Corynebacterium pseudotuberculosis infection in goats. III. The influence of age

Serological examinations for Gorynebacterium pseudotuberculosis in-fection were carried out in 14 known positive herds and in 1 herd that had beeni recently established through purchase of animals from different herds. Serum samples were collected sequentially on up to 4 occasions from animals during their first year of life. In most herds, these animals were examined clinically for superficial swellings once or twice during the same period.The adult goats in 8 infected herds were examined both clinically and serologically. Age distribution was similar in each of these herds. All serum samples were examined for antibodies against Gorynebac-terium pseudotuberculosis in both the bacterial agglutination test (BAT) and the hemolysis inhibition test (HIT).The proportion of kids which were seropositive in both tests de-creased to zero at 4 months of age. At the age of 11–12 months, the proportion of seropositive animals was about 50 % in the BAT and 30 % in the HIT.When examined when housed for the winter at the age of about 8 months (mean age), the prevalence of animals with superficial swellings was 7 %. At the age of about 1 year (mean age), 29 % of the examined animals had such lesions. In the recently established herd, only a few of the yearlings were seropositive.Titre values in BAT and HIT increased until 6 years of age. Anti-body titre values were significantly lower in yearlings than in older goats in both tests, P < 0.005. No significant difference in the propor-tion of goats with superficial swellings was seen at the different ages.     

Corynebacterium pseudotuberculosis infection in goats. VIII. The effect of vaccination against experimental infection

The effect of an inactivated vaccine against C. pseudotuberculosis infection was tested on castrated male kids from a herd free from caseous lymphadenitis. The animals were divided into 3 groups with 8 animals in each. Group 1 was immunized with crude filtrated C. pseudotuberculosis toxoid and whole killed organisms, while Group 2 in addition was given levamisole. The kids were vaccinated twice at an interval of 4 weeks. Group 3 consisted of unvaccinated animals. All groups were challenged subcutaneously with live bacteria 4 weeks after the last vaccination. Unvaccinated animals showed the most severe course of illness after challenge. Development of abscesses in the regional lymph nodes (Inn. subiliaci) was significantly more common in unvaccinated than in vaccinated kids at necropsy 2 months after challenge. There was, however, no such difference between the vaccinated groups, and there was no difference between any of the groups as regards abscess formation at the inoculation site. In each of the 2 vaccinated groups, there was a titre rise following vaccination in the hemolysis inhibition test, whereas no such rise was seen in the bacterial agglutination test. The titre values in both tests increased significantly after challenge in all the groups, the increase being most rapid in the vaccinated animals. The present investigation indicates that development of caseous lesions in lymph nodes in goats, following subcutaneous inoculation with C. pseudotuberculosis, can be reduced by an inactivated vaccine containing whole organisms and crude toxin.     

Corynebacterium pseudotuberculosis infection in goats. IX. The effect of vaccination against natural infection

A vaccination trial was carried out in 10 infected herds. The trial included 247 female kids, the number of animals in each herd varying from 15 to 38. About half of the animals in each herd were vaccinated twice at 3 to 4 week intervals, the first vaccination being carried out before the age of 4 months. A combination of a crude filtrate of C. pseudotuberculosis toxoid with whole organisms, was used. Overall, the prevalence of animals with superficial swellings was higher in the unvaccinated than in the vaccinated group during the first 1-2 years following immunization. However, in some herds superficial swellings were as common in vaccinated as in unvaccinated animals. An antibody response following vaccination was demonstrated in the hemolysis inhibition test, but not in the bacterial agglutination test. Superficial swellings were more common in vaccinated animals which were negative than in animals which were positive in the hemolysis inhibition test at 1 1/2 months after vaccination. The vaccine used in the present study, was not sufficiently efficacious to be recommended as the only protective measure against caseous lymphadenitis in Norwegian goat herds.     

