Estimates of within-herd incidence rates of Mycobacterium bovis in Canadian cattle and cervids between 1985 and 1994

We analysed the individual-animal data from six of the nine outbreaks of tuberculosis in Canadian cattle and cervids from 1985 to 1994. A “positive/reactor” animal was one which had either a positive culture or a positive or suspicious reaction on a mid-cervical, comparative cervical, or gross or histopathological test for tuberculosis. Individual-animal data were collected only for herds which had one or more positive/reactor animals. Data were collected from the outbreak records in the Regional or District offices of Agriculture and Agri-food Canada’s Animal and Plant Health Directorate. The within-herd spread of Mycobacterium bovis was studied by determining the most-likely date at which the herd was first exposed to M. bovis and the number of reactions which had developed by the time the herd was investigated. The animal-time units at risk in the herd were probably overestimated, resulting in conservative estimates of the within-herd incidence rates. Negative-binomial regression was used to investigate factors which might have influenced the within-herd spread of tuberculosis. Increasing age appeared to be a risk factor for being a positive/reactor animal. When compared to animals 0–12 months old, animals 13–24 months old had an incidence rate ratio (IRR) of 7.6, while animals >24 months old had an IRR of 10.4 (p=0.009). Actual and predicted incidence rates for tuberculosis in mature (>24 months old) animals were calculated. Actual and predicted incidence rates were similar for cervids, within an outbreak. There was more variability between actual and predicted rates in the dairy and beef animals. In the one outbreak (Ontario) where there were positive/reactor cervid, dairy and beef herds, the actual incidence rate for cervids (IR=9.3 cases per 100 animal-years) was almost twice that of dairy cattle (IR=5.0) and three times that of beef cattle (IR=3.1).     

Estimates of within-herd incidence rates of Mycobacterium bovis in Canadian cattle and cervids between 1985 and 1994

We analysed the individual-animal data from six of the nine outbreaks of tuberculosis in Canadian cattle and cervids from 1985 to 1994. A “positive/reactor” animal was one which had either a positive culture or a positive or suspicious reaction on a mid-cervical, comparative cervical, or gross or histopathological test for tuberculosis. Individual-animal data were collected only for herds which had one or more positive/reactor animals. Data were collected from the outbreak records in the Regional or District offices of Agriculture and Agri-food Canada’s Animal and Plant Health Directorate. The within-herd spread of Mycobacterium bovis was studied by determining the most-likely date at which the herd was first exposed to M. bovis and the number of reactions which had developed by the time the herd was investigated. The animal-time units at risk in the herd were probably overestimated, resulting in conservative estimates of the within-herd incidence rates. Negative-binomial regression was used to investigate factors which might have influenced the within-herd spread of tuberculosis. Increasing age appeared to be a risk factor for being a positive/reactor animal. When compared to animals 0–12 months old, animals 13–24 months old had an incidence rate ratio (IRR) of 7.6, while animals >24 months old had an IRR of 10.4 (p=0.009). Actual and predicted incidence rates for tuberculosis in mature (>24 months old) animals were calculated. Actual and predicted incidence rates were similar for cervids, within an outbreak. There was more variability between actual and predicted rates in the dairy and beef animals. In the one outbreak (Ontario) where there were positive/reactor cervid, dairy and beef herds, the actual incidence rate for cervids (IR=9.3 cases per 100 animal-years) was almost twice that of dairy cattle (IR=5.0) and three times that of beef cattle (IR=3.1).     

