Characterization and membrane gene-based phylogenetic analysis of avian infectious bronchitis virus Chinese strain HH06

A Chinese isolate of avian infectious bronchitis virus (IBV) designated HH06 was isolated from the kidney tissues of a chicken flock experiencing an outbreak of nephritis. In vivo pathogenicity of the IBV isolate HH06 was determined by inoculating specific pathogen-free (SPF) chickens. The clinical signs and related gross lesions of HH06 infected chickens were similar with those of the field-infected chickens. SPF embryonated eggs were inoculated with virus suspension for serial passage and their genomic RNA was extracted. RT-PCR technique was utilized to amplify the M gene sequence encoding membrane protein of IBV. Recombinant plasmid named T-vector-M was constructed via inserting the M gene into the TA cloning vector, pMD 18-T. The sequenced M gene and its deduced amino acid (aa) sequences were compared with the published sequences of reference strains. The M gene is of 687 bp in length encoding the M protein of 228 amino acids with a predicted molecular weight of 25.4 kDa. The sequences of the M gene and M protein share 83.9–97.9% and 83.6–96.5% homologous identities, respectively, compared with 29 IBV reference strains derived from different regions or countries, which revealed that there are still significant variations between strains. Furthermore, a phylogenetic tree based on these M DNA sequences was generated, and the tree topology suggests that some Chinese IBV strains may have a common ancestor; however, HH06 is a new local IBV isolate that is responsible for the field outbreak of nephritis.