Development of recombinant BCG vaccine and complementary diagnostics for TB control in cattle.

开发用于牛结核病控制的重组卡介苗疫苗和补充诊断。

• 批准号: BB/L004569/1
• 负责人: Johnjoe McFadden
• 金额: $ 69.12万
• 依托单位: University of Surrey
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2014
• 资助国家: 英国
• 起止时间: 2014 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FL004569%2F1
• 关键词: Development; recombinant; BCG; vaccine; complementary

Bovine tuberculosis (TB) caused by Mycobacterium bovis remains a major problem in both the developed world and in developing countries such as India. As well as being a cause of huge economic loss in livestock farming, M. bovis infection can be spread to humans from infected cattle by aerosol or by consumption of contaminated dairy products to cause zoonotic tuberculosis. Control of bovine TB in the west is by detection and subsequent killing of infected animals. Detection of infection is primarily by the tuberculin skin test (PPD) in which a preparation of crude denatured antigen from the tubercle bacillus (tuberculin) is injected under the skin of the animal and is then examined several days later, by which time infected animals have generated a strong visible inflammatory reaction at the site of injection. The test is cheap and effective but it suffers from a major drawback: animals that have been vaccinated with the standard TB vaccine, BCG (an attenuated strain of the bovine TB bacillus Mycobacterium bovis), report positive in the test. This cross-reactivity means that the PPD test cannot be used to diagnose TB in vaccinated animals. So, in order to retain the PPD test, BCG vaccination is not used to control bovine tuberculosis, despite its established efficacy in cattle. An additional problem for any diagnosis and killing control strategy in India is that it is culturally unacceptable to cull infected cattle. The same problem of PPD cross-reactivity with BCG besets programmes for control of TB in humans and is the reason why BCG vaccination is not used in several countries, most notably the USA. However, new diagnostic tests have recently been developed that can distinguish between BCG vaccination and natural infection with the TB bacillus. These tests are based on immunogenic proteins whose genes were spontaneously lost from the genome of M. bovis during the development into the BCG vaccine and so can be used to Differentiate between natural Infection with M. bovis and VAccination with BCG (DIVA antigens). These tests, applied as blood-based interferon-gamma release assays (IGRA), are now widely used to diagnose TB in humans; but they are not as sensitive as PPD for detecting M. bovis infection in cattle. An additional drawback is that the tests are laboratory-based requiring blood samples to be taken from infected animals, transported to the laboratory within a tight timeline and under temperature controlled transport conditions, the test performed and result reported back to the field. This makes current IGRA DIVA tests expensive and inappropriate for countries, such as India, with limited technological and scientific infrastructure. However, over recent years significant progress has been made in applying these DIVA antigens in skin tests. These studies demonstrated that DIVA skin testing constitutes a realistic and practical alternative to IGRA tests with sensitivities approaching, but not yet reaching, those of PPD. Importantly, their specificity, when applied in skin tests, appears to be superior compared to their use in IGRA (>99 %) Therefore, these studies provided proof of principle that a much more cost-effective control strategy for bovine tuberculosis, particularly in India, would be to implement a combination of a cheap and effective vaccine (such as BCG) together with a cheap and effective skin test diagnostic reagent to identify and segregate infected animals from the rest of the herd. It is the aim of this project to develop such a system. The project will have two complementary components. First we will delete genes encoding additional dispensable antigens from the current BCG vaccine, effectively convert non-DIVA antigens present in both in M. bovis and BCG into DIVA antigens present in M. bovis but absent from BCG. In parallel, we will develop a standardized skin test reagent containing a cocktail of both current and new DIVA antigens (those removed from the new BCG vaccine).

