Investigating Farm Fragmentation as a Risk Factor for Bovine Tuberculosis in Cattle Herds: A Matched Case-Control Study from Northern Ireland.

Bovine tuberculosis remains a challenging endemic pathogen of cattle in many parts of the globe. Spatial clustering of Mycoacterium bovis molecular types in cattle suggests that local factors are the primary drivers of spread. Northern Ireland's agricultural landscape is comprised of highly fragmented farms, distributed across spatially discontinuous land parcels, and these highly fragmented farms are thought to facilitate localised spread. We conducted a matched case control study to quantify the risks of bovine tuberculosis breakdown with farm area, farm fragmentation, fragment dispersal, and contact with neighbouring herds. Whilst our results show small but significant increases in breakdown risk associated with each factor, these relationships were strongly confounded with the number of contiguous neighbours with bovine tuberculosis. Our key finding was that every infected neighbour led to an increase in the odds of breakdown by 40% to 50%, and that highly fragmented farms were almost twice as likely to have a bTB positive neighbour compared to nonfragmented farms. Our results suggest that after controlling for herd size, herd type, spatial and temporal factors, farm fragmentation increasingly exposes herds to infection originating from first-order spatial neighbours. Given Northern Ireland's particularly fragmented landscape, and reliance on short-term leases, our data support the hypothesis that between-herd contiguous spread is a particularly important component of the region's bovine tuberculosis disease system.

Quantifying intraherd cattle movement metrics: Implications for disease transmission risk.

There are a paucity of data quantifying on-farm management practices such as the frequency of intraherd cattle movements, use of consolidated or spatially fragmented grazing pastures, and duration of time cattle spend at grass with respect to biosecurity and disease transmission. Such movement dynamics are important when attempting to understand the maintenance of chronic infectious disease, such as bovine tuberculosis (bTB). We captured empirical data on daily cattle movements for a sample of eighteen farms throughout one complete grazing season (n = 18,988 grazing days) and assessed these attributes in relation to herd bTB risk. Dairy herds were stocked at significantly higher densities compared to beef production systems (6.6 animals/ha, 95 % confidence intervals (CI) 6.5-6.7 and 4.1 animals/ha, 95 %CI 4.1 - 4.1 respectively, p < 0.001). Most notably milking cows, were grazed at higher densities than other life stages (e.g. calves, heifers and bullocks) (p < 0.001) and experienced four times the number of movements between pastures. Beef cattle were more likely to be grazed across multiple (rather than single) fields (p < 0.001), with greater time spent on fragmented land away from the main/home farm (p < 0.001). None of the farm or herd attributes analysed (e.g. stocking density, frequency of movement, movement distances or land fragmentation) were associated with herd bovine tuberculosis (bTB) breakdowns during this study. However, there was a weak positive association between bTB breakdowns during the 3 years prior to the study and cattle movement distances (p = 0.05) and time spent on fragmented land (p = 0.08). After a bTB breakdown occurs, restrictions on animals moving out of these herds are implemented to control disease spread, yet we argue that more attention is needed on the role of intraherd grazing patterns in modelling disease transmission risk between herds.

Mycobacterium bovis Population Structure in Cattle and Local Badgers: Co-Localisation and Variation by Farm Type.

Bovine tuberculosis surveillance in Northern Ireland includes Multiple-Locus Variable number tandem repeat Analysis (MLVA) to determine the Mycobacterium bovis genetic type present in both cattle and the predominant wildlife host, the European badger (Meles meles). These data are useful for investigating clusters of infection and understanding the scale at which interspecific transmission may occur. We utilised a comprehensive dataset of routinely sampled isolates from infected cattle and from badgers killed in road-traffic accidents to investigate the spatial co-location of MLVA types in, and between, the badger and cattle populations. Furthermore, we investigated the hypothesis that the type of farming enterprise might explain some variation in this relationship. MLVA types were spatially co-localised in cattle and road-traffic accident (RTA) badger hosts, indicative of a shared epidemic. Dairy herds were more likely to have at least one MLVA type in common with nearby RTA badgers, compared to non-dairy herd types. Marginally more MLVA spatial clustering was observed in non-dairy herds, which may be a consequence of relatively more between-herd movements. For the cattle population, local transmission mechanisms such as infection from contiguous herds, infectious wildlife and short-range between-herd cattle movements appear primarily to drive the epidemic: there appears to be a more limited role for long-range movements. Animal management practices are likely to be the driving force behind this observation, as beef rearing is associated with elevated numbers of animal movements compared to dairy herds.

