Bacteriophage-Based Methods for Detection of Viable subsp. and Their Potential for Diagnosis of Johne's Disease.

Bacteriophage-based methods for detecting subsp. paratuberculosis (MAP) are a potential new approach for diagnosis of Johne's disease (JD). The basis of these tests is a mycobacteriophage (D29) with a lytic lifecycle that is able to infect a range of spp., not just MAP. When added to a test sample, the phages will bind to and infect mycobacterial cells present. If the host mycobacterial cells are viable, the phages will take over the metabolic machinery of the cells to replicate and produce multiple copies of themselves (phage amplification), before weakening the host cell walls by enzyme action and causing cell lysis. Cell lysis releases the host cell contents, which will include ATP, various enzymes, mycobacterial host DNA and progeny D29 phages; all of which can become the target of subsequent endpoint detection methods. For MAP detection the released host DNA and progeny phages have principally been targeted. As only viable mycobacterial cells will support phage amplification, if progeny phages or host DNA are detected in the test sample (by plaque assay/phage ELISA or qPCR, respectively) then viable mycobacteria were present. This mini-review will seek to: clearly explain the basis of the phage-based tests in order to aid understanding; catalog modifications made to the original plaque assay-based phage amplification assay (FASTPlaqueTB™) over the years; and summarize the available evidence pertaining to the performance of the various phage assays for testing veterinary specimens (bovine milk, blood and feces), relative to current JD diagnostic methods (culture, fecal PCR, and blood-ELISA).