The immunological basis of protection obtained by prime-boost immunisation against bovine tuberculosis

牛结核病(BTB)和人类结核病(HTB)的病原体、牛分枝杆菌和结核分枝杆菌分别是导致类似疾病的密切相关的生物。牛分枝杆菌在传播给人类时也会引起疾病，这导致在20世纪50年代引入了基于牛群测试的国家根除计划，以移除受感染的牛。虽然最初在减少羊群发病率方面取得了成功，但在过去的25年里，这种疾病再次成为英国的一个主要问题。这导致考虑将接种疫苗作为另一种控制选择。自20世纪初以来，牛分枝杆菌实验室减毒株(BCG)已被广泛用于接种HTB疫苗。然而，这种疫苗显示出不同的疗效，特别是对成年人的疾病。接种卡介苗可以降低牛的发病率和病理严重程度，但只能防止一定比例的疫苗接种建立感染。因此，它的使用被认为与目前的btb控制方案不相容。因此，HTB和BTB都需要改进的疫苗。最近在人和牛身上的研究表明，与单独接种卡介苗相比，表达分枝杆菌抗原的病毒载体亚单位疫苗增强了卡介苗的免疫效果。例如，与仅接种卡介苗的动物相比，卡介苗和表达85A抗原的重组腺病毒(Rad)的组合减少了牛分枝杆菌攻击后出现可见病变的牛的数量。该项目旨在确定与增强免疫相关的免疫反应的特征。尽管众所周知，对牛分枝杆菌的免疫是由细胞免疫反应介导的，主要涉及T细胞的CD4T细胞亚群，但决定免疫的保护性CD4T细胞的确切特性尚不清楚。卡介苗诱导的免疫可以通过85A抗原的增强而增强，这一证明清楚地表明对这种抗原的反应有助于免疫。因此，该项目将重点分析用卡介苗免疫的牛对85A的免疫反应，然后用85A免疫，然后用毒力牛分枝杆菌攻击。CD4T细胞利用大量抗原特异性受体来识别出现在感染细胞表面的外来微生物蛋白的短片段。该项目将利用新开发的方法来分析这些受体的序列，以跟踪免疫和挑战后响应的85A特异性CD4T淋巴细胞群体。这种方法将与测量频率的生物测试结合使用，--85A增强免疫后，与85A反应的CD4T细胞的频率在多大程度上增加了？-85A增强免疫是否改变了应答的CD4淋巴细胞反应的精细特异性和/或其抗原特异性受体的功能？-与仅BCG免疫相比，85A增强是否增强了特定CD4T细胞的功能能力？-85A特异性CD4 T细胞反应的特定成分在识别和反应牛分支杆菌感染细胞方面比其他成分更有效吗？在牛分枝杆菌免疫动物的攻击下，85A特异性的CD4T细胞群体中的哪些成分反应最快？这些研究的结果将确定与免疫相关的特定CD4T细胞反应的特性，从而提供可测量的参数，这些参数将有助于评估对其他候选疫苗的免疫反应。尽管该项目主要与牛结核病疫苗的开发有关，但该项目的成果也将为其他物种的结核病疫苗开发研究提供信息。

The causal agents of bovine (bTB) and human tuberculosis (hTB), Mycobacterium bovis and M. tuberculosis respectively, are closely related organisms that result in similar diseases. M. bovis can also cause disease when transmitted to humans, and this led in the 1950s to the introduction of national eradication programmes based on testing of cattle herds to remove infected cattle. Although initially successful in reducing herd incidence, the disease has re-emerged as a major problem in the UK over the last 25 years. This has led to consideration of vaccination as an additional control option. A laboratory attenuated strain of M. bovis (BCG) has been used extensively since the early 20th century to vaccinate against hTB. However the vaccine shows variable efficacy, particularly against disease in adults. BCG vaccination of cattle can reduce the incidence and severity of pathology, but only prevents establishment of infection in a proportion of vaccinates. Therefore, its use has been considered incompatible with current bTB control programmes. An improved vaccine is therefore required for both hTB and bTB. Recent studies in humans and cattle have demonstrated that BCG vaccination boosted by virally vectored subunit vaccines expressing mycobacterial antigens results in enhanced protection compared to BCG alone. For example, combination of BCG and a recombinant adenovirus (rAd) expressing the 85A antigen reduced the number of cattle that presented with visible lesions after M. bovis challenge, compared to animals vaccinated with BCG only. This project aims to identify those features of the immune response induced by boosting with 85A that are associated with the enhanced immunity.Although it is well established that immunity against M. bovis is mediated by cellular immune responses, primarily involving the CD4 subset of T cells, the precise properties of the protective CD4 T cells that determine immunity are poorly understood. The demonstration that immunity induced by BCG can be enhanced by boosting with the 85A antigen clearly shows that responses to this antigen contribute to immunity. The project will therefore focus on analysing immune responses to 85A in cattle immunised with BCG followed by 85A and subsequently challenged with virulent M. bovis. CD4 T cells utilise a large library of antigen-specific receptors to recognise short fragments of foreign microbial proteins presented on the surface of infected cells. The project will utilise newly developed methods for analysing the sequences of these receptors to track the responding 85A-specific CD4 T lymphocyte populations following immunisation and challenge. This methodology will be used in conjunction with biological assays that measure the frequency, fine specificity and functional potency of the 85A-specific response to address the following questions:- To what extent is the frequency of CD4 T cells reactive with 85A increased following boost immunisation with 85A? - Does boost immunisation with 85A alter the fine specificity of the responding CD4 -lymphocytes response and/or the repertoire of their antigen-specific receptors?- Does boosting with 85A enhance the functional potency of the specific CD4 T cells compared to BCG immunisation only? - Are particular components of the 85A-specific CD4 T cell response more effective than others at recognising and responding to M. bovis-infected cells?- Which components of the of the 85A-specific CD4 T cell population respond most rapidly following challenge of immunised animals with M. bovis? The results of these studies will identify properties of the specific CD4 T cell response that are associated with immunity and thus provide measurable parameters that will be useful for assessing the immune responses to other candidate vaccines. Although primarily of relevance to vaccine development for tuberculosis in cattle, the outputs of the project will also inform studies of TB vaccine development for other species.