Defective viral genomes (DVGs), which are generated by the viral polymerase in error during RNA replication, can trigger innate immunity and are implicated in altering the clinical outcome of infection. Here, we investigated the impact of DVGs on innate immunity and pathogenicity in a BALB/c mouse model of influenza virus infection. We generated stocks of influenza viruses containing the internal genes of an H5N1 virus that contain different levels of DVGs (indicated by different genome to PFU ratios). In lung epithelial cells, the high DVG stock was immunostimulatory at early time points post infection. DVGs were amplified during virus replication in myeloid immune cells and triggered pro-inflammatory cytokine production. In the mouse model, infection with the different virus stocks produced divergent outcomes. The high DVG stock induced an early type I IFN resonse that limited viral replication in the lungs resulting in minimal weight loss. In contrast, the virus stock with low levels of DVGs replicated to high
titres and amplified DVGs over time resulting in elevated pro-inflammatory cytokines accompanied by rapid weight loss and increased morbidity and mortality. Our results suggest that the timing and levels of immunostimulatory DVGs generated duringinfection contribute to H5N1 pathogenesis.