DIET-SAO Baltic-Consortia- Bothnian Bay methanogenic zone Rotarulab, Danijel Jovicic Metagenomic assembly

Acetate is a key intermediate in mineralizing organic matter, especially in deep subsurface environments with limited electron acceptors. Here, methanogenic archaea can split acetate to CH4 and CO2, while acetate-oxidizing bacteria can oxidize it to CO2 providing reducing equivalents for methanogens.In sediment enrichments from Bothnian Bay, we investigated how conductive particles affect marine acetate metabolism. Granular-activated-carbon (GAC) and magnetite were crucial for linking syntrophic acetate oxidation (SAO) with CO2-reductive methanogenesis, known as CIET (Conductive Particle-mediated Interspecies Electron Transfer). Our previous observations showed a fourfold increase in methanogenesis with conductive particles compared to controls (no conductive particles). Labeled acetate was primarily taken up by a Geobacterales-like organism, converting the 13CH3-group to 13CO2. Inhibition experiments confirmed a metabolic co-dependency between Archaea and Bacteria, using Methyl-CoM inhibitors or antibiotics.Metagenomic analyses identified core CIET organisms exclusively present in incubations with conductive particles. Metagenome-assembled genomes (MAGs) from these particles included the main players: a Geobacterales and a Methanosarcina, found only in such incubations.The Methanosarcina, typical of marine environments, harbors an Rnf-operon flanked by a membrane-bound methanogenesis multiheme cytochrome capable of bidirectional Extracellular Electrons Transfer (EET). The Geobacterales-MAG showcases over 40 multiheme c-type cytochromes, acetate transporters, and a complete acetate-oxidation pathway, confirming their potential for acetate-oxidation, and EET-capabilities.While these two organisms are key in acetate oxidation, other groups in the consortia may have secondary functions. Our findings highlight complex microbial interactions and diverse strategies in CIET, providing valuable insights into the metabolic capabilities of marine sediment consortia related to methane emissions.This study presents, for the first time, a well-characterized environmental DIET consortium using conductive particles, which appears to employ new electron exchange transfer methods. This consortium depends on conductive particles to maintain and process syntrophic acetate oxidation.