Evaluation of single reactor bovine tuberculosis breakdowns based on analysis of reactors slaughtered at an Irish export meat plant

The 'Singleton Protocol' was adopted by the Irish Department of Agriculture Fisheries and Food (DAFF) in 1996 to address the incomplete specificity of the single intra-dermal comparative tuberculin test (SICTT) used in Ireland for the detection of animals infected with bovine tuberculosis (bTB). The protocol allows the early restoration of disease-free status to herds with a single reactor breakdown, where the herd was not confirmed as infected with Mycobacterium bovis by epidemiological investigation, by postmortem examination or by further test. The current study examines the ability of the Singleton Protocol to identify false-positive reactors. It investigates the subsequent herd-reactor rate following single reactor removal and analyses the factors leading to a positive postmortem lesion outcome and a positive reactor retest result. Postmortem lesion results were obtained for 371 reactor animals from single reactor breakdowns that were killed at an export meat plant over a 19-month period. Epidemiological and test data for these animals and their herds were obtained from DAFF databases and analysed by univariate and multivariate statistical analysis. Singleton candidates had an 18.7 per cent lower lesion rate than single animal breakdowns not meeting the singleton criteria. No significant difference was found between Singletons and non singletons in the subsequent reactor retest results. Skin thickness at the SICTT is the most significant determinant of a positive lesion result. The area bTB history was shown to be a significant variable in producing a positive reactor retest result.     

A Cross-sectional Study of Bovine Tuberculosis in Dairy Farms in Asmara,Eritrea

Bovine tuberculosis in dairy cattle in Asmara, Eritrea, was studied using a cross-sectional study to describe its prevalence and to identify factors associated with it. A total of 72 randomly selected herds were included in the study. The comparative intradermal tuberculin test was used for the diagnosis. Of 1813 individual animals tested, 14.5% were reactors. Thirty herds (41.7%) had at least one reactor but, by defining a reactor herd as any herd with two or more reactors, only 19 (26.4%) herds were classified as reactor herds. Based upon individual animal specificity of 98.5%, the calculated herd specificity was >99%. A multiple logistic model showed that the presence of exotic breeds was associated with a high risk (odds ratio = 5.70; 95% confidence interval 1.13–28.8). An increased risk was also linked to large herds. Keeping the animals always indoors reduced the risk, but could not be fitted to the model owing to empty cells.     

Fluorescence polarization assay for the detection of antibodies to Mycobacterium bovis in bovine sera

The performance of a fluorescence polarization assay (FPA) that detects antibodies to Mycobacterium bovis in bovine sera is described. The FPA reported here is a direct binding primary screening assay using a small polypeptide derived from the M. bovis MPB70 protein. A secondary inhibition assay confirms suspect or presumed positive samples. Specificity studies involved five different veterinary laboratories testing 4461 presumed negative bovine samples. FPA specificity was 99.9%. The FPA was used to identify herd status as either M. bovis infected or non-infected. Herd surveillance studies (nine herds) were performed in Mexico and South Africa. The FPA had a specificity of 100% (two negative herds), and correctly identified six of seven infected herds. Finally, sera from 105 slaughter animals that had gross lesions in lymph nodes similar to those seen with bovine tuberculosis were tested by the FPA. Thin sections from the associated formalin-fixed paraffin-embedded samples of lymph nodes were stained using hematoxylin and eosin (H&E) for morphologic examination and using the Ziehl-Neelsen (ZN) method for detection of acid-fast bacilli. Of the 105 animals, 78 were classified as TB suspect based on lesion morphology, 21 were positive by ZN, 9 were positive by FPA and 13 were positive by PCR for the tuberculosis group of Mycobacterium. Among the 21 ZN positives, 11 (52.4%) were PCR positive. Among the 9 FPA positives, 8 (88.9%) were PCR positive. For the 13 PCR positives, 8 (61.5%) were FPA positive and 11 (84.6%) were ZN positives. These results show that use of the FPA for detection of M. bovis infection of cattle has value for bovine disease surveillance programs.     

