Bilateral BBSRC-FAPESP: Defining the Genetic and Semiochemical Basis of Tick Resistance in Cattle

Ticks and tick-borne diseases have major impacts on livestock, humans and companion animals. In particular, the cattle tick is found worldwide in areas with high temperature and humidity. Tick infestations lead to weight loss, anaemia and secondary infections, as well as transmitting diseases such Babesiosis, Theileriosis and Anaplasmosis. Traditionally, chemicals (acaricides) are the main tick control strategy, but this approach is increasingly problematic, mainly due to the evolution acaricide-resistant ticks and generation of chemical residues. Because vaccines have only had mixed success, there is an urgent requirement for new control measures. Cattle vary greatly in the tick loads, and much of this variation is known to be controlled by the genetic composition of the host, therefore breeding for increased tick resistance is possible. We hypothesize that the primary means by which host cattle differ in the tick resistance is through their production of attractant/repellent volatile chemicals on the skin surface, i.e. semiochemicals, which ultimately determine how many ticks remain attached to the skin surface. Identification of these chemicals and the genetic factors that lead to different chemicals being produced by different animals would give us powerful new control opportunities. We could either breed animals that will be more resistant to ticks or we could devise repellents to reduce tick load. This project is designed to achieve this: the goal is to understand the factors underlying genetic variation in the resistance of Brazilian cattle to tick infestations, at both the genetic and biochemical levels.

Firstly, on more than 1000 commercial Girolando cattle in Brazil we will collect detailed data on tick load, biological samples and factors which conceivably influence tick load. These data will provide an overview of factors influencing tick burden. After extracting DNA from the blood, we will genotype each animal using a tool known as a high density SNP chip. This allows us to genotype animals simultaneously for genetic markers at more than 750,000 locations throughout the genome. Analyses of these data will tell us where in the genome are the genes which have a large influence on tick burden, and hence tick resistance. But it doesn't tell us which genes.

To work out which genes are important, we will then examine the extreme animals in much more detail. Using skin rubbings obtained from the animals with the consistently highest and lowest tick burdens we will characterise the behaviour-modifying chemicals (semiochemicals) produced by these two groups of animals using chromatography and spectroscopic analysis. We will test the hypothesis that ticks respond to the same semiochemicals as for biting flies (e.g. 6-methyl-5-hepten-2-one). We will also purchase animals with genotypes that we predict to represent the extremes of susceptibility and resistance, and challenge these animals with ticks. We will then look at the RNA profile of these animals before and after challenge, using skin biopsies. The RNA profiles will tell us which genes are being expressed, and we can compare these profiles between resistant and susceptible animals. This will tell us which genes and biochemical pathways are predictive of resistance.

Using bioinformatics we will then bring all our results together. We will combine the gene expression and the semiochemistry studies to determine which genes are most likely to underpin variation in resistance. We will then relate these results to the SNP chip results: if likely genes fall in regions of the genome that have been shown to influence tick resistance then we have very strong evidence for genes (and genetic markers) that influence resistance. We will then validate our results by genotyping a further 200 phenotyped cows, and devise breeding strategies to improve resistance. We will also investigate semiochemical based or husbandry interventions from our data.

Ticks have major impacts on animals and humans, worldwide. As well as transmitting disease, ticks lead to weight loss, anaemia and secondary infections. Acaricides are used for tick control, but this is problematic due to acaricide-resistance and chemical residue issues. But animals differ substantially in tick load, and this is genetically controlled. We hypothesize that the primary means by which host cattle differ in tick resistance is via their semiochemicals profiles, i.e. attractant/repellent volatile chemicals on the skin surface, with 6-methyl-5-hepten-2-one (a generic stress compound) being our primary candidate. We will perform a genome-wide association study (GWAS) for tick resistance, characterize semiochemicals which differ between cattle with high and low tick infestations (resistant and susceptible animals), identify genes differentially expressed between resistant and susceptible animals, integrate the results to obtain insights into the genetic and biochemical basis of tick resistance, and devise control options. The GWAS will be performed on >1000 Girolando cattle in Brazil, which will have been intensively phenotyped for tick burden with detailed epidemiological data (to identify risk factors). GWAS will be performed using data from the high density bovine SNP chip, giving >750,000 genotypes per animal, and analysed with current state-of-the-art techniques. From skin rubbings from animals with extreme tick counts, semiochemical profiles will be characterised using high resolution chromatography (GC, HPLC) and spectroscopic analysis. Gene expression will be performed using RNAseq on skin biopsies from extreme animals, before and after infestation, and pathways co-expressed with resistance determined. Together these data will inform on true extent of genetic control, underlying mechanisms and indicate actual loci contributing to variation. Validated SNPs for resistance will be identified, as will potential semiochemicals to be used as repellents.

The impact of this project will be upon 1) Brazilian and UK academia and R&D, specifically research institutes and university departments undertaking animal health and agricultural/food production research, 2) business, particularly dairy industries worldwide, 3) Government and inter-Governmental policy makers, particularly those involved in tick control and livelihoods of animal keepers, 4) animal welfare, 5) the environment and 6) the general public. Currently, world food production is characterized by rising commodity prices and concerns over environmental impacts of livestock production. These are difficult challenges at a time when there is increased world-wide demand for animal protein yet continual pressure on animal producers.

Tick infestations are an intractable problem in many parts of the world, worsening with reduced efficacy of acaricides and pressures to limit acaricide usage. Ticks affect animal health, directly and through tick-borne diseases, animal welfare, productivity and the environmental. Sustainable solutions are urgently sought: those provided by this project potentially include the use of host genetics, refined repellents and improved management strategies - and hence will be appropriate for livestock production systems varying greatly in sophistication and technological capabilities. For example, this project will identify SNPs that can be used to breed animals for enhanced resistance. This is a simple, sustainable, cumulative and long term solution, exploiting the fact that dairy industries world-wide now select animals using SNP chip assays. However, it requires a sophisticated breeding infrastructure to work, making it an appropriate technology for most but not all cattle breeders. However, if semiochemicals are identified that can be used as tick repellents we will have a control technique that can be implemented in most situations, particularly where breeding solutions are challenging or immediate control is required. Lastly, if improved management practices can be identified from the epidemiological analyses, these can be applied in any situation, alongside (and enhancing) other control strategies.

The project will impact considerably at the environmental level. The cattle industry has a large carbon footprint and is responsible for a significant proportion of the gaseous emissions that impact on global warming. Switching away from cattle production is not an option in the short term, as the major products (milk and beef) are highly valued by society. Thus, the most straightforward way of reducing these environmental impacts is through increasing the efficiency of the production system, whilst respecting animal welfare; it can be demonstrated that this will lead to significant reductions in environmental impact. Outputs of this project will lead towards this goal. Secondly, tick control has traditionally been through acaricide usage. In addition to acaricide resistance, such chemicals carry a high environmental and human health impact, with chemical residue concerns, and their usage should be minimized. This project will move towards providing the solutions to substitute these chemicals by more sustainable, environmentally friendly solutions.

The project will impact on the general public through more efficient and productive ruminant livestock industries, as this will lead to a more sustainable supply of high quality dairy and beef products. The public will also take comfort knowledge that animals are being farmed in a more welfare-friendly way. The public will also benefit in the medium term from decreased chemical usage, and in the long term from the contributions to decreased environmental impact from cattle production.

In summary, this project will make major contributions to animal health, welfare and productivity, hence to food security, with impacting on Government, through industry, to the general public. The return on investment for the research is potentially enormous.