Development and evaluation of a CRISPR/Cas12a-based diagnostic test for rapid detection and genotyping of HR-HPV in clinical specimens
ABSTRACT Persistent infection with high-risk human papillomavirus (HR-HPV) is the principal etiological factor of cervical cancer. Considering the gradual progression of cervical cancer, the early, rapid, sensitive, and specific identification of HPV, particularly HR-HPV types, is crucial in halting...
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| Format: | Article |
| Language: | English |
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American Society for Microbiology
2025-01-01
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| Series: | Microbiology Spectrum |
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| Online Access: | https://journals.asm.org/doi/10.1128/spectrum.02253-24 |
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| author | Lijuan Yin Ziqian Zhao Chunhua Wang Caihong Zhou Xiuzhen Wu Baoxue Gao Liangyuan Wang Shuli Man Xinkuan Cheng Qiankun Wu Siqi Hu Hongxia Fan Long Ma Hui Xing Liang Shen |
| author_facet | Lijuan Yin Ziqian Zhao Chunhua Wang Caihong Zhou Xiuzhen Wu Baoxue Gao Liangyuan Wang Shuli Man Xinkuan Cheng Qiankun Wu Siqi Hu Hongxia Fan Long Ma Hui Xing Liang Shen |
| author_sort | Lijuan Yin |
| collection | DOAJ |
| description | ABSTRACT Persistent infection with high-risk human papillomavirus (HR-HPV) is the principal etiological factor of cervical cancer. Considering the gradual progression of cervical cancer, the early, rapid, sensitive, and specific identification of HPV, particularly HR-HPV types, is crucial in halting the advancement of the illness. Here, we established a rapid, highly sensitive, and specific HR-HPV detection platform, leveraging the CRISPR/Cas12a assay in conjunction with multienzyme isothermal rapid amplification. Our platform enables the detection and genotyping of 14 types of HR-HPV by using type-specific crRNAs. The outcomes of the detection can be interpreted either through a fluorescence reader or visually. Furthermore, we achieved one-tube multiplex detection of 14 HR-HPV types through the use of multiple amplifications and a crRNA pool. The detection sensitivity of this method is 2 copies/μL with no cross-reactivity, and the results can be obtained within 30 minutes. This method exhibited 100% clinical sensitivity and 100% clinical specificity when applied to 258 clinical specimens. Based on these findings, our CRISPR/Cas-based HR-HPV detection platform holds promise as a novel clinical detection tool, offering a visually intuitive and expedited alternative to existing HPV infection diagnostics and providing fresh perspectives for clinical cervical cancer screening.IMPORTANCEThis study developed a novel high-risk human papillomavirus (HR-HPV) detection platform based on CRISPR/Cas12a technology. This platform not only enables the rapid, highly sensitive, and specific detection and genotyping of 14 types of HR-HPV but also achieves single-tube multiplex detection of 14 HR-HPV types through ingenious design. The outcomes of the detection can be interpreted either through a fluorescence reader or visually. To the best of our knowledge, this is the first paper to utilize CRISPR/Cas diagnostic technology for the simultaneous detection of 14 types of HPV and to evaluate its feasibility in clinical sample detection using a large number of clinical samples. We hope that this work will facilitate the rapid and accurate detection of HPV and promote the broader application of CRISPR/Cas diagnostic technology. |
| format | Article |
| id | doaj-art-5c936e7538db43168d2f42529f6835d7 |
| institution | Kabale University |
| issn | 2165-0497 |
| language | English |
| publishDate | 2025-01-01 |
| publisher | American Society for Microbiology |
| record_format | Article |
| series | Microbiology Spectrum |
