Excess airway iron during severe SARS-CoV-2 infection impairs neutrophil antifungal activity enabling 2° mold infection

Abstract COVID-19-Associated Pulmonary Aspergillosis (CAPA) is an invasive 2° mold infection caused by Aspergillus species affecting ~5% of patients with severe SARS-CoV-2 (SARS-2) infection. In this study, we hypothesized that excess airway iron released during SARS-2 infection promotes 2° infection via enhancement of fungal growth and impairment of neutrophil antifungal activity. We show that SARS-2 bronchoaveolar lavage samples (n=48) have higher levels of total iron, ferric iron, and oxyhemoglobin relative to bacterial pneumonia (n=40). Likewise, FFPE lung tissues from SARS-2 infection exhibited increased extracellular ferric iron, ferrous iron, ferritin, and hemoglobin. We show that both iron and hemoglobin enhance the growth rate of Aspergillus fumigatus via siderophore and reductive iron assimilation dependent mechanism. Utilizing lung epithelial cell transmigration model, we show that iron enhances neutrophil intracellular ROS production resulting in enhanced conidiacidal activity. In contrast, transmigrated neutrophils exposed to both heme and hemoglobin exhibit decreased ROS production and NET formation with significant impairment in hyphal killing. When both neutrophils and fungi are co-cultured with iron, heme, or hemoglobin, fungi consistently win the battle characterized by uncontrolled hyphal proliferation. These data identify both microbial and host mechanisms which may be targeted pharmacologically to mitigate the iron-dependent growth of 2° mold infection.