5' copyback defective viral genomes are major component in clinical and non-clinical influenza samples

Clinical samples from people with influenza disease have been analyzed to assess the presence and abundance of Defective Viral Genomes (DVGs), but these have not been assessed using the same bioinformatic pipeline. The type of DVG most described for influenza infections (deletion DVGs) differs from the most commonly described DVGs from non-segmented negative stranded viruses (5' copyback). This could be attributed to either differences between viruses or the tools used to detect and characterize DVGs. Here we analyze several NGS datasets from people infected with different types of influenza virus using the same bioinformatic pipeline. We observe that 5' copyback DVGs are prevalent in all human clinical samples but not in the cultured samples. To address this discrepancy between clinical and laboratory cultures, we infected cell culture and ferrets with an H5N8 influenza A virus (FLUAV) and analyzed the DVG composition. The results demonstrate that the DVG population is skewed toward 5' copyback DVGs in the in vivo infections and deletion DVGs in the in vitro infections. This demonstrates that there are differences in vivo genome production and in vitro genome production, and this has implications for how the role of DVGs in clinical disease is studied. We also investigate the role the host cofactor ANP32B has in DVG production.