Low nutrient and energy availability has led to the evolution of numerous strategies for overcoming these limitations, of which symbiotic associations represent a key mechanism. Particularly striking are the associations between chemosynthetic bacteria and marine animals that thrive in nutrient-poor environments such as the deep sea because the symbionts allow their hosts to grow on inorganic energy and carbon sources such as sulfide and CO 2 . Remarkably little is known about the physiological strategies that enable chemosynthetic symbioses to colonize oligotrophic environments. In this study, we used metaproteomics and metabolomics to investigate the intricate network of metabolic interactions in the chemosynthetic association between Olavius algarvensis, a gutless marine worm, and its bacterial symbionts. We propose previously undescribed pathways for coping with energy and nutrient limitation, some of which may be widespread in both freeliving and symbiotic bacteria. These pathways include (i) a pathway for symbiont assimilation of the host waste products acetate, propionate, succinate and malate; (ii) the potential use of carbon monoxide as an energy source, a substrate previously not known to play a role in marine invertebrate symbioses; (iii) the potential use of hydrogen as an energy source; (iv) the strong expression of high-affinity uptake transporters; and (v) as yet undescribed energy-efficient steps in CO 2 fixation and sulfate reduction. The high expression of proteins involved in pathways for energy and carbon uptake and conservation in the O. algarvensis symbiosis indicates that the oligotrophic nature of its environment exerted a strong selective pressure in shaping these associations.3-hydroxypropionate bi-cycle | Calvin cycle | proton-translocating pyrophosphatase | pyrophosphate dependent phosphofructokinase | metagenomics G rowth in nutrient-limited environments presents numerous challenges to organisms. Symbiotic and syntrophic relationships have evolved as particularly successful strategies for coping with these challenges. Such nutritional symbioses are widespread in nature and, for example, have enabled plants to colonize nitrogen-poor soils and animals to thrive on food sources that lack essential amino acids and vitamins (1). Chemosynthetic symbioses, discovered only 35 years ago at hydrothermal vents in the deep sea, revolutionized our understanding of nutritional associations, because these symbioses enable animals to live on inorganic energy and carbon sources such as sulfide and CO 2 (2, 3). The chemosynthetic symbionts use the energy obtained from oxidizing reduced inorganic compounds such as sulfide to fix CO 2 , ultimately providing their hosts with organic carbon compounds. Chemosynthetic symbioses thus are able to thrive in habitats where organic carbon sources are rare, such as the deep sea, and the symbionts often are so efficient at providing nutrition that many hosts have reduced their digestive systems (4).The marine oligochaete Olavius algarvensis is a particularly extre...

Sulfur-cycling microorganisms impact organic matter decomposition in wetlands and consequently greenhouse gas emissions from these globally relevant environments. However, their identities and physiological properties are largely unknown. By applying a functional metagenomics approach to an acidic peatland, we recovered draft genomes of seven novel Acidobacteria species with the potential for dissimilatory sulfite (dsrAB, dsrC, dsrD, dsrN, dsrT, dsrMKJOP) or sulfate respiration (sat, aprBA, qmoABC plus dsr genes). Surprisingly, the genomes also encoded DsrL, which so far was only found in sulfur-oxidizing microorganisms. Metatranscriptome analysis demonstrated expression of acidobacterial sulfur-metabolism genes in native peat soil and their upregulation in diverse anoxic microcosms. This indicated an active sulfate respiration pathway, which, however, might also operate in reverse for dissimilatory sulfur oxidation or disproportionation as proposed for the sulfur-oxidizing Desulfurivibrio alkaliphilus. Acidobacteria that only harbored genes for sulfite reduction additionally encoded enzymes that liberate sulfite from organosulfonates, which suggested organic sulfur compounds as complementary energy sources. Further metabolic potentials included polysaccharide hydrolysis and sugar utilization, aerobic respiration, several fermentative capabilities, and hydrogen oxidation. Our findings extend both, the known physiological and genetic properties of Acidobacteria and the known taxonomic diversity of microorganisms with a DsrAB-based sulfur metabolism, and highlight new fundamental niches for facultative anaerobic Acidobacteria in wetlands based on exploitation of inorganic and organic sulfur molecules for energy conservation.

SummarySymbiotic bivalves at hydrothermal vents and cold seeps host chemosynthetic bacteria intracellularly in gill cells. In bivalves, the gills grow continuously throughout their lifetime by forming new filaments. We examined how newly developed gill tissues are colonized in bivalves with horizontal and vertical symbiont transmission (Bathymodiolus mussels versus a vesicoymid clam) using fluorescence in situ hybridization and transmission electron microscopy. Symbiont colonization was similar in mussels and clams and was independent of the transmission modes. Symbionts were absent in the growth zones of the gills, indicating that symbionts colonize newly formed gill filaments de novo after they are formed and that gill colonization is a continuous process throughout the host's lifetime. Symbiont abundance and distribution suggested that colonization is shaped by the developmental stage of host cells. Selfinfection, in which new gill cells are colonized by symbionts from ontogenetically older gill tissues, may also play a role. In mussels, symbiont infection led to changes in gill cell structure similar to those described from other epithelial cells infected by intracellular pathogens, such as the loss of microvilli. A better understanding of the factors that affect symbiont colonization of bivalve gills could provide new insights into interactions between intracellular bacteria and epithelial tissues.

The level of integration between associated partners can range from ectosymbioses to extracellular and intracellular endosymbioses, and this range has been assumed to reflect a continuum from less intimate to evolutionarily highly stable associations. In this study, we examined the specificity and evolutionary history of marine symbioses in a group of closely related sulphur-oxidizing bacteria, called Candidatus Thiosymbion, that have established ecto-and endosymbioses with two distantly related animal phyla, Nematoda and Annelida. Intriguingly, in the ectosymbiotic associations of stilbonematine nematodes, we observed a high degree of congruence between symbiont and host phylogenies, based on their ribosomal RNA (rRNA) genes. In contrast, for the endosymbioses of gutless phallodriline annelids (oligochaetes), we found only a weak congruence between symbiont and host phylogenies, based on analyses of symbiont 16S rRNA genes and six host genetic markers. The much higher degree of congruence between nematodes and their ectosymbionts compared to those of annelids and their endosymbionts was confirmed by cophylogenetic analyses. These revealed 15 significant codivergence events between stilbonematine nematodes and their ectosymbionts, but only one event between gutless phallodrilines and their endosymbionts. Phylogenetic analyses of 16S rRNA gene sequences from 50 Cand. Thiosymbion species revealed seven well-supported clades that contained both stilbonematine ectosymbionts and phallodriline endosymbionts. This closely coupled evolutionary history of marine ecto-and endosymbionts suggests that switches between symbiotic lifestyles and between the two host phyla occurred multiple times during the evolution of the Cand. Thiosymbion clade, and highlights the remarkable flexibility of these symbiotic bacteria.