Metabolic and Population Profiles of Active Subseafloor Autotrophs in Young Oceanic Crust at Deep-Sea Hydrothermal Vents

Abstract

At deep-sea hydrothermal vents, magmatically driven rock-water reactions in the crust generate gases and other reduced compounds that subseafloor microorganisms use for chemolithoautotrophy. In this study, microbial autotrophs from three diffuse flow hydrothermal vents at Axial Seamount in 2013 and 2014 were isotopically labeled using RNA Stable Isotope Probing (RNA-SIP), targeting subseafloor autotrophic mesophiles (30{degrees}C), thermophiles (55{degrees}C), and hyperthermophiles (80{degrees}C). We constructed taxonomic and functional profiles of active chemolithoautotrophs, examined population distributions across sites, and linked primary producers to their specific metabolic strategies within the subseafloor community. Dominant autotrophs exhibited hydrogen-dependent dissimilatory metabolisms such as sulfur and nitrate reduction and methanogenesis, as well as microaerophilic sulfide oxidation even at 80{degrees}C, consistent with fluid chemistries at each site. While hydrogenotrophic methanogenic archaea such as Methanothermococcus were restricted in their distribution and activity, hydrogenotrophic sulfur and nitrate reducers from the Aquificota (Thermovibrio), Campylobacterota (Nautiliaceae, Hydrogenimonas, and Desulfurobacteriaceae) were consistently active and present at all sites and years at both the population and community levels. Hydrogenase transcripts were significantly differentially expressed, and diverse hydrogenases were found in metagenome-assembled genomes of Aquificota members, highlighting the importance and versatility of their hydrogen utilization strategies which likely contribute to their cosmopolitan distribution across geochemically disparate subseafloor sites. Together, this study provides new insights into the functional dynamics and distribution of key subseafloor autotrophic microbial communities in young oceanic crust at deep-sea hydrothermal vents. IMPORTANCEDeep-sea hydrothermal vents are hotspots for life in the dark ocean, where rich animal ecosystems are supported by microbial primary producers utilizing the abundant chemical energy supplied by high-temperature water-rock reactions. Despite increasing knowledge about the geochemistry and microbiology of deep-sea hydrothermal vents, there is still a gap in our understanding of the key microbial players who fix much of the carbon at these sites, especially in the productive subseafloor. In this study, stable isotope probing was used to label active microbial autotrophs in diffuse flow venting fluids from three sites over two years and was combined with metatranscriptomic sequencing to identify their specific metabolic strategies. This research highlights the microbial community composition, function, gene regulation, and population dynamics that enable hydrothermal ecosystems to persist.