The core seafloor microbiome in the Gulf of Mexico is remarkably consistent and shows evidence of recovery from disturbance caused by major oil spills.

The microbial ecology of oligotrophic deep ocean sediments is understudied relative to their shallow counterparts, and this lack of understanding hampers our ability to predict responses to current and future perturbations. The Gulf of Mexico has experienced two of the largest accidental marine oil spills, the 1979 Ixtoc-1 blowout and the 2010 Deepwater Horizon (DWH) discharge. Here, microbial communities were characterized for 29 sites across multiple years in > 700 samples. The composition of the seafloor microbiome was broadly consistent across the region and was well approximated by the overlying water depth and depth within the sediment column, while geographic distance played a limited role. Biogeographical distributions were employed to generate predictive models for over 4000 OTU that leverage easy-to-obtain geospatial variables which are linked to measured sedimentary oxygen profiles. Depth stratification and putative niche diversification are evidenced by the distribution of taxa that mediate the microbial nitrogen cycle. Furthermore, these results demonstrate that sediments impacted by the DWH spill had returned to near baseline conditions after 2 years. The distributions of benthic microorganisms in the Gulf can be constrained, and moreover, deviations from these predictions may pinpoint impacted sites and aid in future response efforts or long-term stability studies.

Decadal Assessment of Polycyclic Aromatic Hydrocarbons in Mesopelagic Fishes from the Gulf of Mexico Reveals Exposure to Oil-Derived Sources.

This study characterizes a decadal assessment of polycyclic aromatic hydrocarbons (PAHs) in the muscle tissues of mesopelagic fish species as indicators of the environmental health of the Gulf of Mexico (GoM) deep-pelagic ecosystem. Mesopelagic fishes were collected prior to the Deepwater Horizon (DWH) oil spill (2007), immediately post-spill (2010), 1 year after the spill (2011), and 5-6 years post-spill (2015-2016) to assess if the mesopelagic ecosystem was exposed to, and retained, PAH compounds from the DWH spill. Results indicated that a 7- to 10-fold increase in PAHs in fish muscle tissues occurred in 2010-2011 (4972 ± 1477 ng/g) compared to 2007 (630 ± 236 ng/g). In 2015-2016, PAH concentrations decreased close to the levels measured in 2007 samples (827 ± 138 ng/g); however, the composition of PAHs still resembles a petrogenic source similar to samples collected in 2010-2011. PAH composition in muscle samples indicated that natural sources (e.g., Mississippi River and natural seeps) or spatial variability within the GoM do not explain the temporal variability of PAHs observed from 2007 to 2016. Furthermore, analysis of different fish tissues indicated the dietary intake and maternal transfer of PAHs as the primary mechanisms for bioaccumulation in 2015-2016, explaining the elevated levels and composition of PAHs in ovarian eggs.

Hydrocarbon-Degrading Bacteria Exhibit a Species-Specific Response to Dispersed Oil while Moderating Ecotoxicity.

The Deepwater Horizon blowout in April 2010 represented the largest accidental marine oil spill and the largest release of chemical dispersants into the environment to date. While dispersant application may provide numerous benefits to oil spill response efforts, the impacts of dispersants and potential synergistic effects with crude oil on individual hydrocarbon-degrading bacteria are poorly understood. In this study, two environmentally relevant species of hydrocarbon-degrading bacteria were utilized to quantify the response to Macondo crude oil and Corexit 9500A-dispersed oil in terms of bacterial growth and oil degradation potential. In addition, specific hydrocarbon compounds were quantified in the dissolved phase of the medium and linked to ecotoxicity using a U.S. Environmental Protection Agency (EPA)-approved rotifer assay. Bacterial treatment significantly and drastically reduced the toxicity associated with dispersed oil (increasing the 50% lethal concentration [LC50] by 215%). The growth and crude oil degradation potential of Acinetobacter were inhibited by Corexit by 34% and 40%, respectively; conversely, Corexit significantly enhanced the growth of Alcanivorax by 10% relative to that in undispersed oil. Furthermore, both bacterial strains were shown to grow with Corexit as the sole carbon and energy source. Hydrocarbon-degrading bacterial species demonstrate a unique response to dispersed oil compared to their response to crude oil, with potentially opposing effects on toxicity. While some species have the potential to enhance the toxicity of crude oil by producing biosurfactants, the same bacteria may reduce the toxicity associated with dispersed oil through degradation or sequestration.

An integrated evaluation of bioenhanced in situ LNAPL dissolution.

