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Journal of Clinical Microbiology, May 2000, p. 1723-1730, Vol. 38, No. 5
0095-1137/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Identification of a Novel DNA Probe for Strain Typing Mycobacterium bovis by Restriction Fragment Length Polymorphism Analysis

Rory O'Brien,1,2,* Orla Flynn,3 Eamon Costello,3 Don O'Grady,3 and Mark Rogers2,4

National Agricultural and Veterinary Biotechnology Centre1 and Department of Zoology,2 University College Dublin, Belfield, Dublin 4, and Tuberculosis Investigation Unit, Faculty of Veterinary Medicine, University College Dublin, Ballsbridge,4 Dublin 4, and Central Veterinary Research Laboratory, Abbotstown, Dublin 15,3 Ireland

Received 21 October 1999/Returned for modification 27 December 1999/Accepted 28 January 2000

Bovine tuberculosis caused by Mycobacterium bovis remains a significant disease of farmed cattle in many countries despite ongoing tuberculosis eradication programs. Molecular typing methods such as restriction fragment length polymorphism (RFLP) analysis and spoligotyping have been used to identify related herd breakdowns in an attempt to identify more precisely the route of infection into cattle herds and to trace the transmission of bovine tuberculosis. A recent geographical survey of Irish M. bovis isolates demonstrated that a significant proportion of isolates (~20%) exhibit a common strain type, limiting the value of current strain typing methods as an epidemiological tool. We have identified and cloned a region of the M. bovis genome, pUCD, which generates a clear, highly polymorphic banding pattern when used as an RFLP probe on AluI restriction-digested M. bovis genomic DNA and which effectively subdivides this common strain type. When used to type 60 Irish M. bovis isolates, pUCD exhibited greater discriminatory power than the commonly used mycobacterial RFLP probes IS6110, PGRS, and DR and detected an equivalent number of strain types to a combination of these three probes. pUCD also detected significantly more strain types than the spoligotyping technique, while maintaining a high level of concordance between epidemiologically related and unrelated herd breakdowns. The polymorphic element within pUCD remains to be fully characterized, however the potential for this probe to greatly decrease the workload necessary to genotype M. bovis by RFLP analysis is compelling.

* Corresponding author. Mailing address: National Agricultural and Veterinary Biotechnology Centre, University College Dublin, Belfield, Dublin 4, Ireland. Phone: (353) 1 7062344. Fax: (353) 1 7061152. E-mail: rory.obrien{at}ucd.ie.

Journal of Clinical Microbiology, May 2000, p. 1723-1730, Vol. 38, No. 5
0095-1137/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


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