CBM progression in a Zymosan-induced inflammation model.

<p>After 15 days post infection with fungal propagules (FP), animals were treated intra lesionally (i.l.) in the footpad with 20μl of a suspension containing 5 mg/ml of zymosan (ZYM) or PBS, until 15 days post treatment start (d.p.t) (A). Animals treated with ZYM displayed intense inflammatory response up to 30 days post infection (d.p.i) (B). Animals facing prolonged inflammation showed no reduction in fungal load over time, as observed for those animals treated with PBS (C). **P<0.01 and ***P<0.001.</p

Quantification of <i>F</i>. <i>pedrosoi</i> fungal cells in the tissue and production of pro-inflammatory cytokines in the course of murine CBM.

<p>Fungal cells were counted on twenty fields chosen at random in histopathological slides with the aid of a counting reticle (A-D). In all groups muriform cells (red arrows) could be identified after 15 days after infection. At the same time, hyphal fragments (brown arrow) were also present in animals infected either with hyphae (FH) (B) or muriform cells (MC) (C). Few conidia (black arrow) are observed after 15 days of infection in animals infected with <i>F</i>. <i>pedrosoi</i> conidia (FC) (A). Cell counts were expressed as cells per mm² of injured tissue. 1000x magnification (A-D). After 15 days of infection, high levels of TNF-α (E), IL-1β (F), IL-6 (G) and MCP-1 (H) were observed in groups with higher numbers of hyphae and muriform cells in the tissue. Cytokine production was measured by ELISA from homogenized footpad tissue. *P<0.05, **P<0.01 and ***P<0.001 compared to FC group.</p

Phagocytosis gene expression in peritoneal macrophages co-culturing with conidia.

<p>Peritoneal macrophages (PM) co-culturing with conidia (FC) did not induce much gene expression related to phagocytosis.</p

<i>In vitro</i> cytokine and chemokine production.

<p>The production of cytokines and chemokines by macrophage in conidia (FC) or muriform cell (MC) co-culture supernatants was assessed by ELISA (A-E). NO₂ concentration in culture supernatants was used as an indicator of NO generation and measured using Griess reagent (F). High levels of TNF-α (A), IL-1β (B) and IL-6 (C) were observed in co-culture with muriform cells, but not with conidia. To assess IL-1β production after 24 hours, peritoneal macrophages required LPS co-stimulation (B). Muriform cells also induced higher levels of MCP-1 after 48h when compared to macrophages infected with conidia (E). Production of IL-12 (D) and NO₂ (F) was strongly inhibited by muriform cells. **P<0.01 and ***P<0.001.</p

Progression of murine chromoblastomycosis induced by different <i>F</i>. <i>pedrosoi</i> fungal forms.

<p>BALB/c mice were infected in the footpad with 1x10⁶ conidia (FC), hyphae (FH), muriform cells (MC) or a combination of hyphal fragments and conidia (FP) in the ratio of 3:1 (A). Morphometric (B) and CFU data (C) showed a fast clearance of inoculated conidia, while infection with MCs was reflected in the persistence of the fungus in the tissue up to 45 days after infection. 400x magnification (A). *P<0.05 and ***P<0.001 compared to FH group.</p