A multiplex reverse transcriptase-PCR assay for the genotyping of avian infectious bronchitis viruses.
Infectious bronchitis viruses (IBVs) in Taiwan have been divided into two genogroups, Taiwan group I (TW-I) and Taiwan group II (TW-II). Heterologous Mass-type strains are widely used as vaccines in the field. This work reports on a rapid and reliable multiplex reverse transcriptase-polymerase chain reaction (mRT-PCR) assay for the genotyping of IBVs. Multiplex primer sets were designed to amplify the region covering hypervariable regions 1 and 2 of the S1 glycoprotein gene. Several local strains and commercially available vaccines were used for evaluating the viral genotyping assay, and a number of field isolates were examined for clinical application. The results showed that all of the examined IBVs were accurately genotyped by identifying the corresponding bands on agarose gels (TW-I: 322 bp, TW-II: 161 bp, Mass type: 256 bp) after the mRT-PCR, in agreement with the viral genome sequence data. The mRT-PCR assay was able to detect viral RNA copies as low as 10(3), 10(50, and 10(3) for the TW-I, TW-II, and Mass-type strains, respectively. The mRT-PCR assay accurately detected and discriminated vaccine viruses from wildtype strains in the field. This assay may be beneficial for virus identification and differentiation in routine disease surveillance.
Chen
Hui-Wen
HW
School of Veterinary Medicine, National Taiwan University, No. 1, Sec. 4, Roosevelt Rd., Taipei 10617, Taiwan.
Wang
Ching-Ho
CH
D005838:Genotype; D001351:Infectious bronchitis virus / Q000235:genetics*; D012367:RNA, Viral / Q000235:genetics; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity
D012367:RNA, Viral