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Volume 40 Issue 4
August 2025

    . doi: 10.1016/j.virs.2025.06.006
    Citation: Zixiao Yang, Xinrong Zhou, Xikui Sun, Liu Cao, Tiefeng Xu, Kun Li, Hongchao Liu, Yanxi Ji, Lihong Liu, Konstantin I. Ivanov, Zhonghan Yang, Deyin Guo, Chun-Mei Li. Rapid and accurate detection of infectious SARS-CoV-2 by viral receptor capture combined with loop-mediated isothermal amplification .VIROLOGICA SINICA, 2025, 40(4) : 613-623.  http://dx.doi.org/10.1016/j.virs.2025.06.006
    shu

    基于病毒受体捕获结合环介导等温扩增法快速精准检测感染性SARS-CoV-2病毒

    • a 中山大学医学院, 感染与免疫中心, 深圳, 518107, 中国;

    • b 广州国家实验室, 广州国际生物岛, 广州, 510320, 中国;

    • c 中山大学中山医学院病原生物学与生物安全学系, 人类病毒学研究所, 热带病防治研究教育部重点实验室, 广州, 510080, 中国;

    • d 中山大学附属第三医院感染性疾病科, 广州, 510630, 中国;

    • e 广州医科大学附属第一医院, 广州呼吸健康研究院, 国家呼吸系统疾病临床医学研究中心, 呼吸疾病国家重点实验室, 广州, 510182, 中国;

    • f 罗蒙诺索夫·莫斯科国立大学, 生物工程与生物信息学系, 莫斯科 119234, 俄罗斯。

    • 快速、准确地检测出具有传染性的病毒颗粒，而不仅仅是病毒核酸，对于避免不必要的隔离以及有效控制诸如2019冠状病毒病（COVID-19）、严重急性呼吸综合征（SARS）和中东呼吸综合征（MERS）等病毒引起的疾病传播至关重要。实时荧光定量聚合酶链反应（RT-qPCR）是COVID-19疫情期间最广泛应用的主要检测技术。然而，它无法区分完整的传染性病毒与表面损伤的非传染性病毒颗粒或游离的病毒RNA。本研究提出了一种结合病毒受体捕获与逆转录环介导等温扩增（RT-LAMP）来特异性检测传染性冠状病毒的策略。我们成功地将该方法应用于检测SARS-CoV-2替代病毒和人冠状病毒NL63（HCoV-NL63）的传染性颗粒。病毒颗粒首先被包被了重组人血管紧张素转化酶2（hACE2）受体的ELISA板捕获。随后，从捕获的颗粒中获取病毒RNA，并使用病毒特异性引物通过RT-LAMP进行检测。在我们的实验条件下，该方法的检测限（LOD）最低可达90 PFU/mL，灵敏度为96.2%，特异性为100%。我们的研究提供了概念验证，即病毒受体捕获结合RT-LAMP能够区分传染性冠状病毒与非传染性病毒颗粒或游离病毒RNA。这为该病毒检测策略成为管理和防控流行性冠状病毒疾病的主流工具提供了基础。

    Rapid and accurate detection of infectious SARS-CoV-2 by viral receptor capture combined with loop-mediated isothermal amplification

    • a. Center for Infection and Immunity, School of Medicine, Shenzhen Campus of Sun Yat-sen University, Shenzhen, 518107, China;

    • b. Guangzhou National Laboratory, Guangzhou International Bio-Island, Guangzhou, 510320, China;

    • c. Institute of Human Virology, Department of Pathogen Biology and Biosecurity, and Key Laboratory of Tropical Disease Control of Ministry of Education, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, 510080, China;

    • d. Department of Infectious Diseases, Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, 510630, China;

    • e. State Key Laboratory of Respiratory Disease, National Clinical Research Center for Respiratory Disease, Guangzhou Institute of Respiratory Health, The First Affiliated Hospital of Guangzhou Medical University, Guangzhou, 510182, China;

    • f. Faculty of Bioengineering and Bioinformatics, Lomonosov Moscow State University, Moscow, 119234, Russia

