA, flmA, and flnA) was constructed and developed by Joensen et al. (). Using the GeneFinder tool (Doumith unpublished), FASTQ reads were mapped to the genes in the SerotypeFinder database using Bowtie 2 (Langmead and Salzberg, ) and the best match to each of the O and H determinants was reported with metrics including coverage, depth, mixture and homology in an XML format for quality assessment. Only in silico predictions of serotype that matched to a gene determinant at >80% nucelotide identity over >80% length were accepted. Stx subtyping was performed as described by Ashton et al. (). FASTQ sequences were deposited in the National Center for Biotechnology Information Short Read Archive under the bioproject PRJNA248064. […]

Pipeline specifications

Software tools SerotypeFinder, Bowtie
Organisms Escherichia coli, Homo sapiens
Diseases Gastrointestinal Diseases, Infection, Sprains and Strains, Genomic Instability