Marine Microbiology Research from the Gordon and Betty Moore Foundation Marine Microbiology Initiative
Jed Fuhrman’s lab studies the distribution, diversity, and activities of a wide range of microorganisms living in marine systems, including viruses, bacteria, archaea, protists. Despite great strides in our understanding about the importance and diversity of marine microbial communities, most of what we know tends to show how the system works in a piecemeal fashion rather than as a big picture. Marine microorganisms are controlled by many influences, which include externally-imposed features like fundamental characteristics of their physical and chemical surroundings, as well as complex interactions among the organisms, such as cross-feeding, competition, cooperation, predation, and infection. This project examines these multiple levels of control, with a wide and integrated viewpoint rather than a narrow focus. Specifically, we aim to generate a fundamental understanding of principal interactions and feedback loops among major players of marine microbial communities. To achieve this, we intend to simultaneously address how nutrient inputs, microbial predators, and viral infection work together in complex ways to control microbial communities and their activities. Most of the fieldwork for this project will be done at our long-standing USC Microbial Observatory site at the San Pedro Ocean Time Series, studied since 2000 with support from the USC Wrigley Institute for Environmental Studies and the National Science Foundation, and at USC’s Wrigley Marine Science Center on Santa Catalina Island.
Specific project goals include the following:
- Characterize the cascading influence of predators on their prey through the links in the microbial food web, via field observations and experiments, using a combination of statistical & network analysis and single cell genomic and metagenomic data.
- Characterize how viral infection of microbial hosts influences food web processes, via a combination of statistical & network analysis and metagenomic and single cell genomic data.
- Connect bacterial and archaeal growth with specific potential sources of nutrition, such as dissolved chemicals or particular phytoplankton taxa, and specific environmental conditions.
- Evaluate interactions among bacteria and archaea, such as competition and possibly cooperation, via network analysis, metagenomic and single cell genomic data.
- Characterize and model the connections between predators, viruses, and bacteria, as well as competition or cooperation among these microbes, to integrate the complex interactions.
Dimensions in Biodiversity Project (NSF supported 2011-2015).
Pattern and Process in Marine Bacterial, Archaeal, and Protistan Biodiversity, and Effects of Human Impacts.
PIs: Jed Fuhrman, David Caron, John Heidelberg, William Nelson, Fengzhu Sun, Ting Chen.
Bacteria, Archaea, and Protists dominate global elemental cycling and are immensely diverse genetically, taxonomically, and functionally. Yet the extent of marine microbial diversity, its patterns, and relationships among genetic, taxonomic, and functional diversity are very poorly characterized, even though the ocean covers 70% of the planet’s surface. Among the least well known variables is the effect of human impacts on native marine microbial systems, although it is recognized that impacted systems are more prone to events like harmful algal blooms. Knowledge of these relationships and impacts are necessary to anticipate the responses of biota to global changes and feedback mechanisms that may alter the extents, rates, and even pathways of such changes. This project will expand upon an existing NSF-funded 10+-year monthly ocean time series (Microbial Observatory) that has focused on a single site midway between Los Angeles and Santa Catalina Island, to also include quarterly sampling adjacent to the impacted LA Harbor region to the barely-impacted Catalina coast. USC already runs facilities in LA Harbor and Catalina, with daily boats between (no cost). Measurements include (1) Genetic diversity: high throughput DNA sequences of “housekeeping” and functional genes. (2) Taxonomic diversity: high throughput tag sequences of small subunit ribosomal RNA genes, flow cytometry, automated image analysis (3) Functional Diversity: (a) Functional measurements (carbon fixation and respiration rates, microbial growth and grazing rates, cell size, morphology, and biomass variations), (b) distribution and expression of particular target functional genes involved with processes central to the cycles of carbon, nitrogen, and sulfur, (c) exploratory metatranscriptomics (evaluation of whole-community RNA) to explore functionalities that were not anticipated. (4) Integrating these: Multivariate statistical and network approaches including newly developed techniques (e.g. Bayesian networks to examine cause-effect relationships), and high speed computational approaches to assess the relationships among the genetic, taxonomic, and functional aspects of biodiversity observed. We will also examine the collected data for signatures and specific effects (on organism identity and functions) associated with human impacted harbor site vs. the relatively pristine one.
Integration: We will use network and time series analysis, along with other statistical tools to integrate “classical” microbial and oceanographic rate process measurements, flow cytometric and microscopic characterizations of communities, along with targeted as well as untargeted metagenomics (whole community DNA analysis) and metatranscriptomics to relate genetic & taxonomic diversity with specific functions (at organismal, food web, and system levels). For example, our results should be able to determine how different variants of particular taxa (e.g. at resolution levels ranging from what might be considered near the subspecies to genus levels) would differ in their association with particular measured functions, functional genes, or particular other taxa – or we might see how particular clusters of related organisms “behave” similarly or differently in their associations. This project offers an unprecedented and potentially transformative opportunity to combine and integrate measurements of genetic, taxonomic, and functional diversity along with direct measurements of system function in a well studied marine system that includes a gradient from one of the world’s busiest harbors to a largely pristine ocean habitat. Far beyond just describing the distributions of organisms and functions (itself a necessary first step), we will specifically link spatial and temporal variations in a variety of functions with variations in genetic and taxonomic community composition.
