The detection of Mycobacterium paratuberculosis in bovine faeces by isolation and the comparison of isolation with the examination of stained smears by light microscopy.

The detection of Mycobacterium paratuberculosis organisms in bovine faeces by isolation was compared with that by the microscopical examination of Ziehl-Neelsen stained faecal smears for the presence of clumps of acid-fast M. paratuberculosis organisms. Faeces were obtained from cattle naturally or experimentally infected with M. paratuberculosis as well as from uninfected cattle. Microscopical examination was an unreliable method for the detection of M. paratuberculosis organisms, since the organisms were only detected in 99 (=55.9%) of 177 culturally positive faecal samples. 1111 addition, clumps of acid-fast organisms indistinguishable from M. paratuberculosis were also observed iin three of 18 samples from cattle free from Johne's disease and in 18 of 37 culturally negative samples from paratuberculous cattle. When M. paratuberculosis organisms were added to faeces from an uninfected cow, results showed that isolation attempts should be positive when 15 or more M. paratuberculosis organisms per gram of faeces are present.

Use of two polysaccharide antigens in ELISA for the detection of antibodies to Echinococcus granulosus in sheep sera.

Polysaccharide antigens were obtained from either the secretions produced during in vitro cultivation of Echinococcus granulosus protoscoleces or from mouse hydatid cyst membranes by phenol extraction. When either of these antigens was used in an enzyme-linked immunosorbent assay antibody activities were detected in sera from sheep infected 27 or more weeks earlier with at least 100 E granulosus eggs. These antibody responses were significantly higher (P less than 0.05) than those of sheep infected with Taenia hydatigena or T ovis and tested with the E granulosus antigens. Very high cross-reacting antibody responses in sera from sheep recently infected with T hydatigena were only detected with the protoscoleces secretions antigen. Neither antigen was sufficiently sensitive or specific for serodiagnostic use. However, when sera were first tested with one antigen and then with the other, and only sera that were positive in both tests were regarded as positive, the overall sensitivity and specificity of this two antigen method increased to about 80 per cent.

Comparison of an enzyme-linked immunospecific assay (ELISA) with the cold complement fixation test for the serodiagnosis of Brucella ovis infection.

An enzyme-linked immunospecific assay (ELISA) for the serodiagnosis of Brucella ovis infection in sheep is described and compared with the cold complement fixation (CF) test. ELISA was performed in microtiter plates, using horse-radish peroxidase conjugated to anti-normal sheep serum globulins, and hydrogen peroxide plus o-phenylenediamine as substrate. A heated, cell-free B. ovis extract was used as antigen in both tests. ELISA was easier to perform, distinguished better between positive and negative sera, and did not need heat-inactivated sera.

An unusual serological response in cattle after use of a commercial leptospiral combined hardjo-pomona vaccine.

Vaccination of four calves with Leptavoid (Wellcome New Zealand Limited) gave rise to Leptospira interrogans serovars hardjo and pomona microscopic agglutination test titres that could not he distinguished in magnitude from post-infection titres. Vaccination of four calves with Lepto-3 (ICI Tasman Limited) gave rise to much lower titres. Revaccination of cows with Leptavoid caused a rise in hardjo titres which was significantly greater than after use of Lepto-3. The possibility that titres were due to the simultaneous infection with serovars pomona and hardjo of only the animals vaccinated with Leptavoid must be discounted.

The use of a cathodic antigen in the immunoelectrophoretic serodiagnosis of Echinococcus granulosus in sheep.

An antigen was prepared from purified sheep hydatid-cyst fluid by gel-filtration on Sephadex G-200 followed by chromatography on DEAE-cellulose. In each case the first-peak material was used. This antigen, which migrated cathodically, was concentrated and used in immunoelectrophoretic analyses of 4X concentrated sera from sheep experimentally and naturally infected with Echinococcus granulosus and from uninfected sheep. Of 34 sheep with E. granulosus infection 31 were positive with the cathodic antigen while of 85 sheep with E. granulosus 8 were (falsely) positive. Many false positives appeared to be associated with heavy infections of Taenia ovis or T. hydatigena larvae. The "arc 5' immunoelectrophoresis test, which is the most specific immuno-diagnostic test for echinococcus infection in humans, was not able to specifically identify E. granulosus infections in sheep.

Use of hamster antisera in the preliminary differentiation of Leptospira interrogans servars hardjo and balcanica.

Preliminary differentiation of Leptospira interrogans serovars hardjo and balcanica can be made with antisera that are collected from hamsters after experimental infection with serovar hardjo and selected for high homologous and low or nil heterologous titres, as measured by the microscopic agglutination test. Rabbit antisera against serovars hardjo and balcanica, cattle antisera against hardjo and sheep antisera against the serogroup Hebdomadis each agglutinated hardjo and balcanica to a similar titre.

Limitations of the use of 5-fluorouracil as a selective agent for the isolation of leptospirae.

It has been shown that some component of Fletcher medium is able to annul the inhibitory action of 5-fluorouracil (400 mug/ml) on bacteria other than leptospirae. The most likely ingredient which can be implicated in this context appears to be beef extract.