A case-control study on the association of selected risk factors with the occurrence of bovine tuberculosis in the Republic of Ireland

We did a case-control study on the association between the occurrence of bovine tuberculosis in herds in the East Offaly area and a number of possible risk factors: herd size, the purchasing of cattle, the herd history, the status of contiguous herds, distance to the nearest badger sett, and animal factors such as age, breed and animal type. The characteristics of 100 herds that had a confirmed outbreak of tuberculosis at a herd test in 1988 were compared with those from an equal number of herds that passed at least one tuberculin test during that year. The study base consisted of 1379 herds mainly from the eastern part of County Offaly. The data were analysed at both herd and animal levels. Factors that were associated with an increased risk of tuberculosis breakdowns at the herd level were (i) the total number of animals in the herd and (ii) the restriction of a contiguous herd for tuberculosis within a period of 6 months prior to or following the herd under study. A number of factors were associated with an animal having an increased risk of failing a tuberculin test. After adjusting for all other variables in the model, cows, heifers and bullocks were 10.3, 14.8 and 8.0 times, respectively, more likely to fail a tuberculin test than calves. Cattle that had teen purchased since the previous herd test were less likely to fail a tuberculin test compared with cattle that had been present at the time of the preceding herd test.     

Tuberculosis in goats on a farm in Ireland: epidemiological investigation and control

This paper describes an outbreak of tuberculosis (TB) caused by Mycobacterium bovis in a dairy goat herd on a farm in Ireland, where 66.3 per cent of the herd tested positive to the single intradermal comparative tuberculin test (SICTT) at initial detection. An epidemiological investigation was conducted to determine the origin of the outbreak, considering issues such as animal movements and herd management practices. Infection was introduced with a consignment of goats, as determined by the variable number tandem repeat profile. Infection was eradicated using a test and cull programme involving the SICTT, the interferon-γ assay and a multiplex immunoassay (Enferplex TB).     

Paratuberculosis sero-status and milk production,SCC and calving interval in Irish dairy herds

The objective of this study was to investigate the impact of paratuberculosis sero-status on milk yield, fat, protein, somatic cell count and calving interval in Irish dairy herds. Serum from all animals over 12 months of age (n = 2,602) in 34 dairy herds was tested for antibodies to Mycobacterium avium subsp. paratuberculosis using an ELISA. Herds were categorised by sero-status into positive, non-negative and negative, where a positive herd contained two or more positive cows, a non-negative herd contained only one positive cow and a negative herd contained no positive cows. Data at animal, parity and herd-level were analysed by multiple regression using general linear models. Positive herds (mean herd size = 129 cows) and non-negative herds (81 cows) were larger than negative herds (72 cows) (P < 0.01). Negative herds had the highest economic breeding index (EBI), while positive herds had the highest estimated breeding value (EBV) for milk yield. There was no significant effect of paratuberculosis sero-status at animal, parity or herd-level on milk yield, milk fat or protein production, somatic cell count score (SCCS) or calving interval. Negative herds tended to have a lower SCCS than positive and nonnegative herds (P = 0.087). This study only examined the effects of paratuberculosis sero-status but did not examine the clinical effects of Johne's disease at the farm or dairy industry levels.     

Effect of paratuberculosis on the diagnosis of bovine tuberculosis in a cattle herd with a mixed infection using interferon-gamma detection assay

Interferon-gamma (IFN-gamma) detection assay is being applied as an ancillary test to tuberculin tests in the diagnosis of bovine tuberculosis to detect the maximum number of infected animals. Among possible factors influencing the performance of tuberculosis-diagnostic tests, paratuberculosis, a widespread disease in Spain and other European countries, has been pointed out as a cause of false positive reactions. Still, its effect on the sensitivity of these tests in cattle has yet to be fully characterized. The impact of paratuberculosis in the apparent sensitivity of IFN-gamma assay was studied in a bullfighting cattle herd with a mixed tuberculosis-paratuberculosis infection, using culture of Mycobacterium bovis and Mycobacterium avium paratuberculosis as the gold standard to determine the infection status of every animal. A total of 218 animals were slaughtered and sampled for bacteriology after blood sampling. IFN-gamma assay showed a lower apparent sensitivity in animals with a mixed infection (50%) compared to all animals suffering tuberculosis (78.3%). This finding indicates that the presence of paratuberculosis in tuberculosis-infected herds could imply a serious impairment in the sensitivity of IFN-gamma detection test.     

