James O’Keeffe

Trial design to estimate the effect of vaccination on tuberculosis incidence in badgers

The principal wildlife reservoir of Mycobacterium bovis in Ireland is the European badger. Studies in the Republic of Ireland (RoI) have shown that badgers culled in association with cattle herd tuberculosis breakdowns (focal culling) have a higher prevalence of infection than the badger population at large. This observation is one rationale for the medium term national strategy of focal badger culling. A vaccination strategy for the control of bovine tuberculosis (bTB) in badgers is a preferred long-term option. The Bacillus Calmette-Guérin (BCG) vaccine has been shown to decrease disease severity in captive badgers under controlled conditions. As the vaccine has been tested in a controlled environment with precise information on infection pressure, it cannot be assumed a priori that the effects of vaccination are similar in the wild, where other environmental and/or ecological factors prevail. For this reason we have designed a vaccine field trial to assess the impact of vaccination on the incidence of TB infection in a wild badger population. The selected study area for the vaccine trial (approximately 755 square kilometers) is divided into three zones each of which has similar characteristics in terms of size, number of main badger setts, cattle herds, cattle and land classification type. Three vaccination levels (100%, 50% and 0%) will be allocated to the three zones in a way that a gradient of vaccination coverage North to South is achieved. The middle zone (zone B) will be vaccinated at a 50% coverage but zone A and C will be randomly allocated with 100% or 0% vaccination coverage. Vaccination within zone B will be done randomly at individual badger level. The objective of this paper is to describe the design of a field tuberculosis vaccination trial for badgers, the epidemiological methods that were used to design the trial and the subsequent data analysis. The analysis will enable us to quantify the magnitude of the observed vaccination effect on M. bovis transmission in badgers under field conditions and to improve our knowledge of the biological effects of vaccination on susceptibility and infectiousness.

The risk of a positive test for bovine tuberculosis in cattle purchased from herds with and without a recent history of bovine tuberculosis in Ireland.

A retrospective cohort study was conducted to assess if cattle sold from Irish dairy herds within 7 months of herd de-restriction (clearance to trade) from a bovine-tuberculosis (BTB) episode had an excess risk of testing positive for BTB during the following 2 years, and to determine other risk factors associated with this outcome. If possible, a predictive metric for herds at high risk of selling future BTB-positive cattle would be generated. The unexposed cohort included all cattle sold within 7 months of the annual herd test in a random sample of dairy herds that did not test positive for BTB in 2003. The exposed cohort consisted of all cattle sold within 7 months of the date of de-restriction in all dairy herds that cleared a BTB episode in 2003. Only cattle sold from herds that were initially found to test positive for BTB using the single intradermal comparative tuberculin test (SICTT)-and not due to discovery of a BTB-positive animal at slaughter-were included as exposed cattle. To aid in the development of a predictive metric, the exposed cohort was subcategorized based on the number of reactors to the SICTT in the herd of origin during the BTB episode immediately prior to sale. The final exposure categories of 0 (unexposed), 1-7, and >or=8 total reactors were considered the unexposed, mildly exposed, and severely exposed cohorts, respectively. A multivariable logistic regression model was fit to the final BTB status of the animal using a generalized estimating equation method (GEE), assuming an exchangeable correlation structure of animals within herds, and using robust standard errors. Exposure level and the other available herd- and animal-level information were modeled. After controlling for other risk factors including the size of the herd of origin and the sex and age of the animal, the three-level exposure variable significantly improved the model (based on a change in Quasi-Akaike Information Criteria of 2.2) and demonstrated a trend of increasing risk of a future positive BTB test with increasing exposure category. The severely exposed cohort of animals had significantly higher risk of a future positive BTB test than the unexposed cohort (OR=1.78, p=0.030).

Population Estimation and Trappability of the European Badger (Meles meles): Implications for Tuberculosis Management

Estimates of population size and trappability inform vaccine efficacy modelling and are required for adaptive management during prolonged wildlife vaccination campaigns. We present an analysis of mark-recapture data from a badger vaccine (Bacille Calmette–Guérin) study in Ireland. This study is the largest scale (755 km2) mark-recapture study ever undertaken with this species. The study area was divided into three approximately equal–sized zones, each with similar survey and capture effort. A mean badger population size of 671 (SD: 76) was estimated using a closed-subpopulation model (CSpM) based on data from capturing sessions of the entire area and was consistent with a separate multiplicative model. Minimum number alive estimates calculated from the same data were on average 49–51% smaller than the CSpM estimates, but these are considered severely negatively biased when trappability is low. Population densities derived from the CSpM estimates were 0.82–1.06 badgers km−2, and broadly consistent with previous reports for an adjacent area. Mean trappability was estimated to be 34–35% per session across the population. By the fifth capture session, 79% of the adult badgers caught had been marked previously. Multivariable modelling suggested significant differences in badger trappability depending on zone, season and age-class. There were more putatively trap-wary badgers identified in the population than trap-happy badgers, but wariness was not related to individual’s sex, zone or season of capture. Live-trapping efficacy can vary significantly amongst sites, seasons, age, or personality, hence monitoring of trappability is recommended as part of an adaptive management regime during large–scale wildlife vaccination programs to counter biases and to improve efficiencies.