Bovine tuberculosis breakdown duration in cattle herds: an investigation of herd, host, pathogen and wildlife risk factors.

BACKGROUND: Despite rigorous controls placed on herds which disclose ante-mortem test positive cattle to bovine tuberculosis, caused by the infection of Mycobacterium bovis, many herds in Northern Ireland (NI) experience prolonged breakdowns. These herds represent a considerable administrative and financial burden to the State and farming community. METHODS: A retrospective observational study was conducted to better understand the factors associated with breakdown duration, which was modelled using both negative binomial and ordinal regression approaches. RESULTS: Six explanatory variables were important predictors of breakdown length in both models; herd size, the number of reactors testing positive in the initial SICCT test, the presence of a lesioned animal at routine slaughter (LRS), the count of M. bovis genotypes during the breakdown (MLVA richness), the local herd-level bTB prevalence, and the presence of herds linked via management factors (associated herds). We report that between 2008 and 2014, mean breakdown duration in NI was 226 days (approx. seven months; median: 188 days). In the same period, however, more than 6% of herds in the region remained under movement restriction for more than 420 days (13 months); almost twice as long as the mean. The MLVA richness variable was a particularly important predictor of breakdown duration. We contend that this variable primarily represents a proxy for beef fattening herds, which can operate by purchasing cattle and selling animals straight to slaughter, despite prolonged trading restrictions. For other herd types, the model supports the hypothesis that prolonged breakdowns are a function of both residual infection within the herd, and infection from the environment (e.g. infected wildlife, contiguous herds and/or a contaminated environment). The impact of badger density on breakdown duration was assessed by including data on main sett (burrow) density. Whilst a positive association was observed in the univariate analysis, confounding with other variables means that the contribution of badgers to prolonged breakdowns was not clear from our study. We do not fully reject the hypothesis that badgers are implicated in prolonging bTB breakdowns via spillback infection, but given our results, we posit that increased disease risk from badgers is unlikely to simply be a function of increasing badger density measured using sett metrics.

Genomic epizootiology of a Brucella abortus outbreak in Northern Ireland (1997-2012).

BACKGROUND: In the recent past (1997-2012), Northern Ireland in the United Kingdom suffered an outbreak of Brucella abortus, which at its height affected over 200 cattle herds. Initially, isolates were characterized using multi-locus variable number tandem repeats analysis (MLVA). While informative in this setting, hyper-variability in some loci limited the resolution necessary to infer fine-scale disease transmission networks. Consequently, we applied whole-genome sequencing to isolates from this outbreak to evaluate higher resolution markers for disease epizootiology. RESULTS: Phylogenetic analysis revealed that the B. abortus outbreak in Northern Ireland was caused by two distinct pathogen lineages. One contained isolates consistent with the 1997-2012 outbreak being linked to a previous endemic infection thought eradicated. The dominant second lineage exhibited little genetic diversity throughout the recrudescent outbreak, with limited population sub-structure evident. This finding was inconsistent with prior MLVA molecular characterizations that suggested the presence of seven clonal complexes. Spatio-temporal modeling revealed a significant association of pairwise SNP differences between isolates and geographic distances. However, effect sizes were very small due to reduced pathogen diversity. CONCLUSIONS: Genome sequence data suggested that hyper-variability in some MLVA loci contributed to an overestimate of pathogen diversity in the most recent outbreak. The low diversity observed in our genomic dataset made it inappropriate to apply phylodynamic methods to these data. We conclude that maintaining data repositories of genome sequence data will be invaluable for source attribution/epizootiological inference should recrudescence ever re-occur. However genomic epizootiological methods may have limited utility in some settings, such as when applied to recrudescent/re-emergent infections of slowly-evolving bacterial pathogens.

