Survival of Bovine Mycobacterium avium Subsp. Paratuberculosis during Grass Silage Forage Fermentation.

Mycobacterium avium subsp. paratuberculosis (MAP) causes Johne’s disease, a chronic, enteric infection that is transmitted by ingestion of the organism. Eradication of MAP from infected farms has been difficult and is likely due to long-term survival of the organism in the environment. Manure from an infected herd applied to growing grasses for forage could be a potential route of infection. Information is needed to determine if proper forage fermentation will kill MAP reducing the risk of transmission. The objective of this study was to evaluate the survival of MAP in infected manure inoculated onto grass forage and fermented for silage. Quantitative, real-time PCR (QRT-PCR) was used to target the IS900 sequence of MAP in rectal samples taken from the infected animals to identify an animal with a high shedding rate to use for inoculation. Forage was gathered at chopping in September 2007 and frozen for the project. The manure from the selected animal (2.0±0.19 X 10⁶ MAP cells g^-1 manure)was then diluted and applied to thawed fresh chopped forage (1.0±0.19 X 10⁵ MAP cells g^-1 manure:700 g of forage), mixed and placed in vacuum sealed bags for fermentation. Six replicate bags and a control (no inoculum) were sampled at 30, 45, 60, 75, and 90 days after sealing. Sub-samples from day 0 (inoculated forage prior to fermentation) and from each time point were analyzed for MAP cell concentration by QRT-PCR. Forage dry matter, pH, and volatile fatty acid profile were also analyzed for each time point. Preliminary work done to evaluate the forage fermentation resulted in a pH of 4.4 and volatile fatty acid concentrations of 4.67%, 2.44%, 0.07%, and 0.02% of lactic, acetic, propionic, and butyric acids, respectively.