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Spoligotyping was used to examine IS6110-positive DNA of 26 Mycobacterium bovis, M. bovis BCG and M. bovis subsp. caprae non-viable isolates stored up to 10 years. All of these isolates were previously identified by biochemical tests and all 17/17 tested isolates were earlier found virulent for guinea pigs. In total seven spoli- gotypes, designated S1-S7, were detected and compared with the spoligotypes of 3 176 isolates in the database of the National Institute of Public Health and the Environment (RIVM) in Bilthoven, the Netherlands. A Neotype M. bovis strain, isolated in 1965 in the USA and thereafter stored in e Czechoslovak National Collection of Type Cultures (My 310/87) since 1987 was of an identical spoligotype S4 with the original reference M. bovis strain from the USA. e M. bovis isolates from capybara's (Hydrochoerus hydrochaeris) imported from Germany to the Czech Republic in 1989, as well as cattle isolates from 1966, 1991 and 1994, were of the most common type S1. Also a human isolate from 1981, a M. bovis BCG vaccine strain and clinical M. bovis BCG isolates from three children with post-vaccinal complications were of this most predominant spoligotype. e four unique spoligotypes S2, S3, S5 and S6 were identified in M. bovis isolates from cattle in the years 1965, 1996 and 1967 in the Czech Republic, respectively, but also in isolates from farmed red deer (Cervus elaphus) from 1991 and in cattle isolates from Slovakia from the year 1992. e scarcely occurring spoligotype S7, which is typical for M. b. caprae was detected in the Czech Republic from farmed red deer (1999), cattle isolates (1966, 1991, 1995) and in a strain isolated from an 80-year-old man (1999). Several strains isolated in each of three outbreaks in cattle herds were examined. Identical spoligotypes were detected in two outbreaks and different causal agents ( M. bovis of spoligotype S1 and M. b. caprae of spoligotype S7) were identified in two cows from the third outbreak. e results confirm an effective control of bovine tuberculosis in the Czech Republic and Slovakia during 1959-1968, because previously circulating spoligotypes were successfully eradicated. e data also suggest other reservoirs of bovine tuberculosis may exist among free-living wild animals.

Molecular epidemiology of bovine tuberculosis in the Czech Republic and Slovakia in the period 1965-2001 studied by spoligotyping