Benjamin A. S. Van Mooy

Tracking Hydrocarbon Plume Transport and Biodegradation at Deepwater Horizon

Diving into Deep Water The Deepwater Horizon oil spill in the Gulf of Mexico was one of the largest oil spills on record. Its setting at the bottom of the sea floor posed an unanticipated risk as substantial amounts of hydrocarbons leaked into the deepwater column. Three separate cruises identified and sampled deep underwater hydrocarbon plumes that existed in May and June, 2010—before the well head was ultimately sealed. Camilli et al. (p. 201; published online 19 August) used an automated underwater vehicle to assess the dimensions of a stabilized, diffuse underwater plume of oil that was 22 miles long and estimated the daily quantity of oil released from the well, based on the concentration and dimensions of the plume. Hazen et al. (p. 204; published online 26 August) also observed an underwater plume at the same depth and found that hydrocarbon-degrading bacteria were enriched in the plume and were breaking down some parts of the oil. Finally, Valentine et al. (p. 208; published online 16 September) found that natural gas, including propane and ethane, were also present in hydrocarbon plumes. These gases were broken down quickly by bacteria, but primed the system for biodegradation of larger hydrocarbons, including those comprising the leaking crude oil. Differences were observed in dissolved oxygen levels in the plumes (a proxy for bacterial respiration), which may reflect differences in the location of sampling or the aging of the plumes. In late June 2010, the Deepwater Horizon oil plume stretched more than 35 kilometers at a depth of 1100 meters. The Deepwater Horizon blowout is the largest offshore oil spill in history. We present results from a subsurface hydrocarbon survey using an autonomous underwater vehicle and a ship-cabled sampler. Our findings indicate the presence of a continuous plume of oil, more than 35 kilometers in length, at approximately 1100 meters depth that persisted for months without substantial biodegradation. Samples collected from within the plume reveal monoaromatic petroleum hydrocarbon concentrations in excess of 50 micrograms per liter. These data indicate that monoaromatic input to this plume was at least 5500 kilograms per day, which is more than double the total source rate of all natural seeps of the monoaromatic petroleum hydrocarbons in the northern Gulf of Mexico. Dissolved oxygen concentrations suggest that microbial respiration rates within the plume were not appreciably more than 1 micromolar oxygen per day.

Phytoplankton in the ocean use non-phosphorus lipids in response to phosphorus scarcity

Phosphorus is an obligate requirement for the growth of all organisms; major biochemical reservoirs of phosphorus in marine plankton include nucleic acids and phospholipids. However, eukaryotic phytoplankton and cyanobacteria (that is, ‘phytoplankton’ collectively) have the ability to decrease their cellular phosphorus content when phosphorus in their environment is scarce. The biochemical mechanisms that allow phytoplankton to limit their phosphorus demand and still maintain growth are largely unknown. Here we show that phytoplankton, in regions of oligotrophic ocean where phosphate is scarce, reduce their cellular phosphorus requirements by substituting non-phosphorus membrane lipids for phospholipids. In the Sargasso Sea, where phosphate concentrations were less than 10 nmol l-1, we found that only 1.3 ± 0.6% of phosphate uptake was used for phospholipid synthesis; in contrast, in the South Pacific subtropical gyre, where phosphate was greater than 100 nmol l-1, plankton used 17 ± 6% (ref. 6). Examination of the planktonic membrane lipids at these two locations showed that classes of sulphur- and nitrogen-containing membrane lipids, which are devoid of phosphorus, were more abundant in the Sargasso Sea than in the South Pacific. Furthermore, these non-phosphorus, ‘substitute lipids’ were dominant in phosphorus-limited cultures of all of the phytoplankton species we examined. In contrast, the marine heterotrophic bacteria we examined contained no substitute lipids and only phospholipids. Thus heterotrophic bacteria, which compete with phytoplankton for nutrients in oligotrophic regions like the Sargasso Sea, appear to have a biochemical phosphorus requirement that phytoplankton avoid by using substitute lipids. Our results suggest that phospholipid substitutions are fundamental biochemical mechanisms that allow phytoplankton to maintain growth in the face of phosphorus limitation.

