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Journal of Clinical Microbiology, June 2002, p. 2126-2133, Vol. 40, No. 6
0095-1137/02/$04.00+0     DOI: 10.1128/JCM.40.6.2126-2133.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Development of Variable-Number Tandem Repeat Typing of Mycobacterium bovis: Comparison of Results with Those Obtained by Using Existing Exact Tandem Repeats and Spoligotyping

Solvig Roring,1* Alistair Scott,1 David Brittain,2 Ian Walker,2 Glyn Hewinson,3 Sydney Neill,2 and Robin Skuce2

Department of Veterinary Science, Queen's University Belfast,1 Veterinary Sciences Division, Department of Agriculture and Rural Development for Northern Ireland, Stormont, Belfast, Northern Ireland,2 Veterinary Laboratories Agency, Weybridge, New Haw, Surrey, England3

Received 3 December 2001/ Returned for modification 16 February 2002/ Accepted 21 March 2002

Various genetic markers have been exploited for fingerprinting the Mycobacterium tuberculosis complex (MTBC) in molecular epidemiological studies, mainly through identifying restriction fragment length polymorphisms (RFLP). In large-scale studies, RFLP typing has practical processing and analysis limitations; therefore, attempts have been made to move towards PCR-based typing techniques. Spoligotyping (spacer oligotyping) and, more recently, variable-number tandem repeat (VNTR) typing have provided PCR-derived typing techniques. This study describes the identification and characterization of novel VNTR loci, consisting of tandem repeats in the size range of 53 to 59 bp in the MTBC, and their assessment as typing tools in 47 Mycobacterium bovis field isolates and nine MTBC strains. Spoligotyping and the previously described set of exact tandem repeats (ETRs) (R. Frothingham and W. A. Meeker-O'Connell, Microbiology 144:1189-1196, 1998) were also applied to the same panel of isolates. The allelic diversity of the individual VNTR loci was calculated, and a comparison of the novel VNTRs was made against the results obtained by spoligotyping and the existing set of ETRs. Eleven unique spoligotypes were discriminated in the panel of 47 M. bovis isolates. Greater resolution was obtained through the combination of the most-discriminating VNTRs from both sets. Considerable discrimination was achieved, with the 47 M. bovis isolates resolved into 14 unique profiles, while all nine MTBC isolates were uniquely differentiated. The novel VNTR markers described increased the discrimination possible in strain typing of M. bovis, with the added benefit of an intuitive digital nomenclature, with the allele copy number of the individual VNTRs providing a profile. VNTR typing was shown to be a valuable technique with great potential for further development and application to epidemiological tracing of tuberculosis transmissions.

* Corresponding author. Mailing address: Department of Veterinary Science, Queen's University Belfast (QUB), Stoney Rd., Stormont, Belfast BT4 3SD, Northern Ireland. Phone: 44 28 90525719. Fax 44 28 90525745. E-mail: S.Roring{at}qub.ac.uk.

Journal of Clinical Microbiology, June 2002, p. 2126-2133, Vol. 40, No. 6
0095-1137/02/$04.00+0     DOI: 10.1128/JCM.40.6.2126-2133.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



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