2008 Center for Microbial Oceanography Research and Education (CMORE) Summer Course “Microbial Oceanography Genomes to Biomes” Student Symposium, June 6th, 2008 University of Hawaii

2008 Center for Microbial Oceanography: Research and Education (CMORE) Summer Course “Microbial Oceanography: Genomes to Biomes” Student Symposium, June 6th, 2008 University of Hawaii Rebecca Case Sophie Clayton Alejandro Andrés Murillo Córdova Li Deng Caroline Fortunato Melissa Laghdass Christopher Reisch Claudia Hernandez Rondón Sandra Schưttner Christine Shulse Yuna Seo Nellie Shaul Danielle Morgan-Smith Elena Stoica Frank K Thomson, III Laure-Anne Ventouras Harvard University MIT/WHOI Joint Program Pontificia Universidad Católica de Chile The University of Bristol University of Maryland CNRS/UMR-France University of Georgia Estación de Fotobiología Playa Unión-Argentina Max Planck Institute for Marine Microbiology Scripps Institution of Oceanography The University of Tokyo Scripps Institution of Oceanography Old Dominion University University of Liverpool Old Dominion University Massachusetts Institute of Technology REBECCA CASE AFFILIATION: Harvard University TITLE: Bacterial-algal interactions and the sulfur cycle ABSTRACT: Algae produce and store the major biogenic source of sulfur in the marine environment, dimethylsulfoniopropionate (DMSP) DMSP is an osmolyte in algae and is made accessible for microbial degradation in the ocean through lysis of algal cells DMSP is degraded by bacteria through the demethylation/demethiolation pathway or the cleavage pathway The former pathway degrades DMSP to sulfate, while the latter pathways product is dimethyl sulfide (DMS) DMS is an important intermediate in the sulfur cycle and influences climate through the formation of cloud condensation nuclei (CCN) Model calculations have shown a (theoretical) negative feedback between DMS producing communities and global mean temperature, suggesting these communities could play a role in climate SOPHIE CLAYTON AFFILIATION: MIT/WHOI Joint Program TITLE: Investigating controls on the seasonal cycle of /Synechococcus/ biomass at MVCO using a simple box model ABSTRACT: It is generally thought that the interaction of physical and biological processes in the open ocean results in a relatively constant background of small phytoplankton cells This has not, however, been fully investigated in coastal ecosystems Recent observations from the Martha's Vineyard Coastal Observatory (MVCO), made using an automated submersible flow cytometer, have shown that this does not seem to be the case there (Sosik & Olson, 2006) Time series of picoplankton biomass, comprised mainly of /Synechococcus/ and picoeukaryotes, from 2003 to the present, show that there is an increase in picoplankton biomass in the spring, sustained throughout the summer, followed by a decrease in the autumn Temperature has been suggested to play a role in limiting growth during the winter months, but the combination of physical and biological processes operating to regulate picoplankton biomass during the spring and autumn transition periods is not well understood The aim of this study, therefore, is to use a simple box model which incorporates temperature-dependent growth, nutrient limitation and a closure term representing the sum of all loss processes to Synechococcus (e.g grazing, mortality, advection,etc ) to investigate the controls on these seasonal variations at MVCO The model is optimized to find the parameters which best fit the observations, and the model-data misfit assessed quantitatively The structure of the model is modified, and the subsequent model-data misfits compared, to test hypotheses about which environmental factors control the seasonal cycle of /Synechococcus/ biomass ALEJANDRO ANDRÉS MURILLO CÓRDOVA AFFILIATION: Pontificia Universidad Católica de Chile TITLE: Cylindrospermopsis raciborskii has model organism for PSP toxins pathway synthesis study ABSTRACT: In Chile, annually in summer an algal bloom (red tide) occur in the sea in austral regions and the microorganism responsible is the dinoflagellate Alexandrium Catenella a eukaryotic microalga that produce saxitoxin (STX) and their analogs, this neurotoxins act blocking the voltage gate sodium channel and can be accumulated by bivalves, producing the “Paralytic Shellfish Poisoning” (PSP) syndrome In the case of fresh water sources the algal blooms with de concomitant toxin contamination are produced by cyanobacteria Cyanobacteria are photosynthetic bacteria that are capable of produce a wide variety of injurious compounds toxics or irritants The species of cyanobacteria that are STX producers are Aphanizomenon flos-aquae, Anabaena circinalis, Lyngbya wollei, Cylindrospermopsis raciborskii and Planktothrix agardhii The pathway of synthesis of STX or genes related with the toxin remains unknown for all the producer organisms, but we know that the precursors are the same for cyanobacteria and dinoflagellate Among the STX producers microorganisms the cyanobacteria are de best model of study, especially de specie Cylindrospermopsis raciborskii, because it has strains that produce toxin; STX and their analogs or cylindrospermopsin and strains non toxin producers and all the strain are genetically homogeneous Our objective