3' Plasma Membrane Phosphoinositides Sustain ROS Production at Persistent Non-canonical Phagocytic Cups that Promote Macrophage Control of Aspergillus fumigatus Hyphae

Aspergillus fumigatus is the leading cause of aspergillosis in immunocompromised individuals. Because A. fumigatus hyphae rapidly outgrow macrophages, they are generally thought to evade macrophage-mediated control once they exceed the size limits of phagocytosis, enabling invasive growth within host tissues. Here, we show that macrophages circumvent this limitation by capturing hyphal tips within persistent tubular phagocytic cups. Rather than resolving into complete phagosomes or disengaging from their targets, these long-lived open compartments maintain key features of phagocytic cups while progressively acquiring characteristics of maturing phagosomes. Their persistence sustains localized NOX2-dependent reactive oxygen species production, leading to hyphal-tip damage and restriction of fungal growth. Sustained PI(3,4)P2 signaling downstream of class I PI3K promotes prolonged NOX2 activity and is required for efficient hyphal restriction. Together, our findings reveal a previously unrecognized antifungal mechanism in which persistent phagocytic cups transform incomplete phagocytosis into sustained antimicrobial activity, extending macrophage control beyond the physical limits of engulfment.