1. A CRISPR/Cas12a-assisted rapid detection platform by biosensing the apxIVA of Actinobacillus pleuropneumoniae
- Author
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Luan, Tian, Wang, Lu, Zhao, Jiyu, Luan, Hui, Zhang, Yueling, Wang, Chunlai, Langford, Paul R., Liu, Siguo, Zhang, Wanjiang, Li, Gang, and Biotechnology and Biological Sciences Research Council (BBSRC)
- Subjects
Microbiology (medical) ,0502 Environmental Science and Management ,0503 Soil Sciences ,Microbiology ,0605 Microbiology - Abstract
Actinobacillus pleuropneumoniae is an important respiratory pig pathogen that causes substantial losses in the worldwide swine industry. Chronic or subclinical infection with no apparent clinical symptoms poses a challenge for preventing transmission between herds. Rapid diagnostics is important for the control of epidemic diseases. In this study, we formulated an A. pleuropneumoniae species-specific apxIVA-based CRISPR/Cas12a-assisted rapid detection platform (Card) that combines recombinase polymerase amplification (RPA) of target DNA and subsequent Cas12a ssDNase activation. Card has a detection limit of 10 CFUs of A. pleuropneumoniae, and there is no cross-reactivity with other common swine pathogens. The detection process can be completed in 1 h, and there was 100% agreement between the conventional apxIVA-based PCR and Card in detecting A. pleuropneumoniae in lung samples. Microplate fluorescence readout enables high-throughput use in diagnostic laboratories, and naked eye and lateral flow test readouts enable use at the point of care. We conclude that Card is a versatile, rapid, accurate molecular diagnostic platform suitable for use in both laboratory and low-resource settings.
- Published
- 2022