Shorter-term risk of Mycobacterium bovis in Irish cattle following an inconclusive diagnosis to the single intradermal comparative tuberculin test

In Ireland, new cases of bovine tuberculosis (bTB) are detected using both field and abattoir surveillance (More and Good, 2006). Field surveillance is conducted through annual testing of all cattle using the single intradermal comparative tuberculin test (SICTT). An animal may be deemed a 'standard inconclusive reactor' (SIR) to the SICTT if the bovine response is >2mm and between 1 and 4mm>the avian response. The herdowner then has three choices for the management of the SIR: option 1 is to have the animal retested after a minimum period of 42 days (an inconclusive reactor retest, IRR), option 2 is to slaughter the SIR and, provided the animal has no visible lesions, have a full herd test 42 days after the SIR leaves the herd, option 3 is to slaughter the SIR and have the lymph nodes examined using histology and/or culture for bTB. In the current study, we examine the bTB risk for SIRs both at slaughter prior to the IRR and at the IRR, and the future bTB risk of TIR animals (so-called 'transient SIRs'; SIR animals with a negative SICTT result at the subsequent IRR) that moved from the herd of disclosure within 6 months of the IRR. We also investigate factors associated with the future bTB status of SIRs at slaughter prior to the IRR and at the IRR. The study population included all SIRs identified in Ireland between 2005 and 2009 inclusive in a herd otherwise Officially TB free (OTF). Between 11.8% and 21.4% of SIRs slaughtered prior to the IRR were confirmed bTB positive at post mortem (using histology or culture if histology was not definitive), compared to 0.13-0.22% of SICTT -ve cohort animals. The post mortem bTB lesion rate of SIRs is lower than the lesion rate reported for reactor animals between 2005 and 2009 of between 34% and 39%, reflecting the doubtful infection status of these animals. Between 20.3% and 27.9% of herds were restricted at the IRR. The herd restriction rate amongst the national herd between 2005 and 2009 varied from 5.09% to 6.02%. TIRs that moved out of the disclosing herd within 6 months of the IRR were 12 times more likely to be bTB positive at the next test/slaughter compared to all animals in the national herd. The same increased risk did not apply to the SICTT -ve cohort animals that moved out of the same herds at the same time. Based on a range of measures, SIRs and TIRs are each at increased bTB risk into the future. Consequently, differential treatment of TIR animals would be justified.     

Bovine tuberculosis and udder health in Irish dairy herds

The association between bovine tuberculosis (TB) infection status based on results from the single intradermal comparative tuberculin test (SICTT) and milk production has been described in dairy cows in TB-infected herds in Ireland. The biological basis was uncertain, but could be related to increased TB susceptibility among lower producing dairy cows. In this study, the relationship between somatic cell count (as an objective measure of udder health) and SICTT reactivity (as a proxy for TB infection status) was investigated. Somatic cell counts of TB infected cows, both during and prior to the lactation of diagnosis of TB infection, were examined and compared to non-infected cows. All Irish dairy herds restricted from trading between June 2004 and May 2005 as a result of two or more TB reactors (test positive) to the SICTT were considered for study. Data were collected on 4340 cows from 419 herds. Previous lactation data for the cows were taken into consideration and all lactations on a cow were analysed together with the years of lactations. There was an inherent hierarchical structure in the data, with lactations nested within cows and cows within herds and so a linear mixed model with two random effects was used to describe the data. Milk production (305-day milk yield) was also included in the model as a fixed effect. The results of the study showed that for all lactations and years under investigation, somatic cell counts for SICTT reactor cows when compared to the non-reactor cows were not significantly different. In this study population, TB infection status was not associated with udder health.     

