Detection of Clostridium perfringens Using Novel Methods Based on Recombinase-Aided Amplification Assay-Assisted CRISPR/Cas12a System
Clostridium perfringens is a highly versatile pathogen of humans and animals. Rapid and sensitive detection methods for C. perfringens are urgently needed for the timely implementation of control. In this study, to provide novel promising methods for the detection of C. perfringens, two rapid, sensi...
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| Veröffentlicht in: | Transboundary and emerging diseases 2023-11, Vol.2023, p.1-11 |
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| creator | Xiao, Xingxing Zhang, Qingxun Wu, Sihong Li, Yi Zhong, Zhenyu Guo, Qingyun Li, Junfang Meng, Qinghui Cheng, Zhibin Duan, Jianbin Wang, Xiaoqiong He, Hongxuan Bai, Jiade Lou, Yongliang |
| description | Clostridium perfringens is a highly versatile pathogen of humans and animals. Rapid and sensitive detection methods for C. perfringens are urgently needed for the timely implementation of control. In this study, to provide novel promising methods for the detection of C. perfringens, two rapid, sensitive, and instrument-free C. perfringens detection methods based on recombinase-aided amplification (RAA) assay and clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated protein 12a (CRISPR/Cas12a) system were developed depending on fluorescence signal (RAA-CRISPR/Cas12a-FL) and lateral flow strip (RAA-CRISPR/Cas12a-LFS), respectively. The limit of detection of the RAA-CRISPR/Cas12a-FL and RAA-CRISPR/Cas12a-LFS methods is 2 copies and 20 copies of C. perfringens genomic DNA per reaction, respectively, and the whole process can be completed in 1 hr. Moreover, these two methods show no cross-reactivity with nontarget bacteria, which were used as a negative control to evaluate the specificity of two developed methods in the detection of C. perfringens and have 100% consistent with real-time polymerase chain reaction tests for 12 clinical samples collected from 2 Chinese Milu at Beijing Milu Ecological Research Center and 6 spiked samples from human blood and stool. Overall, the constructed C. perfringens detection methods, RAA-CRISPR/Cas12a-FL and RAA-CRISPR/Cas12a-LFS, have great potential as a novel detection scheme for the early diagnosis of C. perfringens infection in humans and animals. |
| doi_str_mv | 10.1155/2023/6667618 |
| format | Article |
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Rapid and sensitive detection methods for C. perfringens are urgently needed for the timely implementation of control. In this study, to provide novel promising methods for the detection of C. perfringens, two rapid, sensitive, and instrument-free C. perfringens detection methods based on recombinase-aided amplification (RAA) assay and clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated protein 12a (CRISPR/Cas12a) system were developed depending on fluorescence signal (RAA-CRISPR/Cas12a-FL) and lateral flow strip (RAA-CRISPR/Cas12a-LFS), respectively. The limit of detection of the RAA-CRISPR/Cas12a-FL and RAA-CRISPR/Cas12a-LFS methods is 2 copies and 20 copies of C. perfringens genomic DNA per reaction, respectively, and the whole process can be completed in 1 hr. Moreover, these two methods show no cross-reactivity with nontarget bacteria, which were used as a negative control to evaluate the specificity of two developed methods in the detection of C. perfringens and have 100% consistent with real-time polymerase chain reaction tests for 12 clinical samples collected from 2 Chinese Milu at Beijing Milu Ecological Research Center and 6 spiked samples from human blood and stool. Overall, the constructed C. perfringens detection methods, RAA-CRISPR/Cas12a-FL and RAA-CRISPR/Cas12a-LFS, have great potential as a novel detection scheme for the early diagnosis of C. perfringens infection in humans and animals.