Composition and fate of gas and oil released to the water column during the Deepwater Horizon oil spill

Quantitative information regarding the endmember composition of the gas and oil that flowed from the Macondo well during the Deepwater Horizon oil spill is essential for determining the oil flow rate, total oil volume released, and trajectories and fates of hydrocarbon components in the marine environment. Using isobaric gas-tight samplers, we collected discrete samples directly above the Macondo well on June 21, 2010, and analyzed the gas and oil. We found that the fluids flowing from the Macondo well had a gas-to-oil ratio of 1,600 standard cubic feet per petroleum barrel. Based on the measured endmember gas-to-oil ratio and the Federally estimated net liquid oil release of 4.1 million barrels, the total amount of C1-C5 hydrocarbons released to the water column was 1.7 × 1011 g. The endmember gas and oil compositions then enabled us to study the fractionation of petroleum hydrocarbons in discrete water samples collected in June 2010 within a southwest trending hydrocarbon-enriched plume of neutrally buoyant water at a water depth of 1,100 m. The most abundant petroleum hydrocarbons larger than C1-C5 were benzene, toluene, ethylbenzene, and total xylenes at concentrations up to 78 μg L-1. Comparison of the endmember gas and oil composition with the composition of water column samples showed that the plume was preferentially enriched with water-soluble components, indicating that aqueous dissolution played a major role in plume formation, whereas the fates of relatively insoluble petroleum components were initially controlled by other processes.

Impact of suboxia on sinking particulate organic carbon: Enhanced carbon flux and preferential degradation of amino acids via denitrification

Fluxes of particulate organic carbon (POC) through the oxygen deficient waters in the eastern tropical North Pacific were found to be relatively less attenuated with depth than elsewhere in the eastern North Pacific. The attenuation coefficient (b) for the flux was found to be 0.40 versus the composite value of 0.86 determined by Martin et al. (1987). To examine this further, sinking POC was collected using sediment traps and allowed to degrade in oxic and suboxic experiments. Using a kinetic model, it was found that degradation proceeded at similar rates (roughly 0.8 day−1) under oxic and suboxic conditions, but a greater fraction of bulk POC was resistant to degradation in the suboxic experiments (61% vs. 23%). Amino acids accounted for 37% of POC collected at 75m, but following degradation the value dropped to 17% and 16% in the oxic and suboxic experiments respectively. POC collected from 500m was 10% amino acids. The non-AA component of POC collected at 75m was not degraded under suboxic conditions, while under oxic conditions it was. These results suggest that microbes degrading OC under suboxic conditions via denitrification preferentially utilize nitrogen-rich amino acids. This preferential degradation of amino acids suggests that 9% more nitrogen may be lost via water column denitrification than is accounted for when a more “Redfieldian” stoichiometry for POC is assumed.

Viral Glycosphingolipids Induce Lytic Infection and Cell Death in Marine Phytoplankton

The Death of Cocco Emiliania huxleyi is a coccolithophore, a class of unicellular phytoplankton that forms vast blooms mediating the oceanic carbon cycle through shedding of its calcium carbonate scales. E. huxleyi is routinely infected and killed by lytic viruses that can abruptly halt a bloom. Vardi et al. (p. 861) have found that in E. huxleyi strains that are sensitive or resistant to infection, a sphingolipid-based “arms race” appears to regulate cell fate during host-virus interactions. The lipid also serves as a biomarker for active infection that may help to quantify the role and activity of viruses and virus-mediated processes in the oceans. This information will help in assessing the biogeochemical impact of these plankton species. A specific virus encodes membrane components that broadcast cell death and population demise of its coccolithophore host. Marine viruses that infect phytoplankton are recognized as a major ecological and evolutionary driving force, shaping community structure and nutrient cycling in the marine environment. Little is known about the signal transduction pathways mediating viral infection. We show that viral glycosphingolipids regulate infection of Emiliania huxleyi, a cosmopolitan coccolithophore that plays a major role in the global carbon cycle. These sphingolipids derive from an unprecedented cluster of biosynthetic genes in Coccolithovirus genomes, are synthesized de novo during lytic infection, and are enriched in virion membranes. Purified glycosphingolipids induced biochemical hallmarks of programmed cell death in an uninfected host. These lipids were detected in coccolithophore populations in the North Atlantic, which highlights their potential as biomarkers for viral infection in the oceans.

