A highly multiplexed convection flow qPCR assay and platform delivering rapid, comprehensive identification and predictive AST of pathogens causing UTIs.

项目摘要 尿路感染是最常见的社区和医院获得性感染之一。 (每年就诊约1000万人次)，占相当大的发病率和美国每年的医疗支出 超过60亿美元。虽然尿路感染的病原体很好理解，但患者经常得到经验性的治疗。 缺乏能够识别尿路感染病原体的快速、护理点分子诊断方法导致 抗菌药物管理和高UTI耐药率。 我们建议开始开发一种快速诊断分析和工作流程，以同时检测 所有UTI病原体及其相关耐药基因，适用于临床点临床应用。 这项测试预计将允许更有效的个体化治疗尿路感染，降低 抗菌素耐药性，并改善患者预后。 正如即将发表的一篇自然生物医学工程论文中所描述的那样，Torus生物系统的qPCR系统， Synstia™，已经证明了可重复性的30-plex能力，准确的定量和单核苷酸多态性检测， 灵敏度降至10个基因组拷贝，周转时间不到30分钟。消耗品是 配备嵌入式预淬式微阵列，允许实时检测扩增 无需打开反应室即可生产产品。Synstia系统的灵敏度和动态范围 已经在一系列DNA输入浓度(101到105个拷贝)和阈值范围内进行了评估 值被发现依赖于输入DNA浓度的对数。该平台还拥有 使用针对15种细菌的多重面板成功地进行了快速细菌鉴定 物种。 此第一阶段应用程序建议开发一个60多路UTI面板，用于全面评估 在Synstia平台上30分钟内检测到尿路病原体DNA。在随后的第二阶段，我们将开发， 优化和验证完全集成的消耗品，包括尿样准备(裂解和提取) Synestia平台，并与选定的临床站点协作验证临床性能。该队 预期所展示的化验结果将构成进一步开发 尿路感染的商业护理点诊断测试。

Project Summary Urinary tract infections (UTIs) are among the most prevalent community- and hospital-acquired infections (~10M annual medical visits), accounting for considerable morbidity and annual US healthcare spend of greater than $6B. While UTI pathogens are well understood, patients are frequently treated empirically. The lack of rapid, point of care molecular diagnostics that can identify UTI pathogens results in poor antimicrobial stewardship and high rates of UTI drug resistance. We propose to begin development of a rapid diagnostic assay and workflow that will concurrently detect all UTI pathogens and their associated resistance genes, suitable for point of care clinical applications. This assay is anticipated to allow for more effective personalized treatment of UTIs, reducing rates of antimicrobial resistance, and improving patient outcomes. As described in an upcoming Nature Biomedical Engineering paper, the Torus Biosystems’s qPCR system, Synestia™, has demonstrated reproducible 30-plex capability, accurate quantitation and SNP detection, sensitivity down to 10 genomic copies, and turnaround times of less than 30 minutes. The consumable is equipped with an embedded pre-quenched microarray that allows for real-time detection of amplification products without opening the reaction chamber. The sensitivity and dynamic range of the Synestia system has been evaluated across a range of input concentrations of DNA (101 to 105 copies) and the threshold value was found to be dependent on the log of the input DNA concentration. The platform has also successfully demonstrated rapid bacteria identification using a multiplex panel specific for 15 bacteria species. This Phase 1 application proposes to develop a 60-plex UTI panel for comprehensive assessment of uropathogen DNA within 30 minutes on the Synestia platform. In the subsequent Phase 2, we will develop, optimize, and validate a fully integrated consumable including urine sample prep (lysis and extraction) on the Synestia platform and validate clinical performance in collaboration with select clinical sites. The team anticipates that the demonstrated assay will constitute a solid basis for further development of a commercial point of care diagnostic test for urinary tract infections.