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Summary

Use of a dot-elisa with serum adsorbed with Mycobacterium phlei or with nonadsorbed serum was compared. In addition, results attained using visual observation were compared with those obtained using a densitometer. Infection status of cattle was determined by results of culture of feces from a number of cattle with various degrees of exposure (low prevalence and test-negative) and disease manifestation (clinical suspect vs subclinical infection). Two paratuberculosis-negative herds, fecal culture-confirmed clinically suspect cases of paratuberculosis, and cows from 2 paratuberculosis-infected herds with diagnosis confirmed on the farm (low infection rate) were tested.

Significant (P < 0.05) increase in the dot-elisa response was found in cattle with heavy M paratuberculosis shedding when nonadsorbed and adsorbed sera were used, compared with the response in cattle that were fecal culture-negative or were shedding M paratuberculosis at lower amounts. Paratuberculosis was diagnosed by visual determination in 29 of 44 (65.9%) of fecal culture-positive, clinically suspect cattle when nonadsorbed serum was used. Results of the visual test were negative in 85 of 93 (91.4%) of the fecal culture-negative cattle when nonadsorbed serum was used. However, when using M phlei-adsorbed serum, the sensitivity of the visual determination decreased to 34.1% (15/44), and the specificity increased to 97.8% (91/93). When taking into consideration the amount of bacterial shedding (ie, colonies per gram of feces isolated by culture of feces), 22 of 29 (75.9%) cattle were heavy shedders, with > 1,500 colonies/g of feces at the time of serologic testing when nonadsorbed serum was used, and 11 of 15 (73.3%) cattle were heavy shedders when adsorbed serum was used. Thus, about 75% of paratuberculosis-positive cattle, detected by use of the dot-elisa, were heavy shedders.

Effects on sensitivity and specificity of using various cut-off points for the various test groups were determined by use of video densitometric measurement, because sera were not discretely segregated into distinct groups of positive and negative results. Specificity of the elisa in the 2 fecal culture-negative herds was 96.2% at elisa cut-off optical density value ≥ 0.4 (OD) for adsorbed serum. For nonadsorbed serum, specificity was 78.8% at similar OD cut-off value.

Comparison of dot-elisa results determined by visual vs objective densitometric measurement indicated compatible results for test specificity; adsorption of serum with M phlei increased test specificity. Test sensitivity using visual evaluation was 66%, and was 87.5% using the objective densitometric evaluation for nonadsorbed serum at elisa cut-off OD value of 0.2. This difference was even more pronounced when adsorbed sera were used—34.1 and 82.5%, respectively.

Free access
in American Journal of Veterinary Research

Summary

To establish whether Mycobacterium paratuberculosis could be cultured from Dulbecco phosphate-buffered saline solution (dpbss) and to test 3 sampling methods, dpbss supplemented with 2% fetal bovine serum was inoculated with M paratuberculosis at concentrations of 104, 103, 102, 101, and 100 colony-forming units/ml. The inoculated media was sampled after mixing, after centrifugation, and after centrifugation and decontamination with 0.75% hexadecylpyridinium chloride. The samples were inoculated onto 3 slants of Herrolds egg yolk medium supplemented with sodium pyruvate and mycobactin J and 1 slant without mycobactin J. Mycobacterium paratuberculosis was isolated following all 3 sampling methods for all concentrations. Treatment with hexadecylpyridinium chloride decreased the number of colonies isolated.

To test the efficacy of a 10-step wash procedure for removing M paratuberculosis from bovine ova, washed zona pellucida intact bovine ova were incubated in dpbss supplemented with 2% fetal bovine serum containing concentrations of 104, 103, 102, 101, and 100 colony-forming units of M paratuberculosis/ml for 12 hours at 22 C. Ten zona pellucida intact ova were removed from each concentration and washed by passing through 10 changes of dpbss supplemented with 15% fetal bovine serum. The media from each wash step was inoculated onto slants of Herrolds egg yolk medium. The ova were included with the tenth wash step. Mycobacterium paratuberculosis was isolated from 1 of 10 tenth-wash steps at the 104 concentration and 5 of 10 tenth-wash steps at 103.

Free access
in American Journal of Veterinary Research

SUMMARY

An agar gel immunodiffusion (agid) test was used over a 3-year period to examine 1,871 serum samples from sheep representing 5 Mycobacterium paratuberculosis — infected flocks and 4 flocks presumed to be uninfected. Of 1,032 sheep, 31 had positive agid test results (scoring 1 to 5), and 23 of these 31 were necropsied. Infection with M paratuberculosis was confirmed by 1 or more of the following findings: observation of typical lesions on histologic examination of sections of ileum or ileocecal lymph nodes, observation of clumps of acid-fast bacteria in mucosal smears of ileum, and isolation of the organism from feces or tissue. False-positive results on agid testing were not found in sheep from flocks known to have exposure to Corynebacterium pseudotuberculosis. Diarrhea in infected sheep was observed infrequently; chronic, severe weight loss was the most common sign observed. On histologic examination of tissues from 20 infected sheep, 16 (80%) had diffuse lesions of the ileum and 13 (65%) had acid-fast bacteria in areas of ileal inflammation; 4 had discrete granulomas and peripheral lymphocytic infiltrates in the ileum. Sheep with diffuse lesions tended to have higher mean scores on agid testing and examination for acid-fast bacteria, compared with those from sheep with more discrete lesions. Bacteriologic culture yielded M paratuberculosis from only 3 sheep with paratuberculosis. On the basis of results of this study, we suggest that the nature of the response to infection with M paratuberculosis may influence the results of diagnostic tests for paratuberculosis, and that agid testing may be useful to identify M paratuberculosis infection in sheep with chronic weight loss and in flock-screening programs.

