Single-step multiplex reverse transcription-polymerase chain reaction for the detection and differentiation of QX-like infectious bronchitis virus from the Thai variant and vaccine strains H120, Ma5, and 4/91

Research (Published online: 27-05-2023)

25. Single-step multiplex reverse transcription-polymerase chain reaction for the detection and differentiation of QX-like infectious bronchitis virus from the Thai variant and vaccine strains H120, Ma5, and 4/91

Sucheeva Junnu and Tawatchai Pohuang

Veterinary World, 16(5): 1109-1113

Sucheeva Junnu: Division of Livestock Medicine, Faculty of Veterinary Medicine, Khon Kaen University, Khon Kaen 40002, Thailand; Research Group for Emerging and Re-emerging Infectious Diseases in Animals and Zoonotic Diseases, Faculty of Veterinary Medicine, Khon Kaen University, Khon Kaen 40002, Thailand.

Tawatchai Pohuang: Division of Livestock Medicine, Faculty of Veterinary Medicine, Khon Kaen University, Khon Kaen 40002, Thailand; Research Group for Emerging and Re-emerging Infectious Diseases in Animals and Zoonotic Diseases, Faculty of Veterinary Medicine, Khon Kaen University, Khon Kaen 40002, Thailand.

doi: 10.14202/vetworld.2023.1109-1113

Article history: Received: 01-01-2023, Accepted: 19-04-2023, Published online: 27-05-2023

Corresponding authors: Tawatchai Pohuang

E-mail: ptawat@kku.ac.th

Citation: Junnu S and Pohuang T (2023) Single-step multiplex reverse transcription-polymerase chain reaction for the detection and differentiation of QX-like infectious bronchitis virus from the Thai variant and vaccine strains H120, Ma5, and 4/91, Veterinary World, 16(5): 1109-1113.

Abstract

Background and Aim: QX-like infectious bronchitis virus (IBV) is a highly infectious avian coronavirus that causes respiratory and kidney disease. It is linked to increased mortality and loss of performance in infected chickens worldwide, including Thailand. Thus, a simple and rapid diagnostic method for the diagnosis of QX-like IBV is needed. This study aimed to develop a single-step multiplex reverse transcription-polymerase chain reaction (mRT–PCR) assay to detect and differentiate QX-like IBV from Thai IBV and vaccine strains used in the poultry industry (H120, Ma5, and 4/91).

Materials and Methods: Primer sets specific for QX-like and Thai IBV were designed to target the S1 gene. The specificity of the technique was verified using nine isolates of QX-like IBV, four isolates of Thai IBV, and other avian viral respiratory pathogens. The detection limit was evaluated using a serial ten-fold dilution of QX-like and Thai IBV.

Results: The results showed that single-step mRT–PCR could detect QX-like IBV and differentiate it from Thai IBV and the vaccine strains H120, Ma5, and 4/91. The limit of detection of the developed assay was 10^2.2 embryo infectious dose (EID)₅₀/mL for QX-like IBV and 10^1.8 EID_₅₀/mL for Thai IBV. Interestingly, the developed assay could identify mixed infection by both IBVs in a single sample.

Conclusion: The single-step mRT–PCR assay developed in this study can potentially discriminate QX-like IBV from Thai IBV and the vaccine strains H120, Ma5, and 4/91 in a single reaction. It is also suitable for use in all laboratories with access to conventional PCR equipment.

Keywords: detection, QX-like infectious bronchitis virus, single-step multiplex reverse transcription-polymerase chain reaction, specificity.