Diversity and Ecology of Chemosynthetic Symbioses in Deep-Sea Invertebrates

Abstract

The energy sources that drive biological processes at deep-sea hydrothermal vents and cold seeps include methane and reduced inorganic compounds such as sulfide and hydrogen. These compounds are unavailable to metazoan life, but can be used by chemoautotrophic or methanotrophic bacteria to fuel chemosynthetic primary production. In these habitats, symbioses between invertebrates and chemoautotrophic or methanotrophic bacteria form the basis of ecosystems that thrive in the absence of sunlight. This thesis is made up of two thematic parts. In the first part, two reviews on methane-oxidizing symbionts are presented. Methanotrophic symbiotic bacteria have so far been found in invertebrate animals at hydrothermal vents and cold seeps in the deep sea, and in a plant host in a terrestrial wetland. No methane-oxidizing symbiont has yet been cultured. Therefore, the evidence for methanotrophy in these bacteria has come from ultrastructural, enzymatic, physiological, stable isotope, and molecular biological studies of the symbiotic host tissues. Despite being found in a wide range of hosts, all marine methanotrophic symbionts for which 16S rRNA sequences are available belong to a single lineage within the Gammaproteobacteria. Methane-oxidizing symbionts in the terrestrial habitat belong to the Alphaproteobacteria. However, many of these symbionts have not yet been characterized with molecular methods, and may be more diverse than currently recognized. These reviews summarize the current knowledge of methane-based symbioses, and identify topics for future research.In the second part of this thesis, I present two studies on the ectosymbiosis of Rimicaris exoculata, an alvinocaridid shrimp from hydrothermal vents on the Mid-Atlantic Ridge (MAR). These shrimp have filamentous ectosymbionts on modified appendages and the inner surfaces of the gill chamber. The symbionts were previously assumed to be epsilonproteobacterial sulfur oxidizers. Using the full-cycle 16S rRNA approach, I identified a second filamentous ectosymbiont that belongs to a novel symbiotic lineage within the Gammaproteobacteria. To investigate the biogeography of the symbiosis, I compared the 16S rRNA gene sequences of ectosymbionts from four MAR vent fields, Rainbow, TAG, Logatchev, and South MAR, which are separated by up to 8500 km along the MAR. Differences in the 16S rRNA gene for both the gammaproteobacterial and epsilonproteobacterial symbionts were correlated with geographic distance. In contrast, the phylogeny of the free-living relatives of the epsilonproteobacterial symbiont showed no obvious geographic trend. Host-symbiont recognition could explain the observed symbiont distribution. The metabolism of the symbionts from the Rainbow vent field was investigated by thermodynamic modelling, sequencing of key metabolic genes, and immunohistochemical labelling of proteins in single epibiont cells. Key genes for carbon fixation by the reductive TCA and CBB cycles indicate that both symbionts can fix CO2. Key genes for hydrogen, methane, and sulfur oxidation indicate that the epibionts have the potential to use these three energy sources. In addition, immunohistochemistry showed that particulate methane monooxygenase is expressed by the gammaproteobacterial symbiont, although it does not belong to any of the currently known methanotrophic lineages. These studies show that the phylogenetic and functional diversity of the shrimp epibiosis is greater than previously recognized.