Marine Microbial Diversityy:Deep-sea Sediments And Symbiosis

The ocean is the largest habitat on Earth, harboring diverse niches and tremendous biodiversity, and is closely related to global biogeochemical cycling, climate change, as well as human well-being. However, the unseen majority, i.e., bacteria and archaea, in marine environments is mostly unexplored. To understand diverse microbial communities in the deep sea, this thesis investigated marine microbial diversity in multiple habitats, from phylogenetic, functional and metabolic aspects, as well as in symbiotic association with invertebrates.Deep-sea sediments, the second largest habitat on Earth after the seawater column, harbor diverse microorganisms that are mostly unknown. The first part of this thesis (chapters2and3) focused on investigation of phylogenetic diversity of microorganisms in sediments from gas hydrates and cobalt-rich crust regions. Relatively high bacterial diversity, but low archaeal diversity was found in both studies. Special habitats may play a role in structuring microbial communities,﹕pecially their abnormal amount of chemicals, e.g., high methane concentrations in gas hydrate regions and enriched metals in cobalt-rich crust region. Interesting discoveries were found by examining the microbial communities, which have laid the foundation for future research into topics such as functional genes, transcriptomes and ecological associations studied through in situ hybridization.Microbial diversity can also be investigated through their functional differences, which are carried out by enzymes. To utilize agar, the cell wall component of algae, agarolytic microorganisms produce agarase to digest the neutral chain (agarose) and yield oligosaccharides, which is finally converted to galactose for utilization. The fourth chapter described cloning, expression and characterization of a new agarase from Vibrio strain, CN41. It is a GH50agarase, and has special characteristics that are promising for commercial applications.The last part of this thesis moved from free-living marine microorganisms to symbiotic microorganisms within tubeworms at deep-sea hydrothermal vents. It has been shown that tubeworm symbionts are acquired from the environment (horizontal transmission), rather than inherited from their parents (vertical transmission). However, it is unknown how abundant and diverse the symbionts are in the environment, critical for the acquisition and selection of symbionts from the environment. Hence, the fifth chapter aimed to answer these questions. To retrieve microorganisms in the vent environment, basaltic blocks were deployed for one year at four locations, i.e., among, adjacent and away from tubeworm aggregations, as well as off axis. Quantitative PCR was used to determine the copies of symbiont16S rRNA and ITS, which occur as single copy per genome. Results showed that the abundance of free-living symbionts was highest among tubeworm aggregation, but decreased dramatically with increasing distance. This indicated either replenishment of symbionts through hosts or proliferation of free-living symbionts. The symbiont diversity was investigated through pyrosequencing of ITS region from trophosome tissue and seawater filter. Surprisingly, results suggested tubeworm symbiont population was quite homogeneous within and outside tubeworms at vents, with only one dominant phylotype and three others were found.Another important question is how the symbionts utilize chemicals and produce nutrients for themselves and their hosts. Since these tubeworms are gutless and mouthless, they depend on their chemosynthetic symbionts for nutrients including carbon, nitrogen and sulfur etc. The source of nitrogen is of particular interest, since it is indispensible and various, but underemphasized. Tubeworm species, Ridgeia piscesae, was used as a model to examine nitrogen metabolism at various environmental nitrogen concentrations and of different Ridgeia phenotypes, i.e., long skinny (LS) and short fat (SF) Ridgeia. Gene expression analyses suggested that ammonium was assimilated by symbionts of both phenotypes at sites with different ammonium concentrations, but minor amounts of nitrate respiration and no nitrate assimilation were detected. Differences were found between phenotypes and vent sites, suggesting nitrogen utilization was probably influenced by environmental nitrogen availability and may be different between phenotypes.To summarize, this thesis was developed with a focus on microbial diversity from phylogenetic, metabolic, and functional perspectives. From free-living microorganisms to symbionts of vent invertebrates, this thesis demonstrated a broad view of marine microorganisms. Results promoted our understanding of marine microorganisms and updated the current knowledge. Interesting hypotheses proposed here can be developed into future projects.