Macrophages inhibit Aspergillus fumigatus germination and neutrophil-mediated fungal killing

In immunocompromised individuals, Aspergillus fumigatus causes invasive fungal disease that is often difficult to treat. Exactly how immune mechanisms control A. fumigatus in immunocompetent individuals remains unclear. Here, we use transparent zebrafish larvae to visualize and quantify neutrophil and macrophage behaviors in response to different A. fumigatus strains. We find that macrophages form dense clusters around spores, establishing a protective niche for fungal survival. Macrophages exert these protective effects by inhibiting fungal germination, thereby inhibiting subsequent neutrophil recruitment and neutrophil-mediated killing. Germination directly drives fungal clearance as faster-growing CEA10-derived strains are killed better in vivo than slower-growing Af293-derived strains. Additionally, a CEA10 pyrG-deficient strain with impaired germination is cleared less effectively by neutrophils. Host inflammatory activation through Myd88 is required for killing of a CEA10-derived strain but not sufficient for killing of an Af293-derived strain, further demonstrating the role of fungal-intrinsic differences in the ability of a host to clear an infection. Altogether, we describe a new role for macrophages in the persistence of A. fumigatus and highlight the ability of different A. fumigatus strains to adopt diverse modes of virulence.
Author summary Immunocompromised patients are susceptible to invasive fungal infections, including aspergillosis. However, healthy humans inhale spores of the fungus Aspergillus fumigatus from the environment every day without becoming sick, and how the immune system clears this infection is still obscure. Additionally, there are many different strains of A. fumigatus, and whether the pathogenesis of these different strains varies is also largely unknown. To investigate these questions, we infected larval zebrafish with A. fumigatus spores derived from two genetically diverse strains. Larval zebrafish allow for visualization of fungal growth and innate immune cell behavior in live, intact animals. We find that differences in the rate of growth between strains directly affect fungal persistence. In both wild-type and macrophage-deficient zebrafish larvae, a fast-germinating strain is actually cleared better than a slow-germinating strain. This fungal killing is driven primarily by neutrophils while macrophages promote fungal persistence by inhibiting spore germination. Our experiments underline different mechanisms of virulence that pathogens can utilize—rapid growth versus dormancy and persistence—and inform future strategies for fighting fungal infections in susceptible immunocompromised patients.