|Ewalt, Darla - APHIS|
Submitted to: American Association of Veterinary Laboratory Diagnosticians
Publication Type: Abstract Only
Publication Acceptance Date: October 21, 2004
Publication Date: October 21, 2004
Citation: Bricker, B.J., Ewalt, D.R. 2004. Strain typing brucella abortus isolated from California and Florida cattle herds by using "hoof-prints"-a new DNA fingerprinting technique for brucella [abstract]. American Association of Veterinary Laboratory Diagnosticians. p. 91. Technical Abstract: Techniques for subtyping Brucella abortus strains by conventional means are very limited. Currently, a large panel of metabolic and phenotypic tests is needed to subdivide B. Abortus strains into 7 biovars, only three of which occur in the USA. Furthermore, about 85% of B. Abortus strains isolated from cattle are categorized as biovar-1, limiting the epidemiological value of subtyping. We have developed a new technique called 'HOOF-Prints', which subtypes Brucella strains on the basis of DNA sequence differences. These differences arise from tandem repeats of an 8-bp sequence. Multiple tandem repeats of short DNA sequences (also known as microsatellites) are known to form hotspots for accelerated mutation. The Brucella genome has eight of these hotspots dispersed throughout its tow chromosomes. Analysis of the DNA sequence at each of the eight hotspots produces a very distinctive genetic fingerprint. The procedure is based of PCR technology which is both rapid and very sensitive. In this study, we examined a collection of 46 Brucella strains cultured from cattle in the early 1990's. Among these samples were 27 isolates obtained from 5 different infected herds in California, and 19 isolates obtained from 12 different infected herds in Florida. The samples were subtyped by biovar and by DNA fingerprint. Typing by metabolic and phenotypic analyses showed that all three of the U.S. biovars were present in the study group in the following proportions: 89.1% biovar-1, 8.7% biovar-2 and 2.2% biovar-4. Two strains were identified as the vaccine strainS19. DNA analyses of the 46 strain collection revealed 28 different HOOF-Print patterns. Within most herds the fingerprint patterns were identical or very similar, differing at 2 or less of the 8 loci. Comparison of DNA fingerprints across different herds showed that the strains collected from Florida herds were different and probably genetically unrelated. The Florida strains were also different from the California strains. Comparison of the five California herds, however, showed surprisingly similar DNA patterns suggesting a common source of infection. The data show that HOOF-Print DNA fingerprinting is a highly discriminating method for characterizing Brucella strains, far surpassing the capabilities of biotyping. This technique could be a valuable tool fro epidemiologist trying to study the origins and spread of Brucella outbreaks.