Wein, Tanita (2016) Analysis of Curvibacter sp. colonization dynamics in Hydra and construction of a mobilizable gene knockout system. (Master thesis), Christian-Albrechts-Universität zu Kiel, Kiel, Germany, 77 pp.

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Abstract

All plants and animals host a microbiota, an assemblage of diverse microbes whose symbiotic species often have vital functions for health, development and reproduction of their hosts. Although the microbiota displays degrees of host-specificity, genetic mechanisms underlying colonization and symbiosis formation remain to be understood. The freshwater polyp Hydra vu/garis strain AEP, a member the early-branching Cnidaria, has been shown to host a distinctive bacterial microbiota, comprising mainly members of the Proteobacteria and Bacteroidetes. The main colonizer was found to be Curvibacter sp., a Gram negative β-proteobacterium found in fresh water systems. One clone, Curvibacter AEP 1.3, was maintained in pure culture, sequenced and automatically annotated. Given these prerequisites, a definition of requirements to establish Curvibacter as a model for the study of the Hydra holobiont was necessary. In the first step, conditions for transfer of exogenous DNA by electroporation and conjugation were explored and defined replicative plasmids tested for heterologous gene expression. Subsequently colonization of germfree Hydra polyps with Curvibacter equipped with fluorophore-encoding plasmids was successfully monitored. Furthermore, colonization related candidate genes were identified and knockout plasmids were constructed to create mutants to be screened for aberrant colonization patterns in Hydra. Results give first insights into a yet native model organism and will allow to experimentally elucidate host-microbe interactions within Hydra in the future.

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