Human Neutrophils Are Primed by Chemoattractant Gradients for Blocking the Growth ofAspergillus fumigatus

Abstract

The contribution of human neutrophils to the protection against fungal infections by Aspergillus fumigatus is essential but not fully understood. Whereas healthy people can inhale spores of A. fumigatus without developing disease, neutropenic patients and those receiving immunosuppressive drugs have a higher incidence of invasive fungal infections. To study the role of neutrophils in protection against A. fumigatus infections, we developed an in vitro assay in which the interactions between human neutrophils and A. fumigatus were observed in real time, at single-cell resolution, in precisely controlled conditions. We measured the outcomes of neutrophil-fungus interactions and found that human neutrophils have a limited ability to migrate toward A. fumigatus and block the growth of A. fumigatus conidia (proportion with growth blocked, 69%). The blocking ability of human neutrophils increased to 85.1% when they were stimulated by uniform concentrations of fMLP and was enhanced further, to 99.4%, in the presence of chemoattractant gradients. Neutrophils from patients receiving immunosuppressive treatment after transplantation were less effective against the fungus than those from healthy donors, and broader heterogeneity exists between patients, compared with healthy individuals. Further studies using this microfluidic platform will help understand the relevance of innate immune deficiencies responsible for the higher risk of fungal infections in patients with immunosuppressive disease.

Neutrophil Interactions Stimulate Evasive Hyphal Branching by Aspergillus fumigatus

“…fumigatus growth [27]. Introduction of an fMLP gradient in the current study prior to delivery of neutrophils resulted in increased recruitment of neutrophils to sites of interaction (S1B Fig), an increase in hyphal exposure to NETs (Fig 4A), and increased fungal branching (Fig 4B).…”

“…Although a direct effect of immunosuppressant therapy on neutrophil activity remains controversial, our previous studies indicate that neutrophils from these patients exhibit reduced activity against A . fumigatus [27]. Prophylactic treatment of immunosuppressed patients with broad-spectrum antibiotics, aimed at reducing risk of bacterial infection, is standard practice [52].…”

“…Recent work has begun to explore the applicability of these devices to investigating host-pathogen interaction [26,27]. For example, work by our group recently demonstrated that migration along chemotactic gradients acts to prime neutrophils for antifungal activity against A .…”

“…For example, work by our group recently demonstrated that migration along chemotactic gradients acts to prime neutrophils for antifungal activity against A . fumigatus [27], while others have used microfluidic techniques to identify immunosuppressive fungal metabolites [28]. For microbiologists, microfluidics has provided a useful platform for imaging active Candida hyphal growth at high resolution, revealing that internal polarity protein complexes are asymmetrically skewed toward softer substratum [29].…”

Innate Lung Defense during Invasive Aspergillosis: New Mechanisms

“…In contrast, the killing of conidia is achieved by nonoxidative lactoferrin-mediated iron sequestration. An additional study has recently examined interactions between human neutrophils and A. fumigatus conidia and hyphae in the presence of various chemoattractants and neutrophil activators [13] . Using a nanoliter culture well system, this study demonstrated that less than a third of conidia form hyphae when incubated with pure neutrophils, an amount that continues to decrease when the cocultures contain a uniform amount of leukotriene B4, but not fMLP.…”