Rapid ID and AST Directly from Whole Blood Using Single Molecule Detection

使用单分子检测直接从全血中快速进行 ID 和 AST

• 批准号: 10632827
• 负责人: Alfredo Andres Celedon
• 金额: $ 86.68万
• 依托单位: SCANOGEN, INC.
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-01-30 至 2027-12-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10632827
• 关键词: Acinetobacter baumannii; Agreement; Antibiotics; Antimicrobial Resistance; Antimicrobial susceptibility; Bacteria; Biological Assay; Blood; Blood Circulation; Blood specimen; Categories; Clinical; Detection; Development; Diagnosis; Diagnostic; Enterobacter; Escherichia coli; Goals; Hour; Incubated; Klebsiella pneumoniae; Liquid substance; Methods; Organism; Patients; Performance; Pharmaceutical Preparations; Phenotype; Physicians; Predisposition; Process; Protocols documentation; Pseudomonas aeruginosa; Rapid diagnostics; Reagent; Reference Standards; Resistance; Ribosomal RNA; Sampling; Sepsis; Specificity; Staphylococcus aureus; System; Techniques; Testing; Time; Whole Blood; antimicrobial; cost effective; detection limit; detection method; diagnostic platform; effective therapy; instrument; microorganism; multiplex assay; novel; operation; pathogen; prototype; rapid test; single molecule; targeted treatment

Project Summary Standard identification (ID) and antimicrobial susceptibility testing (AST) of bloodstream pathogens are conducted using slow methods that require blood culture. Diagnostic results are available three or more days after the diagnostic process is started. The lack of rapid diagnostic results forces physicians to treat patients with broad-spectrum antibiotics which are not effective against a growing number of resistant organisms and can induce the development of more antimicrobial resistance. Here, we propose the development of a rapid diagnostic platform that, if successful, will enable effective and targeted therapy of bloodstream infection patients within hours. The new platform will directly analyze whole blood samples using a simple and robust single molecule detection approach called Single MOlecule Tethering or SMOLT. The platform will be rapid (ID in one hour and AST in 4- 6 hours), easy-to-use, and cost-effective. Our preliminary results show that a SMOLT multiplex assay can detect bacteria in blood with limits of detection (LOD) between 1-10 CFU/mL with a turn-around-time of about an hour. The same technique can be used for AST. Preliminary results show that a brief incubation with an antibiotic follow by SMOLT detection can be used to determine if a strain is susceptible, intermediate or resistance in high agreement with the category assigned be a standard reference method. In aim 1, we propose the development of a SMOLT multiplex ID assay capable of detecting and identifying the rRNA of six highly relevant pathogens directly in whole blood. The goal of aim 2 is to develop a SMOLT phenotypic AST assay for combinations of the six ID species and 14 relevant drugs. Our third aim is to develop a prototype instrument for the SMOLT ID and AST assays that is easy-to-use with minimum hands-on-time. The last aim is to evaluate the ID and AST assays using blood samples seeded with a combination of fresh and stock clinical isolates as well as type strains.

项目概要 血流病原体的标准鉴定 (ID) 和抗菌药物敏感性测试 (AST) 使用需要血培养的缓慢方法进行。诊断结果三天或更长时间即可获得 诊断过程开始后。缺乏快速诊断结果迫使医生对患者进行治疗 广谱抗生素对越来越多的耐药微生物无效，并且可以 诱导更多抗菌药物耐药性的发展。在此，我们建议快速发展 诊断平台，如果成功，将为血流感染患者提供有效和有针对性的治疗 数小时内。 新平台将使用简单而强大的单分子检测直接分析全血样本 称为单分子束缚或 SMOLT 的方法。该平台将是快速的（一小时内完成 ID，四小时内完成 AST） 6小时），简单易用，性价比高。我们的初步结果表明，SMOLT 多重检测可以检测 血液中细菌的检测限 (LOD) 介于 1-10 CFU/mL 之间，周转时间约为一小时。 相同的技术可用于 AST。初步结果表明，与抗生素短暂孵育 随后进行 SMOLT 检测，可用于确定菌株是否为易感菌株、中度菌株或高耐药菌株。 与分配的类别一致是标准参考方法。在目标1中，我们建议发展 SMOLT 多重 ID 测定能够检测和识别六种高度相关病原体的 rRNA 直接在全血中。目标 2 的目标是开发 SMOLT 表型 AST 检测，用于组合 6个ID品种和14个相关药物。我们的第三个目标是为 SMOLT ID 开发原型仪器 AST 检测易于使用，操作时间最少。最后一个目标是评估 ID 和 AST 检测 使用混合了新鲜和库存临床分离株以及典型菌株的血液样本进行接种。