Dual-gene detection in a single-tube system based on CRISPR-Cas12a/Cas13a for severe fever thrombocytopenia syndrome virus

被引:11
作者
Zhu, Yating [1 ,2 ]
Xing, Chen [1 ]
Yang, Li [1 ]
Li, Qian [1 ,2 ]
Wang, Xiaofeng [1 ,2 ]
Zhou, Jing [3 ]
Zhang, Cong [1 ]
Ren, Cuiping [1 ,2 ]
Liu, Fahu [4 ]
He, Jun [5 ]
Shen, Bing [1 ]
Du, Yinan [1 ,2 ]
Liu, Yan [1 ]
机构
[1] Anhui Med Univ, Sch Basic Med Sci, Hefei, Peoples R China
[2] Anhui Med Univ, Dept Microbiol & Parasitol, Anhui Prov Lab Microbiol & Parasitol, Hefei, Peoples R China
[3] Anhui Med Univ, Affiliated Hosp 4, Dept Clin Lab, Hefei, Peoples R China
[4] Wannan Med Coll, Affiliated Yijishan Hosp, Dept Clin Lab, Wuhu, Peoples R China
[5] Anhui Ctr Dis Control & Prevent, Microbiol Lab, Hefei, Peoples R China
基金
中国国家自然科学基金;
关键词
SFTSV; CRISPR; Cas12a; Cas13a; multiplex detection; point-of-care testing; BUNYAVIRUS; TRANSMISSION; INFECTION; CLUSTER;
D O I
10.3389/fmicb.2022.977382
中图分类号
Q93 [微生物学];
学科分类号
071005 ; 100705 ;
摘要
Severe fever with thrombocytopenia syndrome (SFTS) is an emerging infectious disease, which is caused by severe fever with thrombocytopenia syndrome virus (SFTSV). The disease results in high mortality and increased morbidity and threatens global public health. Rapid detection of SFTSV is crucial for epidemic prevention in low-resource settings. Here we developed deployable, sensitive and rapid detection methods based on CRISPR/Cas12a or Cas13a technologies. The CRISPR/Cas12a-based detection assay could stably detect the SFTSV L or M genes at 10 cp/mu l. The Cas13a-based method could detect the L gene as low as 0.75 cp/mu l. For point-of-care testing, we combined fluorescence visualization and lateral flow detection with CRISPR/Cas-based assays. Furthermore, using the orthogonal DNA/RNA collateral activity of the Cas12a/Cas13a system, we present the dual-gene detection platform for SFTSV, which can simultaneously detect the L and M genes in a single tube. Based on the dual-gene detection, we designed multiplexed test strips to detect SFTSV. All our methods were initially validated using 52 clinical samples, showing 100% sensitivity and specificity. These new CRISPR/Cas-based detection methods are promising candidates for on-site detection of SFTSV.
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页数:12
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