Submitted to: Molecular and Cellular Probes
Publication Type: Peer reviewed journal
Publication Acceptance Date: 1/3/2005
Publication Date: 5/17/2005
Citation: Ellingson, J.E., Stabel, J.R., Radcliff, R.P., Whitlock, R.H., Miller, J.M. 2005. Detection of mycobacterium avium subspecies paratuberculosis in free-ranging bison (bison bison) by PCR. Molecular and Cellular Probes. 19(3):219-225. Interpretive Summary: Johne's disease is a chronic, debilitating intestinal disorder in cattle,sheep and wild ruminants, characterized by diarrhea, reduced feed intake, weight loss and death. Animals usually become infected when they are young by ingesting feces containing the causative bacteria. However, symptoms of disease do not usually present themselves until the animals reach 3 to 5 years of age or even older. During this time the animal is infected and may be shedding the organism in its feces without showing any clinical signs of disease. In addition to reduced production by these animals through reduced milk production, they also present a potential infective threat to the rest of the herd. Johne’s disease is difficult to diagnose and therefore to control. Development of accurate and sensitive diagnostic tests is essential for controlling the spread of this disease. In this paper, we present results comparing the detection of infection in bison by a gene detection method. Results of this study suggest that the gene detection method is a very sensitive method for identification of infected bison. Being able to accurately detect infected animals will help allay the spread of this disease in wildlife and domesticated animals.
Technical Abstract: Bacterial culture is the 'gold standard' for detecting Mycobacterium avium subspecies paratuberculosis (MAP) infection, but is time consuming, laborious, and recovery of organism varies with species of animal tested. PCR has been used for detection of MAP DNA in feces and tissues. We used PCR to detect MAP DNA isolated from tissues from 25 free ranging North American bison (Bison bison), each with clinical signs compatible with Johne's disease. We report the performance of PCR to detect MAP DNA in both frozen and paraffin embedded ileum, jejunum, and ileocecal lymph node samples collected at the time of slaughter. Specific oligonucleotide primers for PCR amplification were derived from 16S rRNA sequence M. avium subspecies (MAs) and insertion elements IS1245 (MAs avium), IS901 (MAs avium), IS900 (MAP), and hspX (MAP). Genomic DNA samples were prepared three different ways; crude DNA from frozen tissues, crude DNA from paraffin embedded tissues, and purified DNA from paraffin embedded tissues. An animal was considered infected if MAP DNA was detected in at least two separate tissues using the IS900 primer set. Using these criteria, 25 of 25 bison tested were positive for MAP. The data indicate that these free ranging bison have been infected by MAP.