Viruses influence the number and types of organism that compose microbial communities, biogeochemical cycles and the flux and chemical character of carbon and nutrients in marine surface waters. Though it is increasingly feasible to make quantitative measures of the impact of virus activity on microbial processes, our understanding of the interplay between the multitude of host-virus interactions that give rise to ecological scale properties remains in its infancy. This is especially true for lysogenic (temperate) viruses, i.e., viruses with the ability to integrate into their host's genome and remain "dormant" until prompted, by external or internal cues, to initiate a lytic cycle leading to cell rupture and virus release. Recent evidence suggests lysogenic viruses are the dominate type of marine viruses. Thus, indicating that the decision to "lyse or not to lyse" is a prevalent, but poorly understood, process in marine microbial communities. Despite the recognized and profound influence of viruses on marine systems, relatively little effort has been placed on characterizing host-virus interactions of environmental relevance. This project will study the lysogenic viruses that infect members of the ecologically important Roseobacter clade of heterotrophic marine bacteria. Roseobacter abundance is greatest in coastal environments, phytoplankton blooms and in association with organic particles, where these microbes frequently grow in a gel-like matrix, called biofilms, with multiple other species. Indeed, biofilms are hot spots for both microbial and virus activity, and provide an opportunity to better define the impact that viruses have on host physiology and function. This research will address how genome-integrated viruses influence host metabolism and behavior and, in turn, how these host-viruses interactions influence marine microbial biofilm structure and function. The described project will directly support the interdisciplinary training of four graduates, one of which will be involved in training by international scientists. Training opportunities described under currently funded REU programs in both Microbiology and Chemistry at the University of Tennessee will be leveraged and extended to undergraduates participating in the research presented here. All PIs will continue to disseminate their research results in presentations, papers and other forms on a timely basis and data will be shared using public repositories.
The research plan is organized into three objectives: (i) Characterize the role of growth conditions on the lysogenic-lytic switch, (ii) identify genetic determinants that dictate the lysogenic-lytic switch, (iii) quantify the effect of host-virus interactions on microbial biofilm development and function in single and mixed species biofilms. To achieve these objectives, the investigators will use mass spectrometry based high-throughput metabolomics and lipidomics approaches to test the hypothesis that growth substrate will differentially influence host metabolism, particularly intracellular cAMP concentrations, and result in different rates of lysis vs. lysogeny. The project will use a classic, yet highly effective, technique to isolate random mutations in phage (clear plaque mutants) that render them incapable of integration into the host genome (i.e., they will be fixed in the lytic state). Phage mutants will be confirmed by physiological studies and characterized by sequence analysis. To better understand the influence of host-virus interactions on microbial activities that are relevant to ocean carbon cycling vis-a-vis the Biological Pump, the investigators will perform a series of experiments with our host lysogens in single and mixed species biofilms grown on lysate from the phytoplankton species Emiliana huxleyi. The investigators will quantify differences in biofilm biomass, morphological structure, microbial activity and chemical composition of the extracellular matrix. The results from this research will provide the foundational steps necessary to develop a broader understanding of virus effects on an ecologically relevant component of oceanic bacterial communities. From this, it will be possible to begin to extrapolate and quantify the role of this interaction in regional scale carbon cycling.