由牛分枝杆菌（Mycobacterium bovis）引起的牛结核病（TB）在发达国家和发展中国家如印度仍然是一个主要问题。除了造成畜牧业的巨大经济损失外，M。牛传染病可透过气溶胶或食用受污染的奶制品而由受感染的牛传染给人类，从而引起人畜共患结核病。在西方，控制牛结核病的方法是检测并随后杀死受感染的动物。感染的检测主要通过结核菌素皮肤试验（PPD）进行，其中将来自结核杆菌的粗变性抗原（结核菌素）的制剂注射到动物的皮肤下，然后在几天后进行检查，此时感染的动物在注射部位已产生强烈的可见炎症反应。该测试是廉价和有效的，但它有一个主要的缺点：动物接种了标准的结核疫苗，卡介苗（牛结核杆菌牛分枝杆菌的减毒株），在测试中呈阳性。这种交叉反应性意味着PPD试验不能用于诊断接种动物的结核病。因此，为了保留PPD试验，BCG疫苗接种不用于控制牛结核病，尽管它在牛中已确定有效。在印度，任何诊断和宰杀控制策略的另一个问题是，从文化上讲，宰杀受感染的牛是不可接受的。PPD与BCG交叉反应的同样问题困扰着人类结核病控制计划，这也是BCG疫苗接种在几个国家（尤其是美国）未使用的原因。然而，最近开发了新的诊断测试，可以区分卡介苗接种和结核杆菌的自然感染。这些试验是基于免疫原性蛋白质，其基因从M基因组中自发丢失。牛支原体在BCG疫苗研制过程中的特异性，因此可用于区分天然感染与M. Bovis和BCG（DIVA抗原）的VAccination。这些测试，作为基于血液的干扰素-γ释放测定（IGRA），现在被广泛用于诊断人类结核病;但它们不如PPD检测M敏感。牛的感染。另一个缺点是测试是基于实验室的，需要从受感染的动物中采集血样，在严格的时间轴内并在温度受控的运输条件下运输到实验室，进行测试并将结果报告给现场。这使得目前的IGRA DIVA测试昂贵，不适合印度等技术和科学基础设施有限的国家。然而，近年来在皮肤试验中应用这些DIVA抗原方面取得了重大进展。这些研究表明，DIVA皮肤试验构成了IGRA试验的现实和实用的替代品，其敏感性接近但尚未达到PPD的敏感性。重要的是，当应用于皮肤试验时，它们的特异性似乎优于它们在IGRA中的使用（> 99%）。因此，这些研究提供了原则证据，即用于牛结核病的更具成本效益的控制策略，特别是在印度，将一种廉价有效的疫苗（如BCG）与廉价有效的皮肤测试诊断试剂一起使用，以识别受感染的动物并将其与畜群中的其他动物隔离。本项目的目标就是开发这样一个系统。该项目将有两个相辅相成的组成部分。首先，我们将从目前的BCG疫苗中删除编码额外DIVA抗原的基因，有效地转化存在于M. bovis和BCG转化为存在于M. bovis，但不含BCG。与此同时，我们将开发一种标准化的皮试试剂，其中含有现有和新DIVA抗原（从新BCG疫苗中去除的抗原）的混合物。

项目成果

Transposon libraries identify novel Mycobacterium bovis BCG genes involved in the dynamic interactions required for BCG to persist during in vivo passage in cattle.

转座子文库鉴定出新的牛分枝杆菌 BCG 基因，这些基因涉及 BCG 在牛体内传代过程中持续存在所需的动态相互作用。

• DOI: 10.1186/s12864-019-5791-1
• 发表时间: 2019
• 期刊: BMC genomics
• 影响因子: 4.4
• 作者: Mendum TA

Point of Care Tuberculosis Sero-Diagnosis Kit for Wild Animals: Combination of Proteins for Improving the Diagnostic Sensitivity and Specificity.

野生动物床旁结核病血清诊断试剂盒：蛋白质组合可提高诊断敏感性和特异性。

• DOI: 10.1007/s12088-017-0688-7
• 发表时间: 2018
• 期刊: Indian journal of microbiology
• 影响因子: 3
• 作者: Veerasami M

Genetic screening for the protective antigenic targets of BCG vaccination.

BCG 疫苗接种保护性抗原靶点的基因筛查。

• DOI: 10.1016/j.tube.2020.101979
• 发表时间: 2020
• 期刊: Tuberculosis (Edinburgh, Scotland)
• 作者: Smith AA