An outbreak of Mycobacterium bovis infection in fallow deer (Dama dama)

In an outbreak of Mycobacterium bovis infection in fallow deer in South Australia, 3 herds related by recent movement of deer were infected. From these 3 infected herds, 47 of 51 animals were tuberculosis at necropsy. A range of lesions was seen most of which differed from classical bovine tuberculosis in that pus was a white liquid, fibrous encapsulation was not marked and calcification was rare. Histopathology was of classical tuberculosis. M. bovis was cultured from lesions and M. avium-intracellulare was cultured from one deer with no visible lesions. The source of M. bovis infection has not been determined.     

Prevalence of Mycobacterium bovis infection in cervids on privately owned ranches

OBJECTIVE: To determine prevalence of tuberculosis caused by infection with Mycobacterium bovis in cervids on privately owned ranches in northeastern lower Michigan. DESIGN: Epidemiologic survey. ANIMALS: Cervids on 96 privately owned ranches. PROCEDURES: A combination of slaughter and skin tuberculin testing was used to collect data. Infection with M. bovis was confirmed by use of standard necropsy and bacteriologic culture techniques. RESULTS: Cervids with tuberculosis were detected on 1 of the 96 ranches. The apparent prevalence of tuberculosis in cervids from the 96 ranches was 1.1 cases/100 cervids (21 cases/1,867 cervids tested). For the ranch with infected cervids, prevalence of infection with M. bovis was 12.1 cases/100 cervids (21 cases/174 cervids tested). No obvious gross lesions were seen in 8 of 21 white-tailed deer and 1 coyote with culture-confirmed M. bovis infection. CONCLUSIONS AND CLINICAL RELEVANCE: The lack of visible lesions in a substantial proportion of infected animals should be taken into consideration in studies involving detection and prevalence of tuberculosis.     

Controlling tuberculosis in a llama (Lama glama) herd using clinical signs, tuberculin skin testing and serology

An outbreak of tuberculosis (TB), caused by Mycobacterium bovis, was investigated in a small herd of llamas (Lama glama). Based on three ante-mortem diagnostic methods (clinical signs, tuberculin skin test reactions, and 'Rapid Test' serology), 12 llamas were selected for examination post-mortem. Grossly visible lesions suspicious of TB were observed in eight animals, four of which had exhibited clinical signs, one was a skin test 'reactor', and three had been seropositive. M. bovis was isolated from seven of these eight animals. Clinical signs combined with serology were found to be useful in identifying infected animals, but tuberculin skin testing had limited negative predictive value as four llamas that were subsequently confirmed as infected were not detected using this assay.     

A field study of Mycoplasma bovis infection in cattle

After an outbreak of mastitis in cattle caused by Mycoplasma bovis a study was made in 5 herds with recent cases (principal herds) and in 4 control herds. In the principal herds, M. bovis was isolated from milk samples, nasal swabs, and from one vaginal swab. M. bovis was also isolated from nasal swabs of calves in 2 of the 4 control herds, whereas all milk samples and vaginal swabs from the control herds were negative. Evaluation of serum antibody titres to M. bovis among non-mastitic animals of 3 principal herds and 1 control herd showed no difference in distribution of the titre values, which generally were low. However, cows excreting M. bovis in the milk had high antibody titres. The way of introduction to the herds and the spread of the infection within the herds could not be established by the study, which was supplemented by a DNA restriction fragment analysis of a number of M. bovis isolates.     