Using latent class analysis to estimate the test characteristics of the γ-interferon test, the single intradermal comparative tuberculin test and a multiplex immunoassay under Irish conditions

Considerable effort has been devoted to improving the existing diagnostic tests for bovine tuberculosis (single intradermal comparative tuberculin test [SICTT] and γ-interferon assay [γ-IFN]) and to develop new tests. Previously, the diagnostic characteristics (sensitivity, specificity) have been estimated in populations with defined infection status. However, these approaches can be problematic as there may be few herds in Ireland where freedom from infection is guaranteed. We used latent class models to estimate the diagnostic characteristics of existing (SICTT and γ-IFN) and new (multiplex immunoassay [Enferplex-TB]) diagnostic tests under Irish field conditions where true disease status was unknown. The study population consisted of herds recruited in areas with no known TB problems (2197 animals) and herds experiencing a confirmed TB breakdown (2740 animals). A Bayesian model was developed, allowing for dependence between SICTT and γ-IFN, while assuming independence from the Enferplex-TB test. Different test interpretations were used for the analysis: SICTT (standard and severe interpretation), γ-IFN (a single interpretation), and a range of interpretations for the Enferplex-TB (level-1 [high sensitivity interpretation] to level-5 [high specificity interpretation]). The sensitivity and specificity (95% posterior credibility intervals; 95% PCI) of SICTT[standard] relative to Enferplex-TB[level-1] and γ-IFN were 52.9-60.8% and 99.2-99.8%, respectively. Equivalent estimates for γ-IFN relative to Enferplex-TB[level-1] and SICTT were 63.1-70.1% and 86.8-89.4%, respectively. Sensitivity of Enferplex-TB[level-1] (95% PCI: 64.8-71.9%) was superior to the SICTT[standard], and specificity of the Enferplex-TB[level-5] was superior to γ-IFN (95% PCI: 99.6-100.0%). These results provide robust measures of sensitivity and specificity under field conditions in Ireland and suggest that the Enferplex-TB test has the potential to improve on current diagnostics for TB infection in cattle. The extent of that potential will be assessed in further studies.     

Application of the Enfer chemiluminescent multiplex ELISA system for the detection of Mycobacterium bovis infection in goats

A study was conducted to optimise a multiplex serological immunoassay for use in identification of goats infected with Mycobacterium bovis. To assess assay specificity, 31 goats with a history of being free from M. bovis infection were used. To determine assay sensitivity, 180 Single Intradermal Comparative Tuberculin test (SICTT) positive goats were recruited. Additionally, 286 SICTT negative goats classed as potentially exposed animals present in the same positive herds were also included in the study. The results of the assay demonstrated a specificity of 100%. The multiplex assay detected 57/60 SICTT (95.0%) positive animals in one M. bovis infected herd and 120/120 (100%) in a second herd. In a separate experiment, 28 M. caprae culture confirmed infected goats from Spain were assayed, of which 24 (85.7%) were found positive in the test. The results show that inclusion of an antibody based assay can improve the ability to identify M. bovis and M. caprae infected goats. With further development and validation the multiplex assay may prove to be a useful tool for control of M. bovis and M. caprae infection in goats.     

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.     

Modification of the QuantiFERON-TB Gold (In-Tube) assay for the diagnosis of Mycobacterium bovis infection in African buffaloes (Syncerus caffer)

African buffaloes (Syncerus caffer) are the most significant wildlife maintenance hosts of Mycobacterium bovis, the causative organism of bovine tuberculosis (BTB). Current diagnostic tests for the detection of M. bovis infection in free-ranging buffaloes have numerous limitations and we wished to evaluate a modification to a human TB assay, the QuantiFERON-TB Gold (In-Tube) assay (QFT), as a practical diagnostic test for BTB in buffaloes. One hundred and seventy-five buffaloes were tested using the single intradermal comparative tuberculin test (SICTT) and a modified QFT (mQFT). An appropriate cut-off point for the mQFT was derived from SICTT results using receiver operator characteristic curve analysis. Twenty-six SICTT-positive buffaloes were killed and subjected to necropsy, and selected tissues were processed for mycobacterial culture and speciation. An optimal cut-off point for the mQFT was calculated as 66pg/ml. The assay correctly detected 39/40 SICTT-positive buffaloes and 129/134 TST-negative buffaloes and M. bovis was cultured from 21/26 slaughtered SICTT/mQFT-positive animals. The mQFT shows promise as a practical test for M. bovis infection in buffaloes and shows a sensitivity and specificity at least similar to that of the TST.     

