Abstract:
The aims of this study were firstly to establish a nested RT-PCR assay for detection of infectious bronchitis virus (IBV) including IBV isolated in Thailand, secondly to determine the molecular characterization of the recent Thai IBV by analysis of the S1 genes, and lastly to evaluate the levels of protection generated by 2 live attenuated vaccine strains against an IBV isolated in Thailand. In the development of nested RT-PCR, we designed the new primer sets for this assay and the results showed that the sensitivity of the nested PCR was increased 10 fold from virus isolation and 10-100 folds from non-nested RT-PCR. There were no cross-reactions with other avian viruses. These results suggest that the nested RT-PCR can be a sensitive and specific method for the diagnosis of IBV infection. In the molecular characterization, we collected thirty-two Thai IBV isolates from the outbreaks of disease in commercial chicken farms during 2008-2009. After sequencing of the S1 gene, phylogenetic analysis was performed and this found that the Thai IBV isolates were divided into three distinct groups, unique to Thailand (group I), QX-like IBV (group II) and Massachusetts type (group III). Moreover, the recombination events were found in groups I and II, but not in group III Thai IBV. Based on these facts, the field IBV in Thailand has undergone genetic recombination and evolution. In the protection study by using live attenuated vaccine strains H120 and 4/91, the chickens were vaccinated at 1 and 14-day-old with different vaccination programs and challenged with Thai IBV isolate THA80151 at 28-day-old. The results showed that the body weight gains of the vaccinated chickens were higher than the infected but non vaccinated chickens (p<0.05). Furthermore, the morbidity rates and tracheal histopathologic lesion scores of vaccinated chickens were lower than the infected chickens that were not vaccinated (p<0.05) although the infection rates of the tracheas were similar. These suggested that the live attenuated vaccines used in this study could induce clinical protection when administered at an interval of 2 weeks but could not protect against the infection of a challenge strain.
