Considerable evidence has implicated respiratory tract virus potentiation of bacterial adherence, colonization, and superinfection as a significant factor contributing to the pathogenesis of otitis media (OM). Influenza A and B viruses, adenovirus, and respiratory syncytial virus are the primary respiratory tract viruses associated with this disease. Investigations have established a dramatic increase in the development of experimental OM in chinchillas co-inoculated with influenza A virus and Streptococcus pneumoniae (Spn). The mechanism underlying this phenomenon was suggested to involve, in part, viral compromise of eustachian tube mucosal integrity and function. This study was designed to assess and compare the effect of adenovirus and influenza A virus infection on adherence, the kinetics of colonization, and invasion of the middle ear by Spn in the chinchilla model of OM. Cohorts were inoculated intranasally with adenovirus type 1 or influenza A virus, and then inoculated intranasally 7 days later with Spn 6A. All cohorts were observed over a 14-day period after challenge with Spn, and the incidence and severity of OM were assessed by several methods, including culture of the nasopharynx and middle ear effusions. The data indicated that influenza A virus promotes a significant increase in nasopharyngeal colonization by Spn, an increased incidence and severity of OM, and a sustained presence of Spn in the effusions. Adenovirus infection, however, did not enhance colonization by Spn or result in an increased incidence or severity of OM.