The USC Microbial Observatory (NSF supported 2000-2012)
The USC Microbial Observatory (Jed Fuhrman and David Caron, PIs) focuses on exploratory investigation of prokaryotic and unicellular eukaryotic diversity in the San Pedro Channel , California, with an initial focus on time-dependent changes in community composition in relation to environmental parameters. It also includes focused studies of particular microbial groups.
We are currently in 12th year of this project. Our primary sampling site is located midway between Los Angeles and the USC Wrigley Marine Laboratory on Santa Catalina approximately 900 m of water. This site is visited monthly by ship for sampling to 500 m depth. Additional sampling is conducted on an ad hoc basis in coastal water near the lab on Catalina Island.
Viruses in Marine Food Webs (NSF support 2010-2013).
Viruses are the most abundant biological agents in marine plankton and have a tremendous potential impact on biological oceanographic processes, including material & energy flow as well as plankton diversity. Molecular genetic methods to investigate diversity of host bacteria and viruses (including molecular fingerprinting and next generation sequencing of 16S rRNA and other genes for bacterial hosts, fingerprinting and sequencing of marker genes for specific viral groups such as T4-like phage ) permit us to begin unraveling the relationships between virus and host diversity in marine plankton, and how it may affect overall rate processes. This project uses such approaches to study natural and experimental marine microbial systems and address basic hypotheses on the relationships between bacterial and viral communities. Study sites include the San Pedro Ocean Time Series (SPOT) site, midway between Los Angeles and Catalina Island , run by the USC Wrigley Institute for Environmental Studies, and also richer coastal waters near Los Angeles and Catalina Island . Part of the project is continuation of a monthly time series at SPOT of virus abundance and virus community composition at 5 depths to 890 m, to augment other microbiological and oceanographic observations being made at that location as part of the Microbial Observatory Project. In addition to the general time series databasing component which continues to provide valuable microbial community data in its own right, we also use this sampling to investigate hypotheses about viral control of specific common groups of marine bacteria, such as cyanobacteria, marine alpha proteobacteria, and the SAR11 cluster. Short-term, more frequent sampling and experimental mesocosms are also used to examine community and specific subgroup dynamics on the generation time scale, and to examine virus-bacteria association networks. The project has the added value of taking advantage of a unique time series of microbial diversity at an easily accessible marine site, suitable for further analysis such testing hypotheses relating diversity and stability in marine microbes. Samples (e.g. preserved virus concentrates, microbial DNA) are also available for retrospective analysis. Exploratory investigation of virus diversity and activity in sediments is also included as a component of this project
Human Pathogens in the Coastal Zone
Ongoing research in our lab on viruses at Southern California beaches has been funded for several years by USC Sea Grant. This has included measurement of human pathogenic viruses, comparison to bacterial indicators, and development of rapid and convenient molecular biological tests suitable for standardized testing. Such work has been coordinated with local water quality agencies (e.g. Orange County Sanitation District) and user groups (e.g. Surfrider, Santa Monica Baykeeper). Coordination has been facilitated by USC Sea Grant and Southern California Coastal Water Research Project (SCCWRP)
Global Survey of Marine Prokaryotes
Prokaryotes are organisms that lack a membrane-bound nucleus, and they consist of two very different groups of microorganisms, Bacteria and Archaea. They are ubiquitous and immensely important in all global elemental cycles as well as all biological systems in general. The focus of our research is the marine environment. In that environment, like many others, the diversity of naturally-occurring prokaryotes is poorly known. It has only been in the last decade or so, with the advent of molecular biological techniques in this field, that we have the ability to say what kinds of bacteria and archaea live in seawater. We have been using such techniques, originally pioneered by Norman Pace and his colleagues, to study naturally-occurring marine microbes by means of 16S rRNA gene sequences. This approach has provided a valuable and extremely broad phylogenetic framework of classification, and has the huge benefit that it can be done without culturing the organisms. The results are in the form of sequence data, and interpretation within the context of a large existing database is via phylogenetic analysis computer programs.
In our study, we obtained seawater from numerous locations around the world (see map link), collected the organisms on a filter, extracted their DNA, amplified partial 16S rRNA genes, cloned those PCR products, sequenced them, and analyzed the sequences phylogenetically. Detailed scientific protocols can be found in our research publications (e.g. Fuhrman, J.A., and A.A. Davis.1997. Widespread Archaea and novel Bacteria from the deep sea as shown by 16S rRNA gene sequences. Mar. Ecol. Prog. Ser. 150: 275-285). This research has discovered the existence of numerous groups of organisms that were previously unknown. Perhaps the most exciting discovery was that archaea are common in the deep sea (Fuhrman, J.A., K. McCallum, and A.A. Davis. 1992. Novel major archaebacterial group from marine plankton. Nature 356: 148-149). Previously, the archaea were thought to include only “extremophiles” such as hyperthermophiles, halophiles, and methanogens. We have also found numerous novel types of bacteria by this approach.
Phylogenetic trees that include the various sequences we have obtained from seawater samples around the world can be accessed by the accompanying links. There is also a key to clone identifiers. All sequences are available from GenBank. Examination of the trees shows that many groups, such as the SAR 11 cluster, are globally distributed, with some distinct subgroups in warmer surface waters and others in deeper and polar waters. However, not all of the samples had members of the SAR 11 cluster. The cyanobacteria are ubiquitous in warm surface waters, but not found in polar areas. All the cyanobacterial clones, with the exception of a few clones from near Singapore, wererelated to cultivated marine Prochlorococcus and Synechococcus. Numerous other themes are evident in the data.