A visual rapid detection method for bovine infectious rhinotracheitis virus

Abstract:

[Background] Bovine infectious rhinotracheitis virus (IBRV) represents a common pathogenic threat to cattle, causing substantial economic losses in the livestock industry. Therefore, it is urgent to establish a rapid detection method for IBRV. [Objective] To establish a visual rapid detection method for IBRV by leveraging recombinase polymerase amplification (RPA) in conjunction with microfluidic chip. [Methods] The ul52gene of IBRV was selected as the target fragment for molecular detection. Pseudorabies virus, herpesvirus of turkey, and duck plague herpesvirus were selected as the controls. The DNA extract dilutions of IBRV were used to evaluate the sensitivity of the established method. Thirty-four serum and nasal swab specimens from the cattle exhibiting symptoms of acute IBRV infection were used to evaluate the applicability of the established method. [Results] The size of ul52 gene amplicon was 403 bp. Sequencing results confirmed the amplicon as the ul52 gene (NC_063268.1) of IBRV, with the homology of 100%, and no amplification was observed in the control groups. The limit of detection of the established method was 6.9×10³ copies/µL. Furthermore, the established method detected IBRV in cattle with the identification number of SD0213, SD0518, and SD0701, which were consistent with the strain isolation results from both serum and nasal swab specimens. The entire process, from sample preparation to result acquisition, required 110 min. [Conclusion] We successfully developed a visual rapid molecular detection method for IBRV by combining RPA, magnetic probe capture, and microfluidic chip. The established method demonstrates good specificity and sensitivity.

Key words:infectious bovine rhinotracheitis virus;recombinase polymerase amplification;magnetic capture;microfluidic chip