A delayed-melting cryomicroneedle system for probiotic delivery against invasive Candida albicans infection.

The resilience of Candida albicans (C. albicans) biofilms against conventional antifungals necessitates innovative therapeutic strategies. This study presents a delayed-melting cryomicroneedle (cryoMN) platform that enables dermal delivery of Lacticaseibacillus rhamnosus LA3, a probiotic strain exhibiting superior anti-C. albicans activity among the five screened candidates. LA3 achieved a 1.01-lg reduction in planktonic C. albicans viability within 48 h of co-culture while maintaining self-proliferation. Its cell-free supernatant (CFS) displayed concentration-dependent antifungal efficacy, with 50 % CFS resulting in 62.6 % inhibition of planktonic C. albicans and 98 % reduction in biofilm biomass, as confirmed by confocal laser scanning microscopy, which revealed structural biofilm collapse. In infected keratinocyte models, LA3 restored cell viability to approximately 90 % (compared with ≤79.3 % in controls). A biofilm-mimicking gelatin-hyaluronic acid hydrogel scaffold was developed, preserving >90 % probiotic viability for 5 days at 4 °C, relaxing frozen-chain requirement while maintaining microneedle integrity. In murine models of invasive candidiasis, LA3-loaded cryoMNs demonstrated superior therapeutic efficacy compared with both topical LA3 gel and clotrimazole cream, with excellent biosafety (> 90 % cell viability, < 5 % hemolysis, and no detectable organ toxicity). By integrating strain-optimized probiotics with biomimetic delivery technology, this platform represents a minimally invasive strategy for treating deep-seated fungal infections.