is to characterize the toxin producer strains genetically and functionally to identified de enzymatic machinery involved in STX production LI DENG AFFILIATION: The University of Bristol TITLE: The role of cyanophages in regulating bloom-forming freshwater cyanobacteria ABSTRACT: Cyanobacteria are important and diverse members of aquatic systems both in marine and freshwater environments Cyanobacterial viruses (cyanophages) in marine systems have been shown to play roles in mortality of their hosts, as well as driving evolution of core metabolic functions even photosynthesis Thus I wondered what roles cyanophages might play in freshwater systems where cyanobacterial genera were predominately filamentous and bloomforming, rather than unicellular non-bloom-forming cells To this end, I used multiple genetic loci (T4-phage portal protein, major capsid protein) to gain an overall understanding of the phylogenetic diversity of these genes within natural phage communities in a couple of European lakes I next isolated 35 cyanophages that infect bloom-forming freshwater cyanobacteria (Microcystis, Anabaena and Planktothrix) using viral-fraction waters from Lake Zurich, Switzerland and lakes in the Cotswold Water Park, U.K Morphological characterization revealed a variety of morphotypes, including commonly observed tailed phage morphotypes (e.g., podo-, myo-, siphoviruses), as well as the first filamentous cyanophage Host range characterization showed that some cyanophages were capable of infecting cyanobacterial hosts across three genera The ability to infect a wide range of host taxa which dominant water bodies in variable depth and variable time extends the potential reproductive period for lytic propagation of cyanophages, and also has implications for the transfer of genetic information between phylogenetically disparate cyanobacterial lineages A primary example of such viralmediated HGT might be the discovery of a phycocyanin gene previously thought to be horizontally transferred in the host cells and was found in a cyanophage isolate A-CP1 by PCR screen with substantive controls to minimize the probability of amplifying a host gene Phycocyanin is ubiquitous accessory photosynthesis gene thought to be important in host metabolism during phage infection, but also previously hypothesized to have been horizontally transferred is my phage a smoking gun for a mechanism? CAROLINE FORTUNATO AFFILIATION: University of Maryland TITLE: Microbial community observation and prediction in the Columbia River, its estuary and the adjacent coastal ocean ABSTRACT: Microbial communities are dynamic and vary over many environmental gradients The goal of this study is to determine the composition of the microbial communities of the Columbia River system and how the different populations of this community are distributed from river to estuary and across the coastal ocean A 16S rRNA gene based microbial community record will be created using the community fingerprinting technique, denaturing gradient gel electrophoresis (DGGE) Once a map of microbial communities is established, shifts in the distribution of individual microbial populations over space and time will then be connected with environmental factors and linked to forecast models of coastal margin environmental conditions Currently, cruise data from August 2007 is being used to construct a system-wide map of microbial population distributions, with future plans to examine seasonal and interannual changes in these populations MELISSA LAGHDASS AFFILIATION: CNRS/UMR France TITLE: External sources of organic carbon in marine oligotrophic environment: degradation by different functional and phylogenetic groups of the heterotrophic prokaryotes and impact on the metabolic balance ABSTRACT: The metabolic balance (ration between respiration and production of the organic matter) depends directly of the ecosystem functioning Studies suggest that the respiration exceed the production in the oligotrophic ocean causing a visible deficit of organic carbon This deficit can be explain either by a decoupling between production and respiration or by an external source of organic matter that would provide the respiration A temporal monitoring of this deficit is also essential in order to understand the origin of this deficit Besides, the heterotrophic prokaryotes contribute to the essential of the respiration in the oligotrophic environment Their metabolic activity plays also a key role in the metabolic balance The variability of the prokaryotic respiration depends essentially of the bioavailability of organic matter and the nutrient concentration Furthermore, the nature of organic matter would play a role in the presence and activity of phylogenetic groups The identification of these groups is very important in order to understand their role in the metabolic balance In situ observations started in the NW Mediterranean Sea (MOLA station) on February 2007 Samples are carrying out once a month to determine the metabolic balance and identify the phylogenetic prokaryotes of heterotrophic prokaryotes The metabolic balance is evaluated by winckler method Using molecular biology, oligonucleotidic probes specific of bacterial groups or target probes will create So, by combining microautoradiography