title: Multiple-locus variable-number tandem repeat analysis for strain discrimination of non-O157 Shiga toxin-producing Escherichia coli.
abstract: Non-O157 Shiga toxin-producing Escherichia coli (STEC) are foodborne pathogens of growing concern worldwide that have been associated with several recent multistate and multinational outbreaks of foodborne illness. Rapid and sensitive molecular-based bacterial strain discrimination methods are critical for timely outbreak identification and contaminated food source traceback. One such method, multiple-locus variable-number tandem repeat analysis (MLVA), is being used with increasing frequency in foodborne illness outbreak investigations to augment the current gold standard bacterial subtyping technique, pulsed-field gel electrophoresis (PFGE). The objective of this study was to develop a MLVA assay for intra- and inter-serogroup discrimination of six major non-O157 STEC serogroups-O26, O111, O103, O121, O45, and O145-and perform a preliminary internal validation of the method on a limited number of clinical isolates. The resultant MLVA scheme consists of ten variable number tandem repeat (VNTR) loci amplified in three multiplex PCR reactions. Sixty-five unique MLVA types were obtained among 84 clinical non-O157 STEC strains comprised of geographically diverse sporadic and outbreak related isolates. Compared to PFGE, the developed MLVA scheme allowed similar discrimination among serogroups O26, O111, O103, and O121 but not among O145 and O45. To more fully compare the discriminatory power of this preliminary MLVA method to PFGE and to determine its epidemiological congruence, a thorough internal and external validation needs to be performed on a carefully selected large panel of strains, including multiple isolates from single outbreaks.
authors.affiliation: National Institute for Microbial Forensics & Food and Agricultural Biosecurity, Department of Entomology and Plant Pathology, Oklahoma State University, Stillwater OK 74078, United States.
authors.forename: Chris
authors.initials: C
authors.lastname: Timmons
authors.affiliation: Centers for Disease Control and Prevention, Atlanta, GA 30329, United States.
authors.forename: Eija
authors.initials: E
authors.lastname: Trees
authors.affiliation: Centers for Disease Control and Prevention, Atlanta, GA 30329, United States.
authors.forename: Efrain M
authors.initials: EM
authors.lastname: Ribot
authors.affiliation: Centers for Disease Control and Prevention, Atlanta, GA 30329, United States.
authors.forename: Peter
authors.initials: P
authors.lastname: Gerner-Smidt
authors.affiliation: Centers for Disease Control and Prevention, Atlanta, GA 30329, United States.
authors.forename: Patti
authors.initials: P
authors.lastname: LaFon
authors.affiliation: Centers for Disease Control and Prevention, Atlanta, GA 30329, United States.
authors.forename: Sung
authors.initials: S
authors.lastname: Im
authors.affiliation: National Institute for Microbial Forensics & Food and Agricultural Biosecurity, Department of Entomology and Plant Pathology, Oklahoma State University, Stillwater OK 74078, United States. Electronic address: li.ma@okstate.edu.
authors.forename: Li Maria
authors.initials: LM
authors.lastname: Ma
mesh_terms: D015373:Bacterial Typing Techniques / Q000379:methods; D004269:DNA, Bacterial / Q000032:analysis* / Q000235:genetics*; D016521:Electrophoresis, Gel, Pulsed-Field; D004927:Escherichia coli Infections / Q000382:microbiology*; D029968:Escherichia coli Proteins / Q000235:genetics; D006801:Humans; D018598:Minisatellite Repeats; D058889:Molecular Typing / Q000379:methods*; D065288:Serogroup; D022621:Shiga Toxin / Q000096:biosynthesis / Q000235:genetics; D054323:Shiga-Toxigenic Escherichia coli / Q000145:classification / Q000235:genetics* / Q000302:isolation & purification
chemical_list: D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; D022621:Shiga Toxin
keywords: Escherichia coli O157; MLVA; Non-O157 STEC; Outbreak investigation; Strain discrimination