Andres J. Gonzalez, Emmanuel C. Ijezie, Onesmo B. Balemba, Tanya A. Miura

Influenza viruses and rhinoviruses are responsible for a large number of acute respiratory viral infections in human populations and are detected as co-pathogens within hosts. Clinical and epidemiological studies suggest that co-infection by rhinovirus and influenza virus may reduce disease severity and that they may also interfere with each other's spread within a host population. To determine how co-infection by these two unrelated respiratory viruses affects pathogenesis, we established a mouse model using a minor serogroup rhinovirus (RV1B) and mouse-adapted influenza A virus (PR8). Infection of mice with RV1B two days before PR8 reduced the severity of infection by a low or medium, but not high, dose of PR8. Disease attenuation was associated with an early inflammatory response in the lungs and enhanced clearance of PR8. However, co-infection by RV1B did not reduce PR8 viral loads early in infection or inhibit replication of PR8 within respiratory epithelia or in vitro Inflammation in co-infected mice remained focal, in comparison to diffuse inflammation and damage in the lungs of mice infected by PR8. The timing of RV1B co-infection was a critical determinant of protection, suggesting that sufficient time is needed to induce this response. Finally, disease attenuation was not unique to RV1B: dose-dependent co-infection by a murine coronavirus (MHV-1) also reduced the severity of PR8 infection. Unlike RV1B, co-infection with MHV-1 reduced early PR8 replication, which was associated with up-regulation of IFN-β expression. This model is critical for understanding the mechanisms responsible for influenza disease attenuation during co-infection by unrelated respiratory viruses.Importance Viral infections in the respiratory tract can cause severe disease and are responsible for a majority of pediatric hospitalizations. Molecular diagnostics have revealed that approximately 20% of these patients are infected by more than one unrelated viral pathogen. To understand how viral co-infection affects disease severity, we inoculated mice with a mild viral pathogen (rhinovirus or murine coronavirus) followed two days later by a virulent viral pathogen (influenza A virus). This model demonstrated that rhinovirus can reduce the severity of influenza A virus, which corresponded with an early but controlled inflammatory response in the lungs and early clearance of influenza A virus. We further determined the dose and timing parameters that were important for effective disease attenuation and showed that influenza disease is also reduced by co-infection with a murine coronavirus. These findings demonstrate that co-infecting viruses can alter immune responses and pathogenesis in the respiratory tract.