Diagnostic Innovations for Pediatric Tuberculosis in Bolivia

儿童结核病（TB）继续在低收入和中等收入国家构成诊断挑战， 结核病的高发病率，由于少杆菌病在儿童中的广泛影响，以及目前的结核病 培养和聚合酶链反应测试由于成本、有限的可获得性以及 从年幼儿童获得的样本的灵敏度较差。我们的专家团队来自杜兰，约翰 霍普金斯大学、秘鲁卡耶塔诺·埃雷迪亚大学和贝内菲卡·普利斯马协会 通过在秘鲁和玻利维亚超过25年的合作，应对所有这些挑战。我们的目标是直接 通过评估MPI Tony Hu开发的新诊断方法，应对儿童结核病的挑战 在杜兰大学使用CRISPR介导的TB检测（CRISPR-TB）优化检测循环 结核分枝杆菌无细胞DNA（Mtb-cfDNA），并用于分析飞行员冻存血清 对成人和儿童疑似结核病患者、无症状家庭接触者以及一组 有症状的艾滋病毒感染儿童（CLHIV）结核病高危人群。有症状成人队列的结果 合并敏感性为93%，特异性为93%，阳性预测值为95%，阴性预测值为95%。 价值92%。在CLHIV的有限试点研究中，CRISPR-TBD结果准确识别了所有确诊的结核病 (13大多数未确诊结核病的儿童（80%; 52/65）。我们建议招募200名疑似结核病患者 在2个研究群体（试验群体和 验证人群），通过与“Dr. Mario Ortiz苏亚雷斯”儿童医院相关的诊所确定 在玻利维亚的圣克鲁斯。我们将在“测试”中确定cfDNA浓度在外周血中的分布。 由两个年龄组的儿童（2个月-6岁，7-14岁）组成的“人群”， 根据NIH共识病例定义（确诊结核病、未确诊结核病），按结核病可能性对疾病进行分组 TB和不太可能的TB）和年龄匹配的对照组，按潜伏性TB感染（LTBI）的存在分组， 在接受抗生素治疗的TB病例中及时连续测量cfDNA。我们还将验证标准 定量cfDNA的范围建立了结核病或LTBI儿童的临床亚组， 独立验证队列。另一个目的将确定定量cfDNA和定量cfDNA之间的相关性。 基于肺（结核病的主要靶器官）疾病证据的定量成像结核评分 疾病，通过（1）胸片，使用CAD 4 TB v7系统通过计算机辅助分析测量， (2)肺部超声，使用便携式/低成本探头进行，由机器学习算法辅助， 自动解释这些生物标志物将作为潜在的辅因子进行测试， 外周血cfDNA水平，以提高儿童结核病的检测。本研究结果 这将是找到一条检测许多“未确诊”结核病病例的途径的第一步， 理想情况下，在资源匮乏的环境中诊断儿科结核病是非常必要的。

Pediatric tuberculosis (TB) continues to pose diagnostic challenges in low- and middle-income countries with high rates of TB disease, due to the well-described impact of paucibacillary disease in children, and current TB culture and polymerase-chain reaction tests are of limited usefulness due to cost, restricted availability, and poor sensitivity in specimens available from younger children. Our team of experts from Tulane, Johns Hopkins University, Universidad Peruana Cayetano Heredia, and Asociación Benéfica Prisma have confronted all of these challenges through more than 25 years of collaboration in Peru and Bolivia. Our goal is to directly address the challenges of TB in children by evaluating a new diagnostic approach developed by MPI Tony Hu at Tulane University using a CRISPR-mediated TB assay (CRISPR-TB) optimized to detect circulating Mycobacterium tuberculosis cell-free DNA (Mtb-cfDNA), and used to analyze cryopreserved serum in pilot studies from adults and children with presumptive TB, their asymptomatic household contacts, and a cohort of symptomatic children living with HIV (CLHIV) at high risk for TB. Results from symptomatic adult cohorts yielded a pooled sensitivity of 93%; specificity of 93%; positive predictive value of 95%; and negative predictive value of 92%. In limited pilot studies in CLHIV CRISPR-TBD results accurately identified all confirmed TB (13/13) and most children with unconfirmed TB (80%; 52/65). We propose to enroll 200 presumptive TB cases and an equal number of well control subjects in each of 2 study populations (test population and validation population) identified through clinics associated with the “Dr. Mario Ortiz Suarez” Children's Hospital in Santa Cruz, Bolivia. We will determine the distribution of cfDNA concentrations in peripheral blood in a “test population” composed of two age-based groups of children (2 months-6 years, 7-14 years) with respiratory disease grouped by likelihood of TB based on the NIH consensus case definitions (confirmed TB, unconfirmed TB, and unlikely TB) and in age-matched controls grouped by presence of latent TB infection (LTBI), with cfDNA measured serially in time among TB cases receiving antibiotic therapy. We will also validate standard ranges of quantitative cfDNA established for clinical subgroups of children with TB disease or LTBI in an independent validation cohort. An additional aim will determine the correlation between quantitative cfDNA and quantitative imaging-based TB scores based on evidence of disease in the lung, the primary target organ in TB disease, by (1) chest radiograph, measured by computer-aided analysis using the CAD4TB v7 system, and by (2) lung ultrasound, performed with a portable/low-cost probe assisted by machine learning algorithms for automatic interpretation. These biomarkers will be tested as potential cofactors that may be combined with cfDNA levels in peripheral blood, to improve the detection of TB disease in children. The results of this study will be the first step in a process to find a path to allow detection of the many “unconfirmed” TB cases and ideally make the diagnosis of pediatric TB in reach for low resource settings where it is so critically needed.