Marine sponges are hosts to complex microbial communities that may comprise up to 60% of the sponge biomass. Little is known about the roles of these symbiotic bacteria and their contributions to the sponge and thereby indirectly to the coral reef ecosystem in which the sponges live. The hypothesis driving the project is that diverse assemblages of nitrogen-fixing bacteria play an important role in providing sponges with nitrogen, taking nitrogen gas and converting it to organic forms of nitrogen that can be used by sponges. The role of bacterial symbionts of two sponges from Key Largo, Florida, Ircinia strobilina and Mycale laxissima in nitrogen fixation will be determined. The project will include biochemical approaches to demonstrate nitrogen fixation in whole sponges and bacterial isolates, and molecular approaches to study the pattern of where and when the nifH gene is turned on and makes the NifH protein, a key protein in nitrogen fixation. Training will be provided in marine microbiology to graduate students and undergraduate minority students will be encouraged to enter careers in scientific research by participating in a summer course in Marine Microbiology. The project will advance our understanding of microbial diversity and function in complex sponge symbioses, promoting the study of sponge symbioses as models of the complex symbiotic relationships found between microbes and many higher organisms. The project will provide new information on sources of nitrogen for fragile coral reef environments.