Agricultural Microbes Genome 2 Conference

Poster: Functional Genomics
P05_04.html

IDENTIFICATION OF UNIQUE REGIONS WITHIN TWO GENETICALLY SIMILAR MYCOBACTERIAL GENOMES

¹ National Animal Disease Center, 2300 N. Dayton Ave., Ames, Iowa 50010, USA

² University of Minnesota, Dept. of Vet. Pathobiology, 1971 Commonwealth Ave., St Paul, MN 55108, USA

Among the Mycobacterium avium species are two subspecies, avium and paratuberculosis, that are >98% conserved at the nucleotide level. Yet, these two pathogens cause very distinct clinical diseases in diverse hosts. Random sequencing of the M. avium subspecies paratuberculosis (M. paratuberculosis) genome has confirmed the high degree of homology among these subspecies. However, at least eight M. paratuberculosis sequences were identified that do not align with M. avium subspecies avium (M. avium) sequences. Two of these M. paratuberculosis specific sequences, designated #7 and #481, were evaluated further. Southern hybridization and PCR amplification of sequences designated #7 and #481 with several strains experimentally confirm the lack of computer alignments. Using these sequences as probes to screen a genomic library of M. paratuberculosis, ten positive clones were obtained. Four of these clones aligned with the #481 sequence and the other six aligned with #7. Assembly of these overlapping library clone sequences yielded contigs of 4266 and 6118 bp respectively. BLAST analysis of open reading frames contained within the contig #7 sequence identified reasonable homologies to proteins in M. tuberculosis, whereas, BLAST analysis of the #481 sequence revealed no significant homologies. RT-PCR analysis is being used to confirm the ORF predictions in the novel #481 sequence. The unusual bidirectional ORF arrangement of the #481 sequence combined with very weak homology to viral genes may indicate the presence of a mycobacteriophage. These findings may be applicable to the development of new diagnostic tests to distinguish these genetically similar subspecies.