and in situ hybridization from created probes, activity of these groups could determine An experimental approach based on the chemostat culture is too envisaged to know the response of different bacterial groups according to the sources of organic matter CHRISTOPHER REISCH AFFILIATION: University of Georgia TITLE: Purification and Characterization of DMSP Dependent Demethylase (DmdA) from icMarine Bacteria ABSTRACT: The algal metabolite dimethylsulfoniopropionate (DMSP) is ubiquitous in marine environments, ranging in concentration from 5-200 nM in surface waters DMSP is the precursor of the volatile compound dimethylsulfide, whh is the primary natural source of sulfur in the atmosphere Degradation of DMSP occurs through two competing pathways, yielding DMS in the cleavage pathway or methanethiol through the demethylation pathway Recently, the enzyme responsible for the demethylation of DMSP, designated DmdA, was discovered in two marine isolates, Silicibacter pomeroyi DSS-3 and Pelagibacter ubique HTCC1062 To further our understanding of DMSP demethylation, recombinant DmdA from P ubique was purified The purified enzyme exhibited strict substrate specificity for DMSP The apparent Vmax of DmdA was 7.6µmols min-1 mg-1 with an apparent Km for DMSP of 11.5 mM In cell extracts of S pomeroyi DSS-3 the apparent Km for DMSP was 10.2 mM, similar to that of purified recombinant DmdA The intracellular concentration of DMSP in chemostat-grown S pomeroyi DSS-3 was found to range from 66-93 mM, depending on growth conditions These results suggest that cells actively accumulate DMSP as an osmolyte Moreover, initiation of DMSP demethylation by DmdA only occurs at very high intracellular concentrations CLAUDIA HERNÁNDEZ RONDĨN AFFILIATION: Estación de Fotobiología Playa Unión-Argentina Title: Solar radiation effects on bacterioplankton of Patagonia ABSTRACT: The decrease on stratospheric Ozone levels and the rise of incident solar radiation (mainly on UVB range) affects aquatic systems Even under normal levels, solar and UVB radiation generates variety of responses (typically deleterious) on metabolic activity, physiology and diversity of microorganisms Therefore, the bacterioplankton response is related with light penetration and with biogeochemical composition of each ecosystem Chubut area (43° S, Argentinean Patagonia) possesses strong winds that turn in low cloud cover and consequently high levels of solar radiation along the year A decade of punctual studies at this zone evidenced the impact of solar radiation (PAR and UVR).At the Chubut River “mouth” there is high primary and fishery production present Photoinhibition and repair processes related to cell size were reported mainly for phytoplankton In the case of heterotrophic bacteria, there are not internal protection mechanisms to deal with solar stress I developed punctual experiments at the pier with river and seawater (according to tides) along summer Previous results showed mainly UVB effects on bacterioplankton abundance, and growth At the same time samples to check bacterial diversity changes by solar stress are a work on progress SANDRA SCHÖTTNER AFFILIATION: Max Plank Institute for Marine Microbiology TITLE: Microbial habitats in cold-water coral reef ecosystems ABSTRACT: The potential of key ecosystems like cold-water coral reefs to foster enhanced species diversity in the ocean is undisputed However, little is known about the putative role that cold-water coral reefs play as biodiversity hotspots for the “unseen majority” in the oceans: prokaryotic communities Our current efforts evolve around the central question, if cold-water corals act as ecosystem engineers in their reef environment by shaping microbial diversity, activity and functionality via habitat differentiation and release of organic matter Pilot studies in 2006/2007 revealed that bacterial communities may associate very specifically with distinct microbial habitats generated by the reef-building coral Lophelia pertusa, such as the coral skeleton surface, mucus and tissue when compared to ambient sediments and seawater In particular, coral mucus was characterized by distinct and very diverse bacterial patterns, and has shown to act as possible energy vector from corals to microbes in reef waters Whether these trends have a general validity for cold-water coral reef ecosystems is currently under investigation in more extensive studies targeting the diversity, dynamics and potential function of prokaryotes along horizontal and vertical gradients in different reef systems off Northern Norway and Chile CHRISTINE SCHULSE AFFILIATION: Scripps Institution of Oceanography TITLE: Evolutionary origins and genetic regulation of omega-3 polyunsaturated fatty acid synthase gene clusters in marine bacteria ABSTRACT: Bacterial production of long-chain omega-3 polyunsaturated fatty acids (PUFAs), such as EPA (20:5) and DHA (22:6), is constrained to a narrow subset of marine Ỵ³-proteobacteria The availability of multiple genomes from both PUFA+ and closely related PUFA- strains allows for a comparative investigation as to the evolutionary origins of PUFA synthase gene clusters (pfaAE) It is possible that the presence of the pfa gene cluster in different organisms that share the same environment came about