Lipid-based ELISA test for detection of dairy cattle with Johne`s disease

Johne?s disease (JD) is a chronic granulomatous inflammation of the intestine in domestic and wild ruminants usually resulting after along incubation period in constant diarrhea with tremendous weight and ultimately death. This disease is caused by the intracellular pathogen Mycobacterium avium subspecies paratuberculosis (MAP). Johne?s disease currently ranks as one of the most costly infectious diseases of dairy cattle in the USA and of sheep in Australia and New Zealand and has a global distribution especially in developed countries. There are roughly 10 millions of dairy cattle in the US but only 8% of them are tested for Johne?s disease (http://www.nass.usda.gov/QuickStats/index2.jsp). The infection occurs usually after birth and calves that are infected have a short period with mild or no symptoms and more importantly will shed the bacteria. This short usually not recognized period is followed by a long incubation period of two to five years in which the animals are presumably healthy, usually non-shedding, and without any symptoms directing toward the development of Johne?s disease. After the incubation period cattle develop Johne?s disease with the characteristic symptoms of loss of milk production, chronic diarrhea, chronic weight-loss, and finally death of the animal. There are no tools for prevention of treatment for this disease. The only effective treatment is culling the infected animals. Furthermore, there are only two diagnostic tests commercially available for Johne?s disease with questionable efficiency of detecting animals with the disease. Those tests lack reproducibility due to strong batch-to-batch variation determining animals from ?strong positive? with one batch and ?suspect/negative? with a different batch. Both tests exhibit high specificity but insufficient sensitivity. Thus, a more precise test using indirect or direct techniques to identify animals with Johne?s disease is necessary and required by the dairy industry. Two commercially available diagnostic tests for Johne?s disease are in use within the United States. Both tests are based on crude aqueous antigens and exhibit a reasonably specificity of about 97% but have an unacceptable low sensitivity of about 28%. To avoid cross-reactivity, both tests require pre-absorption with crude antigen extracts from M. phlei. In addition, the crude antigens extraction usually results in strong batch-to-batch variation. This variation leads to classifying sera with one batch as positive but with another batch as negative. There is a strong demand from the diagnostic laboratories for a test for Johne?s disease with no batch-to-batch variation, no pre-absorption, and a higher sensitivity. In this project we will identify lipids that serve as the base for an innovative diagnostic test for Johne?s disease in dairy cattle.