Genetic evaluation of IS900 partial sequence of Mycobacterium avium subsp. paratuberculosis Brazilian isolates from bovine milk
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Mycobacterium avium subsp. paratuberculosis (MAP) is the causative agent of paratuberculosis. Insertion sequence IS900 is used for the identification of MAP. The objective of this study was to verify the genetic conservation of IS900 sequences in raw milk samples. To evaluate genetic conservation, 206 quarter milk samples and 16 bulk-tank milk samples were collected. DNA extraction and IS900 PCR were performed in all samples. Six samples amplified the expected fragment. To confirm the identity of the amplified fragments, PCR products were cloned and sequenced. The resulting sequences were compared with other MAP sequences from GenBank, and it was possible to identify eight polymorphic regions and to form five distinct haplotypes. The number of mutations in each haplotype was verified. IS900 sequence is a very well-conserved sequence that could be used as tool for the molecular detection of this agent and epidemiological purposes. The results showed the first genetic analysis on Brazilian isolates of MAP.
KeywordsMycobacterium avium subspecies paratuberculosis Johne’s disease IS900-PCR Milk
The authors would like to thank FAPEMIG (Fundação de Amparo à Pesquisa do Estado de Minas Gerais) for financing this study; CNPq (Conselho Nacional de Desenvolvimento Científico e Tecnológico) and CAPES (Corrdenação de Aperfeiçoamento de Pessoal de Nível Superior) for financing the doctorate fellowship; and Prof. Marcos Jose Pereira Gomes, from UFRGS (Federal University of Rio Grande do Sul), for providing a wild certified MAP strain.
Nucleotide sequence accession numbers
The GenBank accession numbers for IS900 partial sequences from M. avium subsp. paratuberculosis are HM015763, HM015764, HM015765, HM015766, HM015767, and HM015768. Sequences will be available for public access in June 2012.
Conflict of interest
The authors declare that they have no conflict of interest.
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