Management-related risk factors for M. paratuberculosis infection in Michigan, USA, dairy herds

A cross-sectional study was conducted from June through December 1996 to identify management-related risk factors for herd-level M. paratuberculosis infection. Data were collected from 121 participating herds. A two-part questionnaire was administered to gather data on current and previous management practices and herd productivity. A random sample of cows aged ≥24 months was selected from each herd and tested for antibodies to M. paratuberculosis using the IDEXX Antibody ELISA (sensitivity 64%, specificity 96%). A positive herd was one in which ≥2 animals tested positive for antibodies to M. paratuberculosis. A negative herd was one in which no animal tested positive. Herds in which only one animal tested positive were dropped from statistical analysis to reduce the risk of including false-positive herds in the statistical analyses.

There were 80 herds with one or more positive animals and 41 herds with no positive animals in the sample (66% herd-level prevalence). Twenty-six herds (21%) were dropped from further analyses because they had only one positive cow. Twelve herds (10%) were dropped from analysis because of missing data. The resulting sample used for statistical modeling included 46 positive herds and 37 negative herds (55% herd-level prevalence). A multi-variable logistic-regression model was used to evaluate the results. The variable ‘use of an exercise lot for lactating cows' was associated with a three-fold increase in odds of a herd being positive for M. paratuberculosis infection (O.R.=3.01, C.I.=1.03–8.80); ‘cleaning of maternity pens after each use' was associated with a three-fold reduction in odds of a herd being positive for M. paratuberculosis infection (O.R.=0.28, C.I.=0.08–0.89); ‘application of lime to pasture areas in 1993' resulted in a ten-fold decrease in odds of a herd being positive for M. paratuberculosis infection (O.R.=0.10, C.I.=0.02–0.56).     

Tuberculosis outbreak in a dromedary racing herd and rapid serological detection of infected camels

A recent outbreak of tuberculosis (TB) in a dromedary racing herd of 58 animals involved 3 infected animals. Disease was confirmed at necropsy by finding gross lesions from which Mycobacterium bovis (antelope type) was isolated. Sera collected from the camels in this herd were used to evaluate two new serological methods, Multiantigen Print Immunoassay (MAPIA) and rapid test (RT) developed using the lateral-flow technology, in comparison with the intradermal tuberculin tests. Antibodies were found in all three infected dromedaries by both RT and MAPIA, but not in the remaining 55 animals in the herd. With the limited number of animals tested in this study, the serological assays showed the potential for convenient, rapid, and accurate diagnosis of TB in live camels.     

Assessment of diagnostic tools for eradication of bovine tuberculosis in cattle co-infected with Mycobacterium bovis and M. avium subsp. paratuberculosis

The intradermal tuberculin (IDTB) test and the interferon-gamma (IFN-gamma) assay are used worldwide for detection of bovine tuberculosis in cattle, but little is known about the effect of co-infecting agents on the performance of these diagnostic tests. This report describes a field trial conducted in a cattle herd with dual infection (bovine tuberculosis and paratuberculosis) during 3.5 years. It has been based on a strategic approach encompassing serial parallel testing (comparative IDTB test, the IFN-gamma assay and serology of paratuberculosis) that was repeated 8 times over the period, and segregation of animals into two herds. The IDTB test detected 65.2% and the IFN-gamma test detected 69.6% of the Mycobacterium bovis culture-positive cattle. However, the IDTB test performed better during the first part of the trial, while the IFN-gamma test was the only method that detected infected animals during the following three samplings. The number of false positive reactors with the IDTB and/or the IFN-gamma tests was remarkably high compared to other reports, and could be caused by cross-reactivity with M. avium subsp. paratuberculosis. Also, the M. bovis isolates from cattle and wildlife from the same property were characterised using molecular techniques to disclose an epidemiological link. The IDTB test may not be appropriate to eradicate bovine tuberculosis in herds with dual mycobacterial infections. This report highlights the need to use several diagnostic techniques for the accurate detection of M. bovis infected animals in these herds.     

