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A Dual-mode platform for the rapid detection of Escherichia coli O157:H7 based on CRISPR/Cas12a and RPA

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Abstract

Escherichia coli O157:H7 (E. coli O157:H7) is a foodborne pathogenic microorganism that is commonly found in the environment and poses a significant threat to human health, public safety, and economic stability worldwide. Thus, early detection is essential for E. coli O157:H7 control. In recent years, a series of E. coli O157:H7 detection methods have been developed, but the sensitivity and portability of the methods still need improvement. Therefore, in this study, a rapid and efficient testing platform based on the CRISPR/Cas12a cleavage reaction was constructed. Through the integration of recombinant polymerase amplification and lateral flow chromatography, we established a dual-interpretation-mode detection platform based on CRISPR/Cas12a-derived fluorescence and lateral flow chromatography for the detection of E. coli O157:H7. For the fluorescence detection method, the limits of detection (LODs) of genomic DNA and E. coli O157:H7 were 1.8 fg/µL and 2.4 CFU/mL, respectively, within 40 min. Conversely, for the lateral flow detection method, LODs of 1.8 fg/µL and 2.4 × 102 CFU/mL were achieved for genomic DNA and E. coli O157:H7, respectively, within 45 min. This detection strategy offered higher sensitivity and lower equipment requirements than industry standards. In conclusion, the established platform showed excellent specificity and strong universality. Modifying the target gene and its primers can broaden the platform’s applicability to detect various other foodborne pathogens.

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Acknowledgements

We thank LetPub (www.letpub.com) for its linguistic assistance during the preparation of this manuscript.

Funding

This study was supported by the Scientific and Innovative Action Plan of Shanghai (21N31900800), Shanghai Rising-Star Program (23QB1403500), Ministry of Agriculture and Rural Affairs Dairy Cluster Program (2023–2025), Shanghai Science and Technology Commission, the Belt and Road Project (20310750500), Talent Project of SAAS (2023–2025), and the SAAS Program for Excellent Research Team (2022 (B-16)).

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Author notes

  1. Jiawei Luo and Danhong Xu have contributed equally to this work.

Authors and Affiliations

  1. Lanzhou University of Technology, School of Life Science and Engineering, Lanzhou, China

    Jiawei Luo, Yan Zhang & Xiaofeng Liu

  2. The Biotechnology Research Institute, Shanghai Academy of Agricultural Sciences, Key Laboratory of Agricultural Genetics and Breeding, Shanghai, China

    Jiawei Luo, Danhong Xu, Jinbin Wang, Hua Liu, You Li, Yan Zhang & Haijuan Zeng

  3. Crops Ecological Environment Security Inspection and Supervision Center (Shanghai), Ministry of Agriculture and Rural Affairs, Shanghai, China

    Jinbin Wang & Haijuan Zeng

  4. Shanghai Co-Elite Agricultural Sci-Tech (Group) Co. Ltd., Shanghai, China

    Jinbin Wang & Haijuan Zeng

  5. School of Public Health, Academician Workstation, Changsha Medical University, Changsha, China

    Jinbin Wang & Haijuan Zeng

  6. Shanghai Center of Agri-Products Quality and Safety, Shanghai, China

    Bo Deng

Authors
  1. Jiawei Luo
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  2. Danhong Xu
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  3. Jinbin Wang
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  4. Hua Liu
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  5. You Li
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  6. Yan Zhang
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  7. Haijuan Zeng
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  8. Bo Deng
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  9. Xiaofeng Liu
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Contributions

Jiawei Luo and Danhong Xu designed the study. Jiawei Luo, Danhong Xu, Hua Liu, and Jinbing Wang designed and performed the experiments and analyzed the data. Jiawei Luo, Danhong Xu, Bo Deng, Haijuan Zeng, and Xiaofeng Liu wrote and revised the manuscript. All authors read and approved the final version of the manuscript.

Corresponding authors

Correspondence to Haijuan Zeng, Bo Deng or Xiaofeng Liu.

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Luo, J., Xu, D., Wang, J. et al. A Dual-mode platform for the rapid detection of Escherichia coli O157:H7 based on CRISPR/Cas12a and RPA. Anal Bioanal Chem (2024). https://doi.org/10.1007/s00216-024-05301-0

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  • DOI: https://doi.org/10.1007/s00216-024-05301-0

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