Introduction: Respiratory viruses (RVs) represent a significant global health threat. The prevalence of RVs has surged due to the relaxation of public health measures and reduction in non-pharmaceutical interventions in communal settings. RVs have the capacity to induce concurrent or sequential infections. This study aims to explore potential viral interactions and assess the efficacy of a broadly active antiviral drug against prevalent RVs, including Influenza (IAV), SARS-CoV-2, Respiratory Syncytial Virus (RSV), rhinovirus, human metapneumovirus (HMPV), and parainfluenza virus (PIV).
Methods: A biobank of primary bronchial epithelial cells (pBECs) obtained via bronchoscopy was established, and Air-liquid interface (ALI) cultures were subsequently developed. Single virus infections were performed using strains of SARS-CoV-2 (including ancestral, delta, omicron BA.1, and BA.5 variants), Influenza A virus (pdmH1N1 and H3N2 subtypes), clinical isolates of Respiratory Syncytial Virus (RSV) types A and B, as well as laboratory reference strains of rhinovirus, human metapneumovirus (HMPV), and parainfluenza virus (PIV) in the ALI cultures. Transepithelial electrical resistance (TEER), serving as a proxy for tissue integrity, and cellular damage, indicated by lactate dehydrogenase (LDH) levels, were monitored daily. Additionally, cytokine production and induction of interferon-stimulated genes (ISGs) were assessed at 24 hours and 48 hours post-infection.
Results: TEER measurements for all viruses reached their peak at 24 hours post-infection, likely attributed to increased mucus production. The growth kinetics of all viruses indicated peak titres ranging from 10^1.7 to 10^7.95 TCID50/mL. Notably, despite the decrease in TEER and increased virus growth, the ALI monolayer remained relatively intact up to ten days post-infection. Molnupiravir, an antiviral drug effective against SARS-CoV-2, IAV, and RSV, demonstrated dose-dependent inhibition of SARS-CoV-2 in a proof-of-concept experiment using Calu-3 ALI cells. Quantification of interferon responses (Type I and Type II) and levels of ISG induction following viral infections is underway through optimization of qPCR protocols. Current investigations are focused on characterizing sequential respiratory virus infections.
Conclusion: This study aims to offer valuable insights into viral interactions among epidemiologically relevant respiratory viruses (RVs) and the effects of molnupiravir on both single RV infections and sequential infections.