Citation: Bannantine, J.P., Briggs, R.E., Lamont, E.A., Stabel, J.R. 2010. A Gene Specific to Mycobacterium avium subsp. paratuberculosis, But Only at the Transcription-translation Level [abstract]. American Dairy Science Association. Journal of Dairy Science. 93(E-Supplement 1):182.
Technical Abstract: There is no known antibody that detects M. avium subsp paratuberculosis and does not cross react with other M. avium subspecies. In the present study, a monoclonal antibody was identified from mice immunized with a cell membrane fraction of M. avium subsp paratuberculosis strain K-10. This antibody detected a protein in M. avium subsp paratuberculosis whole cell extracts, but did not bind to any of the 20 non-paratuberculosis subspecies strains tested in immunoblot assays. The antibody was further tested with 15 strains of M. avium subsp paratuberculosis and showed variable expression levels of the target binding protein in select strains. This target binding protein was identified by screening a M. avium subsp paratuberculosis-lambda phage genomic expression library with the monoclonal antibody. The identity of the protein was encoded by a gene that was not annotated in the M. avium subsp paratuberculosis K-10 genome sequence and showed no similarity to other proteins in sequence databases. Furthermore, this gene has extensive overlap with an annotated gene on the opposite strand. The epitope detected by the mAb was precisely mapped to 7 amino acids and served as an anchor point in studies that identified the start and stop locations for this unique gene. Similarity searches reveal that the DNA sequence is present in other MAC complex species. However, expression analysis shows that only M. avium subsp paratuberculosis makes a transcript and expresses the protein in macrophages as well as when cultured in Middlebrooks 7H9. The protein is not a strong antigen, but it is detected in the context of Johne's disease. These findings have new implications for comparative genomics and gene regulation differences among mycobacteria.