Corynebacterium pseudotuberculosis infection in goats. I. Evaluation of two serological diagnostic tests

The bacterial agglutination test (BAT) and the hemolysis inhibition test (HIT) were employed and evaluated.The threshold value between positive and negative antibody titres in BAT and HIT was determined on the basis of titration results for sera from 25 adult goats infected with Corynebacterium pseudotuber-culosis (positive), and sera from 25 adult goats from a herd in which caseous lymphadenitis did not occur (negative).Antibody titres were expressed as logio to the reciprocal value of the highest positive serum dilution in both tests. Positive titre (T) in BAT was stipulated as T ≥ 2.1 and in HIT as T > 0.6. The sensitivity and specificity was 0.96 in both tests in the material used. Twenty-three of the 25 goats infected with G. pseudotuberculosis were positive in both BAT and HIT.The reproducibility in both tests was described by the use of correlation coefficient and coefficient of variation. The values were estimated using duplicate determination of titres of 155 serum samples. Correlation coefficients were 0.87 for BAT and 0.95 for HIT. Coef-ficients of variation were 26.4 % for BAT and 14.1 % for HIT. The coefficient of variation was related to titres. It was highest for BAT at low titres.It was concluded that both tests can be used for seroepidemio-logical investigations of C. pseudotuberculosis infection in goats.     

Corynebacterium pseudotuberculosis infection in goats. IV. Course of the infection in two recently infected goat herds

Adult animals from 2 herds were examined clinically and serologically, 5 (Herd A) and 4 (Herd B) times during the same period of 3% years.Serum samples were examined for antibodies against Gorynebac-terium pseudotuberculosis using the bacterial agglutination test (BAT) and the hemolysis inhibition test (HIT).The results of the first examination showed that no animal in Herd A was seropositive, while in Herd B 1 animal showed a high positive titre in BAT. The prevalence of animals with superficial swellings was then 2 % in Herd A and 4 % in Herd B. In both herds, the prevalence of animals with superficial swellings and seropositive reactions increased during the following 1–2 yeairs. About 30 % of animals had superficial lesions and close to 100 % were seropositive. The proportion, of animals with superficial swellings and seropositive reactions was almost constant on subsequent examinations.In some of the animals, superficial swellings were found during 2 or more of the examinations, a few animals having such lesions at the same site on both or all occasions.Animals in Herd A became infected through grazing together with goats from infected herds. Caseous lymphadenitis was introduced into Herd B by animals obtained from infected herds.     

Corynebacterium pseudotuberculosis infection in goats. V. Relationship between the infection and lesions resulting from vaccination against paratuberculosis

In 2 goat herds, one infected with Gorynebacterium pseudo-tuberculosis and one free from the infection,, goats were examined for superficial swellings on the shoulder and chest. All animals in this study had been vaccinated against paratuberculosis before the age of 4 weeks. The vaccine had been applied subcutaneously behind the shoulder. Twenty-two of 40 (55 %) and 31 of 45 (69 %) goats had such lesions in the infected and non-infected herds, respectively. The difference between the herds was not significant, P > 0.05.Swellings found behind the shoulder in 19 goat carcasses derived from 4 herds in which G. pseudotuberculosis infection occurred were examined bacteriologically. No bacteria could be isolated from such lesions in 15 animals, while G. pseudotuberculosis in pure culture was isolated from 3 carcasses, and a mixed bacterial flora from the re-maining carcass. Bacteria could not be isolated from lesions situated behind the shoulder in 7 carcasses from 3 herds free from G. pseudo-tuberculosis infection.It is concluded that most swellings on the shoulder and chest in goats were granulomas resulting from vaccination against paratuber-culosis.     

A serologic method for the detection of Corynebacterium pseudotuberculosis infections in horses

A serologic technique useful for detecting antibodies formed in horses in response to infection with Corynebacterium pseudotuberculosis is described. The test relies on the ability of C. pseudotuberculosis toxin to produce a wide zone of hemolysis when applied to erythrocytes previously treated with a sterile filtrate of Corynebacterium equi broth culture. The synergistic hemolytic activity can be neutralized by anti-C. pseudotuberculosis serum. This test was used to analyze sera from 616 horses for the presence of C. pseudotuberculosis antitoxin. Of 177 animals (see Table 2) found positive, there were 34 horses with bacteriologically confirmed, active infections and 18 with active but unconfirmed infections. In addition, 13 animals had a history of having had the disease and 112 had no history or evidence of having had the infection. The other 439 horses had negative titers. Statistical treatments confirmed the value of the test as an epidemiological tool but precluded using only titers for the diagnosis of active clinical disease.     

