PDF(Pubmed)
Abstract：
There is a growing demand for rapid, sensitive, field-deployable nucleic acid tests for cholera, which usually occurs in rural areas. In this study, we developed a Cas12a-assisted rapid isothermal detection (CARID) system for the detection of toxigenic V. cholerae serogroups O1 and O139 by combining recombinase-aided amplification and CRISPR-Cas (clustered regularly interspaced short palindromic repeats and CRISPR-associated proteins). The results can be determined by fluorescence signal and visualized by lateral flow dipstick. We identified 154 V. cholerae strains and 129 strains of other intestinal diarrheagenic bacteria with a 100% coincidence rate. The limit of detection of CARID was 20 copies/reaction of V. cholerae genomic DNA, which is comparable to that of polymerase chain reaction (PCR) and qPCR. Multiple-CARID was also established for efficiency and economic considerations with an acceptable decrease in sensitivity. Simulated sample tests showed that CARID is suitable for complex samples. In conclusion, CARID is a rapid, sensitive, economically efficient, and portable method for the detection of V. cholerae, which makes it suitable for field responses to cholera.
摘要：
对快速，敏感,可现场部署的霍乱核酸检测，这通常发生在农村地区。在这项研究中,我们开发了一种Cas12a辅助的快速等温检测(CARID)系统,通过结合重组酶辅助扩增和CRISPR-Cas(成簇的规则间隔的短回文重复序列和CRISPR相关蛋白)来检测产毒霍乱弧菌血清群O1和O139.结果可以通过荧光信号确定，并通过侧流试纸可视化。我们鉴定出154株霍乱弧菌菌株和129株其他肠道致泻菌，符合率为100％。CARID的检测限为20拷贝/反应霍乱弧菌基因组DNA，与聚合酶链反应（PCR）和qPCR相当。出于效率和经济考虑,还建立了具有可接受的灵敏度降低的多重CARID。模拟样品测试表明CARID适用于复杂样品。总之,CARID是一种快速的，敏感,经济高效,和便携式霍乱弧菌检测方法，这使得它适合于对霍乱的野外反应。
