C. albicans incorporates methaemoglobin into surface attached and aggregate biofilms that exhibit increased virulence and antifungal resistance

Candida albicans is a leading cause of invasive fungal infections, yet how host-derived factors shape pathogenic biofilm states remains poorly understood. Here we show that oxidised haemoglobin (methaemoglobin; MetHb) which is readily formed upon red blood cell lysis, promotes the formation of C. albicans biofilms. MetHb also induces the rapid agglutination of C. albicans cells into haemoglobin aggregates, that display hallmark biofilm properties including antifungal tolerance, and enhanced virulence. The interaction between C. albicans and MetHb is enhanced by hyphal growth and requires the adhesins Als1 and Als3. Temporal transcriptomic analysis of C. albicans-MetHb aggregate formation revealed a rapid activation of adhesion and haem-iron acquisition pathways, followed by metabolic remodelling and enrichment of hypoxia-associated genes suggesting the establishment of localised microenvironments that favour fermentative metabolism. Consistent with C. albicans-MetHb aggregates acting as aggregate biofilms, they exhibited increased tolerance to antifungals and increased virulence in a Galleria mellonella infection model. Agent-based modelling of C. albicans-MetHb interactions supports a mechanism by which hypha-mediated adhesion drives aggregation and biofilm formation as observed experimentally, providing a valuable tool for future investigation. Together, these findings identify Methaemoglobin as a host-derived driver of rapid C. albicans community assembly. This raises the possibility that high MetHb levels increase the chance that C. albicans drug-tolerant, hypervirulent biofilm or aggregate biofilms become established in vivo. As MetHb levels are found to be elevated during infection, sepsis, surgery or medical device insertion, our novel findings raise the possibility that C. albicans-MetHb agglutination may play a significant, and yet underappreciated, role in multiple pathologies.