Lab-on-a-Film Multiplexed Test for Respiratory Mycobacterial Infections

呼吸道分枝杆菌感染的胶片实验室多重检测

• 批准号: 10689261
• 负责人: Christopher Gerard Cooney
• 金额: $ 99.94万
• 依托单位: AKONNI BIOSYSTEMS, INC.
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-04-21 至 2025-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10689261
• 关键词: Academy; Acid Fast Bacillae Staining Method; Address; Algorithms; American; Aminoglycoside resistance; Antibiotics; Antimicrobial Resistance; Antimicrobial susceptibility; Azithromycin; Bacillus; Bacteria; Biological Assay; Bronchiectasis; Cause of Death; Chronic; Chronic Obstructive Pulmonary Disease; Clarithromycin; Clinic; Clinical; Clinical Microbiology; Clinical Research; Collaborations; Combination Drug Therapy; Companions; Complex; Computer software; Consumption; DNA; DNA Sequence Alteration; Detection; Development; Diagnosis; Diagnostic; Diagnostic tests; Disease; Drug resistance; Elements; Erythromycin; Etiology; Film; Freeze Drying; General Population; Genes; Genetic Polymorphism; Genomics; Genus Mycobacterium; Glass; Goals; Health; Incidence; Individual; Infection; Journals; Laboratories; Laboratory Diagnosis; Laboratory Study; Life; Liquid substance; Low Prevalence; Lung diseases; Lung infections; Macrolide-resistance; Magnetism; Methodology; Methyltransferase; Microbe; Microbial Biofilms; Microbiology; Microfluidics; Molecular; Molecular Diagnostic Testing; Mycobacterium Infections; Mycobacterium abscessus; Mycobacterium avium Complex; Mycobacterium tuberculosis complex; Nucleic Acids; Organism; Patients; Performance; Pharmaceutical Preparations; Phase; Phenotype; Point Mutation; Polymorphism Analysis; Porosity; Positioning Attribute; Prevalence; Process; Protocols documentation; Pulmonary Emphysema; Pulmonary Tuberculosis; Reagent; Regimen; Relapse; Resistance; Resistance to infection; Respiratory Tract Infections; Ribosomes; Risk Factors; Robotics; Rotation; Sampling; Sensitivity and Specificity; Soil; Specificity; Specimen; Sputum; Step Tests; Testing; Time; Tuberculosis; Veterans; Viscosity; Water; automated analysis; clinical diagnosis; clinically relevant; diagnostic algorithm; drug testing; editorial; follower of religion Jewish; imager; instrument; member; mycobacterial; new epidemic; non-tuberculosis mycobacteria; novel therapeutics; optimal treatments; portability; rRNA Genes; radiological imaging; rapid test; respiratory; test strip; tuberculosis treatment; virtual; working group

ABSTRACT Nontuberculous mycobacteria lung disease (NTM-LD) is a silent and emerging epidemic in the U.S. and many parts of the world.1-3 The incidence and prevalence of NTM-LD is increasing yearly and now far exceeds that of tuberculosis (TB) caused by Mycobacterium tuberculosis complex in the U.S.4 Two of the greatest known risk factors for NTM-LD are chronic obstructive pulmonary disease (COPD) and pre-existing bronchiectasis.3 In the U.S., approximately 12 million individuals have COPD, the third leading cause of death in the U.S.5 In addition, the co-occurrence of bronchiectasis in patients with known COPD is up to ~70%.6 Like TB, the requirement for prolonged combination drug therapy is a central tenet of NTM-LD treatment. Consequently, it is essential that several drugs be administered concurrently to maximize sterilizing activity. While TB has benefited from the development of rapid molecular diagnostic tests to simultaneously detect infection and antimicrobial resistance and from recently approved new drugs, the diagnosis and treatment of NTM-LD have not experienced similar advances.7,8 The diagnosis of NTM-LD is complicated by the fact that clinical manifestations and radiographic findings for pulmonary TB and NTM-LD may be virtually indistinguishable. Thus, it is important when diagnosing NTM-LD to “rule out” TB even in regions of lower prevalence of the disease, such as the U.S., because treatment for TB and NTM-LD are substantially different. To differentiate TB from NTM-LD, clinicians must rely on a combination of phenotypic assays and molecular tests to identify the etiological agent and to detect resistance to key antibiotics. Hence, the algorithm for contemporary NTM diagnostic testing is complex, requiring varied testing methodologies, which are either insensitive (acid-fast bacilli smear), inherently slow to obtain the results (culture; up to 6 weeks), or insufficiently comprehensive (lack of molecular tests). Molecular detection of NTM and its antimicrobial resistance from respiratory samples is challenging. The specimen type (sputum) is viscous and highly heterogeneous; bacterial burden is often low but significant; mycobacteria are difficult to lyse; the number of clinically relevant NTM species is considerable; and the polymorphisms that confer drug resistance are numerous. To address these challenges, we propose to automate and integrate the following into a one user-step test: chaotic mixing of glass beads using a rotating magnetic disc to homogenize sputa and lyse bacilli, a porous disc in a pipette tip to purify and concentrate nucleic acid, and a Lab-on-a-Film test to speciate and detect polymorphisms that confer drug resistance. For Phase 2, we propose to develop a test that can rule in/out TB, speciate clinically-relevant NTM, and detect NTM-LD drug resistance markers. To evaluate this test, we propose to perform clinical studies at Mayo Clinic, National Jewish Health, and Wadsworth Center in collaboration with clinical NTM-LD experts, which includes members of: the Journal of Clinical Microbiology Editorial Board, Clinical Laboratory Standards Institute Working Group, fellows of the American Academy of Microbiology and Board of Governors of the Academy.

