Transdermal *Aspergillus oryzae* induces mild, type 2-skewed immune response and memory in mouse skin lymph nodes

The immune response to non-pathogenic fungi entering the skin is poorly understood, despite their ubiquitous presence. This knowledge gap is critical, especially when considering the severe systemic infections caused by pathogenic fungi like Aspergillus fumigatus in immunocompromised individuals. Understanding how non-pathogenic fungi like Aspergillus oryzae (Ao) interact with the immune system could reveal novel immunomodulatory strategies, potentially leveraging components like beta-glucans to influence innate and adaptive immunity.

Mice were subcutaneously inoculated with Ao conidia, and skin-draining lymph nodes were harvested over time. Immune cell subsets were evaluated via flow cytometry. Cytokine expression (including IFN-γ, IL-4, IL-6, IL-10, IL-12, IL-17, TGF-β, TNF-α) was assessed by quantitative reverse transcription-PCR at 4 or 7 days post-inoculation. Antigen-specific antibody production was measured by ELISA in sera collected at day -1, 14, and 35 after immunization with Ao and/or ovalbumin. Lymph node structural reorganization was visualized using fluorescence immunohistostaining, and active cutaneous anaphylaxis was assessed for allergic response impact.

Transdermal entry of Aspergillus oryzae conidia led to a marked enlargement of skin-draining lymph nodes and increased immune cell numbers within days. This response was characterized by a marked increase in activated B cells and type 2 resident dendritic cells. Ao cell wall components and purified β-glucans partially reproduced these responses, supported by the Ao-dependent downregulation of Dectin-1 expression on dendritic cells. However, live Ao elicited the most pronounced lymph node response, as heat inactivation and the cell wall fraction clearly attenuated it, suggesting other components are also necessary. Ao induced a relatively mild response in lymph nodes, characterized by a marked increase in IL-4 expression, while other cytokines were suppressed or unaltered. In the long term, Ao entry elicited immune memory with antibody production specific to conidial proteins and β-glucan.