Establishment of human adenovirus species B detection method based on CRISPR-Cas13a

Abstract:

[Background] Human adenovirus species B (HAdV-B) can cause respiratory tract infections and severe pneumonia with high mortality rates. Currently, there is a lack of rapid and accurate detection methods for HAdV-B. [Objective] To develop a highly specific, sensitive, and user-friendly detection method for HAdV-B. [Methods] Conserved sequences of HAdV-B were screened in the human adenovirus E4 gene region, and specific recombinase-aided amplification (RAA) primers and CRISPR RNA (crRNA) were selected. We then established a rapid nucleic acid detection method targeting HAdV-B by combining RAA, CRISPR-Cas13a system, and easy-readout and sensitive enhanced (ERASE) nucleic acid test strip. [Results] The established method could detect HAdV-B DNA as low as 10⁰ copy/μL within 35 min, with the sensitivity comparable to real-time PCR. No cross-reactivity with other respiratory pathogens was observed. The testing results with simulated samples showed that the method can detect the weak positive samples with a Ct value of 36.19. [Conclusion] The test strip assay established in this study for HAdV-B can detect the target nucleic acids in a simple, rapid, and accurate manner without the need for specialized nucleic acid detection equipment, which provides a new method for detecting HAdV-B.