The effect of the tuberculin test and the consequences of a delay in blood culture on the sensitivity of a gamma-interferon assay for the detection of Mycobacterium bovis infection in cattle

The strategic use of the gamma-interferon (IFN-gamma) assay (Bovigam) can provide a means for the early identification of Mycobacterium bovis infected cattle, thus ensuring their removal from an infected herd. It has been reported that performance of the test can be influenced by various factors including a recent tuberculin skin test and the length of delay between collection and processing of blood samples. In this study, single intradermal comparative tuberculin test (SICTT) reactor and non-reactor cattle were recruited from herds infected with M. bovis and grouped according to their SICTT responses. Group 1 comprised reactor cattle selected on the basis of their SICTT response to PPD-bovine (purified protein derivative of tuberculin) exceeding that of PPD-avian by at least 12mm. Group 2 animals were selected from herds undergoing routine surveillance for bovine tuberculosis and contained standard SICTT reactor cattle (PPD-bovine exceeding that of PPD-avian by at least 4mm) and non-reactors. We investigated the effects of the SICTT on the assay results by measuring the in vitro IFN-gamma responses of Group 1 reactor cattle at time intervals pre- and post-skin test. No significant differences were measured in the IFN-gamma responses of the reactor animals to PPD-bovine and PPD-avian for up to 65 days. To investigate if a delay in processing of blood affected the performance of the assay, we compared results using duplicate blood samples from Group 1 and Group 2 cattle stimulated with PPD antigen at 8h and at 24h after collection. In both groups of animals the mean optical density (OD) values of the assay at 24h post-collection were significantly lower than those at 8h. Our results demonstrated that a delay in processing of the blood samples from cattle subjected to routine surveillance could significantly impact on the outcome of the IFN-gamma assay resulting in a change of the IFN-gamma status of the animals.     

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 complementary diagnosis of naturally occurring tuberculosis in water buffaloes (Bubalus bubalis) in Rio de Janeiro using a MPB70-ELISA, Brazil

As tuberculosis is still a worldwide infection and buffalo breeding represents an important economic activity in various countries, the purpose of this study was to employ an enzyme-linked immunosorbent assay (ELISA) using MPB70 as a capture antigen for the diagnosis of naturally occurring tuberculosis in water buffaloes in Brazil. After the introduction of newly acquired cattle onto a tuberculosis (TB) free farm, an outbreak of TB was recorded in a mixed herd comprising water buffaloes (21) and cattle (46). The entire herd was tested by intradermal tuberculin injection (ITT) and positive animals were slaughtered and tested by culture, polymerase chain reaction (PCR), and ELISA. From the 21 buffaloes sampled, three were reactive by ITT. All the three had positive culture and ELISA, while PCR was positive in two of them. Besides that, one ITT-negative buffalo was slaughtered and presented positive results by both culture and ELISA, and was considered as anergic. Although there were only few animals, those findings demonstrate the diagnostic usefulness of an MPB70-ELISA to correctly detect Mycobacterium bovis tuberculosis in water buffaloes.     

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.     

