With the concurrent circulations of SARS-CoV-2 omicron and influenza A viruses in the community, there is evidence showing co-infection with both viruses. However, disease severity may vary due to the complex immunity landscape of the patients and the neutralizing antibody waning status. The intrinsic dynamic relationship and pathological significance for such co-infections remain largely unknown. The replication kinetics and innate immune responses from the co-infections of SARS-CoV-2 (Omicron BA.1 and D614G variant) and influenza A viruses (pandemic H1N1, seasonal H3N2 and highly pathogenic avian H5N1) were characterized in human respiratory tissue explants, human airway, and alveolar epithelial cells. SARS-CoV-2 reduced the replication of influenza A viruses, but not vice versa, during co-infections in human bronchial tissues and airway epithelial cells. In lung tissues, the co-infections showed minimal effects on each other, but the viral replications of the two viruses were mutually reduced except for H1N1pdm in the alveolar epithelial cells irrespective of the enhancement of the ACE2 receptor. Notably, the co-infections showed a significant upregulation of the innate immune responses of SARS-CoV-2 in comparison to single infections in both respiratory epithelial cells, suggesting that co-infections of influenza A viruses potentially lead to more severe damage to the host than SARS-CoV-2 single infections.

Recent studies have reported a genotype 4 (G4) reassortant Eurasian avian-like (EA) H1N1 virus in swine, demonstrating a potential pandemic threat in humans. Here, we have compared the tropism, replication competence and pro-inflammatory cytokine and chemokine induction of the two G4 EA H1N1 strains in parallel with 2009 pandemic H1N1 (H1N1/pdm/09) and A/Quail/HK/G1/1997 H9N2 (G1) using ex vivo culture of the human respiratory tract and in vitro culture of human peripheral blood-derived macrophages. Our results showed that G4 strains could replicate in ex vivo cultures of human lung and bronchus with a similar replication competence to H1N1/pdm/09. The cytokine induction levels of G4 were similar to H1N1/pdm/09 in macrophages. Taken together, we could extrapolate that the G4 EA H1N1 swine influenza may pose a notable public health threat towards human and should not underestimate this threat.

Congenital lung malformation (CLMs) is a series of developmental lung abnormalities that may persist into adulthood. Some centres would advocate an observation approach rather than opting for surgical excision. However, the potential impact of environmental factors, particularly cigarette smoke exposure, on CLM-affected lung tissue remains poorly understood. This study aimed to evaluate the effects of cigarette smoke extract (CSE) on paired normal and diseased airway organoids (AOs) derived from pediatric lung tissue diagnosed as intralobar sequestration, using N-[N-(3,5-Difluorophenacetyl)-L-alanyl]-S-phenylglycine t-butyl ester (DAPT), a γ-secretase inhibitor, to modulate airway epithelial differentiation. AOs were established from paired normal and diseased regions of pediatric lung tissue. Organoids were cultured under two conditions: without DAPT or with DAPT pretreatment, followed by 4% CSE exposure. Mucus viscosity, MUC5AC protein expression, PAS staining, and RT-qPCR analysis of airway epithelial cell markers, tight junction proteins (E-cadherin, ZO-1) , and proinflammatory cytokines were measured. Disease-derived AOs exhibited higher mucus secretion compared to normal tissue-derived AOs. CSE exposure enhanced mucus viscosity and MUC5AC expression in both groups, with more pronounced effects in diseased organoids. CSE-induced upregulation of mucins and pro-inflammatory cytokines was particularly evident in disease-derived organoids without DAPT treatment. DAPT pretreatment attenuated these CSE-induced changes, while tight junction protein expression remained stable across conditions. Our findings demonstrate that diseased airway organoids show enhanced susceptibility to cigarette-smoke -induced mucus hypersecretion and inflammatory responses, suggesting that patients with congenital airway malformations may be particularly vulnerable to the effects of cigarette smoke exposure.

The emergence of animal influenza viruses circulating in poultry and human populations poses a significant public health threat, yet current risk assessment tools that connect surveillance data to human transmission risk and disease severity are lacking. To address this, we employed a semi-quantitative approach to analyze virus tropism and replication competence, conducting risk assessments of influenza and coronavirus adaptation to human transmission in an ex vivo model, and evaluating virus-induced impairment of alveolar fluid clearance (AFC) in vitro as a correlation of disease severity. Our results showed that seasonal influenza A H1N1, H3N2, influenza B, MERS-CoV, and SARS-CoV exhibited productive viral replication and tissue infection in bronchial tissues, whereas wild bird surveillance isolates such as H5N3 and H7N1 showed minimal replication when compared to pandemic H1N1 and highly pathogenic avian influenza (HPAI) H5N1. Notably, differential lung viral replication and tissue tropism were detected for H5N6 and H9N2. HPAI H5N1, H7N9, MERS-CoV, and SARS-CoV caused more severe AFC impairment than seasonal H1N1, H3N2, and influenza B viruses, correlating with their clinical severity. Overall, these findings revealed an important association between viral tropism and human transmissibility in ex vivo explants, as well as the impairment of AFC in vitro , which aligns with the clinical manifestations of disease severity across different viral strains.