Neutrophil extracellular traps (NETs) and Th-2 dominant immune responses in chronic granulomatous chromobalstomycosis.

Chromoblastomycosis (CBM), a chronic, granulomatous, suppurative mycosis of the skin and subcutaneous tissue, is caused by several dematiaceous fungi. The formation of granulomas, tissue proliferation, and fibrosis in response to these pathogenic fungi is believed to be intricately linked to host immunity. To understand this complex interaction, we conducted a comprehensive analysis of immune cell infiltrates, neutrophil extracellular traps (NETs) formation, and the fibrosis mechanism in 20 CBM lesion biopsies using immunohistochemical and immunofluorescence staining methods. The results revealed a significant infiltration of mixed inflammatory cells in CBM granulomas, prominently featuring a substantial presence of Th2 cells and M2 macrophages. These cells appeared to contribute to the production of collagen I and III in the late fibrosis mechanism, as well as NETs formation. The abundance of Th2 cytokines may act as a factor promoting the bias of macrophage differentiation toward M2, which hinders efficient fungal clearance while accelerates the proliferation of fibrous tissue. Furthermore, the expression of IL-17 was noted to recruit neutrophils, facilitating subsequent NETs formation within CBM granulomas to impede the spread of sclerotic cells. Understanding of these immune mechanisms holds promise for identifying therapeutic targets for managing chronic granulomatous CBM.

Deletion C-terminal thioesterase abolishes melanin biosynthesis, affects metabolism and reduces the pathogenesis of Fonsecaea monophora.

Dematiaceous Fonsecaea monophora is one of the major pathogens of chromoblastomycosis. It has been well established that melanization is catalyzed by the type I polyketide synthase (PKS) in F. monophora. Multidomain protein Type I PKS is encoded by six genes, in which the last enzyme thioesterase (TE) catalyzes the cyclization and releases polyketide. Two PKS genes AYO21_03016 (pks1) and AYO21_10638 have been found in F. monophora and both PKS loci have the same gene arrangement but the TE domain in AYO21_10638 is truncated at 3'- end. TE may be the key enzyme to maintain the function of pks1. To test this hypothesis, we constructed a 3'-end 500 bp deletion mutant of AYO21_03016 (Δpks1-TE-C500) and its complemented strain. We profiled metabolome of this mutant and analyzed the consequences of impaired metabolism in this mutant by fungal growth in vitro and by pathogenesis in vivo. Compared with wild-type strain, we found that the mutant repressed pks1 expression and other 5 genes expression levels were reduced by more than 50%, perhaps leading to a corresponding melanin loss. The mutant also reduced sporulation and delayed germination, became vulnerable to various environmental stresses and was less resistance to macrophage or neutrophil killings in vitro, and less virulence in mice footpad model. Metabolomic analysis indicated that many metabolites were remarkably affected in Δpks1-TE-C500, in particular, an increased nicotinamide and antioxidant glutathione. In conclusion, we confirmed the crucial role of C-terminal TE in maintaining fully function of pks1 in F. monophora. Deletion of TE negatively impacts on the synthesis of melanin and metabolites that eventually affect growth and virulence of F. monophora. Any potential inhibitor of TE then could be a novel antifungal target for drug development.