Experimental Corynebacterium pseudotuberculosis infection in lambs

Lambs were inoculated IV with 3.2 X 10(3) colony forming units (CFU) to 3.2 X 10(6) CFU of Corynebacterium pseudotuberculosis from a 6-hour broth culture supplemented with 0.1% sorbitan monooleate. After 28 days, multiple abscesses were observed in the lungs and lymph nodes. The number of abscesses in the lungs correlated with the inoculation dose. Two lambs given 10(5) CFU and 10(6) CFU died. Multiple abscesses occurred in other lambs given 10(6) CFU to 10(4) CFU and few abscesses occurred in lambs given 10(3) CFU. Corynebacterium pseudotuberculosis was isolated from lung abscesses, inoculation site abscesses, and lymph node abscesses, but not from normal tissues. Because this procedure consistently induced abscesses in the lungs, we believe it will be a suitable challenge system for studies on the prevention of caseous lymphadenitis in sheep.     

Corynebacterium pseudotuberculosis infection in goats. II. The prevalence of caseous lymphadenitis in 36 goat herds in Northern Norway

The precalen-ce of caseous lymphadenitis was surveyed in 36 goat herds in Northern Norway. In each herd, information concerning the occurrence of the disease was obtained from the farmer. Adult animals (1 year of age or older) in 35 herds were examined for superficial swellings, and serum samples were collected from most animals in the herds. The sera were examined for antibodies to Corynebacterium pseudotuber-culosis using the bacterial agglutination test (BAT) and the hemolysis inhibition test (HIT).Gaseous lymphadenitis was diagnosed with certainty in 19 herds. Information from the farmers indicated that the disease indeed oc-curred in these herds, and that the majority had been infected with the disease for many years. The herds had apparently become infected through contact with animals from infected herds. Clinical examina-tions were carried out in 18 of these herds and superficial swellings were found in 26 % of the examined animals. The prevalence of ani-mals with lesions varied from 11 to 40 % among the herds. Of the animals in these herds, 81 % were positive in BAT and 84 % in HIT. The prevalence of positive animals varied from 26 to 99 % in BAT and 28 to 99 % in HIT. The prevalence of seropositive animals was lowest in a herd in which animals were kept separately in stalls.Caseous lymphadenitis could not be diagnosed in 16 herds. In-formation from the farmers indicated that the disease indeed seemed to be absent in 14 of these herds. These 14 herds had no history of contact with animals from herds considered to be infected. However, in the remaining 2 herds, the farmers were somewhat uncertain about the occurrence of the disease. One of these 2 herds had a history of contact with infected herds through participation in a goat “breeding circle”. Only a few of the animals were, however, seropositive and all these had low antibody titres.In 1 newly established herd, a single animal showed a high posi-tive titre in BAT only. All the other animals were negative in both tests. This particular herd consisted of animals obtained both from herds with caseous lymphadenitis and from herds in which the disease was not considered to occur.     

Development of an ELISA for the diagnosis of Corynebacterium pseudotuberculosis infections in goats

An ELISA was developed for the diagnosis of Corynebacterium pseudotuberculosis infections in goats. A bacterial whole cell extract was used as solid-phase antigen, and serum from a culture-positive animal served as the internal reference standard. The well-to-well and assay-to-assay variations were determined to be 12.7 and 33.0%, respectively. A cut off value was determined by parallel testing of 142 sera (112 ELISA-positive, 30 ELISA-negative) in a Western blot, and the correlation between both tests was highly significant (K=0, 93). In addition, the reliability of the ELISA for the detection of infected herds was proven in a double blind study testing 910 sera from 74 goat herds.     

Post-mortem examination in the diagnosis of Johne's disease in goats.