</description><identifier>ISSN: 1865-1674</identifier><identifier>EISSN: 1865-1682</identifier><identifier>DOI: 10.1155/2023/6667618</identifier><language>eng</language><publisher>Berlin: Hindawi</publisher><subject>Amplification ; Animals ; Bacteria ; Biotechnology ; Clostridium perfringens ; CRISPR ; Cross-reactivity ; Deoxyribonucleic acid ; Design ; DNA ; Ecological research ; Enzymes ; Fluorescence ; Polymerase chain reaction ; Recombinase ; Research facilities ; Software ; Statistical analysis</subject><ispartof>Transboundary and emerging diseases, 2023-11, Vol.2023, p.1-11</ispartof><rights>Copyright © 2023 Xingxing Xiao et al.</rights><rights>Copyright © 2023 Xingxing Xiao et al. This is an open access article distributed under the Creative Commons Attribution License (the “License”), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Notwithstanding the ProQuest Terms and Conditions, you may use this content in accordance with the terms of the License. https://creativecommons.org/licenses/by/4.0</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c337t-984d96982907f09a963a157da48c1b1a8bb26923713596fab644fd17e403a55a3</citedby><cites>FETCH-LOGICAL-c337t-984d96982907f09a963a157da48c1b1a8bb26923713596fab644fd17e403a55a3</cites><orcidid>0000-0002-3854-4631 ; 0000-0002-4893-1993 ; 0000-0002-4543-2080 ; 0000-0003-4198-0001</orcidid></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://www.proquest.com/docview/2893644545/fulltextPDF?pq-origsite=primo$$EPDF$$P50$$Gproquest$$Hfree_for_read</linktopdf><linktohtml>$$Uhttps://www.proquest.com/docview/2893644545?pq-origsite=primo$$EHTML$$P50$$Gproquest$$Hfree_for_read</linktohtml><link.rule.ids>315,782,786,12752,12781,21395,21396,21397,21398,21399,23263,27931,27932,33459,33537,33710,33751,34012,34321,34341,36272,43623,43666,43794,43812,43960,44074,44080,44411,64392,64396,72476,74058,74111,74290,74309,74480,74597,74603,74943</link.rule.ids></links><search><contributor>Al Salihi, Karima</contributor><contributor>Karima Al Salihi</contributor><creatorcontrib>Xiao, Xingxing</creatorcontrib><creatorcontrib>Zhang, Qingxun</creatorcontrib><creatorcontrib>Wu, Sihong</creatorcontrib><creatorcontrib>Li, Yi</creatorcontrib><creatorcontrib>Zhong, Zhenyu</creatorcontrib><creatorcontrib>Guo, Qingyun</creatorcontrib><creatorcontrib>Li, Junfang</creatorcontrib><creatorcontrib>Meng, Qinghui</creatorcontrib><creatorcontrib>Cheng, Zhibin</creatorcontrib><creatorcontrib>Duan, Jianbin</creatorcontrib><creatorcontrib>Wang, Xiaoqiong</creatorcontrib><creatorcontrib>He, Hongxuan</creatorcontrib><creatorcontrib>Bai, Jiade</creatorcontrib><creatorcontrib>Lou, Yongliang</creatorcontrib><title>Detection of Clostridium perfringens Using Novel Methods Based on Recombinase-Aided Amplification Assay-Assisted CRISPR/Cas12a System</title><title>Transboundary and emerging diseases</title><description>Clostridium perfringens is a highly versatile pathogen of humans and animals. Rapid and sensitive detection methods for C. perfringens are urgently needed for the timely implementation of control. In this study, to provide novel promising methods for the detection of C. perfringens, two rapid, sensitive, and instrument-free C. perfringens detection methods based on recombinase-aided amplification (RAA) assay and clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated protein 12a (CRISPR/Cas12a) system were developed depending on fluorescence signal (RAA-CRISPR/Cas12a-FL) and lateral flow strip (RAA-CRISPR/Cas12a-LFS), respectively. The limit of detection of the RAA-CRISPR/Cas12a-FL and RAA-CRISPR/Cas12a-LFS methods is 2 copies and 20 copies of C. perfringens genomic DNA per reaction, respectively, and the whole process can be completed in 1 hr. Moreover, these two methods show no cross-reactivity with nontarget bacteria, which were used as a negative control to evaluate the specificity of two developed methods in the detection of C. perfringens and have 100% consistent with real-time polymerase chain reaction tests for 12 clinical samples collected from 2 Chinese Milu at Beijing Milu Ecological Research Center and 6 spiked samples from human blood and stool. Overall, the constructed C. perfringens detection methods, RAA-CRISPR/Cas12a-FL and RAA-CRISPR/Cas12a-LFS, have great potential as a novel detection scheme for the early diagnosis of C. perfringens infection in humans and animals.