Computational protocol: Characterization of oqxAB in Escherichia coli Isolates from Animals, Retail Meat, and Human Patients in Guangzhou, China

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Protocol publication

[…] The genetic diversity of 50 randomly selected oqxAB-positive E. coli isolates from animals (n = 21), food products (n = 9), and patients (n = 20) was characterized by multilocus sequence typing (MLST; and PFGE (Gautom, ). MLST data were aligned using ClustalW, and a phylogenetic tree was constructed by the neighbor joining algorithm using MEGA 7.0 (Kumar et al., ). PFGE patterns were compared and analyzed using BioNumerics (Applied Maths, Sint-Martens-Latem, Belgium) using a cut-off value of 90% to define PFGE clusters. [...] Two IncHI2 plasmids, namely, pHNLDH400 from a patient with E. coli colonization and pHNYJC8 from chicken meat, were selected for sequencing. Plasmid DNA was purified from the transformants using QIAGEN® Plasmid Midi Kit according to the manufacturer's instructions (Qiagen, Hilden, Germany), and was completely sequenced by PacBio single-molecule real-time sequencing (RSII platform) (Pacific Biosciences, Menlo Park, CA, USA). Raw reads were introduced into the non-hybrid Hierarchical Genome Assembly Process. Analysis and annotation of the resulting sequences were performed using RAC (, ISfinder (, BLAST (, RAST server (Aziz et al., ), and the Gene Construction Kit 4.0 (Textco BioSoftware, Inc., Raleigh, NC, USA). Plasmids pHNLDF400 and pHNYJC8 were compared with similar oqxAB-bearing IncHI2 plasmids pHXY0908 and pHK0653 obtained from S. Typhimurium, as well as our previously reported plasmids pHNSHP45-2 (Zhi et al., ), and pHNAH67 from E. coli isolate AHC67 (Yang X. et al., ) using BLASTn and BRIG (Alikhan et al., ). The IncHI2 plasmid R478 (GenBank accession number BX664015) served as the reference plasmid for annotation. […]

Pipeline specifications

Software tools Clustal W, MEGA, BioNumerics, RAST, BLASTN, BRIG
Databases ISfinder InChI
Applications Phylogenetics, Genome data visualization
Organisms Escherichia coli, Homo sapiens, Sus scrofa, Gallus gallus, Salmonella enterica subsp. enterica serovar Typhimurium
Diseases Colonic Neoplasms