基于微流控LAMP技术的血液中假丝酵母菌和曲霉菌同步快速检测

Invasive fungal infections are associated with high morbidity and mortality in critically ill patients due, in part, to diagnostic difficulties in the early stages. Up to now, there is no optimal assay to detect invasive fungi such as candida and aspergillus simultaneously in blood. New techniques are required for invasive mycoses earlier diagnosis, prognostic information and monitoring outcome. Based on the microfluidic chip for mycobacteria identification which developed in our completed national mega research project, in this study, we focus on the target molecules including C.albicans , C. glabrata,C. krusei,C. tropicalis，C. parapsilosisand A. fumigatus，A. flavus,A. niger. LAMP primers that bind to specific target molecules with high specificity and affinity selected based on ITS, rDNA, IGS fungi conserved DNA fragments. Combined with pathogenic microorganism enrichment microfluidic chip and multi-channel detect microfluidic chip to rapid detect different target pathogens in blood at one assay.And taking LAMP microfluidic chip as the core, develop the automated microfluidic systems integration of sample priming, analysis, report, for a new rapid diagnostic technology of fungemia. The LAMP-based compact microfluidic system for diagnosis may even lead to a point-of-care device. The use of LAMP-Microfluidic systems screening and diagnosis invasive fungal infection is expected to continue growing in the near future and may make a substantial impact on biomedical applications.

侵袭性真菌感染的准确、早期诊断是治疗疾病和抢救生命的关键。作为重要的侵袭性真菌感染病原体,血液中假丝酵母和曲霉菌的同步快速检测具有非常重要的意义。然而, 目前真菌的诊断技术远不能满足临床需求。本研究拟在已完成的国家十一五重大专项课题建立的病原细菌微流体芯片技术基础上，以血流感染常见的5种假丝酵母菌和3种曲霉菌为研究对象，采用微流控LAMP技术，以真菌ITS、rDNA、IGS基因保守区为靶标筛选菌种特异性LAMP引物， 结合芯片病原微生物富集技术和多通道芯片同步检测技术，实现快速直接检测血液中假丝酵母菌和曲霉菌的种特异性分析，进而以LAMP微流控芯片为核心设计制作进样、分析、报告一体化检测系统，发展真菌血症的快速诊断新技术，为建立有效的侵袭性真菌感染早期预警、早期诊断与治疗监测技术平台奠定基础。

侵袭性真菌感染的准确、早期诊断是治疗疾病和抢救生命的关键。快速、多重检测侵袭性真菌感染病原体核酸具有非常重要的意义。本研究采用微流控LAMP技术，以真菌ITS、rDNA、IGS基因保守区为靶标筛选菌种特异性LAMP引物， 结合病原微生物富集核酸抽提技术和多通道芯片同步检测技术，实现快速直接检测血液中白念珠菌、热带念珠菌、近平滑念珠菌、光滑念珠菌和克柔念珠菌的种特异性分析，进而以LAMP微流控芯片为核心设计制作进样、分析、报告一体化检测系统，建立了真菌感染的多重快速诊断新技术。本项目申请专利5项，1项体外诊断试剂获批注册证，研发出1台便携式多重LAMP检测设备。为建立有效的侵袭性真菌感染早期预警、早期诊断与治疗监测技术平台奠定基础。