Comparison of serological tests and faecal culture for the detection of Mycobacterium avium subsp. paratuberculosis infection in cattle and analysis of the antigens involved

Three hundred and forty-one sera from cattle in Western Australia and 106 s era from Mycobacterium paratuberculosis faecal culture positive cattle were used to evaluate the performance of two absorbed enzyme-linked immunosorbe nt assays (ELISA) (one locally produced, the other a commercial test) and a complement fixation test (CFT) for the detection of Johne's disease in cat tle. The diagnostic sensitivity (47.2%) of the local ELISA was significantl y higher than that of the commercial ELISA (31.1%), and significantly highe r than that for the complement fixation test (17.9%) and immunoblot (20.8%) . Diagnostic specificity for the two ELISAs was 99.7% and 97.9% and similar for CFT and immunoblot (97.1% and 97.7%, respectively). The diagnostic sen sitivity rose for both ELISAs and the CFT as the number of M. paratuberculo sis isolated from the faeces increased. The ELISA antigen was characterised by polyacrylamide gel electrophoresis and electrophoretic immunoblotting a nd was found to consist mostly of a carbohydrate-type macromolecule of 32-4 2 kDa. This macromolecule was identified as lipoarabinomannan (LAM) by usin g a LAM-specific monoclonal antibody in immunoblots and purified LAM in abs orption experiments. By applying more complex antigen preparations in immun oblots, serum antibodies against proteins of 47, 37, 30, 24 and 21 kDa. and against the 32-42 kDa carbohydrate component were frequently found in infe cted cattle, and of these the 47 kDa protein and the 32-42 kDa antigen were immune-dominant. Pre-absorption of the sera with M. phlei sonicate indicat ed that the protein antigens contributed markedly to non-specific serologic al cross-reactions, while the 32-42 kDa non-protein macromolecule appeared to be specific. (C) 1999 Elsevier Science B.V. All rights reserved.