| spelling | doaj-art-5c936e7538db43168d2f42529f6835d72025-01-07T14:05:19ZengAmerican Society for MicrobiologyMicrobiology Spectrum2165-04972025-01-0113110.1128/spectrum.02253-24Development and evaluation of a CRISPR/Cas12a-based diagnostic test for rapid detection and genotyping of HR-HPV in clinical specimensLijuan Yin0Ziqian Zhao1Chunhua Wang2Caihong Zhou3Xiuzhen Wu4Baoxue Gao5Liangyuan Wang6Shuli Man7Xinkuan Cheng8Qiankun Wu9Siqi Hu10Hongxia Fan11Long Ma12Hui Xing13Liang Shen14State Key Laboratory of Food Nutrition and Safety, Key Laboratory of Industrial Microbiology, Ministry of Education, Tianjin Key Laboratory of Industry Microbiology, National and Local United Engineering Lab of Metabolic Control Fermentation Technology, China International Science and Technology Cooperation Base of Food Nutrition/Safety and Medicinal Chemistry, College of Biotechnology, Tianjin University of Science & Technology, Tianjin, ChinaState Key Laboratory of Food Nutrition and Safety, Key Laboratory of Industrial Microbiology, Ministry of Education, Tianjin Key Laboratory of Industry Microbiology, National and Local United Engineering Lab of Metabolic Control Fermentation Technology, China International Science and Technology Cooperation Base of Food Nutrition/Safety and Medicinal Chemistry, College of Biotechnology, Tianjin University of Science & Technology, Tianjin, ChinaDepartment of Clinical Laboratory, Xiangyang Central Hospital, Affiliated Hospital of Hubei University of Arts and Science, Xiangyang, Hubei Province, ChinaState Key Laboratory of Food Nutrition and Safety, Key Laboratory of Industrial Microbiology, Ministry of Education, Tianjin Key Laboratory of Industry Microbiology, National and Local United Engineering Lab of Metabolic Control Fermentation Technology, China International Science and Technology Cooperation Base of Food Nutrition/Safety and Medicinal Chemistry, College of Biotechnology, Tianjin University of Science & Technology, Tianjin, ChinaDynamiker Sub-Center of Beijing Key Laboratory for Mechanisms Research and Precision Diagnosis of Invasive Fungal Disease, Tianjin, ChinaState Key Laboratory of Food Nutrition and Safety, Key Laboratory of Industrial Microbiology, Ministry of Education, Tianjin Key Laboratory of Industry Microbiology, National and Local United Engineering Lab of Metabolic Control Fermentation Technology, China International Science and Technology Cooperation Base of Food Nutrition/Safety and Medicinal Chemistry, College of Biotechnology, Tianjin University of Science & Technology, Tianjin, ChinaState Key Laboratory of Food Nutrition and Safety, Key Laboratory of Industrial Microbiology, Ministry of Education, Tianjin Key Laboratory of Industry Microbiology, National and Local United Engineering Lab of Metabolic Control Fermentation Technology, China International Science and Technology Cooperation Base of Food Nutrition/Safety and Medicinal Chemistry, College of Biotechnology, Tianjin University of Science & Technology, Tianjin, ChinaState Key Laboratory of Food Nutrition and Safety, Key Laboratory of Industrial Microbiology, Ministry of Education, Tianjin Key Laboratory of Industry Microbiology, National and Local United Engineering Lab of Metabolic Control Fermentation Technology, China International Science and Technology Cooperation Base of Food Nutrition/Safety and Medicinal Chemistry, College of Biotechnology, Tianjin University of Science & Technology, Tianjin, ChinaState Key Laboratory of Food Nutrition and Safety, Key Laboratory of Industrial Microbiology, Ministry of Education, Tianjin Key Laboratory of Industry Microbiology, National and Local United Engineering Lab of Metabolic Control Fermentation Technology, China International Science and Technology Cooperation Base of Food Nutrition/Safety and Medicinal Chemistry, College of Biotechnology, Tianjin University of Science & Technology, Tianjin, ChinaAcademy of National Food and Strategic Reserves Administration, Beijing, ChinaInstitute of Pediatrics, Faculty of Pediatrics, The Seventh Medical Center of Chinese PLA General Hospital, Beijing, ChinaDepartment of Pathogen Biology, School of Basic Medical Sciences, Tianjin Medical University, Tianjin, ChinaState Key Laboratory of Food Nutrition and Safety, Key Laboratory of Industrial Microbiology, Ministry of Education, Tianjin Key Laboratory of Industry Microbiology, National and Local United Engineering Lab of Metabolic Control Fermentation Technology, China International Science and Technology Cooperation Base of Food Nutrition/Safety and Medicinal Chemistry, College of Biotechnology, Tianjin University