High-throughput multiplex MIRU-VNTR typing of Mycobacterium bovis

Spoligotyping is the most widely used method for genotyping Mycobacterium bovis (M. bovis). However, its discriminatory power varies widely between countries. MIRU-VNTR typing could be a promising alternative, although it generally requires the time consuming and laborious simplex PCR assays using s...

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Bibliographic Details
Published in:Research in veterinary science 2014-06, Vol.96 (3), p.422-425
Main Authors: Navarro, Y., Herranz, M., Romero, B., Bouza, E., Domínguez, L., de Juan, L., García-de-Viedma, D.
Format: Article
Language:English
Subjects:
Animals
DNA, Bacterial - chemistry
DNA, Bacterial - genetics
Genotype
Genotyping
Minisatellite Repeats - genetics
MIRU-VNTRtyping
Multiplex
Mycobacterium bovis
Mycobacterium bovis - genetics
Polymerase Chain Reaction - methods
Polymerase Chain Reaction - veterinary
Spain
Tuberculosis - genetics
Tuberculosis - veterinary
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Summary:Spoligotyping is the most widely used method for genotyping Mycobacterium bovis (M. bovis). However, its discriminatory power varies widely between countries. MIRU-VNTR typing could be a promising alternative, although it generally requires the time consuming and laborious simplex PCR assays using standard agarose gel electrophoresis. The accuracy of this approach depends on good standardization and a certain degree of expertise. This study presents a version of MIRU-VNTR based on three triplex PCRs with automatic allelic assignation of the products analyzed in capillary electrophoresis. The technique was prospectively applied to 44 M. bovis and two Mycobacterium caprae (M. caprae) isolates, and 22 different MIRU-VNTRtypes were obtained; with spoligotyping, only 14 different types were obtained. The proposal makes it possible to shorten response times, automate procedures, and increase accuracy, thus minimizing errors in assigning genotypes. It would enable the switch from a standard limited method of genotyping M. bovis to a high-throughput discriminatory fingerprinting approach.
ISSN:0034-5288
1532-2661
DOI:10.1016/j.rvsc.2014.03.012