Free access
in American Journal of Veterinary Research

SUMMARY

A dot elisa was developed for detection of antibodies to Mycobacterium paratuberculosis. The assay was evaluated by testing sera from cattle that were determined, by bacteriologic culturing of feces, to be infected with M paratuberculosis and were suspected of having clinical disease. Further evaluation involved testing sera from cattle in which M paratuberculosis had not been isolated from feces on several attempts. Results of the dot elisa were positive for sera from 86 of 101 infected cattle, and results were negative for sera from 64 of 64 noninfected cattle. Results of conventional elisa and agar gel immunodiffusion (agid) tests were positive for 79 of 99 and for 51 of 101 infected cattle, respectively.

The dot elisa also was evaluated by comparing results of testing 708 sera with results of bacteriologic culturing of matched fecal samples from 262 cattle in 3 central Ohio dairy herds known to include cattle infected with M paratuberculosis. Results of the dot elisa were positive for 25 of 39 sera from cattle with positive results on culturing of concurrently obtained fecal specimens. The dot elisa results were negative for 661 of 669 sera from cattle with negative results to culturing of concurrently obtained fecal specimens. The 39 sera from cattle with positive results on bacteriologie culturing of matched fecal specimens had positive results for elisa and the agid test 25 and 14 times, respectively. The 669 sera from cattle with concurrently negative results on bacteriologie culturing of feces had negative results to elisa and the agid test 559 and 668 times, respectively.

Free access
in American Journal of Veterinary Research

Abstract

Objective—To evaluate the ability of orally administered aspirin to mitigate 3-methylindole (3MI)-induced respiratory tract disease and reduced rate of gain in feedlot cattle.

Animals—244 beef cattle.

Procedure—In a masked, randomized, controlled field trial, calves were untreated (controls) or received a single orally administered dose of aspirin (31.2 g) on entry into a feedlot. Serum 3MI concentrations were measured on days 0, 3, and 6. Rumen 3MI concentration was measured on day 3. Cattle were observed daily for clinical signs of respiratory tract disease. Lungs were evaluated at slaughter for gross pulmonary lesions.

Results—Mean daily gain (MDG) in cattle treated with aspirin, compared with control cattle, was 0.06 kg greater in the backgrounding unit and 0.03 kg greater for the overall feeding period. Neither serum nor rumen 3MI concentrations appeared to modify this effect. Cattle treated with aspirin were more likely to be treated for respiratory tract disease. Mortality rate, gross pulmonary lesions, and serum and rumen 3MI concentrations were similar between groups. Increased rumen 3MI concentration was associated with a small difference in risk of lung fibrosis.

Conclusions and Clinical Relevance—Cattle given a single orally administered dose of aspirin on feedlot entry had higher MDG in the backgrounding unit and for the overall feeding period, but this finding could not be attributed to mitigation of effects of 3MI. This may have been influenced by low peak 3MI production and slow rates of gain. (Am J Vet Res 2000;61:1209–1213)

Full access
in American Journal of Veterinary Research

Abstract

Objective—To determine whether immunity against bovine respiratory syncytial virus (BRSV) mitigates the effects of 3-methylindole (3MI) on occurrence of bovine respiratory tract disease (BRD) and rate of gain in feedlot cattle.

Animals—254 mixed-breed beef cattle.

Procedure—Cattle were randomly assigned to 1 of 3 groups at the time of arrival at the feedlot. One group was vaccinated with an inactivated BRSV vaccine, another was vaccinated with a modified-live BRSV vaccine, and the third was maintained as unvaccinated control cattle. On days 0 and 28, serum BRSV antibody concentrations were measured, using serum neutralizing and ELISA techniques. Serum 3MI concentrations were measured at feedlot arrival and 3 days later. Cattle were monitored for development of BRD. At slaughter, lungs were evaluated grossly for chronic lesions.

Results—Higher serum 3MI concentrations early in the feeding period were associated with lower mean daily gain. Control cattle were more likely to be treated for BRD after day 3, compared with cattle vaccinated with the modified-live BRSV vaccine. Humoral immunity against BRSV did not appear to modify the effect of 3MI on development of BRD or mean daily gain.

Conclusions and Clinical Relevance—Results suggest that abrogating the effects of 3MI and BRSV infection may improve the health and growth performance of feedlot cattle. However, in this study, immunity against BRSV did not appear to protect against the potential synergism between 3MI and BRSV infection, possibly because of the slow rates of gain of cattle included in the study or timing of sample collection. (Am J Vet Res 2000;61:1309–1314)

Full access
in American Journal of Veterinary Research

Abstract

Objective—To evaluate sensitivity and specificity of a new ELISA for antibodies against Mycobacterium avium subsp paratuberculosis.

Design—Cross-sectional observational survey.

Sample Population—Serum samples from 590 cattle that were infected with M avium subsp paratuberculosis and 723 cattle that were not infected.

Procedure—Serum samples were tested by use of an ELISA for antibodies against M avium subsp paratuberculosis.

Results—Sensitivity of the test varied from 15.4 to 88.1%, depending on the clinical stage and bacterial shedding status of the cattle.

Conclusions and Clinical Relevance—Results obtained with use of the new ELISA agreed favorably with those of a previous ELISA. Practitioners must be aware of variability in the sensitivity of the test, which depends on the clinical and shedding status of the cattle, because this may affect interpretation of test results. (J Am Vet Med Assoc 2001;218:1163–1166)

Full access
in Journal of the American Veterinary Medical Association