A rapid, POC array MDR/XDR-TB genotyping.

快速 POC 阵列 MDR/XDR-TB 基因分型。

• 批准号: 7661599
• 负责人: Heather Koshinsky
• 金额: $ 28.31万
• 依托单位: INVESTIGEN, INC.
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-07-22 至 2010-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7661599
• 关键词: Accident and Emergency department; Alleles; Area; Binding; Biological Assay; Biological Products; Businesses; Cells; Characteristics; Clinic; Clinical; Color; Commit; Conserved Sequence; Country; DNA; Data Collection; Detection; Development; Diagnosis; Diagnostic Procedure; Diagnostic tests; Discrimination; Disease Management; Drug Resistant Tuberculosis; Drug resistance; Dyes; Elements; Evaluation; Extreme drug resistant tuberculosis; Genes; Genomics; Genotype; Goals; Government; Hour; Hybrids; Immigration; International; Lead; Life; Link; Methods; Microscopy; Mission; Multi-Drug Resistance; Multidrug-Resistant Tuberculosis; Mutation; Mutation Detection; Mycobacterium tuberculosis; Nucleic Acids; Patients; Peptide Nucleic Acids; Persons; Pharmaceutical Preparations; Poultry Diseases; Prevention; Protocols documentation; Public Health; Reaction; Research; Research Support; Resources; Sampling; Scheme; Sensitivity and Specificity; Solutions; Specificity; Sputum; Surface; System; Techniques; Technology; Temperature; Testing; Travel; Tuberculosis; United States; United States National Institutes of Health; United States Public Health Service; Visit; Visual; base; burden of illness; design; emergency service responder; genotyping technology; human disease; improved; innovation; interest; nucleic acid detection; point of care; point-of-care diagnostics; promoter; public health relevance; success; theories; transmission process

DESCRIPTION (provided by applicant): The diagnosis of tuberculosis (TB) is the major weak link in global TB control. Drug resistant TB compounds diagnosis and treatment challenges. The most widely used diagnostic test, sputum smear microscopy, is cumbersome, insensitive, involves multiple patient visits and cannot detect multi drug or extensively drug resistant (MDR/XDR-TB). Sputum culture is costly and depending on the method used takes 4-8 weeks to obtain a result and up to 16 weeks to detect MDR/XDR-TB. Consequently, there is an urgent need for a simple, rapid diagnostic test to identify the presence of Mycobacterium tuberculosis (MTB) and perform MDR/XDR genotyping that could be performed at the public health clinic while the patient waits. The long-term objective of this project is to develop a MTB identification and MDR/XDR-TB genotyping test that can be used at the point-of-care (POC). Investigen's "smartDNATM technology" enables DNA detection to occur rapidly at room temperature in homogenous solution with colorimetric readout. The smartDNA MTB identification assay consists of: (1) conventional sputum treatment, (2) sample disruption, (3) MTB target capture by a peptide nucleic acid (PNA)-microparticle, and (4) detection via a `light-activated' color change of a second (detection) PNA-DNA target bound dye. Investigen plans to extend the use of smartDNA-based detection to MDR/XDR- TB genotyping. The proposed protocol for MTB identification and MDR/XDR-TB genotyping involves (1) conventional sputum treatment, (2) sample disruption, (3) PNA-microparticle enrichment capture of MTB conserved sequences in each of the target areas of interest, (4) sequence specific binding of the enrichment captured targets on elements of an allele-specific PNA array and (5) detection of the resulting PNA-DNA hybrids using smartDNA. The successful development of the POC smartDNA MDR/XDR-TB genotyping technology requires that (1) probe sets for the capture, detection and allele discrimination of mutations involved in MDR/XDR-TB be designed, and (2) a practical way is found to apply the smartDNA dye color change reaction to PNA-DNA hybrids bound to a planar surface. There are three specific aims. (1) Demonstrate utility of the conceived system to differentiate single base changes in MTB genomic DNA. (2) Design probe sets for the detection of mutations. (3) Develop smartDNA readout on a PNA array. A smartDNA-based MDR/XDR- TB genotyping assay would greatly improve the state of MTB diagnosis and treatment, and accordingly, is relevant to the NIH mission of reducing the burden of illness through supporting research for the improved diagnosis, prevention and cure of human diseases. PUBLIC HEALTH RELEVANCE While the United States has been largely successful in detecting, treating and controlling TB, in many countries TB, MDR-TB and XDR-TB are a serious and growing public health issue; international travel and immigration have increased transmission. The research to be completed on this project is anticipated to lead to the development of a TB, MDR-TB and XDR-TB identification system that will provide results while the patient waits and can be used in resource constrained settings throughout the world, in clinics and emergency rooms in the United States and potentially even at US borders greatly aiding public health services in the detection, treatment and control of TB, MDR-TB and XDR-TB.

