The ISME Journal (2020) 14:2568–2579 https://doi.org/10.1038/s41396-020-0707-2 ARTICLE Physiological dynamics of chemosynthetic symbionts in hydrothermal vent snails 1 2 2 3 3 2 Corinna Breusing ● Jessica Mitchell ● Jennifer Delaney ● Sean P. Sylva ● Jeffrey S. Seewald ● Peter R. Girguis ● Roxanne A. Beinart 1 Received: 7 November 2019 / Revised: 15 June 2020 / Accepted: 23 June 2020 / Published online: 2 July 2020 © The Author(s) 2020. This article is published with open access Abstract Symbioses between invertebrate animals and chemosynthetic bacteria form the basis of hydrothermal vent ecosystems worldwide. In the Lau Basin, deep-sea vent snails of the genus Alviniconcha associate with either Gammaproteobacteria (A. kojimai, A. strummeri)orCampylobacteria (A. boucheti) that use sulfide and/or hydrogen as energy sources. While the A. boucheti host–symbiont combination (holobiont) dominates at vents with higher concentrations of sulfide and hydrogen, the A. kojimai and A. strummeri holobionts are more abundant at sites with lower concentrations of these reductants. We posit that adaptive differences in symbiont physiology and gene regulation might influence the observed niche partitioning 1234567890();,: 1234567890();,: between host taxa. To test this hypothesis, we used high-pressure respirometers to measure symbiont metabolic rates and examine changes in gene expression among holobionts exposed to in situ concentrations of hydrogen (H2: ~25 µM) or hydrogen sulfide (H2S: ~120 µM). The campylobacterial symbiont exhibited the lowest rate of H2S oxidation but the highest rate of H2 oxidation, with fewer transcriptional changes and less carbon fixation relative to the gammaproteobacterial symbionts under each experimental condition. These data reveal potential physiological adaptations among symbiont types, which may account for the observed net differences in metabolic activity and contribute to the observed niche segregation among holobionts. Introduction thereby fostering both ecological and evolutionary diversi- fication [3–7]. Symbiont-mediated niche expansion, habitat Symbiotic microbes are increasingly recognized as impor- partitioning and local adaptation have been well described tant drivers of animal development, physiology and spe- in insect-microbe and coral-algae symbioses [3–5], but ciation [1, 2]. How symbionts influence the biology of their these processes are less understood in other symbiotic host is particularly evident in nutritional symbiotic rela- systems. tionships, which often broaden the ecological niche of the Mutualistic symbioses between chemosynthetic bacteria animal partner by providing access to novel resources, and invertebrate animals at deep-sea hydrothermal vents offer intriguing opportunities to study the effects of sym- biont physiology on host adaptation and niche segregation in the ocean, given the intricate links between geochemical Supplementary information The online version of this article (https:// environment, symbiont metabolism and host survival. In doi.org/10.1038/s41396-020-0707-2) contains supplementary material, which is available to authorized users. these associations, the symbiotic bacteria oxidize reduced chemicals present in the hydrothermal fluids—e.g., sulfide * Corinna Breusing (H2S), methane (CH4) and/or hydrogen (H2)—to gain [email protected] energy for the fixation of inorganic carbon into organic 1 University of Rhode Island, Graduate School of Oceanography, matter, which provides the bulk of nutrition for the animal Narragansett, RI, USA host (chemosynthesis; [8]). Chemosynthetic symbioses 2 Harvard University, Department of Organismic and Evolutionary have evolved many times: invertebrate hosts come from Biology, Cambridge, MA, USA divergent taxonomic groups within three invertebrate phyla, 3 Woods Hole Oceanographic Institution, Department of Marine while their symbionts have evolved from several distinct Chemistry and Geochemistry, Woods Hole, MA, USA lineages of two bacterial phyla [8]. Host and symbiont taxa Physiological dynamics of chemosynthetic symbionts in hydrothermal vent snails 2569 Table 1 Summary of the six experiments presented here, including the reductant supplied, geographic coordinates and depths from which the snail species were collected, as well as the number (N) of control and treatment animals used in each experiment. Exp. Reductant Vent field Latitude Longitude Depth (m) Species Symbiont Control (N) Treatment (N) 1H2S ABE –20.7630 –176.1913 2155 A. boucheti ε 2 4 A. kojimai γ-1 2 4 2H2STu’i Malila –21.9891 –176.5682 1884 A. strummeri γ-1 3 4 A. kojimai γ-1 3 4 3H2 Tu’i Malila –21.9891 –176.5682 1888 A. strummeri γ-1 3 4 A. kojimai γ-1 3 5 4H2 ABE –20.7617 –176.1917 2130 A. boucheti ε 2 4 A. kojimai γ-1 2 5 5H2 Tahi Moana –20.6838 –176.1834 2214 A. strummeri γ-1 3 3 A. boucheti ε 3 3 6a None Tow Cam –20.3166 –176.1361 2703 A. boucheti ε 0 3 A. kojimai γ-1 0 3 aAs we were unable to include an acclimation control, transcriptomic sequencing was not performed for individuals in this experiment. typically show a strong selectivity towards each other, as species and symbiont 16S ribosomal RNA (rRNA) phylo- each host species associates with only one to a few distinct types appear to be highly selective towards each other: A. bacterial symbiont lineages [8]. Thus, given that symbiont boucheti typically harbors a campylobacterial symbiont phylogenetic diversity can correspond to functional diver- phylotype (called ε), whereas A. kojimai and A. strummeri sity, ecological differences among host taxa may be tied to associate with two distinct lineages of gammaproteo- the specific traits of their symbiont(s). bacterial symbiont phylotypes (called γ-1 and γ-Lau) [13]. Diversity in chemosynthetic traits among symbionts may Beinartetal.[13] suggested that the distribution of be especially important to habitat segregation in these sys- different host–symbiont combinations is determined by tems. Chemosynthetic host–symbiont associations (holo- local and regional variations in vent geochemistry. While bionts) are often characterized by heterogeneous A. boucheti holobionts are predominantly found at distributions at both local and regional scales, with discrete northern sites characterized by high-fluid concentrations patches or zones of animal taxa occurring within a single- of H2SandH2,theA. kojimai and A. strummeri holobionts vent field or entire fields being dominated by one or a few are more frequently observed at southern sites where taxa [9–11]. Since the composition and concentration of concentrations of these reductants are lower (Supple- fluid compounds for chemosynthetic processes can vary mentary Table S1). The authors hypothesized that differ- substantially over temporal and spatial scales [12], the ences in symbiont metabolic potential might drive the distribution of many holobionts seems to be linked to taxon- differential abundance of symbiont and host genotypes. specific associations with particular habitat characteristics. Subsequent genome comparisons between the three Thus, local and regional community composition, and symbiont lineages showed that these symbionts are ultimately the overall biological diversity, may be strongly broadly similar in terms of gene content related to che- influenced by the partitioning of holobionts into distinct molithoautotrophy, although differences exist in the spe- physico-chemical niches. Yet, to date, the mechanistic links cific encoded enzyme types and metabolic pathways [18]. between fluid geochemistry and holobiont community The large overlap in overall metabolic potential but dis- composition are poorly known. crepancy in realized pathways implies that differences in Snails of the genus Alviniconcha provide a unique enzyme kinetics and gene regulation might underlie the opportunity to understand symbiont-meditated habitat par- observed habitat partitioning between snail holobionts titioning at hydrothermal vents [13, 14]. In the Eastern Lau [18]. To address this question, we supplied the ELSC Spreading Center (ELSC), Tonga, three species of Alvini- Alviniconcha holobionts with defined concentrations of concha (A. boucheti, A. kojimai, A. strummeri) co-occur at dissolved sulfide and hydrogen in high-pressure shipboard several vent localities [13, 15], where they establish endo- experiments and measured changes in metabolic rates and symbioses with three different lineages of chemoauto- gene expression. Exposing the symbionts to the same trophic Gammaproteobacteria or Campylobacteria environmental conditions allowed us to compare the (formerly Epsilonproteobacteria;[16]). The bacterial sym- holobionts’ transcriptional responses and chemosynthetic bionts are assumed to be environmentally acquired given activities, thereby providing insights into potential adap- that phylogenetic studies suggest an absence of tations that are likely important to their realized dis- host–symbiont co-evolution [13, 17]. Nevertheless, host tribution across a range of geochemical habitats. 2570 C. Breusing et al. Materials and methods H2 and H2S oxidation and O2 respiration rates were cal- culated as in ref. [19] by comparing the steady-state Sample collection and experimental setup concentrations of H2,H2S, and O2 between effluents from treatment and empty control aquaria (Supplementary Alviniconcha were collected with the ROV ROPOS during Tables S2 and S3; and Supplementary Figs. S3 and S4). In the R/V Falkor expedition FK160409 in 2016 (Table 1). the experiments without empty control aquarium, the Snails were recovered in insulated