Replication Dynamics, Pathogenicity, and Evolution of Influenza Viruses in Caco-2 Cells

Influenza virus is a major etiologic agent for respiratory illnesses, affecting 5-30% of global population annually, leading to high number of deaths and hospitalizations. Influenza virus primarily infects epithelial cells of respiratory tracts and cause respiratory and systemic symptoms. In considerable number of patients, infections with influenza viruses have been associated with gastrointestinal symptoms such as diarrhea, vomiting, and abdominal pain. Moreover, up to 20.6 % of influenza-infected patients shed the virus in feces. Therefore, direct infection of intestinal cells with the influenza virus is suspected, but the mechanism of this infection is not clear. This study examined the adaptation ability of and evolution of two influenza A subtypes (A/H3N2 and A/H1N1pdm 09) and one influenza B virus (B/Yamagata) in human Caucasian colon adenocarcinoma cells (Caco-2). The impact of these viruses in Caco-2 cells expression of cellular hormones and cytokines was assessed. All three influenza viruses used in this study were able to replicate in Caco-2 cells with H3N2 virus adapting faster than H1N1 virus and IBV. The expression of hormone and cytokines in Caco-2 cells was considerably different between the three viruses and among the passages, however a pattern of induction was observed at late phase of infection (48 hr post infection). Deep sequencing analysis revealed number of amino acid substitutions in HA protein of H3N2 and H1N1 viruses, these substitutions were mostly in the antigenic site. Moreover, studying the virus evolution at the quasispecies level based on HA protein revealed that H3N2 and H1N1 harbored more diverse virus populations when compared to IBV, indicating their higher evolution within Caco-2 cells. The findings of this study indicate differential replication, pathogenicity and evolution of H1N1 and H3N2 viruses in intestinal cells. This study was limited to only one cell line and few subsets of viruses and, hence, a further larger study including in vivo experiments is needed to further explain the gastrointestinal complication of influenza infection.