Argonaute-integrated isothermal amplification for rapid, portable, multiplex detection of SARS-CoV-2 and influenza viruses

Xingyu Ye; Haiwei Zhou; Xiang Guo; Donglai Liu; Zhonglei Li; Junwei Sun; Jun Huang; Tao Liu; Pengshu Zhao; Heshan Xu; Kai Li; Hanming Wang; Jihua Wang; Li Wang; Weili Zhao; Qian Liu; Sihong Xu; Yan Feng

doi:10.1016/j.bios.2022.114169

Argonaute-integrated isothermal amplification for rapid, portable, multiplex detection of SARS-CoV-2 and influenza viruses

Biosens Bioelectron. 2022 Jul 1:207:114169. doi: 10.1016/j.bios.2022.114169. Epub 2022 Mar 18.

Authors

Xingyu Ye¹, Haiwei Zhou², Xiang Guo¹, Donglai Liu², Zhonglei Li¹, Junwei Sun¹, Jun Huang³, Tao Liu¹, Pengshu Zhao¹, Heshan Xu¹, Kai Li⁴, Hanming Wang⁵, Jihua Wang⁵, Li Wang⁶, Weili Zhao⁶, Qian Liu⁷, Sihong Xu⁸, Yan Feng⁹

Affiliations

¹ State Key Laboratory of Microbial Metabolism, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai, 200240, China.
² National Institutes for Food and Drug Control, Beijing, 100050, China.
³ School of Biological and Chemical Engineering, Zhejiang University of Science and Technology, Hangzhou, 310023, China.
⁴ GeneTalks Biotechnology Inc., Changsha, 410013, China.
⁵ Guangdong Medical Laboratory Technology (Rapid Diagnostic) Engineering Technology Research Center, Guangzhou, 510641, China; Guangzhou Wondfo Biotech Co., Ltd., Guangzhou, 510641, China.
⁶ Shanghai Institute of Hematology, State Key Laboratory of Medical Genomics, National Research Center for Translational Medicine at Shanghai, Ruijin Hospital, Shanghai Jiao Tong University of Medicine, Shanghai, 200025, China.
⁷ State Key Laboratory of Microbial Metabolism, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai, 200240, China. Electronic address: liuqian1018@sjtu.edu.cn.
⁸ National Institutes for Food and Drug Control, Beijing, 100050, China. Electronic address: xushong@nifdc.org.cn.
⁹ State Key Laboratory of Microbial Metabolism, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai, 200240, China. Electronic address: yfeng2009@sjtu.edu.cn.

Abstract

Isothermal amplification methods are a promising trend in virus detection because of their superiority in rapidity and sensitivity. However, the generation of false positives and limited multiplexity are major bottlenecks that must be addressed. In this study, we developed a multiplex Argonaute (Ago)-based nucleic acid detection system (MULAN) that integrates rapid isothermal amplification with the multiplex inclusiveness of a single Ago for simultaneous detection of multiple targets such as SARS-CoV-2 and influenza viruses. Owing to its high specificity, MULAN can distinguish targets at a single-base resolution for mutant genotyping. Moreover, MULAN also supports portable and visible devices with a limit of detection of five copies per reaction. Validated by SARS-CoV-2 pseudoviruses and clinical samples of influenza viruses, MULAN showed 100% agreement with quantitative reverse-transcription PCR. These results demonstrated that MULAN has great potential to facilitate reliable, easy, and quick point-of-care diagnosis for promoting the control of infectious diseases.

Keywords: Argonaute; Isothermal amplification; Multiplex detection; Point-of-care diagnosis; SARS-CoV-2 detection.

MeSH terms

Biosensing Techniques*
COVID-19* / diagnosis
Humans
Molecular Diagnostic Techniques / methods
Nucleic Acid Amplification Techniques / methods
Orthomyxoviridae* / genetics
RNA, Viral / analysis
RNA, Viral / genetics
SARS-CoV-2 / genetics
Sensitivity and Specificity

Substances

RNA, Viral