Dancer, P. | bioRxiv Science

bioRxiv2022

Mutual inhibition of airway epithelial responses supports viral and fungal co-pathogenesis during coinfection

Awareness that fungal coinfection complicates viral respiratory infections causing worse disease outcome has recently emerged. The environmental fungus Aspergillus fumigatus (Af) has been reported as the main driver of fungal coinfection in patients suffering from viral infections caused by Cytomegalovirus, Influenza or more recently SARS-CoV2. The airway epithelium is the first common point of contact between inhaled pathogens and the host. Aberrant airway epithelial cell (AEC) responses against fungal challenge have been described in patients susceptible to aspergillosis. Therefore, it is likely that a dysregulation of AEC responses during fungal-viral coinfection represents a potent driver for the development of fungal disease. Here we used an in vitro model of Af-viral infection of AECs to determine outcomes of spore internalisation, killing and viral replication during coinfection. Our data indicate that viral stimulation, while boosting Af uptake by AECs, limits Af spore killing by those cells, favouring fungal persistence and growth. Type I viral-induced interferon release was significantly decreased in the presence of Af hyphal forms suggesting a possible role of Af secreted factors in modulating viral pathogenicity. We next explored the impact of Af challenge in SARS-CoV2 replication within airway epithelial cells using nano-luciferase as a measure of viral replication. We found that Af increased SARS-CoV2 pathogenicity in a strain-dependent manner. Collectively, our findings demonstrate a mutual inhibition of antifungal and antiviral AEC responses during Af-viral coinfection and also suggest that some fungal factors might be key regulators of co-pathogenicity during in lung infection.

microbiology↗

bioRxiv2022

Epithelial uptake of Aspergillus fumigatus drives efficient fungal clearance in vivo and is aberrant in Chronic Obstructive Pulmonary Disease (COPD)

Hundreds of spores of the common mould Aspergillus fumigatus (Af) are inhaled daily by human beings, representing a constant, often fatal, threat to our respiratory health. The small size of Af spores suggest that interactions with Airway Epithelial Cells (AECs) are frequent and we and others have previously demonstrated that AECs are able to internalise Af spores. We thus hypothesised that Af spore uptake and killing by AECs is important for driving efficient fungal clearance in vivo and that defective spore uptake and killing would represent major risk factors for Aspergillus-related diseases. In order to test this, we utilised single-cell approaches based on Imaging Flow Cytometry (IFC) and live-cell microfluidic imaging to measure spore uptake and outcomes in vitro, in vivo and using primary human AECs. In vitro, viability of immortalised AECs was largely unaffected by Af uptake and AECs were able to significantly curtail the growth of internalised spores. Applying our approach directly to infected mouse lungs we demonstrated, for the first time, that Af spores are internalised and killed by AECs during whole animal infection, whereby only ~3% of internalised spores remained viable after 8 hours of co-incubation with murine AECs. Finally, in vitro analysis of primary human AECs from healthy and at-risk donors revealed significant alterations in the uptake and consequent outcomes in Chronic Obstructive Pulmonary Disease (COPD), whereby gorging COPD-derived AECs were unable to quell intracellular Af as efficiently as healthy primary AECs. We have thus demonstrated that AECs efficiently kill Af spores upon uptake in vivo and that this process is altered in COPD, a well-known risk factor for debilitating fungal lung disease, thereby suggesting that AECs critically contribute to the efficient clearance of inhaled Af spores and that dysregulation of curative AEC responses represents a potent driver of Aspergillus-related diseases.

microbiology↗