The Xenopus tropicalis response to infection by Batrachochytrium dendrobatidis

冷血脊椎动物对传染病的个体水平反应是一个被忽视的研究领域。然而，最近出现的外温热疾病已实现的和潜在的经济代价是巨大的。丛枝孢子菌(Batrachochytrium Dendrobatidis)于1997年被描述为圈养两栖动物的病原体，目前在全球范围内正在摧毁商业两栖动物养殖场、研究群体和自然种群。目前对这种真菌感染的了解显然是不够的；我们理解的主要空白是两栖动物如何对这种寄生虫做出反应，以及这种寄生虫如何导致宿主死亡。我们缺乏这些信息是因为缺乏可靠的两栖动物模型系统，可以用来识别支撑宿主-病原体相互作用的生物学机制。我们的项目将通过研究使用非洲爪蛙作为模型系统来研究丝状真菌病的可行性来纠正这种不平衡。与其姊妹分类群X.laevis不同的是，X.topicalis非常容易患上壶菌病。热带疟原虫的完整注释基因组序列现在已经完成，这有助于分析两栖动物对感染的反应，这是通过关键感染相关基因的变化来衡量的。我们的初步实验将集中于表征热带X线虫在(I)不同剂量的病原体和(Ii)不同温度下对感染的反应。使用共聚焦显微镜，我们将通过对与非洲爪哇免疫反应的体液和细胞介导的手臂相关的关键抗原进行染色，来可视化非洲爪哇与病原体的相互作用。随后，我们将分析未感染、非致死性感染和致死性感染动物之间的基因表达变化。我们将通过设计可靠的定量PCR分析来监测感染过程中基因的表达，这些基因涉及(I)Th1(细胞介导的)、(Ii)Th2(基于抗体)和(Iii)先天免疫反应。我们将重点分析已知在控制其他脊椎动物感染中起重要作用的基因，并通过使用比较基因组技术从热带X.tropicalis基因组中挖掘数据。这些分析将首次揭示两栖动物对感染的适应性和先天反应的类型和大小(S)。了解这些过程将使我们能够进一步发展热带X线虫作为模式物种，在这个模式物种中，我们可以在拥有发达的免疫学和遗传工具箱的易管理系统中研究宿主-病原体生物学。这将使我们能够将我们的研究应用于针对树枝杆菌的疫苗开发，考虑到这种疾病的全球经济成本，这是一个紧迫的目标。最后，确定两栖动物中与传染病相关的宿主反应的类型，对于理解其他迄今被忽视的冷血物种的宿主反应将具有广泛的相关性。

The individual-level response of cold-blooded vertebrates (ectotherms) to infectious disease is a neglected area of research. However, the realized and potential economic costs of recently emerged diseases of ectotherms are substantial. The chytrid fungal pathogen, Batrachochytrium dendrobatidis, was recently described in 1997 as a pathogen of captive amphibians, and chytridiomycosis is presently devastating commercial amphibian farms, research colonies and natural populations on a global basis. The current state of knowledge regarding this fungal infection is manifestly inadequate; major gaps in our understanding are how amphibians respond to this chytrid and how this chytrid causes the death of its host. We lack this information due to the absence of a reliable amphibian model system that can be used to identify the biological mechanisms underpinning the host-pathogen interaction. Our project will rectify this imbalance by investigating the feasibility of using the African clawed frog Xenopus tropicalis as a model system to research chytridiomycosis. X. topicalis, unlike its sister taxa X. laevis, is highly susceptible to chytridiomycosis. The fully annotated genome sequence for X. tropicalis is now complete which facilitates the analysis of amphibian responses to infection as measured by changes in key infection-related genes. Our initial experiments will focus on characterising the X. tropicalis-response to infection under (i) different dosages of the pathogen and (ii) different temperatures. Using confocal microscopy, we will visualise the Xenopus-pathogen interaction by staining against key antigens that are associated with the humoral and cell-mediated arms of the Xenopus immune-response. Subsequently, we will then analyse the changes in gene-expression between uninfected, non-lethally infected and lethally infected animals. We will do this by designing robust quantitative PCR assays that monitor gene-expression over the time-course of an infection for classes of genes that are involved in (i) the Th1 (cell-mediated), (ii) the Th2 (antibody-based) and (iii) innate immune response. We will focus our analyses on genes that are known to be important in controlling infection in other vertebrate species, and that we have data-mined from the X. tropicalis genome by our use of comparative-genomic techniques. These analyses will reveal, for the first time, the type and magnitude of the amphibian adaptive and innate response(s) to infection. Understanding these processes will allow us to further develop X. tropicalis as a model species within which we can investigate host-pathogen biology within a tractable system that has well-developed immunological and genetic-toolboxes. This will allow us to apply our research to vaccine development against B. dendrobatidis, an urgent goal given the global economic cost of this disease. Finally, determining the type of host response that is associated with infectious disease in amphibians will have a broad relevance for understanding host responses in other, thus far neglected, cold-blooded species.