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

壁虱和壁虱传播的疾病对家畜、人类和同伴动物都有重大影响。特别是，牛虱在世界各地高温高湿的地区都有发现。扁虱感染导致体重减轻、贫血和继发感染，以及传播巴贝斯虫病、泰勒虫病和无浆体病等疾病。传统上，化学物质(杀螨剂)是防治硬蜱的主要策略，但这种方法越来越有问题，主要原因是抗杀螨剂的进化和化学残留物的产生。由于疫苗的效果好坏参半，因此迫切需要新的控制措施。牛的壁虱负荷差异很大，而这种变异很大程度上是由宿主的遗传组成控制的，因此育种可以提高对壁虱的抵抗力。我们推测，宿主牛在抗蚤能力上的不同主要是通过它们在皮肤表面产生吸引/驱避的挥发性化学物质，即信号化学物质，最终决定有多少扁虱附着在皮肤表面。识别这些化学物质以及导致不同动物产生不同化学物质的遗传因素将给我们带来强大的新的控制机会。我们可以培育对虱子更具抵抗力的动物，或者我们可以设计出驱虫剂来减少虱子的载量。该项目旨在实现这一目标：目标是从遗传和生物化学水平上了解巴西牛对壁虱侵染抗性的遗传变异背后的因素。首先，我们将收集巴西1000多头商业吉罗兰多牛的虱子载量、生物样本和可能影响虱子载量的因素的详细数据。这些数据将提供影响扁虱负担的因素的概述。从血液中提取DNA后，我们将使用一种名为高密度SNP芯片的工具对每种动物进行基因分型。这使我们能够同时对动物进行基因分型，在基因组中超过75万个位置进行遗传标记。对这些数据的分析将告诉我们，在基因组中的哪些基因对虱子的负担有很大的影响，从而对虱子的抵抗力有很大影响。但它并没有告诉我们是哪些基因。为了找出哪些基因是重要的，我们将更详细地研究极端动物。使用从扁虱负担持续最高和最低的动物身上获得的皮肤拓片，我们将使用层析和光谱分析来表征这两组动物产生的改变行为的化学物质(信号化学物质)。我们将测试这样一种假设，即扁虱对叮咬苍蝇的信号化学物质(例如6-甲基-5-庚烯-2-酮)的反应是相同的。我们还将购买具有我们预测的代表极端易感性和抵抗力的基因型的动物，并用扁虱挑战这些动物。然后，我们将使用皮肤活检来观察这些动物在挑战前后的RNA图谱。RNA图谱将告诉我们哪些基因正在表达，我们可以在抗性动物和敏感动物之间比较这些图谱。这将告诉我们哪些基因和生化途径是抗药性的预测。利用生物信息学，我们将把我们所有的结果汇集在一起。我们将结合基因表达和信号化学研究来确定哪些基因最有可能支持抗药性的变异。然后，我们将把这些结果与SNP芯片的结果联系起来：如果可能的基因落在基因组中已被证明影响扁虱抗性的区域，那么我们就有非常有力的证据表明影响抗性的基因(和遗传标记)。然后，我们将通过对另外200头表型奶牛进行基因分型来验证我们的结果，并制定提高抵抗力的育种策略。我们还将从我们的数据中调查基于信号化学或畜牧业的干预措施。

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.