In the past five years, inclusion body hepatitis-hydropericardium syndrome caused by fowl adenovirus type 4 (FAdV-4) has been rampant in China and is still an epidemic at present. We had found the contamination of FAdV-4 in Newcastle disease virus (NDV)-attenuated vaccine from a large farm with aforementioned disease, and then conjectured that the use of this contaminated vaccine might be an important reason for the outbreak of the FAdV-4 infection. To assess the pathogenicity of this contaminated vaccine and the interaction between FAdV and LaSota, systemic animal experiments were launched with FAdV infection via the contaminated vaccine, as well as FAdV direct infection. Results showed that no chicks died after FAdV direct infection, while a 16.7% mortality rate appeared after FAdV infection through the contaminated vaccine; the synergistic reaction of FAdV and LaSota aggravated their inhibitory action for weight growth of SPF chickens, enhanced one another's capacity for immune system destruction, promoted their multiplication in vivo, and restrained antibody production. This study demonstrated the intracorporal interaction and enhanced pathogenicity of FAdV-4 and LaSota strain in contaminated NDV-attenuated vaccine, illustrating how the exogenous virus causes infection and induces severe diseases, which reminds us that the damage of attenuated vaccine contaminated with FAdV-4 even in extremely low dose is not insignificant.