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.

项目摘要 血液病原体的标准识别（ID）和抗菌敏感性测试（AST）是 使用需要血液培养的慢方法进行。诊断结果可获得三天或更多天 诊断过程开始后。缺乏快速诊断结果迫使医生治疗患者 广谱抗生素对越来越多的抗性生物无效，并且可以 诱导更多抗菌耐药性的发展。在这里，我们提出了快速的发展 诊断平台，如果成功，将实现有效和有针对性的血液感染患者的治疗 几个小时内。 新平台将使用简单且可靠的单分子检测直接分析全血样品 方法称为单分子绑扎或Smolt。该平台将是快速的（ID在一小时内，AST为4-- 6小时），易于使用且具有成本效益。我们的初步结果表明，Smolt多重测定可以检测到 血液中的细菌在1-10 CFU/mL之间具有检测限制（LOD），转弯时间约为一个小时。 相同的技术可用于AST。初步结果表明，短暂与抗生素孵育 遵循Smolt检测可用于确定应变是易感性，中间还是高的抗性 与分配的类别达成协议是标准参考方法。在AIM 1中，我们提出了发展 能够检测和识别六种高度相关病原体的RRNA的Smolt多路复用ID测定法 直接在全血中。 AIM 2的目的是开发Smolt表型AST测定法以组合 六种ID物种和14种相关药物。我们的第三个目的是为Smolt ID开发原型仪器和 最少动手的AST测定法可以易于使用。最后的目的是评估ID和AST分析 使用与新鲜和库存临床分离株以及类型菌株结合的血液样本。