Elevated carbon dioxide enhances the growth and reduces the antifungal susceptibility of <i>Histoplasma capsulatum</i>

<i>Histoplasma capsulatum</i> is a thermally dimorphic fungal pathogen. It causes approximately 500,000 infections annually in the United States. <i>Histoplasma</i> is present as avirulent mycelia in the soil and transforms into pathogenic yeasts at the human body temperature upon inhalation. This elevated temperature triggers the expression of many virulence factors that enable <i>Histoplasma</i> yeasts to survive and proliferate within immune cells (i.e., macrophages) in the human lungs. In addition to elevated temperature, <i>Histoplasma</i> yeasts also experience other environmental changes within the mammalian host, such as elevated carbon dioxide (CO₂) (ambient air vs host tissues) during infection. However, the impact of elevated CO₂ on <i>Histoplasma</i> yeasts remains completely unknown. In this study, our results showed that elevated CO₂ enhanced <i>Histoplasma</i>'s growth, particularly increasing its ability to utilize certain amino acids (e.g., alanine) as the sole carbon source. We also found that elevated CO₂ reduced <i>Histoplasma</i>'s susceptibility to antifungals <i>in vitro</i>. <i>Histoplasma</i>'s enhanced growth and reduced antifungal susceptibility under elevated CO₂ were not pH-dependent. Our findings suggest that the elevated CO₂ within mammalian hosts could potentially enhance <i>Histoplasma</i>'s virulence. Future antifungal susceptibility tests for <i>Histoplasma</i> should be performed at 5% CO₂ for clinically relevant results.IMPORTANCEThe fungal pathogen <i>Histoplasma capsulatum</i> lives in the soil. <i>Histoplasma</i> spores can be inhaled and cause respiratory infections. The human body is vastly different from the soil. One of the major differences is the carbon dioxide (CO₂) concentration (0.04% in the ambient air vs 5% or above in the human body). Therefore, it is important to understand the impact of elevated CO₂ on <i>Histoplasma</i>. We found that elevated CO₂ promotes <i>Histoplasma</i>'s growth, suggesting that elevated CO₂ could potentially enhance <i>Histoplasma</i>'s virulence during infection. Our results showed that elevated CO₂ reduces <i>Histoplasma</i>'s antifungal susceptibility, suggesting that antifungal susceptibility tests for <i>Histoplasma</i> should be performed at elevated CO₂ for clinically relevant results.