Rapid visual detection of Monkeypox virus by one-step LAMP-CRISPR/Cas12b assay
Abstract Background Monkeypox virus (MPXV) infection has garnered significant global attention due to its rising incidence and substantial public health implications. A rapid, sensitive, and accurate diagnostic method is urgently required to enable early intervention and effective management of MPXV...
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BMC
2025-05-01
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| Online Access: | https://doi.org/10.1186/s12985-025-02780-0 |
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| author | Jinlei Guo Yunfeng Shan Ge Hu Xiaojuan Zhu Kangchen Zhao Tao Wu Qiao Qiao Ying Chi Lunbiao Cui Yiyue Ge |
| author_facet | Jinlei Guo Yunfeng Shan Ge Hu Xiaojuan Zhu Kangchen Zhao Tao Wu Qiao Qiao Ying Chi Lunbiao Cui Yiyue Ge |
| author_sort | Jinlei Guo |
| collection | DOAJ |
| description | Abstract Background Monkeypox virus (MPXV) infection has garnered significant global attention due to its rising incidence and substantial public health implications. A rapid, sensitive, and accurate diagnostic method is urgently required to enable early intervention and effective management of MPXV outbreaks. Results In this study, we developed a novel one-step assay that integrates loop-mediated isothermal amplification (LAMP) with CRISPR/Cas12b in one-pot for the detection of MPXV. The entire detection process did not require opening the lid of the reaction tube and could be completed within 40 min using extracted viral nucleic acids, which is faster than real-time quantitative PCR (qPCR). And the results could be interpreted through either real-time fluorescence or naked-eye visualization. The limit of detection (LOD) of the assay was demonstrated to be 6.5 copies per reaction and no cross-reactivity with other pathogens such as HSV, EBV, CVA16, EV-A71, and MV was found. Furthermore, when evaluated using 113 clinical samples, the assay achieved 100% sensitivity (71/71) and 100% specificity (42/42) compared to the qPCR. Conclusions In resource-limited settings, our method requires only a portable heat block or water bath and a blue light or ultraviolet flashlight for visual detection of MPXV, making it highly accessible. The integration of LAMP and CRISPR/Cas12b provides a robust, user-friendly platform for point-of-care testing, with promising potential for the rapid molecular diagnosis of infectious diseases. |
| format | Article |
| id | doaj-art-52a4b68c44e742b2a2d313895ab90442 |
| institution | DOAJ |
| issn | 1743-422X |
| language | English |
| publishDate | 2025-05-01 |
| publisher | BMC |
| record_format | Article |
| series | Virology Journal |
| spelling | doaj-art-52a4b68c44e742b2a2d313895ab904422025-08-20T03:08:40ZengBMCVirology Journal1743-422X2025-05-0122111010.1186/s12985-025-02780-0Rapid visual detection of Monkeypox virus by one-step LAMP-CRISPR/Cas12b assayJinlei Guo0Yunfeng Shan1Ge Hu2Xiaojuan Zhu3Kangchen Zhao4Tao Wu5Qiao Qiao6Ying Chi7Lunbiao Cui8Yiyue Ge9NHC Key Laboratory of Enteric Pathogenic Microbiology, Jiangsu Provincial Medical Key Laboratory of Pathogenic Microbiology in Emerging Major Infectious Diseases, Jiangsu Provincial Center for Disease Control and PreventionNHC Key Laboratory of Enteric Pathogenic Microbiology, Jiangsu Provincial Medical Key Laboratory of Pathogenic Microbiology in Emerging Major Infectious Diseases, Jiangsu Provincial Center for Disease Control and PreventionNHC Key Laboratory of Enteric Pathogenic Microbiology, Jiangsu Provincial Medical Key Laboratory of Pathogenic Microbiology in Emerging Major Infectious Diseases, Jiangsu Provincial Center for Disease Control and PreventionNHC Key Laboratory of Enteric Pathogenic Microbiology, Jiangsu Provincial Medical Key Laboratory of Pathogenic Microbiology in Emerging Major Infectious Diseases, Jiangsu Provincial Center for Disease Control and PreventionNHC Key Laboratory of Enteric Pathogenic Microbiology, Jiangsu Provincial Medical Key Laboratory of Pathogenic Microbiology in