Characteristics of Northern Irish cattle herds without bovine tuberculosis infection.

BACKGROUND: Despite ongoing eradication efforts, bovine tuberculosis (bTB) is endemic in cattle herds in Northern Ireland (NI). This disease has serious implications for the economy, farming and animal welfare. Previous research identified a population of herds which have remained free from bTB infection for 10 years (2004-2014). Understanding the characteristics of these herds may have important implications for eradication efforts, such as spatially targeted interventions. METHODS: A cluster analysis and a retrospective case-control analysis was conducted to compare bTB- free herds with herds which experienced prolonged infection (ie, bTB breakdowns lasting more than ≥ 365 days). RESULTS: Only small, localised clusters of herds which have remained free from bTB were revealed, thus limiting the potential for spatially targeted interventions. The results illustrated the importance of herd size to disease status; over 27 per cent of the bTB-free herds had up to 10 animals. However, the data also showed that there were no inward movements in the year before the bTB skin test in those herds which remained free from bTB. CONCLUSIONS: Attention should therefore be given to the cattle movement network in NI to better understand the risk associated with cattle purchasing.

Liver fluke (Fasciola hepatica) co-infection with bovine tuberculosis in cattle: A prospective herd-level assessment of herd bTB risk in dairy enterprises.

Co-infection of tuberculosis (TB) and helminths is recognized as a significant problem in regions where such pathogens are endemic and chronic cases exist. Co-infection can modulate the immune system leading to interference with diagnostic tests, increased pathological impacts and pathogen persistence. However, research has found that such interactions between pathogens can be context and species specific. Recent studies have suggested that liver fluke, Fasciola hepatica, infection may impact on immunological responses and diagnostics for bovine tuberculosis (bTB; caused by Mycobacterium bovis) in cattle. Where evidence of such interaction exists, there would be an onus on policy makers to adjust eradication programs to minimize impacts. We assessed the association between herd-level bTB breakdown risk and seasonal variation in liver fluke exposure based on 5,753 bulk tank milk (BTM) samples from 1,494 dairy herds across Northern Ireland. BTM was tested by an IDEXX antibody specific enzyme-linked immunosorbent assay (ELISA) using the 'f2' antigen as a detection agent. The ELISA determined the result based on a sample to (known) positive ratio (S/P%) from which binary status and categories of exposure were derived. Associations were tested using multivariable random effects models. Models predicting bTB risk were not improved with the inclusion of liver fluke exposure levels. Variations in modelling liver fluke exposure (S/P%, binary, categories of exposure) and bTB risk (skin test breakdowns, post-mortem confirmed breakdowns, breakdown size and lag effects) also failed to support associations (neither positive nor negative) between the pathogens at herd-level. These results, along with previously published animal-level data from Northern Ireland, suggest that the nexus between bTB and F. hepatica may have small size effects at the population-level. However, our results also highlight the high prevalence of F. hepatica in cattle in our study population, and therefore we cannot fully discount the potential hypothesis of population-level depression of immune response to M. bovis due to co-infection.

Is There a Relationship Between Bovine Tuberculosis (bTB) Herd Breakdown Risk and Mycobacterium avium subsp. paratuberculosis Status? An Investigation in bTB Chronically and Non-chronically Infected Herds.

