Project description:The endosymbiont population of the hydrothermal vent tube worm Riftia pachyptila consists of a single 16S phylotype of sulfur-oxidizing gammaproteobacteria. The intracellular symbiont exhibits remarkable morphological heterogeneity, from small rod-shaped or coccoid cells to large cocci, which were suggested to be part of a common cell cycle. To assess whether these morphological differences are accompanied by distinct metabolic profiles, we physically enriched individual symbiont cells sizes by density gradient centrifugation and subjected these enrichments to metaproteomic analysis and statistical evaluation using clustering and random forests. Unlike previous molecular studies, which examined the metabolism of the symbiont population as whole, we were thus able to unravel comprehensive protein abundance patterns of individual symbiont subpopulations. Supported by microscopic analyses, our metaproteomic results show that Riftia symbiont cells of different sizes are stages of a physiological differentiation process: Small symbionts actively divide and may establish cellular symbiont-host interaction, as indicated by highest abundance of the cell division key protein FtsZ and highly abundant chaperones and porins in this initial phase. We furthermore present evidence that large symbionts, on the other hand, do not divide, but still replicate DNA, leading to DNA endoreduplication. Highest abundance of enzymes for CO2 fixation, carbon storage and biosynthesis indicates that in its late differentiation stage, the symbiont’s metabolism is efficiently geared on the production of organic material. We propose that this symbiont aging process enhances the productivity of the symbiosis as a whole.
Project description:The deep-sea tubeworm Riftia pachyptila is a model system for a mutualistic association: The adult worm has no digestive system, but completely relies on one phylotype of endosymbiotic chemosynthetic bacteria for nutrition. The bacteria, in turn, are provisioned by the host. Metabolism and physiology of this symbiosis, particularly of the uncultured symbiont, have been subject to various studies. Yet, how both partners interact on the molecular level remains largely unknown. To study these host-symbiont interactions in detail, we sequenced the R. pachyptila host transcriptome de novo, and conducted comprehensive metaproteomic comparisons of symbiont-containing and symbiont-free R. pachyptila tissues under energy-rich and energy-limiting conditions. Our results demonstrate that R. pachyptila invests a considerable part of its proteome to provision the symbionts with inorganic compounds. It acquires symbiont-derived biomass primarily by digesting parts of the symbiont population. The R. pachyptila immune system apparently not only protects the holobiont from pathogens, but is also involved in symbiont population control. The symbiont expresses a repertoire of proteins dedicated to communication with the host, including eukaryote-like proteins that may counteract phagocytosis. During energy limitation, i.e., when reduced sulfur compounds are lacking, the host apparently increases symbiont digestion. We show here an intricate network of interaction pathways that shapes the R. pachyptila holobiont. Together with the metabolic flexibility of the association under varying energy conditions, this probably forms the basis for the success of this tight association under the highly challenging deep-sea conditions.
Project description:The deep-sea tubeworm Riftia pachyptila lacks a digestive system but completely relies on bacterial endosymbionts for nutrition. Although the symbiont has been studied in detail on the molecular level, such analyses were unavailable for the animal host, because sequence information was lacking. To identify host-symbiont interaction mechanisms, we therefore sequenced the Riftia transcriptome, which served as a basis for comparative metaproteomic analyses of symbiont-containing versus symbiont-free tissues, both under energy-rich and energy-limited conditions. Our results suggest that metabolic interactions include nutrient allocation from symbiont to host by symbiont digestion and substrate transfer to the symbiont by abundant host proteins. We furthermore propose that Riftia maintains its symbiont by protecting the bacteria from oxidative damage while also exerting symbiont population control. Eukaryote-like symbiont proteins might facilitate intracellular symbiont persistence. Energy limitation apparently leads to reduced symbiont biomass and increased symbiont digestion. Our study provides unprecedented insights into host-microbe interactions that shape this highly efficient symbiosis.IMPORTANCE All animals are associated with microorganisms; hence, host-microbe interactions are of fundamental importance for life on earth. However, we know little about the molecular basis of these interactions. Therefore, we studied the deep-sea Riftia pachyptila symbiosis, a model association in which the tubeworm host is associated with only one phylotype of endosymbiotic bacteria and completely depends on this sulfur-oxidizing symbiont for nutrition. Using a metaproteomics approach, we identified both metabolic interaction processes, such as substrate transfer between the two partners, and interactions that serve to maintain the symbiotic balance, e.g., host efforts to control the symbiont population or symbiont strategies to modulate these host efforts. We suggest that these interactions are essential principles of mutualistic animal-microbe associations.
Project description:The giant tubeworm Riftia pachyptila lives in symbiosis with the chemoautotrophic gammaproteobacterium Cand. Endoriftia persephone. Symbionts are released back into the environment upon host death in high-pressure experiments, while microbial fouling is not involved in trophosome degradation. Therefore, we examined the antimicrobial effect of the tubeworm's trophosome and skin. The growth of all four tested Gram-positive, but only of one of the tested Gram-negative bacterial strains was inhibited by freshly fixed and degrading trophosome (incubated up to ten days at either warm or cold temperature), while no effect on Saccharomyces cerevisiae was observed. The skin did not show antimicrobial effects. A liquid chromatography-mass spectrometric analysis of the ethanol supernatant of fixed trophosomes lead to the tentative identification of the phospholipids 1-palmitoleyl-2-lyso-phosphatidylethanolamine, 2-palmitoleyl-1-lyso-phosphatidylethanolamine and the free fatty acids palmitoleic, palmitic and oleic acid, which are known to have an antimicrobial effect. As a result of tissue autolysis, the abundance of the free fatty acids increased with longer incubation time of trophosome samples. This correlated with an increasing growth inhibition of Bacillus subtilis and Listeria welshimeri, but not of the other bacterial strains. Therefore, the free fatty acids produced upon host degradation could be the cause of inhibition of at least these two bacterial strains.