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United States Department of Agriculture

Agricultural Research Service

Research Project: EXPLORING GENOMIC DIFFERENCES AND ECOLOGICAL RESERVOIRS TO CONTROL FOODBORNE PATHOGENS

Location: Meat Safety & Quality Research

Title: Phylogenetic classification of Escherichia coli O26 strains from human, animals, and environmental origins using nucleotide polymorphisms

Authors
item Norman, Keri -
item Clawson, Michael
item Strockbine, Nancy -
item Mandrell, Robert
item Johnson, Roger -
item Ziebell, Kim -
item Zhao, Shaohua -
item Fratamico, Pina
item Stones, Robert -
item Allard, Marc -
item Bono, James

Submitted to: American Society for Microbiology General Meeting
Publication Type: Abstract Only
Publication Acceptance Date: February 20, 2013
Publication Date: May 19, 2013
Citation: Norman, K.N., Clawson, M.L., Strockbine, N.A., Mandrell, R.E., Johnson, R., Ziebell, K., Zhao, S., Fratamico, P.M., Stones, R., Allard, M., Bono, J.L. 2013. Phylogenetic classification of Escherichia coli O26 strains from human, animals, and environmental origins using nucleotide polymorphisms. American Society for Microbiology General Meeting. Poster No. 484 p. 40.

Technical Abstract: Background: Shiga toxin-producing Escherichia coli (STEC) O26 strains are food-borne pathogens that were recently classified as adulterants in certain beef products. Little is known about their genetic diversity, including whether or not phylogenetic subtypes within the serogroup vary in their associations with human disease, animals, or the environment. The objectives of this study were to identify nucleotide polymorphisms in a diverse sampling of human, animal and environmental STEC O26 strains that could be used to classify STEC O26 phylogenetic subtypes, and to test the subtypes for associations with source origins and human virulence traits. Materials: DNA was extracted from a discovery panel of 189 epidemiologically-unlinked STEC O26 strains; of which 106 were isolated from humans and 83 were isolated from animals or the environment. Separate DNA pools from human and animal/environmental isolates were created and sequenced on a Roche 454 GS FLX sequencer to a level of 1.3X genome coverage. Polymorphisms were mapped to a reference sequence for STEC O26 and identified using Geneious software (Biomatters Ltd, Auckland, New Zealand). Select polymorphisms defined by frequency and genomic position within conserved regions of the genome were validated and used to genotype a panel of strains consisting of 243 STEC O26, 52 non-STEC O26, and 89 other STEC strains by high-throughput multiplexing assays using the MassARRAY assay design software and iPLEX Gold chemistry on a MassARRAY genotyping system (Sequenom, Inc., San Diego, CA). Results: Sequencing of the pooled DNAs identified 30,850 putative polymorphisms. From those, 283 validated polymorphisms defined 64 unique genotypes, each tagged by a minimum of 43 polymorphisms, among the 295 O26 strains tested. Phylogenetic trees of the genotypes were divided by clades that correlated with the presence or absence of Shiga toxin gene(s), a human virulence determinant. No correlation between genotype and strain source was observed. Additionally, strains with the STEC H11 flagellar antigen group were closely related regardless of their O serogroup. Conclusions: Nucleotide polymorphism discovery of pooled DNA from STEC O26 strains identified an informative set of polymorphisms to classify STEC O26 strains according to their genetic relatedness.

Last Modified: 12/18/2014
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