Rapid Diagnosis of Early TB in HIV+ Patients

印度的结核病诊断依赖于临床症状、直接从痰液中提取的涂片显微镜检查和/或胸部X光检查（CXR）。由HIV感染引起的免疫功能障碍改变了TB的临床和放射学表现，并且TB的微生物确认是有问题的，因为HIV+TB+患者具有增加的涂片阴性（SN）TB和肺外TB（EPTB）的频率。尽管现有药物完全可以治疗，但对HIV+患者不敏感的诊断检测的性能较差，导致结核病进展迅速且往往致命的人群中结核病的诊断严重不足。一种简单、快速的检测方法来识别HIV+患者中的涂阳（SP）和SN症状性或无症状性结核病，将对结核病相关的发病率和死亡率产生重大影响。 M.已经确定了结核病的潜在候选者，以设计用于HIV+患者的结核病的简单诊断测试。它们是MS、MPT 51、PPE 55、ESAT 6和CFP 10。这些抗原的抗体（Ab）已被证明在所有水平的免疫抑制，无论肺部表现和结核病的艾滋病毒+结核病患者的网站，也可检测到无症状的艾滋病毒+患者的血清中发展为症状性结核病在随后的2-6个月。在来自最终未发展为TB的CD 4 T细胞匹配的HIV+患者的回顾性血清中缺乏针对这些抗原的Ab表明，在HIV+患者中，这些Ab应答表明活动性TB，无论是否存在TB的临床症状，并且在细菌负荷达到通过直接显微镜检测的阈值之前。在目前的研究中，我们建议评估这些抗体在前瞻性研究中识别SN和SP症状/无症状HIV+TB+患者的潜力，并绘制印度患者中抗体识别的优势表位。为此，我们将招募一组约200名ART初治HIV+患者，并使用直接显微镜检查以及去污/浓缩痰涂片、固体培养基培养和痰、血液和尿液NAAT评价对他们进行结核病调查（目标1）;评价是否存在针对重组纯化MS、MPT 51、ESAT 6的Ab，招募个体血清标本中CFP 10和PPE 55，并将Ab的存在与TB的细菌学确认相关联（目的2）并在肽微阵列上鉴定由来自印度患者的血清Ab识别的5种抗原上的免疫显性表位，以定义可用于设计肽的肽，在印度艾滋病毒+患者中进行结核病快速检测。

TB diagnosis in India relies on clinical symptoms, microscopic examination of smears made directly from sputum and/or chest X-rays (CXR). The immune dysfunction caused by HIV-infection alters clinical and radiological presentation of TB, and microbiological confirmation of TB is problematic since HIV+TB+ patients have increased frequency of smear-negative (SN) TB and extrapulmonary TB (EPTB). Despite being totally treatable with the existing drugs, the poorer performance of insensitive diagnostic tests in HIV+ patients leads to significant under-diagnosis of TB in the very population where TB progression is rapid and often fatal. A simple, rapid test to identify smear-positive (SP) and SN symptomatic or asymptomatic TB in HIV+ patients would have significant impact on TB-related morbidity and mortality. Five immunogenic proteins of M. tuberculosis that are potential candidates for devising a simple diagnostic test for TB in HIV+ patients have been identified. These are MS, MPT51, PPE55, ESAT6 and CFP10. Antibodies (Abs) to these antigens have been demonstrated at all levels of immune-suppression, regardless of pulmonary manifestations and site of TB in HIV+TB+ patients, and are also detectable in sera of asymptomatic HIV+ patients who progressed to symptomatic TB in the subsequent 2-6 months. Absence of Abs to these antigens in retrospective sera from CD4 T cell-matched HIV+ patients who did not eventually develop TB suggests that in HIV+ patients, these Ab responses signal active TB whether or not clinical symptoms of TB are present and before the bacterial burden reaches the threshold of detection by direct microscopy. In the current studies we propose to evaluate the potential of these Abs to identify SN and SP symptomatic/asymptomatic HIV+TB+ patients in prospective studies and to map their dominant epitopes that are recognized by Abs in Indian patients. For this, we will recruit a cohort of ~200 ART-naive HIV+ patients and investigate them for TB using direct microscopy as well as with decontaminated/concentrated sputum smears, cultures on solid media and NAAT evaluation of sputum, blood and urine (Aim 1); evaluate the presence of Abs to recombinant purified MS, MPT51, ESAT6, CFP10 and PPE55 in serum specimens from the recruited individuals and correlate the presence of Abs with bacteriological confirmation of TB (Aim 2) and identify the immunodominant epitopes on the 5 antigens recognized by serum Abs from Indian patients on peptide microarrays to define peptides that can be used to devise a peptide-based rapid test for TB in Indian HIV+ patients.