Oral Vaccination of Free-Living Badgers (Meles meles) with Bacille Calmette Guérin (BCG) Vaccine Confers Protection against Tuberculosis.

A field trial was conducted to investigate the impact of oral vaccination of free-living badgers against natural-transmitted Mycobacterium bovis infection. For a period of three years badgers were captured over seven sweeps in three zones and assigned for oral vaccination with a lipid-encapsulated BCG vaccine (Liporale-BCG) or with placebo. Badgers enrolled in Zone A were administered placebo while all badgers enrolled in Zone C were vaccinated with BCG. Badgers enrolled in the middle area, Zone B, were randomly assigned 50:50 for treatment with vaccine or placebo. Treatment in each zone remained blinded until the end of the study period. The outcome of interest was incident cases of tuberculosis measured as time to seroconversion events using the BrockTB Stat-Pak lateral flow serology test, supplemented with post-mortem examination. Among the vaccinated badgers that seroconverted, the median time to seroconversion (413 days) was significantly longer (p = 0.04) when compared with non-vaccinated animals (230 days). Survival analysis (modelling time to seroconversion) revealed that there was a significant difference in the rate of seroconversion between vaccinated and non-vaccinated badgers in Zones A and C throughout the trial period (p = 0.015). For badgers enrolled during sweeps 1–2 the Vaccine Efficacy (VE) determined from hazard rate ratios was 36% (95% CI: -62%– 75%). For badgers enrolled in these zones during sweeps 3–6, the VE was 84% (95% CI: 29%– 97%). This indicated that VE increased with the level of vaccine coverage. Post-mortem examination of badgers at the end of the trial also revealed a significant difference in the proportion of animals presenting with M. bovis culture confirmed lesions in vaccinated Zone C (9%) compared with non-vaccinated Zone A (26%). These results demonstrate that oral BCG vaccination confers protection to badgers and could be used to reduce incident rates in tuberculosis-infected populations of badgers.

Factors affecting European badger (Meles meles) capture numbers in one county in Ireland

Understanding factors affecting the number of badgers captured at and around badger setts (burrows) is of considerable applied importance. These factors could be used to estimate probable badger densities for bovine tuberculosis (bTB) control and also for monitoring badger populations from a conservation perspective. Furthermore, badger management and vaccination programs would benefit by increasing the probability of efficiently capturing the target badger populations. Within this context, it was investigated whether badger capture numbers can be estimated from field signs and previous capture histories. Badger capture records (initial and repeated capture numbers at a sett) from a large-scale removal program (405 km(2), 643 setts) were used. Univariable count models indicated that there were a number of significant potential predictors of badger numbers, during initial capture attempts. Using a multivariable zero-inflated Poisson (ZIP) model of initial captures we found that badger capture numbers were significantly affected by sett type, season, year, and the number of sett entrances in active use. Badger capture numbers were also affected by the total previous catch during repeated capture events and by the number of previous capture attempts. There was a significant negative trend in badger captures across events. Measures of the ability of these models to estimate badger captures suggested that the models might be useful in estimating badger numbers across a population; however the confidence intervals associated with these predictions were large.

Spatial and temporal analyses of metrics of tuberculosis infection in badgers (Meles meles) from the Republic of Ireland: Trends in apparent prevalence

Badgers are a wildlife host of Mycobacterium bovis, the causative agent of bovine tuberculosis (bTB), and an important contributor to the epidemiology of bTB in cattle in Ireland and Britain. Repeated culling of badgers in high prevalence cattle bTB areas has been used in the Republic of Ireland as one tool to reduce intra- and interspecific transmission of M. bovis. We assessed factors that influenced infection prevalence of culled badgers from 2009 to 2012 (n=4948) where spatial, temporal and intrinsic factor data were available using multivariable modelling. Prevalence appeared higher in western areas than eastern areas of Ireland and badgers were more likely to be test-positive if caught at a sett (burrow system) which was close to other infected setts (spatial clustering of infection). There was a significant positive association between badger test-status and cattle prevalence of M. bovis infection at a spatial scale of 1km around setts. Badgers were more likely to be deemed test positive if they were male (OR: 1.9) or a parous female (OR: 1.7), compared to a female who had never conceived. Our results are consistent with different groups within badger populations having differential exposures and therefore infection risk (for example, parous vs. non-parous females). Furthermore, bTB clusters within the badger population, with greater risk to badgers in setts that are closest to other infected setts. The effective scale of the association of bTB risk between badger and cattle populations may be relatively large in Ireland. Our data indicate that the overall trend in prevalence of M. bovis infection in badgers has decreased in Ireland (P<0.001) while controlling for significant confounders over the study period, and follows a longer temporal trend from 2007 to 2013, where unadjusted apparent prevalence declined from 26% to 11% during 2007 to mid-2011, followed by a stable trend between 9 and 11% thereafter (n=10,267).