The seroprevalence of Mycoplasma bovis in lactating cows in Switzerland, particularly in the republic and canton of Jura]

Sporadic cases of infection with Mycoplasma bovis have been observed in Switzerland since 1983. However, five severe outbreaks of endemic mastitis in a geographically distinct, small region (Canton du Jura) during the period 1995 to 1997 prompted the present investigation on the seroprevalence of infection with M. bovis among milking cows in Switzerland. A commercially prepared indirect enzyme immunoassay was used. Among a stratified random sample of 118 herds of milking cows in Switzerland, at least one positive animal was detected in 56 (47%) of the herds, whereas 6.1% of the 1816 individual animals tested positive. An epidemiological study was performed in the Canton du Jura region among 51 herds in order to assess the importance of management factors in the spread of M. bovis infection. The herd-level prevalence was 78%, and the seroprevalence at the level of the 1354 individual animals tested was 13.4%. A multivariate analysis of possible risk factors showed purchase of animals to be the only variable significantly associated with serological status of the herd with an "odds ratio" of 10.8.     

Multiple strains of Mycobacterium bovis revealed by molecular typing in a herd of cattle

Mycobacterium bovis isolates from an outbreak of bovine tuberculosis in a herd of cattle in Rio de Janeiro, Brazil, were analysed by spoligotyping and variable-number tandem repeat PCR analysis of the mycobacterial interspersed repetitive unit and exact tandem repeats. Molecular typing revealed a high genetic diversity of strains in the herd. The genetic diversity could be explained by the introduction of infected animals from different sources.     

Effect of skin testing and segregation on the prevalence of bovine tuberculosis, and molecular typing of Mycobacterium bovis, in Ethiopia

In 2002, the prevalence of bovine tuberculosis (tb) among 500 cattle on Holeta Farm, near Addis Ababa, Ethiopia, was 48 per cent, and the farm was divided into positive and negative herds. After three consecutive rounds of skin testing and segregation of skin test-positive and -negative animals, the prevalence of bovine tb was reduced from 14 per cent to 1 per cent in the negative herd in a year. Spoligotyping of 41 isolates from 17 cows gave an identical and unique spoligotype pattern, which can be represented as the binary number 1100000101111110111111100010000000000100000, where 1 indicates the presence of a spacer and 0 represents a loss. This spoligotype pattern had not previously been reported on the Mycobacterium bovis spoligotype database, and it was therefore designated SB1176, Ethiopian M bovis strain 1 (EMbs1). The variable number tandem repeat (VNTR) profile of the strain was 5254(*)33.1, which differed from the VNTR profile of strains reported in Great Britain.     

Blood culture and stimulation conditions for the diagnosis of tuberculosis in cervids by the Cervigam assay

Mitogen- and antigen-induced interferon-gamma (IFN-gamma) responses of peripheral blood leucocytes from cervids were evaluated by a commercial whole-blood assay. The assay was applied to Mycobacterium bovis-infected white-tailed deer and reindeer, M bovis BCG-vaccinated white-tailed deer and elk, and unvaccinated, uninfected white-tailed deer, fallow deer, elk and reindeer. The responses of the M bovis-infected white-tailed deer to pokeweed mitogen (PWM) varied with time and between individuals. The responses of the M bovis-infected reindeer to PWM and M bovis purified protein derivative (PPD) were positively associated. Samples from tuberculosis-free captive herds in various parts of the USA were also evaluated. Four per cent of fallow deer, 20 per cent of elk, 44 per cent of white-tailed deer, and 91 per cent of reindeer had responses to PWM exceeding 0.25 Delta optical density, that is, PWM stimulation minus no stimulation. The specificity of the responses to M bovis PPD and a Mycobacterium tuberculosis complex-specific antigen rESAT-6:CFP-10, excluding animals not responding to PWM, ranged from 78 per cent to 100 per cent and was dependent upon the species and the positive response cut-off value. The results show that the commercial assay is valid for the detection of TB in reindeer; however, further development of the assay will be required before it is used in surveillance programmes for white-tailed deer, fallow deer, and elk.