The Synergistic Pathogenesis Between H9N2Avian Influenza Virus And Infectious Bronchitis Virus

Both H9N2avian influenza virus (AIV) and infectious bronchitis virus (IBV) are very important respiratory tract pathogens in chickens. Co-infection of H9N2AIV and IBV in chickens leads to great economic losses in poultry industry worldwide. But up to now little is known about the synergism pathogenesis between these two agents, and there are no efficient measures for the the prevention and contrll. To improve our understanding of the synergistic pathogenesis of this dual infection, SPF chickens were inoculated with H9N2AIV and/or IBV intranasally. And we determined the morbidity and mortality rates, virus shedding, the distribution of virus and the expression of innate immune related cytokines by clinical observation, microscopic analysis and real-time RT-PCR.The results showed that the mortality rates of chickens inoculated with IBV alone and with H9N2AIV and IBV simultaneously were12%and48%respectively, while the group infected with H9N2or PBS alone was clinically normal and did not led to any death. In addition, co-infection of H9N2AIV with IBV increased the severity of clinical signs and morbidity rates. Therefore, there is a pathogenic synergism between H9N2AIV and IBV.Compared with chickens infected with H9N2or PBS alone, co-infection of H9N2AIV with IBV led to a longer shedding period of these two viruses and higher levels of virus shedding in both oropharyngeal and cloacal swabs especially during the crest-time. The viral load in the trachea, lung, kidney, thymus, spleen and bursa of fabricius of H9N2AIV and/or IBV infected specific pathogen free (SPF) chickens were determined by quantitative real-time RT-PCR, and the results showed that co-infection could promote the replication of both AIV H9N2and IBV. It is noteworthy that high levels of viral RNA of H9N2AIV in the kidneys and bursa of fabricius were found in chickens of co-infection group, while no viral RNA was detected in the kidneys of chickens infected with H9N2AIV alone and only few viral RNA was detected in the bursa of fabricius on7day post inoculation. The mRNA expression levels of IFN-α, IFN-β, IFN-y, IL-1β and IL-18were determined by quantitative real-time RT-PCR. The results showed that expression levels of cytokines varied in different tissue, and the levels of cytokine expression correlated with viral replication. Transcriptional levels of immunity related cytokines revealed that the co-infection group showed stronger innate immune responses than either H9N2AIV or IBV infection alone, which might be one of the mechanisms for the synergistic pathogenicity of the two viruses.In summary, the results in this study indicated that there is a strong pathogenic synergism between H9N2AIV and IBV. Co-infection of these two viruses in chickens results in increased viral replication, and spread to various organs, and stronger production of cytokines related to innate immune and inflammatory response.