Cryptic carbon and sulfur cycling between surface ocean plankton

Significance In the surface ocean, organic matter released by phytoplankton and degraded by heterotrophic bacteria is a key step in the carbon cycle. Compounds important in this trophic link are poorly known, in part because of the thousands of chemicals making up marine dissolved organic matter. We cocultured a Roseobacter clade bacterium with the diatom Thalassiosira pseudonana and used gene expression changes to assay for compounds passed to the bacterium. A C3-sulfonate with no previously known role in the microbial food web was identified and subsequently shown to be an abundant diatom metabolite and actively cycling compound in seawater. This work identifies a missing component of the marine carbon and sulfur cycles. About half the carbon fixed by phytoplankton in the ocean is taken up and metabolized by marine bacteria, a transfer that is mediated through the seawater dissolved organic carbon (DOC) pool. The chemical complexity of marine DOC, along with a poor understanding of which compounds form the basis of trophic interactions between bacteria and phytoplankton, have impeded efforts to identify key currencies of this carbon cycle link. Here, we used transcriptional patterns in a bacterial-diatom model system based on vitamin B12 auxotrophy as a sensitive assay for metabolite exchange between marine plankton. The most highly up-regulated genes (up to 374-fold) by a marine Roseobacter clade bacterium when cocultured with the diatom Thalassiosira pseudonana were those encoding the transport and catabolism of 2,3-dihydroxypropane-1-sulfonate (DHPS). This compound has no currently recognized role in the marine microbial food web. As the genes for DHPS catabolism have limited distribution among bacterial taxa, T. pseudonana may use this sulfonate for targeted feeding of beneficial associates. Indeed, DHPS was both a major component of the T. pseudonana cytosol and an abundant microbial metabolite in a diatom bloom in the eastern North Pacific Ocean. Moreover, transcript analysis of the North Pacific samples provided evidence of DHPS catabolism by Roseobacter populations. Other such biogeochemically important metabolites may be common in the ocean but difficult to discriminate against the complex chemical background of seawater. Bacterial transformation of this diatom-derived sulfonate represents a previously unidentified and likely sizeable link in both the marine carbon and sulfur cycles.

Host–virus dynamics and subcellular controls of cell fate in a natural coccolithophore population

Marine viruses are major evolutionary and biogeochemical drivers in marine microbial foodwebs. However, an in-depth understanding of the cellular mechanisms and the signal transduction pathways mediating host–virus interactions during natural bloom dynamics has remained elusive. We used field-based mesocosms to examine the “arms race” between natural populations of the coccolithophore Emiliania huxleyi and its double-stranded DNA-containing coccolithoviruses (EhVs). Specifically, we examined the dynamics of EhV infection and its regulation of cell fate over the course of bloom development and demise using a diverse suite of molecular tools and in situ fluorescent staining to target different levels of subcellular resolution. We demonstrate the concomitant induction of reactive oxygen species, caspase-specific activity, metacaspase expression, and programmed cell death in response to the accumulation of virus-derived glycosphingolipids upon infection of natural E. huxleyi populations. These subcellular responses to viral infection simultaneously resulted in the enhanced production of transparent exopolymer particles, which can facilitate aggregation and stimulate carbon flux. Our results not only corroborate the critical role for glycosphingolipids and programmed cell death in regulating E. huxleyi–EhV interactions, but also elucidate promising molecular biomarkers and lipid-based proxies for phytoplankton host–virus interactions in natural systems.

Remodeling of intermediate metabolism in the diatom Phaeodactylum tricornutum under nitrogen stress