Emerging Major Infectious Diseases, Jiangsu Provincial Center for Disease Control and PreventionNHC Key Laboratory of Enteric Pathogenic Microbiology, Jiangsu Provincial Medical Key Laboratory of Pathogenic Microbiology in Emerging Major Infectious Diseases, Jiangsu Provincial Center for Disease Control and PreventionNHC Key Laboratory of Enteric Pathogenic Microbiology, Jiangsu Provincial Medical Key Laboratory of Pathogenic Microbiology in Emerging Major Infectious Diseases, Jiangsu Provincial Center for Disease Control and PreventionNHC Key Laboratory of Enteric Pathogenic Microbiology, Jiangsu Provincial Medical Key Laboratory of Pathogenic Microbiology in Emerging Major Infectious Diseases, Jiangsu Provincial Center for Disease Control and PreventionNHC Key Laboratory of Enteric Pathogenic Microbiology, Jiangsu Provincial Medical Key Laboratory of Pathogenic Microbiology in Emerging Major Infectious Diseases, Jiangsu Provincial Center for Disease Control and PreventionNHC Key Laboratory of Enteric Pathogenic Microbiology, Jiangsu Provincial Medical Key Laboratory of Pathogenic Microbiology in Emerging Major Infectious Diseases, Jiangsu Provincial Center for Disease Control and PreventionAbstract Background Monkeypox virus (MPXV) infection has garnered significant global attention due to its rising incidence and substantial public health implications. A rapid, sensitive, and accurate diagnostic method is urgently required to enable early intervention and effective management of MPXV outbreaks. Results In this study, we developed a novel one-step assay that integrates loop-mediated isothermal amplification (LAMP) with CRISPR/Cas12b in one-pot for the detection of MPXV. The entire detection process did not require opening the lid of the reaction tube and could be completed within 40 min using extracted viral nucleic acids, which is faster than real-time quantitative PCR (qPCR). And the results could be interpreted through either real-time fluorescence or naked-eye visualization. The limit of detection (LOD) of the assay was demonstrated to be 6.5 copies per reaction and no cross-reactivity with other pathogens such as HSV, EBV, CVA16, EV-A71, and MV was found. Furthermore, when evaluated using 113 clinical samples, the assay achieved 100% sensitivity (71/71) and 100% specificity (42/42) compared to the qPCR. Conclusions In resource-limited settings, our method requires only a portable heat block or water bath and a blue light or ultraviolet flashlight for visual detection of MPXV, making it highly accessible. The integration of LAMP and CRISPR/Cas12b provides a robust, user-friendly platform for point-of-care testing, with promising potential for the rapid molecular diagnosis of infectious diseases.https://doi.org/10.1186/s12985-025-02780-0Monkeypox virusLoop-mediated isothermal amplificationCRISPR/Cas12bOne-stepVisual detectionPoint-of-care testing |
| spellingShingle | Jinlei Guo Yunfeng Shan Ge Hu Xiaojuan Zhu Kangchen Zhao Tao Wu Qiao Qiao Ying Chi Lunbiao Cui Yiyue Ge Rapid visual detection of Monkeypox virus by one-step LAMP-CRISPR/Cas12b assay Virology Journal Monkeypox virus Loop-mediated isothermal amplification CRISPR/Cas12b One-step Visual detection Point-of-care testing |
| title | Rapid visual detection of Monkeypox virus by one-step LAMP-CRISPR/Cas12b assay |
| title_full | Rapid visual detection of Monkeypox virus by one-step LAMP-CRISPR/Cas12b assay |
| title_fullStr | Rapid visual detection of Monkeypox virus by one-step LAMP-CRISPR/Cas12b assay |
| title_full_unstemmed | Rapid visual detection of Monkeypox virus by one-step LAMP-CRISPR/Cas12b assay |
| title_short | Rapid visual detection of Monkeypox virus by one-step LAMP-CRISPR/Cas12b assay |
| title_sort | rapid visual detection of monkeypox virus by one step lamp crispr cas12b assay |
| topic | Monkeypox virus Loop-mediated isothermal amplification CRISPR/Cas12b One-step Visual detection Point-of-care testing |
| url | https://doi.org/10.1186/s12985-025-02780-0 |
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