Multi-locus variable number tandem repeat analysis for Escherichia coli causing extraintestinal infections

https://doi.org/10.1016/j.mimet.2009.09.006Get rights and content

Abstract

Discriminatory genotyping methods for the analysis of Escherichia coli other than O157:H7 are necessary for public health-related activities. A new multi-locus variable number tandem repeat analysis protocol is presented; this method achieves an index of discrimination of 99.5% and is reproducible and valid when tested on a collection of 836 diverse E. coli.

Introduction

Discriminatory molecular typing methods for non-O157:H7 Escherichia coli are necessary for outbreak investigation, public health surveillance, and molecular epidemiological studies. Pulsed-field gel electrophoresis (PFGE) is the current standard for molecular subtyping, but it is time-consuming, and the results can be difficult to compare across laboratories. Multi-locus variable number tandem repeat analysis (MLVA) has shown promise in differentiating closely related strains of E. coli (Keys et al., 2005, Lindstedt et al., 2004, Noller et al., 2003, Noller et al., 2004). However, these MLVA protocols have been developed specifically for E. coli O157:H7. The objective of this study was to develop a fast and reliable MLVA protocol which could be used alone or in conjunction with other typing methods such as enterobacterial repetitive intergenic consensus 2 (ERIC2) sequence PCR, to genotype large collections of E. coli.

Section snippets

E. coli isolates

E. coli isolates from three sources were included in the study: 417 isolates were from retail chicken, pork, and beef meat that were systematically selected from the Canadian Integrated Program on Antimicrobial Resistance Surveillance (CIPARS) collection; 74 E. coli isolates were from restaurant and ready-to-eat food items collected by the Division de l'inspection des aliments, Ville de Montréal; and 345 E. coli isolates were from human extraintestinal infections. E. coli was grown in

Results and conclusions

An MLVA method was developed to rapidly and unambiguously group diverse E. coli isolates. A total of 803 diverse E. coli and 33 E. coli isolates belonging to known clonal groups were analyzed with the new MLVA method. All 836 isolates were typeable by MLVA. The average call rates for each locus are presented in Table 1. Sequencing results confirmed the presence of tandem repeats at each locus.

Evaluation of the 803 diverse E. coli identified 303 isolates (38%) with unique MLVA profiles and 121

Acknowledgments

We wish to thank members of the surveillance team of the Canadian Integrated Program for Antimicrobial Resistance Surveillance (Lucie Dutil and Danielle Daignault); the Division de l'inspection des aliments, Ville de Montréal (Myrto Mantzavrakos and Annie Laviolette) and the Student Health Services Clinical Technician (Christiaine Lacombe). This study was supported by the Public Health Agency of Canada (A.R.M) and by the Canadian Institutes of Health Research, M.Sc. Award (C.V.).

References (19)

  • B.A. Lindstedt et al.

    Multiple-locus variable-number tandem-repeats analysis of Escherichia coli O157 using PCR multiplexing and multi-colored capillary electrophoresis

    J. Microbiol. Methods

    (2004)
  • J.B. Bender et al.

    Surveillance for Escherichia coli O157:H7 infections in Minnesota by molecular subtyping

    N. Engl. J. Med.

    (1997)
  • M.J. Brownstein et al.

    Modulation of non-templated nucleotide addition by Taq DNA polymerase: primer modifications that facilitate genotyping

    Biotechniques

    (1996)
  • Danoff-Burg, J., Xu, C. Biodiversity Calculator. http://www.columbia.edu/itc/cerc/danoff-burg/MBD_Links.html . 2009....
  • H. Grundmann et al.

    Determining confidence intervals when measuring genetic diversity and the discriminatory abilities of typing methods for microorganisms

    J. Clin. Microbiol.

    (2001)
  • P.R. Hunter

    Reproducibility and indices of discriminatory power of microbial typing methods

    J. Clin. Microbiol.

    (1990)
  • P.R. Hunter et al.

    Numerical index of the discriminatory ability of typing systems: an application of Simpson's Index of Diversity

    J. Clin. Microbiol.

    (1988)
  • J.R. Johnson et al.

    Improved repetitive-element PCR fingerprinting for resolving pathogenic and nonpathogenic phylogenetic groups within Escherichia coli

    Clin. Diagn. Lab. Immunol.

    (2000)
  • C. Keys et al.

    Highly diverse variable number tandem repeat loci in the E. coli O157:H7 and O55:H7 genomes for high-resolution molecular typing

    J. Appl. Microbiol.

    (2005)
There are more references available in the full text version of this article.

Cited by (8)

View all citing articles on Scopus

Ethics committee approval: The study protocol was approved by the McGill University, Institutional Review Board (A01-M04-05A).

View full text