</description><subject>Amplification</subject><subject>Animals</subject><subject>Bacteria</subject><subject>Biotechnology</subject><subject>Clostridium perfringens</subject><subject>CRISPR</subject><subject>Cross-reactivity</subject><subject>Deoxyribonucleic acid</subject><subject>Design</subject><subject>DNA</subject><subject>Ecological research</subject><subject>Enzymes</subject><subject>Fluorescence</subject><subject>Polymerase chain reaction</subject><subject>Recombinase</subject><subject>Research facilities</subject><subject>Software</subject><subject>Statistical 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Methods Based on Recombinase-Aided Amplification Assay-Assisted CRISPR/Cas12a System</atitle><jtitle>Transboundary and emerging diseases</jtitle><date>2023-11-14</date><risdate>2023</risdate><volume>2023</volume><spage>1</spage><epage>11</epage><pages>1-11</pages><issn>1865-1674</issn><eissn>1865-1682</eissn><abstract>Clostridium perfringens is a highly versatile pathogen of humans and animals. Rapid and sensitive detection methods for C. perfringens are urgently needed for the timely implementation of control. In this study, to provide novel promising methods for the detection of C. perfringens, two rapid, sensitive, and instrument-free C. perfringens detection methods based on recombinase-aided amplification (RAA) assay and clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated protein 12a (CRISPR/Cas12a) system were developed depending on fluorescence signal (RAA-CRISPR/Cas12a-FL) and lateral flow strip (RAA-CRISPR/Cas12a-LFS), respectively. The limit of detection of the RAA-CRISPR/Cas12a-FL and RAA-CRISPR/Cas12a-LFS methods is 2 copies and 20 copies of C. perfringens genomic DNA per reaction, respectively, and the whole process can be completed in 1 hr. Moreover, these two methods show no cross-reactivity with nontarget bacteria, which were used as a negative control to evaluate the specificity of two developed methods in the detection of C. perfringens and have 100% consistent with real-time polymerase chain reaction tests for 12 clinical samples collected from 2 Chinese Milu at Beijing Milu Ecological Research Center and 6 spiked samples from human blood and stool. Overall, the constructed C. perfringens detection methods, RAA-CRISPR/Cas12a-FL and RAA-CRISPR/Cas12a-LFS, have great potential as a novel detection scheme for the early diagnosis of C. perfringens infection in humans and animals.</abstract><cop>Berlin</cop><pub>Hindawi</pub><doi>10.1155/2023/6667618</doi><tpages>11</tpages><orcidid>https://orcid.org/0000-0002-3854-4631</orcidid><orcidid>https://orcid.org/0000-0002-4893-1993</orcidid><orcidid>https://orcid.org/0000-0002-4543-2080</orcidid><orcidid>https://orcid.org/0000-0003-4198-0001</orcidid><oa>free_for_read</oa></addata></record> |
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| subjects | Amplification Animals Bacteria Biotechnology Clostridium perfringens CRISPR Cross-reactivity Deoxyribonucleic acid Design DNA Ecological research Enzymes Fluorescence Polymerase chain reaction Recombinase Research facilities Software Statistical analysis |
| title | Detection of Clostridium perfringens Using Novel Methods Based on Recombinase-Aided Amplification Assay-Assisted CRISPR/Cas12a System |
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