Early detection of immune responses against Mycobacterium avium subsp. paratuberculosis infection

早期检测针对鸟分枝杆菌亚种的免疫反应。

• 批准号: 341848-2013
• 负责人: DeBuck, Jeroen
• 金额: $ 2.19万
• 依托单位: University of Calgary
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2013
• 资助国家: 加拿大
• 起止时间: 2013-01-01 至 2014-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=527486
• 关键词: Early; detection; immune; responses; against

Johne's disease (JD), a chronic enteritis caused by Mycobacterium avium subsp. paratuberculosis (Map), is an economically important disease in dairy cattle. For effective control of JD it is important to detect and isolate infected animals before they become infectious (=start shedding). However, current diagnostic tests (fecal culture and antibody ELISA) have poor sensitivity for detecting low shedding animals. It has been well established that a cellular immune response (CIR) against Map infection precedes the production of detectable anti-Map antibodies. This Map specific response can be measured in an interferon-gamma release assay. Blood needs to be analysed in the lab within 12 h of collection, which is a major disadvantage of this test compromising its general use. Modifying this test so that it can be performed easily on-farm would dramatically improve JD control. Because the measurable CIR decreases over time it is important to combine its detection with that of specific antibodies in complementary tests. As currently no on-farm JD diagnostics exist, we aim to develop a set of on-farm molecular assays which can detect immune responses which are associated with all stages of Map infection. 1) A first type of assay will detect the CIR in infected cattle. A biosensor will be produced which converts the presence of interferon-gamma in blood stimulated with Map antigens into a colorimetric output. 2) A second type of assay will measure Map specific antibodies in serum with antibody/antigen binding triggering the activation of an enzymatic colorimetric reaction. 3) Previously, we developed different assays to detect Map infection based on biomarkers identified in experimentally infected calves by transcriptomics. These assays will be further validated on large numbers of naturally infected animals. 4) In parallel, we will use digital droplet PCR to precisely quantify shedding levels in the animals that provide blood samples for the biosensor and biomarker test development to link the test outcomes to levels of infectivity. In summary, the development of a small array of assays, together targeting several stages of infection will significantly increase the sensitivity to detect infected animals and improve JD control programs.

乔恩病（JD），一种由鸟分枝杆菌亚种引起的慢性肠炎。副结核病（MAP）是奶牛上具有经济上重要的疾病。为了有效控制JD，重要的是在传染性之前检测和隔离感染动物（=开始脱落）。但是，当前的诊断测试（粪便培养和抗体ELISA）对于检测低脱落动物的敏感性差。已经很好地确定，针对MAP感染的细胞免疫反应（CIR）先于可检测到的抗MAP抗体的产生。可以在干扰素释放分析中测量该地图的特定响应。需要在收集到12小时内在实验室中分析血液，这是该测试损害其一般用途的主要缺点。修改此测试以便可以轻松地进行农场进行，这将显着改善JD控制。由于可测量的CIR随着时间的流逝而降低，因此在互补测试中将其检测与特定抗体的检测相结合非常重要。由于目前尚无农产品JD诊断，我们旨在开发一组农场分子测定，这些分子测定可以检测与MAP感染所有阶段有关的免疫反应。 1）第一类测定将检测到被感染的牛中的CIR。 将产生一个生物传感器，该生物传感器将用MAP抗原刺激的血液中的干扰素γ转换为比色输出。 2）第二种类型的测定将测量具有抗体/抗原结合的血清中特定抗体，从而触发酶色反应的激活。 3）以前，我们开发了不同的测定方法来检测基于通过转录组学实验感染犊牛中发现的生物标志物的MAP感染。这些测定法将在大量自然感染动物上得到进一步验证。 4）同时，我们将使用数字液滴PCR准确地量化动物的脱落水平，这些动物为生物传感器和生物标志物测试开发提供血液样本，以将测试结果与感染水平联系起来。总而言之，针对多个感染阶段的一小部分测定法的发展将显着提高检测感染动物并改善JD控制程序的敏感性。