End-point diagnostics of Giardia duodenalis assemblages A and B by combining RPA with CRISPR/Cas12a from human fecal samples
Abstract Background Giardia duodenalis is a common enteric protozoan parasite that is categorized into eight assemblages (A–H). In particular, assemblages A and B are zoonotic, capable of infecting both humans and animals worldwide, resulting in significant economic losses and public health challeng...
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BMC
2024-11-01
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| Series: | Parasites & Vectors |
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| Online Access: | https://doi.org/10.1186/s13071-024-06559-0 |
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| author | Yilin Wang Fuchang Yu Yin Fu Qian Zhang Jinfeng Zhao Ziyang Qin Ke Shi Yayun Wu Junqiang Li Xiaoying Li Longxian Zhang |
| author_facet | Yilin Wang Fuchang Yu Yin Fu Qian Zhang Jinfeng Zhao Ziyang Qin Ke Shi Yayun Wu Junqiang Li Xiaoying Li Longxian Zhang |
| author_sort | Yilin Wang |
| collection | DOAJ |
| description | Abstract Background Giardia duodenalis is a common enteric protozoan parasite that is categorized into eight assemblages (A–H). In particular, assemblages A and B are zoonotic, capable of infecting both humans and animals worldwide, resulting in significant economic losses and public health challenges in epidemic regions. Thus, the development of rapid, accurate and non-laboratory-based diagnostic methods for infected animals is crucial for the effective prevention and control of giardiasis. Recent advancements in clustered, regularly interspaced, short palindromic repeats (CRISPR) and CRISPR-associated (Cas) protein (Cas12a) systems allow promising avenues for nucleic acid detection, characterized by their high flexibility, sensitivity and specificity. Methods Combined recombinase polymerase amplification and CRISPR/Cas12a systems were combined and used as end-point diagnostic methods (termed REPORT) to detect G. duodenalis assemblage A and B. The diagnostic results can be observed by fluorescence readouts with the naked eye under blue light or colorimetric signals using a lateral flow strip (LFS). Results The limit of detection (LOD) of the REPORT‑based G. duodenalis assemblage A detection was 2.04 CFU/ml and 10 trophozoites per gram (TPG), and the LOD of assemblage B was 1.1 CFU/ml and 10 cysts per gram (CPG). The REPORT‑based G. duodenalis assemblage A and assemblage B detection methods have strong specificity and no cross-reactivity with other assemblages of G. duodenalis or common enteric parasitic protozoa and have excellent performance in clinical sample detection. Conclusions This study presents a novel strategy for the direct identification of G. duodenalis assemblages A and B, requiring neither highly trained personnel nor costly specialized equipment. Graphical Abstract |
| format | Article |
| id | doaj-art-07bb0e89392b4d3d8a0ae8a60a8c2549 |
| institution | Kabale University |
| issn | 1756-3305 |
| language | English |
| publishDate | 2024-11-01 |
| publisher | BMC |
| record_format | Article |
| series | Parasites & Vectors |
| spelling | doaj-art-07bb0e89392b4d3d8a0ae8a60a8c25492024-11-17T12:12:34ZengBMCParasites & Vectors1756-33052024-11-0117111310.1186/s13071-024-06559-0End-point diagnostics of Giardia duodenalis assemblages A and B by combining RPA with CRISPR/Cas12a from human fecal samplesYilin Wang0Fuchang Yu1Yin Fu2Qian Zhang3Jinfeng Zhao4Ziyang Qin5Ke Shi6Yayun Wu7Junqiang Li8Xiaoying Li9Longxian Zhang10College of Veterinary Medicine, Henan Agricultural UniversityCollege of Veterinary Medicine, Henan Agricultural UniversityCollege of Veterinary Medicine, Henan Agricultural UniversityYebio Bioengineering Co., Ltd of QingdaoCollege of Veterinary Medicine, Henan Agricultural UniversityCollege of Veterinary Medicine, Henan Agricultural UniversitySchool of Medicine, Xinxiang UniversityCollege of Veterinary Medicine, Henan Agricultural UniversityCollege of Veterinary Medicine, Henan Agricultural UniversityCollege of Veterinary Medicine, Henan Agricultural UniversityCollege of Veterinary Medicine, Henan Agricultural UniversityAbstract Background Giardia duodenalis is a common enteric protozoan parasite that is categorized into eight assemblages (A–H). In particular, assemblages A and B are zoonotic, capable of infecting both humans and animals worldwide, resulting in significant economic losses and public health challenges in epidemic regions. Thus, the development of rapid, accurate and non-laboratory-based diagnostic methods for infected animals is crucial for the effective prevention and control of giardiasis. Recent advancements in clustered, regularly interspaced, short palindromic repeats (CRISPR) and CRISPR-associated (Cas) protein (Cas12a) systems allow promising avenues for nucleic acid detection, characterized by their high flexibility, sensitivity and specificity. Methods Combined recombinase polymerase amplification and CRISPR/Cas12a systems were combined and used as end-point diagnostic methods (termed REPORT) to detect G. duodenalis assemblage A and B. The diagnostic results can be observed by fluorescence readouts with the naked eye under blue light or colorimetric signals using a lateral flow strip (LFS). Results The limit of detection (LOD) of the REPORT‑based G. duodenalis assemblage A detection was 2.04 CFU/ml and 10 trophozoites per gram (TPG), and the LOD of assemblage B was 1.1 CFU/ml and 10 cysts per gram (CPG). The REPORT‑based G. duodenalis assemblage A and assemblage B detection methods have strong specificity and no cross-reactivity with other assemblages of G. duodenalis or common enteric parasitic protozoa and have excellent performance in clinical sample detection. Conclusions This study presents a novel strategy for the direct identification of G. duodenalis assemblages A and B, requiring neither highly trained personnel nor costly specialized equipment. Graphical Abstracthttps://doi.org/10.1186/s13071-024-06559-0Giardia duodenalisRecombinase polymerase amplificationCRISPR/Cas12aVisualized detectionOn-site detection |
| spellingShingle | Yilin Wang Fuchang Yu Yin Fu Qian Zhang Jinfeng Zhao Ziyang Qin Ke Shi Yayun Wu Junqiang Li Xiaoying Li Longxian Zhang End-point diagnostics of Giardia duodenalis assemblages A and B by combining RPA with CRISPR/Cas12a from human fecal samples Parasites & Vectors Giardia duodenalis Recombinase polymerase amplification CRISPR/Cas12a Visualized detection On-site detection |
| title | End-point diagnostics of Giardia duodenalis assemblages A and B by combining RPA with CRISPR/Cas12a from human fecal samples |
| title_full | End-point diagnostics of Giardia duodenalis assemblages A and B by combining RPA with CRISPR/Cas12a from human fecal samples |
| title_fullStr | End-point diagnostics of Giardia duodenalis assemblages A and B by combining RPA with CRISPR/Cas12a from human fecal samples |
| title_full_unstemmed | End-point diagnostics of Giardia duodenalis assemblages A and B by combining RPA with CRISPR/Cas12a from human fecal samples |
| title_short | End-point diagnostics of Giardia duodenalis assemblages A and B by combining RPA with CRISPR/Cas12a from human fecal samples |
| title_sort | end point diagnostics of giardia duodenalis assemblages a and b by combining rpa with crispr cas12a from human fecal samples |
| topic | Giardia duodenalis Recombinase polymerase amplification CRISPR/Cas12a Visualized detection On-site detection |
| url | https://doi.org/10.1186/s13071-024-06559-0 |
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