Cattle‐associated risk factors for human tuberculosis in rural livestock‐keeping communities,Uganda

Tuberculosis (TB) is a leading infectious cause of human death worldwide. In cattle, TB infection results in productivity losses, trade barriers and zoonotic transmission via milk, meat or direct contact. We conducted a cross‐sectional study in rural communities in Uganda between 2014 and 2016 to evaluate the association between tuberculosis skin test (TST) positivity in humans and cattle‐associated risk factors: household herd positivity and raw milk consumption. Human and cattle TSTs were performed in communities followed by a survey of household practices. TST data were available on 493 humans, 184 (37.3%) with positive results, and 1,441 cattle, 50 (3.5%) with positive results. We fit separate log binomial generalized estimating equation models for the herd positivity‐human TST positivity association, stratified on sex; and for the raw milk consumption‐human TST positivity association, stratified on frequency of milk consumption. Having at least one TST‐positive bovid in the household's herd was significantly associated with lower risk of TB among men (prevalence ratio [PR] 0.66, 95% CI 0.49, 0.87) but was not significantly associated with TB among women (PR 1.21, 95% CI 0.76, 1.95). This apparent protective effect was contrary to our a priori hypothesis of higher exposure effect among men, the primary caretakers of cattle. This finding may be the result of residual confounding by socioeconomic status; wealthier individuals may be less likely to be TB positive, but more likely to have TST‐positive herds by virtue of larger herd sizes, ability to purchase new and possibly infected stock, and propensity to keep more TB‐susceptible European breeds. For raw milk consumption, effect estimates were close to one and not statistically significant. Thus, in settings where bovine TB prevalence is low, such as Uganda, cattle‐associated zoonotic transmission may be rare, and cattle‐associated risk factors may not be important drivers of human TB burden.     

The comparative performance of the single intradermal test and the single intradermal comparative tuberculin test in Irish cattle, using tuberculin PPD combinations of differing potencies

In national bovine tuberculosis (BTB) control programmes, testing is generally conducted using a single source of bovine purified protein derivative (PPD) tuberculin. Alternative tuberculin sources should be identified as part of a broad risk management strategy as problems of supply or quality cannot be discounted. This study was conducted to compare the impact of different potencies of a single bovine PPD tuberculin on the field performance of the single intradermal comparative tuberculin test (SICTT) and single intradermal test (SIT). Three trial potencies of bovine PPD tuberculin, as assayed in naturally infected bovines, namely, low (1192IU/dose), normal (6184IU/dose) and high (12,554IU/dose) were used. Three SICTTs (using) were conducted on 2102 animals. Test results were compared based on reactor-status and changes in skin-thickness at the bovine tuberculin injection site. There was a significant difference in the number of reactors detected using the high and low potency tuberculins. In the SICTT, high and low potency tuberculin detected 40% more and 50% fewer reactors, respectively, than normal potency tuberculin. Furthermore, use of the low potency tuberculin in the SICTT failed to detect 20% of 35 animals with visible lesions, and in the SIT 11% of the visible lesion animals would have been classified as negative. Tuberculin potency is critical to the performance of both the SICTT and SIT. Tuberculin of different potencies will affect reactor disclosure rates, confounding between-year or between-country comparisons. Independent checks of tuberculin potency are an important aspect of quality control in national BTB control programmes.     

A longitudinal study of Neospora caninum infection on a dairy farm in New Zealand

A 600-cow New Zealand dairy herd experienced an abortion storm in 1997 and was monitored (blood sampling at about 3-month intervals) from May 1997 until January 1999. Abortion risk reached 9% in 1997 and was highest in heifers at 19%. The abortion risk decreased in 1998 to 3.2% (still somewhat higher than during the years prior to the outbreak). The serological reaction pattern for Neospora caninum showed an association with abortion risk only around the time of the 1997 outbreak when seropositive cows were 4.2 times more likely to abort than negative ones. Over the whole study period, only 27% of cows that were sampled on all nine visits always tested negative. Offspring from dams which had positive tests for Neospora caninum were 2.4 times more likely to abort than those from dams testing consistently negative. Controlling for age and breed, seropositive cows produced more milk than those that were consistently negative. Infection might have been present endemically within this herd prior to the epidemic, but in 1997 an additional factor appeared to have triggered the outbreak.     