as a result of lateral gene transfer Despite potential physiological advantages conferred by PUFA production under low temperature or high pressure regimes (membrane fluidity/phase dynamics), results to date suggest alternative selective advantages possibly unrelated to thermal- or piezotolerance Currently, little is known regarding PUFA synthesis in these bacteria other than that it proceeds via a novel polyketide synthesis mechanism In order to explore the genetic regulation and coordinated production of PUFAs, a recombinant EPA producing E coli strain has been engineered and is being utilized to explore the biochemical pathway of PUFA synthesis Multifunctional domain profiling of pfa gene products is revealing the biochemical structure and molecular machinery responsible for PUFA production and comparisons amongst Pfa synthases reveal the basis for EPA versus DHA discrimination Further biochemical exploration of the different functional domains in the Pfa proteins via domain swapping and combinatorial synthesis will provide a recipe of domains necessary for PUFA synthesis Ultimately, the collation of genomic, genetic, and biochemical data will be used to further explore the ecological function of bacterial PUFA production in the marine environment YUNA SEO AFFILIATION: The University of Tokyo TITLE: Particle Capture by Marine Bacteria ABSTRACT: In aquatic environments large numbers of submicron particles are generally present, and their turnover and fate are of considerable ecological importance in geochemical cycles Efficient degradation of submicron particles can be attained if the particles are retained on the surfaces of bacterial cells rather than remaining suspended in the aqueous phase I hypothesized that some bacterial groups possess particle-capture ability To verify this hypothesis, I took two approaches First, paramagnetic particles were added to natural seawater samples to separate out bacterial groups with PC capabilities Method development was carried out in combination with community structure analysis of the separated bacteria by denaturing gradient gel electrophoresis and multidimensional scaling Bacterial groups recovered by this new approach were different from either the representative ones in the original seawater sample or large particle-associated bacterial groups Second, atomic force microscopy was utilized to directly observe the submicron particles within natural bacterial populations The numbers of cells retaining submicron particles were high in the inner part of Tokyo Bay and at depths of 500 to 2000 m in the open ocean I suggest that there are bacterial groups in natural seawater that possess PC ability NELLIE SHAUL AFFILIATION: Scripps Institution of Oceanography TITLE: Investigating the biotic formation and degradation of methoxylated polybrominated diphenyl ethers (MeO-PBDEs) in marine ecosystems ABSTRACT: Many halogenated organic compounds were considered to be of anthropogenic origin, but there is now evidence for their production in nature For example, MeO-PBDEs have been shown to accumulate in whale tissues, and radiocarbon evidence indicates a natural, ubiquitous, marine origin We are currently investigating answers to the following questions: What are the ubiquitous biotic marine source(s)? Are the natural analogs accumulating in the marine environment? Can bacteria in the marine environment degrade these natural MeO-PBDEs? Investigation of these questions will occur in several phases: extraction and analysis of dissolved and particulate organic matter to detect MeO-PBDEs; search GenBank environmental databases for biosynthetic clusters similar to what has been found in sponge systems to identify pelagic biotic sources; and incubate natural assemblages of bacteria with 13C-labeled MeO-PBDEs to investigate incorporation and degradation Examining sources and fates of halogenated aromatic compounds in the environment remains as a vital component to our understanding of contaminant biogeochemistry If we are to understand and quantify the persistence and transformation of industrial analogs, we must first characterize biotic production and distribution of related compounds in environmental systems This research would help to fulfill vital niches in contaminant biogeochemistry, marine microbial ecology, and remediation DANIELLE MORGAN-SMITH AFFILIATION: Old Dominion University TITLE: Quantification and community composition of deep-sea flagellates based on nuclear morphology and fluorescence in situ hybridization ABSTRACT: The pelagic deep ocean is the largest habitat on earth, yet we know very little about the distribution and activity of protists in this environment Flagellates, for instance, which are responsible for controlling bacterial abundance in the surface ocean, and which are major players in the material and energy transfer of pelagic food webs, have not been studied to any significant extent in the deep sea For this project, water samples were taken in the eastern subtropical North Atlantic at depths ranging from 100m to 5000m and were filtered onto membranes which underwent tyramide signal amplification fluorescence in situ hybridization (TSA-FISH) using oligonucleotide probes of eukaryote-specific sequences These filters were then scanned for positive