    • Rapid and accurate detection of infectious virus particles, not just viral nucleic acid, is essential to avoid unnecessary quarantine and effectively control the spread of viral diseases such as coronavirus disease 2019 (COVID-19), severe acute respiratory syndrome (SARS), and Middle East respiratory syndrome (MERS). Real-time quantitative polymerase chain reaction (RT-qPCR) was the most widely used detection technique during the COVID-19 outbreak. However, it cannot discriminate between intact infectious viruses and surface-distorted, non-infectious virus particles or naked viral RNA. In this study, we present a strategy for the specific detection of infectious coronaviruses by combining viral receptor capture and reverse transcription loop-mediated isothermal amplification (RT-LAMP). We successfully applied this strategy to detect infectious virus particles of the SARS-CoV-2 surrogate virus and the human coronavirus NL63 (HCoV-NL63). Virus particles were first captured on ELISA plates coated with the recombinant human angiotensin-converting enzyme 2 (hACE2) receptor. Viral RNA was then extracted from the particles and detected by RT-LAMP using virus-specific primers. In our experimental setting, the proposed method had a minimum detection limit (LOD) of 90 PFU/mL, sensitivity of 96.2%, and specificity of 100%. Our study provides a proof-of-concept that viral receptor capture combined with RT-LAMP can differentiate infectious coronaviruses from non-infectious virions or naked viral RNA. This paves the way for this virus detection strategy to become a mainstream tool for the management, prevention and control of epidemic coronavirus diseases.
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      Rapid and accurate detection of infectious SARS-CoV-2 by viral receptor capture combined with loop-mediated isothermal amplification

        Corresponding author: Deyin Guo, guo_deyin@gzlab.ac.cn
        Corresponding author: Chun-Mei Li, lichm8@mail.sysu.edu.cn
      • a. Center for Infection and Immunity, School of Medicine, Shenzhen Campus of Sun Yat-sen University, Shenzhen, 518107, China;
      • b. Guangzhou National Laboratory, Guangzhou International Bio-Island, Guangzhou, 510320, China;
      • c. Institute of Human Virology, Department of Pathogen Biology and Biosecurity, and Key Laboratory of Tropical Disease Control of Ministry of Education, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, 510080, China;
      • d. Department of Infectious Diseases, Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, 510630, China;
      • e. State Key Laboratory of Respiratory Disease, National Clinical Research Center for Respiratory Disease, Guangzhou Institute of Respiratory Health, The First Affiliated Hospital of Guangzhou Medical University, Guangzhou, 510182, China;
      • f. Faculty of Bioengineering and Bioinformatics, Lomonosov Moscow State University, Moscow, 119234, Russia
      • Received Date: 28 March 2025
      • Accepted Date: 27 June 2025

      Abstract: Rapid and accurate detection of infectious virus particles, not just viral nucleic acid, is essential to avoid unnecessary quarantine and effectively control the spread of viral diseases such as coronavirus disease 2019 (COVID-19), severe acute respiratory syndrome (SARS), and Middle East respiratory syndrome (MERS). Real-time quantitative polymerase chain reaction (RT-qPCR) was the most widely used detection technique during the COVID-19 outbreak. However, it cannot discriminate between intact infectious viruses and surface-distorted, non-infectious virus particles or naked viral RNA. In this study, we present a strategy for the specific detection of infectious coronaviruses by combining viral receptor capture and reverse transcription loop-mediated isothermal amplification (RT-LAMP). We successfully applied this strategy to detect infectious virus particles of the SARS-CoV-2 surrogate virus and the human coronavirus NL63 (HCoV-NL63). Virus particles were first captured on ELISA plates coated with the recombinant human angiotensin-converting enzyme 2 (hACE2) receptor. Viral RNA was then extracted from the particles and detected by RT-LAMP using virus-specific primers. In our experimental setting, the proposed method had a minimum detection limit (LOD) of 90 PFU/mL, sensitivity of 96.2%, and specificity of 100%. Our study provides a proof-of-concept that viral receptor capture combined with RT-LAMP can differentiate infectious coronaviruses from non-infectious virions or naked viral RNA. This paves the way for this virus detection strategy to become a mainstream tool for the management, prevention and control of epidemic coronavirus diseases.

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