Evaluation of the Epidemiological Relevance of Variable-Number Tandem-Repeat Genotyping of Mycobacterium bovis and Comparison of the Method with IS 6110 Restriction Fragment Length Polymorphism Analysis and Spoligotyping

Author:

Allix Caroline1,Walravens Karl2,Saegerman Claude3,Godfroid Jacques2,Supply Philip4,Fauville-Dufaux Maryse1

Affiliation:

1. Institut Pasteur de Bruxelles, Laboratoire Tuberculose et Mycobactéries, rue Engeland 642, 1180 Bruxelles, Belgium

2. Centre d'Etude et de Recherches Vétérinaires et Agrochimiques (CERVA-CODA), Section Maladies Bactériennes et Immunologie, Groeselenberg 99, 1180 Bruxelles, Belgium

3. Agence Fédérale pour la Sécurité de la Chaîne Alimentaire, DG Politique de Contrôle, Secrétariat du Comité Scientifique, Avenue Simon Bolivar 30, 1000 Bruxelles, Belgium

4. Institut Pasteur de Lille, Laboratoire des Mécanismes Moléculaires de la Pathogenèse Bactérienne, INSERM U629, 1, rue du Professeur Calmette, BP 245, 59019 Lille Cedex, France

Abstract

ABSTRACT Sources of Mycobacterium bovis contamination remain unclear for many cases of animal and human disease. A major limitation is the lack of sufficiently informative or epidemiologically well evaluated molecular methods for typing. Here, we report an evaluation of a high-throughput method based on 29 mycobacterial interspersed repetitive unit-variable-number tandem-repeat (MIRU-VNTR) loci to genotype 127 M. bovis isolates from cattle from 77 different Belgian farms, representative of a nationwide collection obtained from 1995 to 2003. MIRU-VNTR stability was demonstrated by analyzing a series of 74 isolates in total, obtained from different animals from a single farm or from different farms with an identified epidemiological link. The genotyping results and the genotypic diversity ( h ) were compared with those obtained by IS 6110 restriction fragment length polymorphism (RFLP) analysis and spoligotyping. Among 68 isolates with no known epidemiological link, MIRU-VNTR typing discriminated better than either RFLP analysis or spoligotyping, with isolates taken individually (32 versus 16 and 17 genotypes; h = 0.91 versus 0.73 and 0.85, respectively) or in combination (32 versus 28 genotypes; h = 0.91 versus 0.92). Maximal resolution was already achieved with a subset of 9 loci. The observed congruence of the genetic relationships based on IS 6110 RFLP analysis, spoligotyping, and MIRU-VNTR markers is consistent with a clonal population structure of M. bovis. These results support MIRU-VNTR typing as a convenient and discriminatory technique for analysis of the population structure of M. bovis in much greater detail and for addressing some still unresolved issues in the epidemiology of the pathogen.

Publisher

American Society for Microbiology

Subject

Microbiology (medical)

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