Comparison of SYBR green I real-time RT-PCR with conventional agarose gel-based RT-PCR for the diagnosis of infectious bronchitis virus infection in chickens in Morocco

Abstract

BackgroundA rapid, sensitive, and specific molecular method for the diagnosis of infectious bronchitis virus (IBV) infection is important in curbing infectious bronchitis outbreaks in Morocco and other countries.MethodsIn this study, an easy-to-perform SYBR green I real-time reverse transcriptase polymerase chain reaction (RT-PCR) targeting the nucleocapsid gene of IBV was developed and compared with conventional agarose gel-based RT-PCR for the detection of IBV infection.ResultsWe found that the SYBR green I real-time RT-PCR was at least 10 times more sensitive than the agarose gel electrophoresis detection method. The assay exhibited high specificity for IBV infection. All negative controls, such as Newcastle disease virus, infectious bursal disease virus, and avian influenza virus, were not detected.ConclusionThe SYBR green I real-time RT-PCR test described herein can be used to rapidly distinguish IBV from other respiratory pathogens, which is important for diagnosis and control of infectious bronchitis outbreaks in Morocco. The test is a valuable and useful method as a routine assay for diagnosis of clinical IBV infection in commercial chickens.

Recombinase polymerase amplification–nucleic acid lateral flow immunoassays for Newcastle disease virus and infectious bronchitis virus detection

“…Several real-time RT-PCRs based tests have been reported for sensitive detection of NDV and IBV [10,12,13,26]. However, they required more than 1 h for test result.…”

“…Confirmative laboratory diagnosis can be carried out by virus isolation, haemagglutination inhibition, enzyme-linked immunosorbent assay (ELISA), immunofluorescence techniques and neutralization test [1,2,[4][5][6][7]. Also, reverse transcription-PCR (RT-PCR) and quantitative real time RT-PCR (qRT-PCR) have been reported for molecular detection of these viruses [8][9][10][11][12][13]. However, the lack of adequate laboratory resources, skilled staff requirement and the time consuming nature of the above mentioned diagnostic methods constitute obstacles to the rapid detection under field condition.…”

A sensitive one-step TaqMan amplification approach for detection of rubella virus clade I and II genotypes in clinical samples

“…The coefficient of variation was calculated according to the following formula: CV = (SD [Ct-value]/overall mean [Ct-value]) 9 100 [9].…”

Identificación de genogrupos del virus de la Enfermedad de Gumboro en granjas avícolas en Colombia

“…Las técnicas moleculares (RT-PCR, RFLP y qRT-PCR) permiten la caracterización más precisa de los cambios en el genoma que se relacionan con modificaciones de la antigenicidad (Ikuta et al, 2001;Jackwood et al, 2001;Fellahi et al, 2016). Como resultado de lo anterior, recientemente Jackwood et al, (2018) propusieron un esquema de clasificación en genogrupos basado en el análisis de la región hvVP2 del IBDV, que intenta brindar más información de los cambios que se han presentado en este virus, buscando una mejor correlación con los cuadros clínicos observados en campo.…”