Bacterial-invertebrate symbioses: from an asphalt cold seep to shallow water

Abstract

Symbiotic associations are complex partnerships that can lead to new metabolic capabilities and the establishment of novel organisms. The diversity of these associations is very broad and there are still many mysteries about the origin and the exact relationship between the organisms that are involved in a symbiosis (host and symbiont). Some of these associations are essential to the hosts, such as the chemosynthetic symbioses occurring in invertebrates of the deep-sea. In others the host probably would rather not be the host, as in the case of parasitic microbes. My PhD research focuses on symbiotic and parasitic associations in chemosynthetic and non-chemosynthetic invertebrates. This thesis describes and discusses three different aspects of associations between bacteria and marine invertebrates. The first aspect focuses on chemosynthetic associations from a unique asphalt seep called Chapopote in the Gulf of Mexico (GoM). Phylogenetic analyses of host genes (cytochrome-c-oxidase subunit I) and bacterial genes (16S rRNA) in two Bathymodiolus mussel species and an Escarpia tubeworm showed that both the hosts and their chemosynthetic symbionts are very similar to their congeners from the northern GoM. Unexpectedly, a novel symbiont most closely related to hydrocarbon degrading bacteria of the genus Cycloclasticus was discovered in B. heckerae. Stable carbon isotope values in B. heckerae tissues of lipids typical for Cycloclasticus spp. were consistently heavier by 2.5permil than other lipids indicating that the novel symbiont might use isotopically heavy hydrocarbons from the asphalt seep as an energy and carbon source. The discovery of a novel symbiont that may be able to metabolize hydrocarbons is particularly intriguing because until now only methane and reduced sulfur compounds have been identified as energy sources in chemosynthetic symbioses. The large amounts of hydrocarbons available at Chapopote would provide these mussel symbioses with a rich source of nutrition. The second aspect of this thesis deals with bacteria that infect the nuclei of marine invertebrates and were recently found to be widespread in deep-sea Bathymodiolus mussels. Because of their potentially lethal effect on bivalve populations, I looked for the presence of intranuclear bacteria in economically important and commercially available bivalve species, i.e. oysters (Crassostrea gigas), razor clams (Siliqua patula and Ensis directus), blue mussels (Mytilus edulis), Manila clams (Venerupis philippinarum), and common cockles (Cerastoderma edule). Fluorescence in situ hybridization (FISH) revealed the presence of intranuclear bacteria in all investigated bivalves except oysters and blue mussels. Preliminary tests with real-time PCR showed massive amounts of intranuclear bacteria in some of the bivalve species, raising the question if these might affect not only the health of the bivalves but possibly also of the humans that eat them. In the third and final aspect of my thesis, I examined the general diversity of bacteria in the gill tissues of deep-sea and shallow-water mussels and clams. Comparative 16S rRNA sequence analysis and cultivation experiments revealed a much higher diversity than previously recognized. This thesis shows that bivalves are ideal models for studying the microbiota of marine invertebrates because of the high diversity of both highly specific and more generalized symbiotic and parasitic bacteria in their gill tissues.