Unraveling the hidden world: Variability and complexity of holopelagic Sargassum biofilms

Holopelagic Sargassum spp. -including S. natans var. natans (SNN), S. natans var. wingei (SNW), and S. fluitans var. fluitans (SFF)-have become a major ecological and socio-economic concern due to recurrent mass strandings in the Caribbean. This study aims to complement the growing knowledge on the holopelagic Sargassum holobiont by incorporating the three-dimensional organization of its biofilms. Here, we provide the first detailed characterization of holopelagic Sargassum biofilms using scanning electron microscopy (SEM) combined with bacterial abundance measurements. Samples were collected from nearshore waters and stranded mats in Guadeloupe. We assessed biofilm elemental composition, maturity, and surface coverage, along with the presence of diatoms and superficial filamentous bacterial coverage (SFBC). Here, we report for the first time in macroalgal biofilms the occurrence of biofilm-coated diatom units (BCDU) with protrusions, as well as Ca-P-O crystals. Biofilm structure and composition varied among taxa, thallus compartments, and environmental contexts. SNN exhibited the lowest surface roughness, the weakest biofilm development in nearshore conditions, and the lowest abundances of Ca-P-O crystals. In contrast, SFF consistently displayed high densities of crystals, diatoms, and BCDU with protrusions, indicating greater microbial and morphological complexity, while SNW maintained a stable biofilm architecture largely independent of thallus part and environment. Stranding increased bacterial abundance across all taxa and enhanced diatom colonization in SNW and SFF. Across taxa, axes were least affected, whereas fronds, particularly in SNN,were most strongly impacted. SFBC, highest in SNN, was negatively correlated with crystal abundance, which was highest in SFF and SNW. Overall, these findings provide a three-dimensional perspective on intra-thallus and interspecific variation in Sargassum biofilms and demonstrate how interactions among host surface traits, microbial components, and environmental context shape biofilm structure, stability, and potentially host resilience.