Monitoring trap-related injury status during large-scale wildlife management programmes: an adaptive management approach

Wildlife management and research programmes often rely on the capturing of live animals. All capture methods for wildlife involve a risk of injury. For that reason, monitoring of injuries should be completed concurrently, and efficiently, throughout the intervention programme, or study period, to uphold welfare standards. Here, we present a practical adaptive management system to monitor trap-related injuries as a component of broader attempts to maintain welfare standards. The monitoring system described is part of a large-scale tuberculosis-control management programme of European badgers (Meles meles) in the Republic of Ireland. Badgers were captured in stopped cable restraints. A standardised operating procedure was developed to allow for the necropsy of large numbers of badgers (N = 18,596) by a veterinary pathologist. As part of this programme, badger injury status was evaluated using a categorical severity scale ranging from 0 (no impact due to capture) to 6 (death) for badgers captured in restraints. No restraint-related deaths were recorded over the study period. Eighty-four percent of captured badgers exhibited no impact or only superficial hair/skin compression (scale 0–1). Logistic models indicated that injury severity was significantly influenced by year, season, age class, gender, weight and the body position where the badger was captured. Despite the low prevalence of severe injuries recorded, the proportion of animals with a minor injury type (skin oedema; scale 2) increased during the study period. This highlights the need for continual sensitive welfare vigilance during prolonged programmes. We suggest improvements to the current programme and additional adjustments for future vaccination campaigns. We contend that this type of monitoring, coupled with adaptive management strategies, is essential for wildlife programmes to maintain high animal welfare standards, at least in terms of injuries, but also highlight that trap injuries are only one component of animal welfare. We suggest that this adaptive management system could serve as a template for other similar wildlife programmes.

Quantification of Mycobacterium bovis transmission in a badger vaccine field trial

In the UK and Ireland, Bacille Calmette-Guérin (BCG) vaccination of badgers has been suggested as one of a number of strategies to control or even eradicate Mycobacterium bovis infection in badgers. In this manuscript, we present the results of a badger field trial conducted in Ireland and discuss how the novel trial design and analytical methods allowed the effects of vaccination on protection against infection and, more importantly, on transmission to be estimated. The trial area was divided into three zones North to South (A, B and C) where vaccination coverages of 0, 50 and 100%, respectively, were applied. Badgers were trapped over a 4year period. Badgers were assigned to either placebo or vaccine treatment, with treatment allocation occurring randomly in zone B. Blood samples were collected at each capture, and serology was performed in these samples using a chemiluminescent multiplex ELISA system (Enfer test). The analysis aimed to compare new infections occurring in non-infected non-vaccinated badgers to those in non-infected vaccinated ones, while accounting for the zone in which the badger was trapped and the infection pressure to which this individual badger was exposed. In total, 440 records on subsequent trappings of individual non-infected badgers were available for analysis. Over the study period, 55 new infections occurred in non-vaccinated (out of 239=23.0%) and 40 in vaccinated (out of 201=19.9%) badgers. A Generalized Linear Model (GLM) with a cloglog link function was used for analysis. Statistical analysis showed that susceptibility to natural exposure with M. bovis was reduced in vaccinated compared to placebo treated badgers: vaccine efficacy for susceptibility, VES, was 59% (95% CI=6.5%-82%). However, a complete lack of effect from BCG vaccination on the infectivity of vaccinated badgers was observed, i.e. vaccine efficacy for infectiousness (VEI) was 0%. Further, the basic reproduction ratio as a function of vaccination coverage (p) (i.e. R(p)) was estimated. Given that the prevalence of M. bovis infection in badgers in endemic areas in Ireland is approximately 18%, we estimated the reproduction ratio in the unvaccinated population as R(0)=1.22. Because VES was now known, the reproduction ratio for a fully vaccinated population was estimated as R(1)=0.50. These results imply that with vaccination coverage in badgers exceeding 30%, eradication of M. bovis in badgers in Ireland is feasible, provided that the current control measures also remain in place.