Development of improved vaccines effective against emerging strains of the fish pathogen Yersinia ruckeri

尽管在疫苗开发和化疗方面取得了进展，但细菌性病原体仍然是水产养殖生产的重大制约因素，也是对野生种群的潜在威胁。除了所谓的“新”细菌性病原体外，威胁往往来自已存在的病原体，它们不断进化，以绕过已制定的控制战略。这种不断进化的病原体的一个例子是革兰氏阴性细菌拉克氏耶尔森氏菌，它引起大西洋鲑鱼（Salmo salar）和虹鳟（Oncorhynchus mykiss）的重要经济感染。三十多年来，在英格兰和威尔士的养殖虹鳟鱼中，ruckeri被确定为肠道红嘴病（ERM）的病原。这种有机体也会在养殖的大西洋鲑鱼中引起重大的疾病爆发，但人们对该物种的拉克氏菌流行病学知之甚少。直到最近，主要由于广泛使用疫苗接种，拉克氏菌对养殖鳟鱼和鲑鱼的风险降低。然而，世界范围内越来越多的实地疫苗失效报告被认为与新的拉克氏耶氏菌致病性菌株的出现有关。传统上，美国和欧洲虹鳟鱼中的大多数ERM感染是由密切相关的O1血清型洛克氏菌菌株的一个亚群（生物型）引起的，以所谓的哈格曼菌株为代表。然而，近年来出现了一种新的、独特的血清型O1生物型，它首先在英国被发现，但现在正日益影响整个欧洲和美国的虹鳟鱼生产。从受感染的大西洋鲑鱼中分离出来的菌株似乎更加多样化，代表了更广泛的o血清型（O1、O2和O5）。初步证据表明，鲑鱼致病性血清型对虹鳟的致病性明显低于对大西洋鲑鱼的致病性。革兰氏阴性菌的外膜（OM）位于病原体和宿主之间的界面，外膜蛋白（OMPs）在宿主与病原体的相互作用中起着关键作用，如粘附和定植、营养吸收（特别是铁的获取）、逃避宿主免疫反应和毒力因子的分泌。由于许多omp受到强大的选择压力（例如来自宿主免疫系统），它们可以在单一细菌物种中表现出不同程度的多样性。这种多样性很可能赋予菌株不同的毒力属性和宿主嗜性。此外，许多omp是表面暴露和免疫原性的，因此可能作为保护性抗原。尽管拉克氏菌是养殖鲑科鱼的重要病原体，但人们对其不同菌株中omp的多样性及其在疾病过程中的作用知之甚少。此外，我们对鲑鱼和鳟鱼体内宿主对拉克氏菌的免疫反应知之甚少，也不知道是哪些细菌因素保护了它们免受感染。这个项目的目的是：1。开展苏格兰鲑鱼鲁克氏菌流行病学调查，并评估该生物在海水中传播的有效性。2. 对不同生长条件下从鲑鱼和鳟鱼中分离的关键拉克氏耶夫氏菌的OM蛋白组进行鉴定，并通过免疫蛋白组学方法鉴定交叉保护抗原（和潜在的候选疫苗）。3. 比较选定的分离株对大西洋鲑鱼和鳟鱼的毒力。4. 比较大西洋鲑鱼和鳟鱼对拉克氏菌分离株的免疫反应动力学。5. 目的研究大西洋鲑鱼和鳟鱼不同分离株鲁克氏耶夫氏菌制备的疫苗免疫的血清型特异性和保护作用。

Despite advances in vaccine development and chemotherapy, bacterial pathogens remain a significant constraint to aquaculture production, as well as a potential threat to wild stocks. In addition to so-called 'new' bacterial pathogens, the threat often comes from established pathogens evolving to circumvent developed control strategies. An example of such an evolving pathogen is the Gram-negative bacterium Yersinia ruckeri which causes economically-important infections of Atlantic salmon (Salmo salar) and rainbow trout (Oncorhynchus mykiss). Y. ruckeri has been identified as the aetiological agent of enteric redmouth disease (ERM) of farmed rainbow trout in England and Wales for more than thirty years. The organism also causes significant outbreaks of disease in farmed Atlantic salmon but very little is known about the epidemiology of Y. ruckeri in this species. Until recently, Y. ruckeri posed a reduced risk to farmed trout and salmon due largely to the extensive use of vaccination. However, increasing worldwide reports of vaccine breakdown in the field are believed to be associated with the emergence of new pathogenic strains of Y. ruckeri. Traditionally, the majority of ERM infections in rainbow trout in the USA and Europe have been caused by a subgroup (biotype) of closely-related serotype O1 Y. ruckeri strains represented by the so-called Hagerman strain. However, a new and distinct serotype O1 biotype has emerged in recent years that was first identified in the UK but which is now increasingly affecting rainbow trout production across Europe and the USA. Isolates recovered from infected Atlantic salmon appear to be more diverse and represent a wider range of O-serotypes (O1, O2 and O5). Preliminary evidence suggests that salmon-pathogenic serotypes are significantly less pathogenic to rainbow trout than they are to Atlantic salmon. The outer membrane (OM) of Gram-negative bacteria is at the interface between pathogen and host and outer membrane proteins (OMPs) play key roles in host-pathogen interactions such as adherence and colonization, nutrient uptake (specifically iron acquisition), evasion of the host immune response and secretion of virulence factors. Because many of these OMPs are under strong selection pressures (e.g. from the host immune system) they can exhibit varying degrees of diversity within a single bacterial species. This diversity is likely to confer upon strains different virulence attributes and host tropisms. In addition, many OMPs are surface-exposed and immunogenic and are therefore likely to serve as protective antigens. Despite the importance of Y. ruckeri as a pathogen of farmed salmonids, very little is known about the diversity of its OMPs in different strains or their roles in the disease process. Furthermore, very little is known about the host immune response to Y. ruckeri in salmon and trout or about the bacterial factors responsible for providing protection against infection. The aims of this project are as follows: 1. To carry out an epidemiological survey of the prevalence of Y. ruckeri in Scottish salmon and assess the effectiveness of transmission of the organism in seawater. 2. To characterize the OM proteomes of key selected isolates of Y. ruckeri recovered from salmon and trout under different growth conditions and identify cross-protective antigens (and potential vaccine candidates) by immunoproteomics. 3. To compare the virulence of selected isolates for Atlantic salmon and trout. 4. To compare the kinetics of the immune response to selected isolates of Y. ruckeri in Atlantic salmon and trout. 5. To study serotype specificity of immunity and the protection afforded by vaccines prepared from different isolates of Y. ruckeri in Atlantic salmon and trout.