Background: Bovine tuberculosis (bTB; Mycobacterium bovis) remains a significant problem in a number of countries, and is often found where M. avium subsp. paratuberculosis (MAP) is also present. In the United Kingdom, bTB has been difficult to eradicate despite long-term efforts. Co-infection has been proposed as one partial mechanism thwarting eradication. Methods: A retrospective case-control study of 4,500 cattle herds in Northern Ireland, where serological testing of cattle for MAP, was undertaken (2004-2015). Blood samples were ELISA tested for MAP; infection of M. bovis was identified in herds by the comparative tuberculin test (CTT) and through post-mortem evidence of infection. Case-herds were those experiencing a confirmed bTB breakdown; control-herds were not experiencing a breakdown episode at the time of MAP testing. A second model included additional testing data of feces samples (culture and PCR results) to better inform herd MAP status. Multi-level hierarchical models were developed, controlling for selected confounders. A sensitivity analysis of the effect of MAP sample numbers per event and the prior timing of tuberculin-testing was undertaken. Results: 45.2% (n = 250) of case observations and 36.0% (3,480) of control observations were positive to MAP by ELISA (45.8% and 36.4% when including ancillary fecal testing, respectively). Controlling for known confounders, the adjusted odds ratio (aOR) for this association was 1.339 (95%CI:1.085-1.652; including ancillary data aOR:1.356;95%CI:1.099-1.673). The size-effect of the association increased with the increasing number of samples per event used to assign herd MAP status (aOR:1.883 at >2 samples, to aOR:3.863 at >10 samples), however the estimated CI increased as N decreased. 41.7% of observations from chronic herds were MAP serology-positive and 32.2% from bTB free herds were MAP positive (aOR: 1.170; 95%ci: 0.481-2.849). Discussion: Cattle herds experiencing a bTB breakdown were associated with increased risk of having a positive MAP status. Chronic herds tended to exhibit higher risk of a positive MAP status than bTB free herds, however there was less support for this association when controlling for repeated measures and confounding. MAP co-infection may be playing a role in the success of bTB eradiation schemes, however further studies are required to understand the mechanisms and to definitively establish causation.

Seasonal variation of Fasciola hepatica antibodies in dairy herds in Northern Ireland measured by bulk tank milk ELISA.

Bovine fasciolosis, caused by the infection of the trematode parasite Fasciola hepatica, remains a problem in dairy herds causing significant production losses. In this study, bulk milk tank samples were utilised to generate a comprehensive survey of the variation in liver fluke exposure over the four seasons of 2016 in Northern Ireland (NI). Samples were tested using an antibody ELISA test; within-herd prevalence levels were categorised relative to sample-to-positive ratio (S/P%). Overall, 1494 herds (~ 50% of all active dairy farms in NI) were sampled. In total, 5750 samples were tested with 91% of herds having a sample result for each season. The proportion of herds with evidence of liver fluke exposure was very high across the year, with 93.03% of all bulk milk samples having some indication of liver fluke antibody presence. A high proportion of samples (2187/5750; 38.03%) fell within the highest infection class (indicating high within-herd prevalence). There was significant seasonal variation in the mean S/P%. A multivariable random effect ordinal logit model suggested that the greatest probability of being in a higher infection class was in winter, whilst the lowest was recorded during summer. There was a significant negative association between increasing herd liver fluke infection class and herd size. Furthermore, there was significant variation in infection levels across regions of Northern Ireland, with higher infection levels in northern administrative areas. This study demonstrates the very high liver fluke exposure in this region of Europe, and that risk is not equally distributed spatially or across seasons in dairy herds.

The influence of pigmentation patterning on bumblebee foraging from flowers of Antirrhinum majus.

Patterns of pigmentation overlying the petal vasculature are common in flowering plants and have been postulated to play a role in pollinator attraction. Previous studies report that such venation patterning is significantly more attractive to bee foragers in the field than ivory or white flowers without veins. To dissect the ways in which venation patterning of pigment can influence bumblebee behaviour, we investigated the response of flower-naïve individuals of Bombus terrestris to veined, ivory and red near-isogenic lines of Antirrhinum majus. We find that red venation shifts flower colour slightly, although the ivory background is the dominant colour. Bees were readily able to discriminate between ivory and veined flowers under differential conditioning but showed no innate preference when presented with a free choice of rewarding ivory and veined flowers. In contrast, both ivory and veined flowers were selected significantly more often than were red flowers. We conclude that advantages conferred by venation patterning might stem from bees learning of their use as nectar guides, rather than from any innate preference for striped flowers.