Fig. 3. Fe accumulation mediates DAB oxidation at CWAs. (A) H₂O₂ accumulation in CWAs revealed by DAB staining 24 hpi. (B) Double staining for H₂O₂ and Fe³⁺ (blue) 24 hpi. (C) Double staining for H₂O₂ and Fe³⁺ at a primary germ tube-associated CWA 5 hpi. (D) Vesicle-like bodies within a CWA double stained for H₂O₂ and Fe³⁺ 24 hpi. (E) Wheat leaves with (right) or without (left) pretreatment of 1.5 mM DFO were stained using DAB and photographed 24 hpi. (F-H) DFO blocks H₂O₂ generation at CWAs. Upper panel: without DFO pretreatment. Lower panel: with 1.5 mM DFO pretreatment. (F) DFO blocks H₂O₂ generation at appressorial germ tube-associated CWAs. (G) DFO blocks H₂O₂ generation at primary germ tube-associated CWAs but not cytoplasmic aggregation (arrowhead). (H) Pretreatment with DFO does not affect nuclear migration (arrows) at CWAs. agt, appressorial germ tube; c, conidium; N, host nucleus; pgt, primary germ tube. Scale bar, 20 µm.