抽象的 非结核分枝杆菌肺病 (NTM-LD) 在美国和许多国家是一种悄然出现的流行病 1-3 NTM-LD 的发病率和患病率逐年增加，目前已远远超过 在美国，由结核分枝杆菌复合体引起的结核病 (TB) 4 两种最大的已知风险 NTM-LD 的因素包括慢性阻塞性肺病 (COPD) 和既往存在的支气管扩张症。 3 在美国，大约有 1200 万人患有慢性阻塞性肺病 (COPD)，这是美国第三大死因5 此外， 已知慢性阻塞性肺病 (COPD) 患者中同时发生支气管扩张的比例高达 70%。6 与结核病一样，需要 长期联合药物治疗是 NTM-LD 治疗的核心原则。因此，至关重要的是 同时施用多种药物以最大限度地发挥灭菌活性。虽然结核病已受益于 开发快速分子诊断测试以同时检测感染和抗菌药物耐药性 而且从最近批准的新药来看，NTM-LD的诊断和治疗还没有经历过类似的情况。 7,8 NTM-LD 的诊断因临床表现和放射学检查而变得复杂 肺结核和 NTM-LD 的检查结果实际上可能无法区分。因此，重要的是当 即使在美国等疾病流行率较低的地区，也能诊断 NTM-LD 以“排除”结核病， 因为 TB 和 NTM-LD 的治疗有很大不同。为了区分 TB 和 NTM-LD，临床医生 必须依靠表型测定和分子测试的结合来识别病原体并 检测对关键抗生素的耐药性。因此，当代 NTM 诊断测试的算法很复杂， 需要不同的测试方法，这些方法要么不敏感（抗酸杆菌涂片），要么本质上缓慢 获得结果（培养；最长 6 周），或不够全面（缺乏分子测试）。 从呼吸道样本中检测 NTM 及其抗菌药物耐药性具有挑战性。这 标本类型（痰）粘稠且高度不均匀；细菌负荷通常较低但很显着； 分枝杆菌难以裂解；临床相关的 NTM 物种数量相当可观；和 赋予耐药性的多态性有很多。为了应对这些挑战，我们建议 将以下内容自动化并集成到一个用户步骤测试中： 使用旋转器对玻璃珠进行混乱混合 用于均化痰液和裂解杆菌的磁盘，移液器吸头中的多孔盘用于纯化和浓缩 核酸，以及薄膜实验室测试，以形成和检测赋予耐药性的多态性。为了 第二阶段，我们建议开发一种测试，可以排除/排除结核病、区分临床相关的 NTM 并检测 NTM-LD耐药标记物。为了评估这项测试，我们建议在梅奥诊所进行临床研究， 国家犹太健康中心和沃兹沃斯中心与临床 NTM-LD 专家合作，其中包括 成员：《临床微生物学杂志》编辑委员会、临床实验室标准协会 工作组、美国微生物学会研究员和该学会理事会。