Post-mortem examinations play an important role in Johne’s disease programmes in Norway. The results of such examinations of samples of viscera from 2997 goats carried out during the 5-year period 1972–1976 are given. The investigations show that the demonstration of macroscopical changes in mesenteric lymph nodes and small intestine has only limited value as a guideline in the post-mortem diagnosis of Johne’s disease in goats. Often macroscopical changes were not seen or they were non-specific. Caseous and/or calcified foci in mesenteric lymph nodes in infected animals were demonstrated quite often whilst observed intestinal changes were strikingly few. Corrugation of the mucosa was rare. However, in sections of macroscopically unchanged intestine marked epithelioid cell infiltrations and abundant acid-fast bacilli were not uncommon. In sporadic cases productive inflammation with tubercle formation was seen in lymph nodes in infected animals.Bacteriological culture was by far the most reliable post-mortem diagnostic method. By this method 92% of the infected goats were detected. The corresponding figures for histological examination and microscopy were 54% and 47%, respectively.     

Identification of Corynebacterium pseudotuberculosis isolates from sheep and goats by PCR

The present study was carried out to estimate the prevalence of caseous lymphadenitis (CL) in sheep and goats slaughtered at the local abattoir in Elazig province located in the east of Turkey, between September and December 2000. A total of 2046 sheep and 2262 goat carcasses were examined during the study period and 118 abscessed lymph nodes, 89 from sheep and 29 from goats, were collected. Corynebacterium spp. strains were isolated from 81.4% of the abscesses, giving an overall prevalence of 2.2%. The prevalence was 3.5 and 1.1% in sheep and goats, respectively. PCR on DNA extracted from 96 suspicious isolates, using a pair of Corynebacterium pseudotuberculosis-specific primers, was positive for 93. Although cross-reaction with C. ulcerans, a human/bovine species, was observed, the PCR assay used in this study may successfully be applied for the diagnosis of CL in goats and sheep as an alternative to conventional methods, owing to its advantages of specificity and speed.     

Comparison of an interferon-gamma to a phospholipase D enzyme-linked immunosorbent assay for diagnosis of Corynebacterium pseudotuberculosis infection in experimentally infected goats

The optimal method of control of caseous lymphadenitis of goats caused by Corynebacterium pseudotuberculosis is eradication of infection by identification and removal of infected carrier animals. The objective of this study was to compare detection of C. pseudotuberculosis experimentally infected goats using a commercially available bovine interferon-gamma (IFN-gamma) whole blood enzyme-linked immunosorbent assay (ELISA) to serological response to a recombinant phospholipase D (PLD) ELISA. The tests were assessed repeatedly over 1 year in three infected and three non-infected goats. Using a IFN-gamma optical density cut-off at 0.10 as positive under the conditions used, the test accurately detected C. pseudotuberculosis experimentally infected goats over a 363 day period with a reliability of 89.2% and non-infected goats with a reliability of 97.1%. Using a cut-off value of the mean for negative samples plus two standard deviations, the PLD ELISA detected C. pseudotuberculosis experimentally infected goats over this period with a reliability of 81.0% and non-infected goats with a reliability of 97.0%. The PLD ELISA was however more predictive than the IFN-gamma ELISA of the presence of lesions observed at postmortem examination of infected goats.     

Determining the sensitivity of abattoir surveillance for ovine Johne's disease

OBJECTIVE: The objective of this study was to determine the sensitivity of abattoir surveillance of intestinal tract lesions for detecting ovine Johne's disease (OJD) under normal meatwork conditions. DESIGN: The design of this study was a diagnostic test validation. The three OJD inspectors were the diagnostic test and follow-up histopathological examination was used for test validation. PROCEDURE: Approximately 1200 sheep were procured from known high prevalence OJD infected farms. The sheep viscera were tagged (numbered) and then examined as they were processed on the abattoir line by three experienced meat inspectors. Their observations were independently recorded on a cassette tape. Specified sections of viscera were prepared and subjected to histopathological examination and these results were compared with the inspector diagnoses. RESULTS: The sensitivity of abattoir inspection for OJD varied between inspectors from 53 percent to 87 percent. The specificity varied from 97 to 100 percent. It appeared that the level of sensitivity for detecting disease was higher in lines of sheep where the disease was more prevalent. It also appeared that formal training was an important aspect in ensuring a high level CONCLUSION: Abattoir surveillance is a very economical and rapid method of assessing the OJD status of sheep. On the basis of these results it is reasonable to suggest that abattoir surveillance has a sensitivity of approximately 70 percent. This technique is useful as an ancillary to other testing regimes for negative assurance programs where a sheep identification system is used.