Persistent cutaneous phaeohyphomycosis caused by Brunneofusispora hyalina in a patient with a heterozygous CARD9 variant: the opportunistic potential of a newly described environmental fungus

Objectives This report documents the first isolation of rare dematiaceous fungus Brunneofusispora hyalina from a cutaneous lesion in a patient with heterozygous CARD9 variant. The study characterizes this novel pathogen and simulates infection and treatment processes to understand this opportunist and explore therapeutic strategies. Methods Direct microscopy and histopathology confirmed the fungal infection. By collecting morphological images of the isolate, assessing temperature-dependent growth, performing multi-locus and whole-genome sequencing, and conducting antifungal susceptibility testing, the new pathogen’s virulence and antifungal resistance were comprehensively analyzed. Using CARD9-deficient mice to establish infection models and administer drug treatments. Results The patient, confirmed by whole-exome sequencing to have a heterozygous CARD9 variant, was diagnosed with phaeohyphomycosis caused by B. hyalina through barcode sequencing. Meanwhile, functional annotation revealed metabolic versatility and enrichment in secondary metabolite pathways, while 395 PHI homologs were identified mainly from plant-pathogenic fungi. The strain exhibited abundant hyphal anastomosis and psychrotolerant growth. Antifungal susceptibility tests using modified agar dilution and broth dilution methods showed low MICs to voriconazole, posaconazole, itraconazole, and terbinafine, with values of 0.5-1 µg/mL, 0.5–2 µg/mL, 1–2 µg/mL, and 0.25–0.5 µg/mL, respectively. However, fluconazole showed a high MIC (128 µg/mL). In contrast, amphotericin B inhibitory concentrations differed substantially between two testing methods, with discrepancies exceeding three twofold dilution titers. In a CARD9 knockout mouse model, B. hyalina caused persistent inflammatory infections without mortality, whereas infection in wild-type mice had self-limiting symptoms. Despite six weeks of combined antifungal therapy, complete eradication of infection was not achieved. Conclusions B. hyalina is a new opportunistic fungus causing phaeohyphomycosis, particularly in individuals with CARD9 variants. Genomic analysis shows that B. hyalina shares many genes with plant pathogens and lacks specialized human virulence factors, suggesting that host immune vulnerability is key to infection. While antifungal drugs such as triazoles and terbinafine demonstrate low inhibitory concentrations in vitro , combination therapy in mouse models reduces fungal burden but does not completely clear the infection, indicating the necessity for prolonged treatment in immunocompromised hosts. Future exploration may focus on immune restoration and hyperthermia, which may complement antifungal therapy and improve outcomes.