The interaction between a low pathogenic avian influenza virus (A/CK/TUN/145/2012), a H9N2 Tunisian isolate, and a vaccine strain (H120) of avian infectious bronchitis, administered simultaneously or sequentially three days apart to chicks during 20 days, was evaluated using ELISA antibody levels, quantitative reverse transcription-polymerase chain reaction (qRT-PCR) analyses and histopathology examination. First, the in vivo replication interference of avian influenza virus (AIV) and infectious bronchitis virus (IBV) was evaluated using qRT-PCR to detect accurately either AIV or IBV genomes or viral copy numbers during dual infections. Second, we have determined the amount of specific antibodies in sera of chick´s infected with AIV alone, IBV alone, mixed AIV?+?IBV, IBV then AIV or AIV IBV 3 days later using an ELISA test. Finally, histopathological analyses of internal organs from inoculated chicks were realized. Quantitative results of AIV and IBV co-infection showed that interferences between the two viruses yielded decreased viral growth. However, in the case of super-infection, the second virus, either AIV or IBV, induced a decrease in the growth of the first inoculated virus. According to our results, vaccine application was safe and do not interfere with AIV H9N2 infection, and does not enhance such infection. In conclusion, co-infection of chicks with AIV and IBV, simultaneously or sequentially, affected the clinical signs, the virus replication dynamics as well as the internal organ integrity. The results proposed that infection with heterologous virus may result in temporary competition for cell receptors or competent cells for replication, most likely interferon-mediated.