{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,27]],"date-time":"2026-08-27T19:25:34Z","timestamp":1787858734040,"version":"build-2784847793"},"reference-count":25,"publisher":"MDPI AG","issue":"3","license":[{"start":{"date-parts":[[2024,2,29]],"date-time":"2024-02-29T00:00:00Z","timestamp":1709164800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100012166","name":"National Key Research and Development Program of China","doi-asserted-by":"publisher","award":["2023YFC3041400"],"award-info":[{"award-number":["2023YFC3041400"]}],"id":[{"id":"10.13039\/501100012166","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Viruses"],"abstract":"<jats:p>Monkeypox virus (MPXV), the pathogen responsible for the infectious disease monkeypox, causes lesions on the skin, lymphadenopathy, and fever. It has posed a global public health threat since May 2022. Highly sensitive and specific detection of MPXV is crucial for preventing the spread of the disease. Pyrococcus furiosus Argonaute (PfAgo) is an artificial DNA-guided restriction cleavage enzyme programmable with 5\u2032-phosphorylated ssDNA sequences, which can be developed to specifically detect nucleic acids of pathogens. Here, a PfAgo-based system was established for the detection of MPXV-specific DNA targeting the F3L gene. A short amplicon of 79 bp could be obtained through a fast PCR procedure, which was completed within 45 min. Two 5\u2032-phosphorylation guide DNAs were designed to guide PfAgo to cleave the amplicon to obtain an 18 bp 5\u2032-phosphorylation sequence specific to MPXV, not to other orthopoxviruses (cowpox, variola, and vaccinia viruses). The 18 bp sequence guided PfAgo to cleave a designed probe specific to MPXV to emit fluorescence. With optimized conditions for the PfAgo-MPXV system, it could be completed in 60 min for the detection of the extracted MPXV DNA with the limit of detection (LOD) of 1.1 copies\/reaction and did not depend on expensive instruments. Successful application of the PfAgo-MPXV system in sensitively detecting MPXV in simulated throat swabs, skin swabs, sera, and wastewater demonstrated the system\u2019s good performance. The PfAgo platform, with high sensitivity and specificity established here, has the potential to prevent the spread of MPXV.<\/jats:p>","DOI":"10.3390\/v16030382","type":"journal-article","created":{"date-parts":[[2024,2,29]],"date-time":"2024-02-29T05:59:19Z","timestamp":1709186359000},"page":"382","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":11,"title":["Fast and Ultrasensitive Detection of Monkeypox by a Pyrococcus furiosus Argonaute System Coupled with a Short Amplification"],"prefix":"10.3390","volume":"16","author":[{"given":"Ping","family":"He","sequence":"first","affiliation":[{"name":"CAS Key Laboratory of Special Pathogens and Biosafety, Center for Biosafety Mega-Science, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan 430207, China"},{"name":"University of Chinese Academy of Sciences, Beijing 100049, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Wenhao","family":"Zhou","sequence":"additional","affiliation":[{"name":"CAS Key Laboratory of Special Pathogens and Biosafety, Center for Biosafety Mega-Science, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan 430207, China"},{"name":"University of Chinese Academy of Sciences, Beijing 100049, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9948-8880","authenticated-orcid":false,"given":"Hongping","family":"Wei","sequence":"additional","affiliation":[{"name":"CAS Key Laboratory of Special Pathogens and Biosafety, Center for Biosafety Mega-Science, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan 430207, China"},{"name":"University of Chinese Academy of Sciences, Beijing 100049, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9024-0793","authenticated-orcid":false,"given":"Junping","family":"Yu","sequence":"additional","affiliation":[{"name":"CAS Key Laboratory of Special Pathogens and Biosafety, Center for Biosafety Mega-Science, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan 430207, China"},{"name":"University of Chinese Academy of Sciences, Beijing 100049, China"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2024,2,29]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","unstructured":"Beer, E.M., and Rao, V.B. 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