Longer-term risk of Mycobacterium bovis in Irish cattle following an inconclusive diagnosis to the single intradermal comparative tuberculin test

In Ireland, new bovine tuberculosis (bTB) cases are detected using both field and abattoir surveillance. During field surveillance, an animal may be deemed a 'standard inconclusive reactor' (SIR) to the single intradermal comparative tuberculin test (SICTT) if the bovine response is >2mm, and from 1 to 4mm greater than the avian response. Little is known about the future infection risk posed by SIR animals that pass a subsequent retest, so-called 'transient SIR' (TIR) animals. The objective of this study was to critically evaluate the future bTB status of TIR animals, by examining the future risk of bTB diagnosis over the 4 years following initial SIR diagnosis and clearance at the subsequent retest. The study included all TIRs that were identified as SIRs in 2005 in otherwise free herds at tests with no other reactors at that test and that were clear at the subsequent retest. The analysis was restricted to cows that were neither sold, other than direct to slaughter, nor exported from the herd during the follow up period (to the end of 2009). Five control cows were randomly selected from each study herd. A parametric survival model with shared frailties, to account for clustering within herds, was developed to model time from passing a retest to future bTB diagnosis. The final parametric survival model contained the variables: TIR status in 2005, inconclusive status during the follow-up period, location, herd restricted during the study, time since last restriction within the herd and age. The time ratio for the TIR status variable was significant (p<0.001) indicating that on average the time to diagnosis with bTB for TIRs was 78% shorter compared to the non-TIRs. The frailty term was significant (p<0.001) indicating that animals within some herds were more likely to become reactors compared to other herds. These results have important implications for national policy and future management of TIR animals. Further, private veterinary practitioners and their clients should be aware of the increased risk associated with TIRs.     

Bovine tuberculosis and milk production in infected dairy herds in Ireland

This study describes the relationship between bovine tuberculosis (TB) and milk yield in TB-infected dairy herds in Ireland. The study had two objectives: to determine whether cows detected as TB reactors (and thus subject to immediate slaughter) were likely to be the higher milk-producing cows, and to determine whether subclinical TB infection was associated with reduced milk production at or around the time of disclosure (detection). All Irish dairy herds restricted from trading between the 1^st June 2004 and the 31^st May 2005 as a result of two or more TB reactors by the Single Intradermal Comparative Tuberculin Test (SICTT) were considered for study. The data consisted of 419 herds. Data were collected on all TB reactors and a random sample of 5 non-reactor cows in these herds: a data set of 4340 cows (2342 TB reactors and 1998 non-reactors). Previous milk data for the cows were taken into consideration and thus all lactations on a cow were analysed together with the years of lactations. There was an inherent hierarchical structure in the data, with lactations nested within cows and cows within herds and thus a linear mixed model with two random effects was used to describe the data. The results of this study showed that for all lactations and years under investigation, milk yield was significantly lower for TB reactor cows, with differences ranging from 120 kg (2003, lactation 3) to 573 kg (2001, lactation 1), when compared to the non-reactor cows.     

Risk factors for epidemic respiratory disease in Norwegian cattle herds

An epidemic of acute respiratory disease associated with bovine respiratory syncytial virus (BRSV) occurred during the winter and spring of 1995 in two neighbouring veterinary districts in the south-eastern part of Norway.The objective of this study was to describe the time course of the outbreak associated with BRSV in the cattle herds, and to determine the association between selected herd factors and the risk of experiencing a herd outbreak of acute respiratory disease.Data from 431 cattle herds on the dates of disease occurrence, location of the farms, herd size, age profile and production type were collected retrospectively for 1995. The risk of acute respiratory disease occurring in a cattle herd was related to the herd size as well as the type of production, with an expressed interaction between these two variables. From the Cox proportional-hazards model, the risk of a herd outbreak in a mixed herd of 20 animals was estimated to be 1.7-times greater than in a dairy herd and 3.3-times greater than a beef herd (reference category) of a comparable size. On increasing the herd size to 50 animals, the risk increased 1.3-fold for a mixed herd, 3.3-fold for a dairy herd, and 2.1-fold for a beef herd, compared to the risk for a corresponding type of herd of 20 animals.     

