Wheat cells show positional responses to invasive Zymoseptoria tritici

The stomatal complex of grasses consists of two guard cells and two adjacent subsidiary cells that cooperate during stomatal closure. Zymoseptoria tritici , the main causal agent of Septoria tritici blotch in wheat, enters the host via stomata. Here we test the hypothesis that the stomatal complex shows focused sub-cellular responses to invading Z. tritici hyphae. We have combined live-cell transmission light microscopy, immunofluorescence and CRS microscopy to identify cell wall modifications triggered by hyphal invasion. Furthermore, we have used confocal fluorescence microscopy and automated quantitative image analysis to assess whether host cells respond to hyphae through spatial redistribution of organelles. We find that subsidiary cells construct papillae that are accurately aligned with hyphal position even when hyphae are occluded by guard cells. These are distinct from those induced by powdery mildew, with callose restricted to a crust that surrounds content with a high-amplitude Raman signal in the CH-band. Peroxisome populations in subsidiary cells show distributions with modes weakly correlated with hyphal position but do not differ significantly between compatible and incompatible interactions. Our data suggest local changes to cell wall architecture and focal accumulation of organelles in subsidiary cells could play roles in crop defence during host leaf penetration by Z. tritici . Molecular strategies to amplify these responses may provide novel routes for crop protection. ### Competing Interest Statement The authors have declared no competing interest.