TSA-FISH and DAPI signals using an epifluorescence microscope with a robotic stage We compared filters across depth and geographic area in terms of abundance of eukaryotes and by community composition based on DAPI nuclear morphology In comparison to prokaryotes, numbers of eukaryotes drop off disproportionately with depth ELENA STOICA AFFILIATION: University of Liverpool TITLE: In situ identification of planktonic ammonia-oxidizing bacteria (AOB) in the NW Black Sea coast ABSTRACT: Ammonia oxidation, the first step in nitrification, is a key process in the marine N cycle that results in the formation of nitrate trough activity of microbial (bacterial and archaeal) ammoniaoxidizers Both aerobic and anaerobic ammonia oxidation have been studied in the central part of the Black Sea, the largest marine anoxic basin in the world However, no data on taxonomic diversity and ecological roles of ammonia-oxidizers community are available for the NW Black Sea, a dynamic region largely influenced by Danube River and anthropogenic factors We investigated the presence and relative abundance of the main prokaryotic taxa for Bacteria and ammonia-oxidizing bacteria (AOB) by a combination of whole-cell fluorescence in situ hybridization with rRNA-targeted oligonucleotide probes and catalyzed reporter deposition (CARD-FISH) With horseradish peroxidase-labeled oligonucleotide probes we specifically detected Bacteria, beta- and Gamma-Proteobacteria and the major groups of ammonia-oxidizing bacteria (Beta-AOB and Gamma-AOB) in the Danube-Black Sea mixing zone during 2007 CARD-FISH analysis indicated that planktonic AOB might constitute a significant fraction of the bacterioplankton in the NW Black Sea coastal system, and show distinct spatial abundance patterns in relation to the prevailing ecological conditions FRANK K THOMSON, III AFFILIATION: Old Dominion University TITLE: Ships Ballast as a Potential Vector for the Transfer of Antibiotic Resistance Genes Among Coastal Environments ABSTRACT: Ships ballast has been implicated as a vector in the dispersal of invasive species into new environments Numerous cases have been documented for the introduction of macroorganisms via ballasting operations, but what is less known is the role of ballast water and residuals in the spread of potentially harmful microbes, especially with reference to novel genotypes Of particular interest are the genes encoding for various forms of antibiotic resistance, many of which are carried on mobile genetic elements A three-year sampling effort has yielded over 300 putative isolates of Vibrio cholerae from ships' ballast tanks and various environmental sources, of which 208 have been profiled for antibiotic susceptibility using twelve diverse antibiotics The results demonstrate widespread resistance to b-lactams (74.6%), with isolated instances of resistance to other antibiotics Plasmid extractions and restriction enzyme analyses have shown the presence of an ostensible plasmid of approximately 38 kbp in many of these isolates Additional work is being done using restriction fragment length polymorphisms and PCR for specific antibiotic resistance genes in an attempt to characterize these plasmids Knowledge from this study and future microcosm experiments will help us to ascertain the potential for horizontal gene transfer in a ballast tank setting LAURE-ANNE VENTOURAS AFFILIATION: MIT ABSTRACT: Despite extensive efforts to understand and predict the effects that recent anthropogenic activity will have on the physico-chemical properties of the ocean, much is still unknown as to how the marine biosphere will be impacted To understand how these unnatural perturbations specifically affect microbial communities, we will use a metagenomics approach to analyze the results of mesocosm perturbation experiments Changes at the gene expression level will be monitored using 454-sequencing technology, while changes at the community structure level will be monitored using a genome-proxy microarray technology that was developed in-house Currently I am working on applying the genome-proxy microarray to study the changes in microbial community structure over time and depths at the ALOHA-HOT station The genome-proxy microarray allows sampling of both the phylogenetic and functional breadth of a community by using probes that target the gen ome sections from a variety of uncultivated microbes After gaining good understanding of the background structure of the microbial assemblages of the North Pacific Tropical Gyre, I will carry out the mesocosm perturbation experiments that will either focus on changing temperature, pH, heavy metal or specific pollutant concentrations  ... hypothesized to have been horizontally transferred is my phage a smoking gun for a mechanism? CAROLINE FORTUNATO AFFILIATION: University of Maryland TITLE: Microbial community observation and prediction... map of microbial communities is established, shifts in the distribution of individual microbial populations over space and time will then be connected with environmental factors and linked to forecast... culture is too envisaged to know the response of different bacterial groups according to the sources of organic matter CHRISTOPHER REISCH AFFILIATION: University of Georgia TITLE: Purification and