A chitin-like component masks the immunorecognition of β-glucans on sclerotic cells by Dectin-1.

<p>(A1–A4) The binding of murine-derived Dectin-1 to the saprophytic <i>F. pedrosoi</i> in PDB before (A1) and after (A4) β-glucanase treatment, or to the 30-day (A2) and 50-day (A3) induced cultures containing sclerotic cells in ATCC medium 830 was detected by PE-anti-murine Dectin-1 mAb using confocal microscope. (B1 and B2) The binding of human-derived Dectin-1 to the induced cultures containing sclerotic cells was detected by PE-anti-human Dectin-1 mAb using confocal microscope. (C) The binding of murine-derived mannose receptor (MR) to the induced cultures containing sclerotic cells and chlamydospores was detected by FITC-anti-murine MR using confocal microscope. (D) FITC-WGA was used to detect the distribution of chitin-like component on the induced cultures containing sclerotic cells before (D1) and after (D2) Chitinase treatment using confocal microscope. (E1 and E2) The binding capacity of FITC-WGA to the saprophytic <i>F. pedrosoi</i> or to the induced cultures before and after chitinase treatment was detected using flow cytometry assay, and represented as the Mean Fluorescence Intensity (MFI). The induced cultures without any treatment were set as blank control. (F1 and F2) SEM was used to observe the surface ultra-structure of saprophytic hypha (F1) and in vitro-transformed sclerotic cells (F2) in the presence or absence of chitinase pretreatment. SEM, Scale bar = 4 µm (left) or 500 nm (middle and right). (G1 and G2) The binding of murine-derived Dectin-1 to the induced cultures containing sclerotic cells with chitinase pretreatment was detected by confocal microscope. (A-D, G) The transformed sclerotic cells and chlamydospores were indicated by red arrows. (H1 and H2) The binding capacity of murine-derived Dectin-1 to the saprophytic <i>F. pedrosoi</i> before and after β-glucanase treatment, or to the induced cultures before and after chitinase treatment was detected by indirect immunofluorescence assay using flow cytometer, and represented as the Mean Fluorescence Intensity (MFI). The fungal cells only incubated with PE-conjugated anti-murine Dectin-1 mAb were set as blank control. (E2 and H2) Data are shown as mean ± SEM from three individual experiments performed in triplicates, and significance tested using LSD-t test.