of Science & Technology, Tianjin, ChinaDepartment of Clinical Laboratory, Xiangyang Central Hospital, Affiliated Hospital of Hubei University of Arts and Science, Xiangyang, Hubei Province, ChinaDepartment of Clinical Laboratory, Xiangyang Central Hospital, Affiliated Hospital of Hubei University of Arts and Science, Xiangyang, Hubei Province, ChinaABSTRACT Persistent infection with high-risk human papillomavirus (HR-HPV) is the principal etiological factor of cervical cancer. Considering the gradual progression of cervical cancer, the early, rapid, sensitive, and specific identification of HPV, particularly HR-HPV types, is crucial in halting the advancement of the illness. Here, we established a rapid, highly sensitive, and specific HR-HPV detection platform, leveraging the CRISPR/Cas12a assay in conjunction with multienzyme isothermal rapid amplification. Our platform enables the detection and genotyping of 14 types of HR-HPV by using type-specific crRNAs. The outcomes of the detection can be interpreted either through a fluorescence reader or visually. Furthermore, we achieved one-tube multiplex detection of 14 HR-HPV types through the use of multiple amplifications and a crRNA pool. The detection sensitivity of this method is 2 copies/μL with no cross-reactivity, and the results can be obtained within 30 minutes. This method exhibited 100% clinical sensitivity and 100% clinical specificity when applied to 258 clinical specimens. Based on these findings, our CRISPR/Cas-based HR-HPV detection platform holds promise as a novel clinical detection tool, offering a visually intuitive and expedited alternative to existing HPV infection diagnostics and providing fresh perspectives for clinical cervical cancer screening.IMPORTANCEThis study developed a novel high-risk human papillomavirus (HR-HPV) detection platform based on CRISPR/Cas12a technology. This platform not only enables the rapid, highly sensitive, and specific detection and genotyping of 14 types of HR-HPV but also achieves single-tube multiplex detection of 14 HR-HPV types through ingenious design. The outcomes of the detection can be interpreted either through a fluorescence reader or visually. To the best of our knowledge, this is the first paper to utilize CRISPR/Cas diagnostic technology for the simultaneous detection of 14 types of HPV and to evaluate its feasibility in clinical sample detection using a large number of clinical samples. We hope that this work will facilitate the rapid and accurate detection of HPV and promote the broader application of CRISPR/Cas diagnostic technology.https://journals.asm.org/doi/10.1128/spectrum.02253-24CRISPR/Cas12amultiplexdetectionHR-HPVgenotypingclinical samples |
| spellingShingle | Lijuan Yin Ziqian Zhao Chunhua Wang Caihong Zhou Xiuzhen Wu Baoxue Gao Liangyuan Wang Shuli Man Xinkuan Cheng Qiankun Wu Siqi Hu Hongxia Fan Long Ma Hui Xing Liang Shen Development and evaluation of a CRISPR/Cas12a-based diagnostic test for rapid detection and genotyping of HR-HPV in clinical specimens Microbiology Spectrum CRISPR/Cas12a multiplex detection HR-HPV genotyping clinical samples |
| title | Development and evaluation of a CRISPR/Cas12a-based diagnostic test for rapid detection and genotyping of HR-HPV in clinical specimens |
| title_full | Development and evaluation of a CRISPR/Cas12a-based diagnostic test for rapid detection and genotyping of HR-HPV in clinical specimens |
| title_fullStr | Development and evaluation of a CRISPR/Cas12a-based diagnostic test for rapid detection and genotyping of HR-HPV in clinical specimens |
| title_full_unstemmed | Development and evaluation of a CRISPR/Cas12a-based diagnostic test for rapid detection and genotyping of HR-HPV in clinical specimens |
| title_short | Development and evaluation of a CRISPR/Cas12a-based diagnostic test for rapid detection and genotyping of HR-HPV in clinical specimens |
| title_sort | development and evaluation of a crispr cas12a based diagnostic test for rapid detection and genotyping of hr hpv in clinical specimens |
| topic | CRISPR/Cas12a multiplex detection HR-HPV genotyping clinical samples |
| url | https://journals.asm.org/doi/10.1128/spectrum.02253-24 |
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