描述（申请人提供）：结核病（TB）的诊断是全球结核病控制的主要薄弱环节。耐药结核病化合物的诊断和治疗挑战。最广泛使用的诊断测试，痰涂片显微镜检查，是繁琐的，不敏感的，涉及多个病人的访问，不能检测多药或广泛耐药（MDR/XDR-TB）。痰液培养费用昂贵，根据所使用的方法，需要4-8周才能获得结果，最长需要16周才能检测出MDR/XDR-TB。因此，迫切需要一种简单、快速的诊断测试来鉴定结核分枝杆菌（MTB）的存在并进行MDR/XDR基因分型，其可以在患者等待的同时在公共卫生诊所进行。本项目的长期目标是开发一种可在床旁（POC）使用的MTB鉴定和MDR/XDR-TB基因分型检测。Investigen的“smartDNATM技术”使DNA检测能够在室温下在均质溶液中快速进行，并具有比色读数。smartDNA MTB鉴定试验包括：（1）常规痰液处理，（2）样品破碎，（3）通过肽核酸（PNA）-微粒捕获MTB靶标，和（4）通过第二（检测）PNA-DNA靶标结合染料的“光激活”颜色变化进行检测。Investigen计划将基于smartDNA的检测扩展到MDR/XDR-TB基因分型。MTB鉴定和MDR/XDR-TB基因分型的拟定方案包括（1）常规痰液处理，（2）样本破碎，（3）在每个目标靶区域中富集捕获MTB保守序列的PNA微粒，（4）富集捕获的靶在等位基因特异性PNA阵列元件上的序列特异性结合，以及（5）使用smartDNA检测所得PNA-DNA杂交体。POC smartDNA MDR/XDR-TB基因分型技术的成功开发需要（1）设计用于MDR/XDR-TB中涉及的突变的捕获、检测和等位基因鉴别的探针组，以及（2）找到将smartDNA染料变色反应应用于结合到平面表面的PNA-DNA杂交体的实用方法。有三个具体目标。(1)证明设想的系统区分MTB基因组DNA中的单碱基变化的实用性。(2)设计用于检测突变的探针组。(3)在PNA阵列上开发smartDNA读数。基于smartDNA的MDR/XDR-TB基因分型测定将极大地改善MTB诊断和治疗的状态，因此，与NIH的使命相关，即通过支持用于改善人类疾病的诊断、预防和治愈的研究来减轻疾病负担。公共卫生相关性虽然美国在检测、治疗和控制结核病方面取得了很大成功，但在许多国家，结核病、耐多药结核病和广泛耐药结核病是一个严重且日益严重的公共卫生问题;国际旅行和移民增加了传播。该项目即将完成的研究预计将导致结核病、耐多药结核病和广泛耐药结核病识别系统的开发，该系统将在患者等待时提供结果，并可用于世界各地资源有限的环境、美国的诊所和急诊室，甚至可能用于美国边境，极大地帮助公共卫生服务部门检测、治疗和控制结核病，耐多药结核和广泛耐药结核。