Analysis of data, curated over 3 years of sediment sampling, improves our understanding of microbiota assembly in marine sediment. Furthermore, we demonstrate the importance of cross-kingdom integration of information in the study of microbial community ecology. Finally, the urgent need to propose an applicable approach for environmental health monitoring is addressed here by establishment of sediment microbiota as a robust and sensitive model.

Summary The symbioses between the vestimentiferan tubeworms and their chemosynthetic partners (Gammaproteobacteria, Chromatiales and Sedimenticolaceae) hallmark the success of these organisms in hydrothermal vent and hydrocarbon seep deep‐sea habitats. The fidelity of these associations varies, as both the hosts and the symbionts can be loose in partner choice. Some tubeworms may host distinct symbiont phylotypes, which often co‐occur in a single host individual. To better understand the genetic basis for the promiscuity of tubeworm symbioses, we assembled and investigated metagenome‐assembled genomes of two symbiont phylotypes (species, based on the average nucleotide identity < 95%) in Lamellibrachia anaximandri, a vestimentiferan endemic to the Mediterranean Sea, in individuals collected from Palinuro hydrothermal vents (Italy) and hydrocarbon seeps (Eratosthenes seamount and Palmahim disturbance). Using comparative genomics, we show that mainly mobilome and genes involved in defence mechanisms distinguish the symbiont genotypes. While many central metabolic functions are conserved in the tubeworm symbionts, nitrate respiration (Nar, Nap and Nas proteins) is modular, yet this modularity is not linked to phylotype, but rather to geographic location, potentially implying adaptation to the local environment. Our results hint that variation in a single moonlighting protein may be responsible for the fidelity of these symbioses.

Bathymodioline mussels rely on thiotrophic and methanotrophic chemosynthetic symbionts for nutrition, yet, secondary heterotrophic symbionts are often present and play an unknown role in the fitness of the organism. The bathymodioline Idas mussels that thrive in gas seeps and on sunken wood in the Mediterranean Sea and the Atlantic Ocean, host at least six symbiont lineages that often co-occur, including the primary, chemosynthetic methane- and sulfur-oxidizing gammaproteobacteria, and the secondary Methylophagaceae, Nitrincolaceae and Flavobacteraceae symbionts, whose physiology and metabolism are obscure. Little is known about whether and how these symbionts interact or exchange metabolites. Here we curated metagenome-assembled genomes of Idas modiolaeformis symbionts and used genome-centered metatranscriptomics and metaproteomics to assess key symbiont functions. The Methylophagaceae symbiont is a methylotrophic autotroph, as it encoded and expressed the ribulose monophosphate and Calvin-Benson-Bassham cycle enzymes, particularly RuBisCO. The Nitrincolaceae ASP10-02a symbiont likely fuels its metabolism with nitrogen-rich macromolecules and may provide the holobiont with vitamin B12. The Flavobacteriaceae Urechidicola symbionts likely degrade glycans and may remove NO. Our findings indicate that these flexible associations allow for expanding the range of substrates and environmental niches, via new metabolic functions and handoffs.

Bathymodioline mussels rely on thiotrophic and/or methanotrophic chemosynthetic symbionts for nutrition, yet, secondary heterotrophic symbionts are often present and play an unknown role in the fitness of the organism. The bathymodioline Idas mussels that thrive in gas seeps and on sunken wood in the Mediterranean Sea and the Atlantic Ocean, host at least six symbiont lineages that often co-occur. These lineages include the primary symbionts chemosynthetic methane- and sulfur-oxidizing gammaproteobacteria, and the secondary symbionts, Methylophagaceae, Nitrincolaceae and Flavobacteriaceae, whose physiology and metabolism are obscure. Little is known about if and how these symbionts interact or exchange metabolites. Here we curated metagenome-assembled genomes of Idas modiolaeformis symbionts and used genome-centered metatranscriptomics and metaproteomics to assess key symbiont functions. The Methylophagaceae symbiont is a methylotrophic autotroph, as it encoded and expressed the ribulose monophosphate and Calvin-Benson-Bassham cycle enzymes, particularly RuBisCO. The Nitrincolaceae ASP10-02a symbiont likely fuels its metabolism with nitrogen-rich macromolecules and may provide the holobiont with vitamin B12. The Urechidicola (Flavobacteriaceae) symbionts likely degrade glycans and may remove NO. Our findings indicate that these flexible associations allow for expanding the range of substrates and environmental niches, via new metabolic functions and handoffs.