|Robbe austerman, Suelee|
Submitted to: International Colloquium on Paratuberculosis
Publication Type: Proceedings
Publication Acceptance Date: 8/13/2005
Publication Date: 7/1/2006
Citation: Paustian, M., Zhu, Z., Robbe Austerman, S., Sreevatsan, S., Kapur, V., Bannantine, J.P. 2006. Comparative genomic analysis of Mycobacterium avium subspecies paratuberculosis isolates obtained from multiple host species. In: Proceedings of the 8th International Colloquium on Paratuberculosis, August 13-17, 2005, Copenhagen, Denmark. p. 393-400. Interpretive Summary: The bacterium Mycobacterium avium subsp. paratuberculosis (M. paratuberculosis) causes Johne’s Disease, an economically important intestinal infection of ruminants such as cattle and sheep. The complete genome sequence for a bovine M. paratuberculosis isolate is available; however it remains unclear whether this sequence is representative of isolates obtained from other species. We used a DNA microarray to compare the genomic DNA of M. paratuberculosis isolates obtained from cattle, sheep, goats, birds, humans, and several other species. Our results indicate that there are several segments of genomic DNA that are missing from sheep isolates. Additionally, M. paratuberculosis sheep isolates contain several segments of DNA that are present in the genome of Mycobacterium avium subspecies avium but not M. paratuberculosis isolates from other species. None of the isolates from the other species examined in this study contained any of these deleted or acquired sequences. Our results indicate that M. paratuberculosis sheep isolates can be distinguished from cattle isolates by the presence or absence of several large pieces of genomic DNA. Other researchers should benefit from the results of this work, as it will enable them to focus on the differences between cattle and sheep isolates in order to determine what causes M. paratuberculosis to infect different host animals.
Technical Abstract: We have designed and built a DNA microarray consisting of 70mer oligonucoeotides representing all of the ORFs identified in the Map K10 genome sequence as well as intergenic regions in order to investigate whether the genome content of Mycobacterium avium subspecies paratuberculosis (Map) K10 is representative of other bovine isolates as well as isolates from other species. In addition, oligonucleotides representing coding sequences from the Mycobacterium avium subspecies avium (Maa) 104 genome that are not present in the Map K10 genome were also included on the microarray. Genomic DNA from Map isolates was fluorescently labeled, mixed with alternately labeled Map K10 DNA, and competitively hybridized on the Map microarray. ORFs were classified as present or divergent based on the relative fluorescent intensities of the experimental samples compared to Map K10 DNA. Map isolates cultured from cattle, bison, sheep, goat, avian, and human sources were hybridized to the Map microarray. Three deleted regions were observed in the genomes of three Map isolates obtained from sheep. Additionally, four clusters of ORFs homologous to sequences in the Maa 104 genome were identified in three of the sheep isolates. One of these clusters encodes glycopeptidolipid biosynthesis enzymes which have not previously been identified in Map. Differences in hybridization across many of the isolates examined were detected for the microarray targets representing the insertion sequence IS_MAP04, the major membrane protein encoded by MAP2121c, and several regions encoding proteins with unknown function suggesting that there may be variation in the sequence or copy number of these regions. One cattle isolate and one sheep isolate of Map were found to contain variable genome content compared to the other isolates examined from these species.