Significance When starved for nutrients, diatoms redirect carbon toward biosynthesis of storage lipids, triacylglycerols (TAGs). We examined how this modification is achieved in the diatom Phaeodactylum tricornutum. Under nitrogen stress, the cells cannibalized their photosynthetic apparatus while recycling intracellular nitrogen and redirecting it to synthesize nitrogen assimilation enzymes. Simultaneously, they allocated newly fixed carbon toward lipids. In contrast, a nitrate reductase knocked-down strain shunted ∼40% more carbon toward TAGs than the wild type without losing photosynthetic capacity. Our results show that diatoms can remodel their intermediate metabolism on environmental cues and reveal that a key signal in this remodeling is associated with nitrogen assimilation. This insight informs a strategy of developing a much more efficient pathway to produce algal-based biofuels. Diatoms are unicellular algae that accumulate significant amounts of triacylglycerols as storage lipids when their growth is limited by nutrients. Using biochemical, physiological, bioinformatics, and reverse genetic approaches, we analyzed how the flux of carbon into lipids is influenced by nitrogen stress in a model diatom, Phaeodactylum tricornutum. Our results reveal that the accumulation of lipids is a consequence of remodeling of intermediate metabolism, especially reactions in the tricarboxylic acid and the urea cycles. Specifically, approximately one-half of the cellular proteins are cannibalized; whereas the nitrogen is scavenged by the urea and glutamine synthetase/glutamine 2-oxoglutarate aminotransferase pathways and redirected to the de novo synthesis of nitrogen assimilation machinery, simultaneously, the photobiological flux of carbon and reductants is used to synthesize lipids. To further examine how nitrogen stress triggers the remodeling process, we knocked down the gene encoding for nitrate reductase, a key enzyme required for the assimilation of nitrate. The strain exhibits 40–50% of the mRNA copy numbers, protein content, and enzymatic activity of the wild type, concomitant with a 43% increase in cellular lipid content. We suggest a negative feedback sensor that couples photosynthetic carbon fixation to lipid biosynthesis and is regulated by the nitrogen assimilation pathway. This metabolic feedback enables diatoms to rapidly respond to fluctuations in environmental nitrogen availability.

Rapid microbial respiration of oil from the Deepwater Horizon spill in offshore surface waters of the Gulf of Mexico

The Deepwater Horizon oil spill was one of the largest oil spills in history, and the fate of this oil within the Gulf of Mexico ecosystem remains to be fully understood. The goal of this study—conducted in mid-June of 2010, approximately two months after the oil spill began—was to understand the key role that microbes would play in the degradation of the oil in the offshore oligotrophic surface waters near the Deepwater Horizon site. As the utilization of organic carbon by bacteria in the surface waters of the Gulf had been previously shown to be phosphorus limited, we hypothesized that bacteria would be unable to rapidly utilize the oil released from the Macondo well. Although phosphate was scarce throughout the sampling region and microbes exhibited enzymatic signs of phosphate stress within the oil slick, microbial respiration within the slick was enhanced by approximately a factor of five. An incubation experiment to determine hydrocarbon degradation rates confirmed that a large fraction of this enhanced respiration was supported by hydrocarbon degradation. Extrapolating our observations to the entire area of the slick suggests that microbes had the potential to degrade a large fraction of the oil as it arrived at the surface from the well. These observations decidedly refuted our hypothesis. However, a concomitant increase in microbial abundance or biomass was not observed in the slick, suggesting that microbial growth was nutrient limited; incubations amended with nutrients showed rapid increases in cell number and biomass, which supported this conclusion. Our study shows that the dynamic microbial community of the Gulf of Mexico supported remarkable rates of oil respiration, despite a dearth of dissolved nutrients.

Phosphorus supply drives rapid turnover of membrane phospholipids in the diatom Thalassiosira pseudonana

In low-phosphorus (P) marine systems, phytoplankton replace membrane phospholipids with non-phosphorus lipids, but it is not known how rapidly this substitution occurs. Here, when cells of the model diatom Thalassiosira pseudonana were transferred from P-replete medium to P-free medium, the phospholipid content of the cells rapidly declined within 48 h from 45±0.9 to 21±4.5% of the total membrane lipids; the difference was made up by non-phosphorus lipids. Conversely, when P-limited T. pseudonana were resupplied with P, cells reduced the percentage of their total membrane lipids contributed by a non-phosphorus lipid from 43±1.5 to 7.3±0.9% within 24 h, whereas the contribution by phospholipids rose from 2.2±0.1 to 44±3%. This dynamic phospholipid reservoir contained sufficient P to synthesize multiple haploid genomes, suggesting that phospholipid turnover could be an important P source for cells. Field observations of phytoplankton lipid content may thus reflect short-term changes in P supply and cellular physiology, rather than simply long-term adjustment to the environment.