Bluetongue virus transmission between ruminant host and Culicoides vectors: the importance of insect saliva proteins

蓝舌病病毒（BTV）是一种节肢动物传播的病毒（一种虫媒病毒），通过吸血吸血的蠓（库蠓属）在反刍动物宿主之间传播。该病毒可感染大多数反刍动物，但在绵羊（特别是欧洲品种）、某些鹿种以及在较小程度上在牛中引起出血热和严重的临床疾病。直到十年前，蓝舌病还是一种气候温暖、有传播能力的昆虫媒介种群的地区的疾病。然而，自1998年以来，BTV已逐步向北进入欧洲，这一过程与当地气候变暖和某些蠓媒介种群分布扩大有关。2006年，蓝舌病的爆发比以往任何时候都更向北，影响了比利时、荷兰、德国、卢森堡和法国东北部。该病毒导致羊甚至牛出现临床疾病，大约50%的羊和10%的牛出现临床症状死亡。这次暴发证实，一些北欧蠓种（与英国相同）可以传播BTV，并表明包括英国在内的整个欧洲现在都面临进一步暴发的威胁。然而，并非所有蠓种都能有效传播BTV，这表明病毒的传播和疾病的分布受昆虫媒介和所涉及的病毒株的影响。吸血昆虫将唾液沉积到宿主的皮肤上。这会引起皮肤的炎症和过敏，从而导致常见的红点和瘙痒，甚至在蚊虫叮咬后出现过敏性囊泡。BTV通过昆虫的唾液传播，必须在炎症反应中存活下来才能感染反刍动物宿主。然而，唾液也含有蛋白酶（有助于防止凝血），它可以修饰病毒，甚至增加其对昆虫载体的传染性。通常认为虫媒病毒是通过喂食昆虫而被注射到宿主的血液中。然而，蠓用它们的口器来切割和损伤皮肤组织，所以它们可以以血液、组织和淋巴液的混合池为食。BTV不注射到血管中，而是留在唾液中沉积在皮肤中。病毒可以在某些皮肤细胞中复制，另外在宿主的炎症反应细胞中复制，这些细胞被吸引到咬伤部位。因此，BTV在皮肤中的含量可能高于血液中的含量，这可能对吸血蠓的吸收很重要。因此，病毒似乎已经进化到可以利用唾液蛋白和宿主的反应，来提高传播效率和生存机会。我们最近开发了大量分离蚊唾液蛋白的方法，以研究其在媒介昆虫与反刍动物宿主之间传播BTV的作用。最初的研究表明，不同种类的蠓有不同的唾液蛋白，这可能有助于解释它们载体“能力”的差异。皮尔布赖特有独特的设施来研究蠓、羊和蓝舌病病毒本身，这将使我们能够探索它们之间的关系以及对传播机制的影响。该项目将比较来自欧洲、非洲和美洲不同蠓类的唾液，以确定唾液蛋白在哺乳动物和昆虫组织培养细胞或绵羊中BTV感染中的作用。该项目将阐明皮肤中本地产生的病毒对BTV传播的重要性，并将确定病毒是否可以在受感染和未受感染的绵羊之间机械传播（通过其他昆虫或针），或在一只蚊子以同一只羊为食时直接从一只蚊子传播到另一只蚊子。这项研究的结果将提高我们对欧洲蓝舌病传播的认识，有助于确定蓝舌病传播的风险并实施适当的控制措施。

Bluetongue virus (BTV) is an arthropod-borne virus (an Arbovirus) transmitted between its ruminant hosts by blood feeding 'biting' midges (Culicoides spp.). The virus can infect most ruminants but causes haemorrhagic fever and severe clinical disease, in sheep (especially European breeds), some species of deer and to a lesser extent in cattle. Until a decade ago bluetongue was a disease of warm climates, areas containing populations of transmission-competent insect vector species. However, since 1998 BTV has moved progressively further north into Europe, a process related to warming of the local climate and an expanding distribution of certain midge vector populations. In 2006, a bluetongue outbreak occurred much further north than ever before, affecting Belgium, the Netherlands, Germany, Luxembourg, and North East France. The virus causes serve clinical disease in sheep and even cattle and approximately 50% of sheep and 10% of cattle developing clinical signs die. This outbreak confirms that some northern European midge species (which are the same as in the UK) can transmit BTV, and demonstrates that the whole of Europe, including the UK, is now under threat of further outbreaks. However, not all species of midge can transmit BTV efficiently, suggesting that transmission of the virus and the distribution of the disease is influenced by the insect vectors and the strain of virus involved. Blood feeding insects deposit saliva into the skin of the host. This causes inflammation and hypersensitivity in the skin, which results in the familiar red spots and itching, or even allergic vesicles, after midge or mosquito bites. BTV is transmitted in the insect's saliva and must survive the inflammation response in order to infect the ruminant host. However, the saliva also contains proteases (to help prevent clotting) which can modify the virus, even increasing its infectivity for the insect vector. It is often assumed that Arboviruses are injected into the blood of the host by feeding insects. However, midges use their mouthparts to cut and damage skin tissues, so they can feed on a mixed pool of blood, tissue and lymphatic fluid. BTV is not injected into blood vessels but left in the saliva deposited into the skin. The virus can replicate in certain skin cells and additionally in cells of the hosts inflammatory response that are attracted to the biting site. Therefore BTV may be present at higher levels in the skin than in the bloodstream, which may be important for uptake by feeding midges. So it appears that the virus has evolved to make use of the saliva proteins and the host's response, to increase the efficiency of transmission, and its chances of survival. We have recently developed methods to isolate midge saliva proteins in large quantities, to investigate their role in BTV transmission between vector insects and ruminant hosts. Initial studies indicate that different midge species have different saliva proteins, which may help explain differences in their vector 'competence'. There are unique facilities at Pirbright to work on midges, sheep and the bluetongue virus itself, that will allow us to explore their relationships and influence on transmission mechanisms. The project will compare saliva from different European, African and American midges, to determine the role of saliva proteins in BTV infection in mammalian and insect tissue culture cells, or in the sheep. The project will elucidate the importance of locally generated virus in the skin for BTV transmission and will also determine if the virus can be transmitted mechanically between infected and non-infected sheep (by other insects or by needle), or directly from one midge to another while they feed on the same sheep. The results of this research will improve our understanding of BTV transmission in Europe, helping to define the risk of bluetongue transmission and to implement appropriate control measures.