Risk factors for the between-herd spread of Mycobacterium bovis in Canadian cattle and cervids between 1985 and 1994.

Microorganisms of the genus Mycobacterium cause tuberculosis in many animal species including humans. Generally, Mycobacterium bovis (M. bovis) infects cattle and cervids, but it has the potential to infect virtually all species of mammals. This study examined and analysed the data from the nine outbreaks of tuberculosis in Canadian cattle and cervids from 1985 to 1994. For the purposes of this study, a positive herd was one with at least one culture-positive animal. A reactor herd had at least one animal which was positive or suspicious on a mid-cervical, comparative cervical, or gross or histopathologic test for tuberculosis. Herd classification was either reactor/positive or negative. Data for the study were collected from the outbreak records in the Regional or District offices of Agriculture and Agri-Food Canada. Logistic regression was used to study spread of tuberculosis between herds. Two risk factors were identified: increasing herd size; and, the reason why a herd was investigated as part of the outbreak. This latter factor was interpreted as a surrogate measure for the nature of contact between the study herd and other potentially infected herds in the outbreak. Increasing herd size was associated with an increased risk of being positive for tuberculosis with herds of 16-35, 36-80, and >80 animals having odds ratios of 2.9, 5.8, and 9.3, respectively, when compared to a herd size of <16 animals (p < 0.001). When compared to perimeter testing (i.e. testing herds within a specified radius of an infected herd), all other reasons for investigation had higher odds ratios (p < 0.001). These odds ratios were 57.8 for traceout herds (i.e. herds which had purchased animal(s) from a reactor/positive herd), 31.8 for herds with pasture or fence-line contact with a reactor/positive herd, and 14.9 for traceback herds (i.e. herds which had been a source of animals for reactor/positive herd(s)).     

Outbreak of bovine tuberculosis featuring anergy to the skin test, udder lesions and milkborne disease in young calves

A severe outbreak of bovine tuberculosis in a 1300-head, multisite dairy herd in Great Britain had several unusual features, including anergy to the tuberculin skin test, milkborne disease in calves and a farm cat, and a risk of human infection. The outbreak was controlled by culling 221 cattle over 15 months, by using the gamma-interferon (gamma-ifn) test and by the examination of milk samples. The gamma-ifn test detected infected animals that were not detected by the skin test.     

Effect of an inactivated paratuberculosis vaccine on the intradermal testing of goats for tuberculosis

The effect of an inactivated paratuberculosis vaccine on the diagnosis of tuberculosis (TB) in goats was investigated in a herd with a history of clinical paratuberculosis but which was free of TB. Cohorts of animals in 2006, 2008 and 2009, were vaccinated once at 1 month of age, and 50% of the 2006 cohort served as unvaccinated controls. The goats were aged 8 months, 20 months and 3.5 years old at the time of the survey. All animals were assessed using a single intradermal injection of bovine tuberculin purified protein derivative (PPD) (SID test), or using both bovine and avian PPD (CID test). An interferon (IFN)-γ assay using both bovine and avian PPD was carried out on the 2006 cohort and was interpreted according to three different 'cut-off' points. No unvaccinated (control) animals tested positive to any of the assays, confirming that the herd was TB-free. The SID test had a low specificity in vaccinated animals at 8 and 20 months of age, whereas the CID test demonstrated 100% specificity in animals ≥20 months-old. The specificity of IFN-γ assay was less than maximal for vaccinated animals 3.5 years old as small numbers of false positives were detected, although this depended on the chosen cut-off point. The study findings demonstrate that the use of an inactivated paratuberculosis vaccine in goats <1 month-old in a TB-free herd does not result in false positives to a CID test for TB when performed in animals ≥20 months-old.