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Journal of Clinical Microbiology, January 1998, p. 168-178, Vol. 36, No. 1
0095-1137/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.

Evaluation of Four DNA Typing Techniques in Epidemiological Investigations of Bovine Tuberculosis

Debby Cousins,1,* Suzette Williams,1 Ernesto Liébana,2 Alicia Aranaz,2 Annelies Bunschoten,3 Jan Van Embden,3 and Trevor Ellis1

Australian Reference Laboratory for Bovine Tuberculosis, Animal Health Laboratories, Agriculture Western Australia, South Perth, Western Australia 6151, Australia1; Departamento de Patologa Animal I (Sanidad Animal), Facultad de Veterinaria, Universidad Complutense de Madrid, s/n, 28100 Madrid, Spain2; and Department of Bacteriology, Research Laboratory for Infectious Diseases, National Institute of Public Health and Environmental Protection, 3720A Bilthoven, The Netherlands3

Received 22 April 1997/Returned for modification 8 July 1997/Accepted 1 October 1997

DNA fingerprinting techniques were used to type 273 isolates of Mycobacterium bovis from Australia, Canada, the Republic of Ireland, and Iran. The results of restriction fragment length polymorphism (RFLP) analysis with DNA probes from IS6110, the direct repeat (DR), and the polymorphic GC-rich sequence (PGRS) were compared with those of a new PCR-based method called spacer oligonucleotide typing (spoligotyping) developed for the rapid typing of Mycobacterium tuberculosis (J. Kamerbeek et al., J. Clin. Microbiol. 35:907-914, 1997). Eighty-five percent of the isolates harbored a single copy of IS6110, and 81.5% of these carried IS6110 on the characteristic 1.9-kb restriction fragment. RFLP analysis with IS6110 identified 23 different types, RFLP analysis with the DR probe identified 35 types, RFLP analysis with the PGRS probe identified 77 types, and the spoligotyping method identified 35 types. By combining all results, 99 different strains could be identified. Isolate clusters were frequently associated within herds or were found between herds when epidemiological evidence confirmed animal movements. RFLP analysis with IS6110 was sufficiently sensitive for the typing of isolates with more than three copies of IS6110, but RFLP analysis with the PGRS probe was the most sensitive typing technique for strains with only a single copy of IS6110. Spoligotyping may have advantages for the rapid typing of M. bovis, but it needs to be made more sensitive.

* Corresponding author. Mailing address: Australian Reference Laboratory for Bovine Tuberculosis, Agriculture Western Australia, 3 Baron-Hay Ct., South Perth, Western Australia 6151, Australia. Phone: 618 9368 3429. Fax: 618 9474 1881. E-mail: dcousins{at}agric.wa.gov.au.

Journal of Clinical Microbiology, January 1998, p. 168-178, Vol. 36, No. 1
0095-1137/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.


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