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Marine bryozoans Bugula neritina and Watersipora spp. and their proteobacterial symbionts


Better understanding the symbioses between the widely spread bryozoans Bugula neritina and Watersipora spp. and their respective bacterial symbionts is the goal of my dissertation research. The symbiosis between B. neritina and its uncultivated gamma-proteobacterial symbiont "Candidatus Endobugula sertula" is the best understood bryozoan-bacteria symbiosis. It had long been hypothesized that bryostatins, potent anti-cancer compounds isolated from B. neritina, were actually produced by "E. sertula," indicating a role in chemical defense for the bacterial symbionts. In this study, in situ hybridizations were used to directly link "E. sertula" to mRNA from a large polyketide synthase (PKS) gene cluster that is hypothesized to encode bryostatin biosynthesis genes. RNase protection assays (RPAs) and reverse transcription- PCR (RT-PCR) were used to detect transcripts from natural B. neritina samples throughout the ̃77 kb PKS cluster. These results suggest that the cluster is functional which is consistent with its hypothesized role in bryostatin biosynthesis. Symbioses between bacteria and bryozoans in the genus Watersipora are less understood. The consistent association between bacteria and Watersipora arcuata was reported in 1983, but after the cultivation-based identification of the bacterial symbionts as mollicutes in 1989, nothing further was reported on the symbiosis until now. In this study PCR, DNA sequencing and fluorescence in situ hybridizations (FISH) of Watersipora larvae with symbiont specific probes were used to identify the uncultivated bacterial symbionts of Watersipora spp. from several locations along the California coast. The bacterial symbionts of W. "subtorquata" and W. "arcuata" turned out to be closely related alpha-proteobacteria, not mollicutes, and were named "Candidatus Endowatersipora palomitas" and "Candidatus Endowatersipora rubus," respectively. The systematics of Watersipora are very confused, so this study included the cataloging of host sample cytochrome C oxidase subunit I (COI) gene sequences, light micrographs and scanning electron micrographs (SEMs). COI gene sequence-based phylogenetic analyses of Watersipora were also conducted. The location of bacterial symbionts in W. "subtorquata" at non-larval developmental stages was also investigated using FISH. A preliminary chemical analysis using liquid chromatography- mass spectrometry (LC-MS) of W. "subtorquata" adult and larval extracts revealed the presence of 1,8- dihydroxyanthraquinone in adults and larvae and genistein in larvae

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