The farm environment: an overlooked source of Mycobacterium bovis?

Bovine tuberculosis (bTB) is a cattle disease which can be transmitted to people (although human infections are rare in the UK). Despite decades of efforts to eradicate it, the disease has spread across much of western Britain. The disease seems to persist because the tests used to detect and remove infected cattle miss some infected animals, and because wildlife (especially badgers) can also catch the disease and pass it on to cattle.
Most cattle catch bTB by breathing in specks of mucus or saliva from other cattle. However, new research shows that badgers avoid cattle, and very seldom come close enough to transmit (or catch) the disease in this way. Because there is very strong evidence that badgers give bTB to cattle, and that cattle give bTB to badgers, this finding strongly suggests that cattle can both transmit and catch bTB without coming into close contact with another animal.
Both cattle and badgers excrete bTB bacteria, which can survive in the environment for weeks or months. We can quantify these bacteria in the environment by using genetic methods which count the number of copies of bacterial DNA in a sample of soil, faeces, or water. Our preliminary studies have detected such bacteria in cattle slurry, and in the soil of fields where cattle graze, as well as in badger faeces collected on cattle farms. It is likely that cattle can catch bTB from these sources; however, current bTB management does little to address the risk posed by bTB in the environment.
Our project aims to address six research questions:
(1) Where in the farm environment are bTB bacteria concentrated?
(2) Are the bTB bacteria found in the environment still alive and hence likely to infect cattle?
(3) To what extent do cattle come into contact with bTB bacteria in the environment?
(4) Does spreading slurry onto fields contribute to bTB bacteria in the environment?
(5) Do the bTB bacteria found in the environment come mostly from cattle or from badgers?
(6) How might farms be managed to avoid cattle catching bTB from the environment?
Our project will seek to answer these six questions by a careful study of 20 farms located in Cornwall, where rates of cattle bTB are very high. Our team will collect thousands of samples of cattle dung, slurry, soil and badger faeces, as well as swabs from inside barns and cattle troughs. These samples will be brought back to our laboratory where we shall extract DNA from them and count the numbers of copies of bTB genetic code from each one. These analyses will allow us to map where on the farms bTB bacteria are most abundant, and to explore whether the numbers and distribution of bacteria vary between seasons. At the same time, we shall track the movements of both cattle and muckspreaders (used to spread slurry) using technology similar to that present in vehicle satnav systems. Tracking muckspreaders will help us to assess whether bTB is more abundant in fields where slurry has been spread recently, and tracking cattle will help us to assess where they are at greatest risk of encountering bTB bacteria in the environment.
We shall test whether bTB found in the environment is still alive and also explore how long bTB bacteria survive in cattle dung and badger faeces held inside small open-air enclosures inaccessible to animals.
To assess where environmental bacteria come from, we plan to compare the DNA of bTB bacteria found in soil, barns and troughs with that of bacteria known to have come from cattle (found in fresh cattle dung) and from badgers (found in fresh badger faeces). If, for example, the DNA strains found in soil were similar to those found in cattle dung, but different from those found in badger faeces, it would suggest that most environmental contamination comes from cattle.
To ensure that our research findings can be translated into practical guidance for farm management, our project steering committee includes representatives from the farming industry and the veterinary profession.

The focus of the research is to better understand how the environment can act as a source of Mycobacteria bovis and provide mitigation strategies to significantly reduce the potential for transmission, as we hypothesise that environmental contamination is a vital component in the transmission of this disease between animals on farms. We aim to provide a definitive study of the prevalence and detection frequency of M. bovis on 20 farms in Cornwall, a high bTB incidence area. This will be the first study to combine detailed molecular detection of a wide range of environmental samples including cattle slurry, totaling 18,000 samples with detailed cattle movement information. This will be carried out contemporaneously with intensive farm monitoring including tracking cattle movements and approaches to slurry handling. The farms have already been the subject of a three year study of cattle and badger movements so there is an extensive database available to augment with environmental prevalence. A heatmap of the location, presence and quantity of M. bovis DNA shed into the environment will be produced and combined with information on with these sources to determine the relative risk of cattle exposure to M. bovis in various locations within the farm environment. Furthermore, the viability of environmental M. bovis detected on these farms will be assessed to establish whether these environmental pools of M. bovis may be potentially infectious. The longevity of environmental M. bovis will be determined using exclosure experiments where the M. bovis load and viability of faeces will be assessed over time. Metagenomic techniques will be developed during this study to retrieve whole genomes directly from environmental samples without the need to culture. This will enable the genome of strains present in the environment on these farms to be determined and potentially attributed to a source as well as information on the longevity of strains.

This proposal has the potential for very significant impacts on both mitigation of risk for exposure of cattle to sources of M. bovis in the farm environment, establish appropriate biosecurity measures and inform policy for suitable eradication measures at the farm scale to reduce the risks of bTB herd breakdowns. The research work will provide data for development of statistical models which can be used to evaluate biosecurity methods and indeed evaluate a wide range of strategies to control risk such as vaccination, culling and isolation.
We aim to identify processes influencing the distribution of sources of M. bovis which provide risk for onward transmission to cattle and other animals and wildlife. Thus we will evaluate appropriate intervention strategies including BCG vaccination, and provide added value to the implementation of molecular detection tools by establishing appropriate sampling regimes and high risk areas on farms. This will provide farmers and vets with vital information for often simple and effective control measures in addition to informing policy makers of the most appropriate measures to develop biosecurity strategies for high risk areas. A primary beneficiary of these results is Defra who have a continuing interested in badger management, and predicted conditions for badger-cattle and cattle-badger transmission, particularly in the edge zone where bTB herd breakdowns are advancin. Vets, farmers, Defra and wildlife enthusiasts are interested in developments of non-invasive molecular tools for environmental monitoring of M. bovis to increase farm biosecurity against bTB in cattle, in surveillance of badger populations to inform badger management, and to measure efficacy of badger BCG vaccination and culling against M. bovis shedding into the environment.

The heat maps, models and strain diversity data coupled with strain dissemination and persistence data will be a valuable resource and influence the decision making process that impacts on livestock farmers' economic activities. Farmers can take responsibility for biosecurity and we aim to assist directly in this process. Stakeholders will benefit from direct interaction with the research and this can be implemented via the steering committee consisting of pathogenicity experts, vets, farmers and commercial kit detection company LGC Group. Methods for environmental monitoring of disease exposure risks are widely sought as we become more aware of the import

Data generated from along the geographical edge of endemic bTB in the UK will provide insights into the processes regulating seeding of M. bovis in advance of bTB becoming established in cattle and/or badgers. We envisage a future bolt-on proposal to conduct whole genome analysis of M. bovis isolates from badger and cattle along this frontier to determine the role played by the environment in disease transmission. In fact our work in Tanzania monitoring shedding into environmental sites of both M. bovis and M. tuberculosis has provided both vets and clinicians with opportunities for disease surveillance and this will apply for bTB in the UK without the need for invasive and expensive animal tests. This will aid in the evaluation of herd breakdown data and also information on wildlife infection status.
The work will contribute to enhancing quality of life and health of the farming community suffering from chronic bTB and economic difficulties. It provides materials for education at all levels, in providing direct access to the scientists and results disseminated through lectures, seminars and talks, and providing the epidemiological data and biological samples for UG and PG projects. The wider public will be informed by unbiased delivery at public meeting, museum talks, encouraging more engagement and public interest in science.