A reverse vaccinology approach to a bTB vaccine

牛结核病是由牛分枝杆菌感染牛引起的。这种细菌在自然界中分布广泛，因为它还感染许多其他野生动物物种，因此，野生动物感染充当细菌的宿主，定期进入家牛。这样做的后果是双重的。首先，BTB阳性的牛必须被扑杀，这会对农民及其维持牛群的能力产生连锁反应。其次，BTB对人类感染构成威胁，主要是通过食用受污染的牛奶。今天，结核杆菌对个人的感染极其罕见，但这种威胁依然存在，因此，结核杆菌感染是不受欢迎的，需要控制或最好是根除。自20世纪90年代中期的历史回顾以来，獾已被确定为BTB向家牛传播的主要途径之一，这反过来又导致了试图通过宰杀獾来打破这一传播途径的尝试。这些措施只取得了有限的成功和公众的极大关注，几乎没有改变对传播路线的物理破坏。另一种方法是接受BTB在环境中广泛传播，但通过以前的疫苗接种来防止牛感染。然而，目前还没有一种有效的牛结核病疫苗，迫切需要开发这种疫苗的新方法。人们普遍认为，疫苗的功能是通过在目标动物体内产生抗体反应来防止细菌确定最初的感染。这些抗体是所有哺乳动物免疫系统的正常产物，通常与细菌的外部结合，因此阻止它与牛细胞结合，通常是肺上皮细胞。因此，可以在细菌表面的外面找到BTB的保护性成分，也就是产生保护性抗体反应的成分。细菌表面有许多成分，这些成分中的任何一个，或者可能是许多成分的组合，由细菌基因组编码的蛋白质，可能对有效免疫的发展至关重要。然而，目前还不清楚到底需要哪些。在这项研究中，我们建议生产BTB的所有表面成分，并批量测试它们诱导有效免疫反应的能力。我们建议在牛身上做这项工作，以便对我们的测试疫苗的反应是典型的，如果最终的疫苗用于典型的牛群。我们的工作分为三个相关部分。首先，我们将从牛分枝杆菌中鉴定出在生物体表面发现的所有蛋白质，并以安全和有效的方式生产每一种蛋白质。我们的初步计算表明，可能需要数百种这样的蛋白质才能找到有效免疫所必需的少数蛋白质。其次，我们将使用我们的表面蛋白作为牛的试验疫苗，为了使这一过程高效，我们将使用混合蛋白进行这项工作，以便使用最少的牛。免疫接种后，我们将从牛身上提取血液样本，并测试它们预防牛分枝杆菌感染的能力。最后，我们将研究保护机制，以及我们所确定的各个组成部分如何协同工作，为牛提供有效的免疫屏障。我们的方法是详尽的，但它有可能为结核病的疫苗发现计划划上句号，并确定最终有效疫苗生产的最佳候选组合。

Bovine tuberculosis, bTB, is the result of the infection of cattle by the bacterium Mycobacterium bovis. The bacterium is distributed widely in nature as it also infects many other wildlife species and as a result of this, wildlife infection acts as a reservoir for the bacterium which periodically get across into domestic cattle. The consequence of this is twofold. First, cattle that are bTB positive must be culled with a knock-on effect on the farmer and his ability to maintain a herd. Second, bTB is a threat for human infection, primarily via the consumption of contaminated milk. Today bTB infection of individuals is extremely rare, but the threat remains and for these reasons bTB infection is unwelcome and needs to be controlled or, preferably, eradicated. Since a historical review in the mid-1990s, the badger has been identified as one of the major routes of transmission of bTB to domestic cattle and this, in turn, has led to attempts to break this transmission route via badger culling. These have met with only limited success and immense public concern leaving the physical breaking of the transmission route mostly unaltered. An alternative approach is to accept that bTB circulates widely in the environment but to prevent cattle infection by previous vaccination. At the present time however an effective vaccine for bTB in cattle is not available and new methods to develop such a vaccine are urgently needed. It is generally accepted that vaccines function by generating an antibody response in the target animal which prevents the bacterium from establishing the initial infection. These antibodies, which are a normal product of the immune system of all mammals, generally bind to the outside of the bacterium and so prevent it from binding to cattle cells, usually the epithelial cells of the lung. It follows that the protective components of bTB, that is, the components that will generate the antibody response that is protective, are to be found on the outside of the bacterium surface. Bacteria have many components on their surface and any, or perhaps a combination of many, of these components, proteins encoded by the bacterium genome, could be essential for the development of effective immunity. However exactly which are required is currently unknown. In this research, we propose to produce all of the surface components of bTB and to test them in batches for their ability to induce an effective immune response. We propose to do this work in cattle so that the response measured to our test vaccines is typical of what will be found if an eventual vaccine is used in typical herds. Our work breaks down into three related components. Firstly, we will identify all of those proteins from the M. bovis bacterium that are to be found on the surface of the organism and produce each of them in a safe and efficient manner. Our initial calculations suggest that several hundred such proteins may be required in order to find the few that are necessary for effective immunity. Secondly, we will use our surface proteins as test vaccines in cattle and to make this process efficient we will carry out this work with mixes of proteins so that the least number of cattle has to be used. Following the immunizations we will take blood samples from the cattle and test them for the ability to prevent M. bovis infection. Finally, we will examine the mechanism of protection and how the individual components so we have identified work together to provide the cattle with an effective barrier of immunity. Our approach is exhaustive but it has the potential to draw a line under the vaccine discovery program for bTB and to identify the best mix of candidates for eventual effective vaccine production.