Avibacterium paragallinarum and Gallibacterium anatis are recognized bacterial pathogens both infecting the respiratory tract of chickens. The present study investigated outcomes of their co-infection by elucidating clinical signs, pathological lesions and bacteriological findings. Additionally, the efficacy of a commercially available vaccine to prevent diseases caused by Av. paragallinarum and G. anatis was evaluated. Birds inoculated with G. anatis alone did not present any clinical signs and gross pathological lesions in the respiratory tract. However, clinical signs of infectious coryza were reproduced in non-vaccinated birds that were challenged with Av. paragallinarum alone or together with G. anatis. Such clinical signs were more severe in the co-infected group including death of four birds. Some of the birds which were vaccinated and challenged showed mild clinical signs at 7 dpi. Inflammation of sinus infraorbitalis was the most prominent gross pathological lesion found in the respiratory tract of non-vaccinated birds inoculated either with Av. paragallinarum and G. anatis or Av. paragallinarum alone. In the reproductive tract, haemorrhagic follicles were observed in non-vaccinated birds that were infected either with G. anatis alone or together with Av. paragallinarum. In vaccinated birds, no gross pathological lesions were found except in one bird which was co-infected with both the pathogens characterized by mucoid tracheitis. Bacteriological investigations revealed that multiplication of G. anatis at 7 dpi was supported by the co-infection with Av. paragallinarum. Altogether, it can be concluded that simultaneous infection of Av. paragallinarum and G. anatis can increase the severities of disease conditions in chickens. In such a scenario, vaccination appears to be an effective tool for prevention of the disease as protection was conferred based on clinical, pathological, bacteriological and serological data.