Neutrophil Interactions Stimulate Evasive Hyphal Branching by Aspergillus fumigatus.

Felix Ellett,Galit H Frydman,Caroline N Jones,Daniel Irimia,Julianne Jorgensen,Xiaorong Lin

doi:10.1371/journal.ppat.1006154

Abstract

Invasive aspergillosis (IA), primarily caused by Aspergillus fumigatus, is an opportunistic fungal infection predominantly affecting immunocompromised and neutropenic patients that is difficult to treat and results in high mortality. Investigations of neutrophil-hypha interaction in vitro and in animal models of IA are limited by lack of temporal and spatial control over interactions. This study presents a new approach for studying neutrophil-hypha interaction at single cell resolution over time, which revealed an evasive fungal behavior triggered by interaction with neutrophils: Interacting hyphae performed de novo tip formation to generate new hyphal branches, allowing the fungi to avoid the interaction point and continue invasive growth. Induction of this mechanism was independent of neutrophil NADPH oxidase activity and neutrophil extracellular trap (NET) formation, but could be phenocopied by iron chelation and mechanical or physiological stalling of hyphal tip extension. The consequence of branch induction upon interaction outcome depends on the number and activity of neutrophils available: In the presence of sufficient neutrophils branching makes hyphae more vulnerable to destruction, while in the presence of limited neutrophils the interaction increases the number of hyphal tips, potentially making the infection more aggressive. This has direct implications for infections in neutrophil-deficient patients and opens new avenues for treatments targeting fungal branching.

Highlights

Aspergillus fumigatus is a filamentous, conidiating fungus ubiquitous to the environment
Conidia grow into elongated hyphal structures, which present a unique challenge for the cellular immune system: Hyphae cannot be phagocytosed by immune cells, requiring deployment of extracellular antifungal mechanisms such as degranulation [5], production of extracellular reactive oxygen species (ROS) [6], secretion of ion-sequestering proteins such as Calprotectin [7] and Lactoferrin [8], and production of neutrophil extracellular traps (NETs) [9,10,11]
We find that A. fumigatus branching after interaction with human neutrophils is independent of neutrophil NADPH oxidase activity and NETosis, but can be phenocopied by iron chelation and mechanical or physiological stalling of hyphal tip extension

Summary

Introduction

Aspergillus fumigatus is a filamentous, conidiating fungus ubiquitous to the environment. It has been generally assumed that controlled branching enhances fungal invasion, as strains that exhibit perturbed branching behavior and irregular growth polarity exhibit reduced virulence in vivo [17,18,19]. Perturbed branching in these mutant strains is driven by altered ROS production or localization [20,21], calcium signalling [18,22], hypoxic stress responses [19], or cell wall synthesis pathways [23,24]. The study of host-hyphae interactions in vitro and in animal models of IA is limited by lack of control over the interactions and lack of methods to study these interactions at single cell resolution

Methods

Results

Discussion

Conclusion