Performance evaluation of in situ bioremediation processes in the field is difficult due to uncertainty created by matrix and contaminant heterogeneity, inaccessibility to direct observation, expense of sampling, and limitations of some measurements. The goal of this research was to develop a strategy for evaluating in situ bioremediation of light nonaqueous-phase liquid (LNAPL) contamination and demonstrating the occurrence of bioenhanced LNAPL dissolution by: (1) integrating a suite of analyses into a rational evaluation strategy; and (2) demonstrating the strategy's application in intermediate-scale flow-cell (ISFC) experiments simulating an aquifer contaminated with a pool of LNAPL (naphthalene dissolved in dodecane). Two ISFCs were operated to evaluate how the monitored parameters changed between a "no bioremediation" scenario and an "intrinsic in situ bioremediation" scenario. Key was incorporating different measures of microbial activity and contaminant degradation relevant to bioremediation: contaminant loss; consumption of electron acceptors; and changes in total alkalinity, pH, dissolved total inorganic carbon, carbon-stable isotopes, microorganisms, and intermediate metabolites. These measurements were integrated via mass-flux modeling and mass-balance analyses to document that in situ biodegradation of naphthalene was strongly accelerated in the "intrinsic in situ bioremediation" scenario versus "no bioremediation." Furthermore, the integrated strategy provided consistent evidence of bioenhancement of LNAPL dissolution through intrinsic bioremediation by a factor of approximately 2 due to the biodegradation of the naphthalene near the pool/water interface.

The southern Gulf of Mexico: A baseline radiocarbon isoscape of surface sediments and isotopic excursions at depth.

The southern Gulf of Mexico (sGoM) is home to an extensive oil recovery and development infrastructure. In addition, the basin harbors sites of submarine hydrocarbon seepage and receives terrestrial inputs from bordering rivers. We used stable carbon, nitrogen, and radiocarbon analyses of bulk sediment organic matter to define the current baseline isoscapes of surface sediments in the sGoM and determined which factors might influence them. These baseline surface isoscapes will be useful for accessing future environmental impacts. We also examined the region for influence of hydrocarbon deposition in the sedimentary record that might be associated with hydrocarbon recovery, spillage and seepage, as was found in the northern Gulf of Mexico (nGoM) following the Deepwater Horizon (DWH) oil spill in 2010. In 1979, the sGoM experienced a major oil spill, Ixtoc 1. Surface sediment δ13C values ranged from -22.4‰ to -19.9‰, while Δ14C values ranged from -337.1‰ to -69.2‰. Sediment δ15N values ranged from 2.8‰ to 7.2‰, while the %C on a carbonate-free basis ranged in value of 0.65% to 3.89% and %N ranged in value of 0.09% to 0.49%. Spatial trends for δ13C and Δ14C were driven by water depth and distance from the coastline, while spatial trends for δ15N were driven by location (latitude and longitude). Location and distance from the coastline were significantly correlated with %C and %N. At depth in two of twenty (10%) core profiles, we found negative δ13C and Δ14C excursions from baseline values in bulk sedimentary organic material, consistent with either oil-residue deposition or terrestrial inputs, but likely the latter. We then used 210Pb dating on those two profiles to determine the time in which the excursion-containing horizons were deposited. Despite the large spill in 1979, no evidence of hydrocarbon residue remained in the sediments from this specific time period.

Tracing the incorporation of carbon into benthic foraminiferal calcite following the Deepwater Horizon event.

Following the Deepwater Horizon (DWH) event in 2010, hydrocarbons were deposited on the continental slope in the northeastern Gulf of Mexico through marine oil snow sedimentation and flocculent accumulation (MOSSFA). The objective of this study was to test the hypothesis that benthic foraminiferal δ13C would record this depositional event. From December 2010 to August 2014, a time-series of sediment cores was collected at two impacted sites and one control site in the northeastern Gulf of Mexico. Short-lived radioisotopes (210Pb and 234Th) were employed to establish the pre-DWH, DWH, and post-DWH intervals. Benthic foraminifera (Cibicidoides spp. and Uvigerina spp.) were isolated from these intervals for δ13C measurement. A modest (0.2-0.4‰), but persistent δ13C depletion in the DWH intervals of impacted sites was observed over a two-year period. This difference was significantly beyond the pre-DWH (background) variability and demonstrated that benthic foraminiferal calcite recorded the depositional event. The longevity of the depletion in the δ13C record suggested that benthic foraminifera may have recorded the change in organic matter caused by MOSSFA from 2010 to 2012. These findings have implications for assessing the subsurface spatial distribution of the DWH MOSSFA event.

Large-scale deposition of weathered oil in the Gulf of Mexico following a deep-water oil spill.

The blowout of the Deepwater Horizon (DWH) drilling rig in 2010 released an unprecedented amount of oil at depth (1,500 m) into the Gulf of Mexico (GoM). Sedimentary geochemical data from an extensive area (∼194,000 km2) was used to characterize the amount, chemical signature, distribution, and extent of the DWH oil deposited on the seafloor in 2010-2011 from coastal to deep-sea areas in the GoM. The analysis of numerous hydrocarbon compounds (N = 158) and sediment cores (N = 2,613) suggests that, 1.9 ± 0.9 × 104 metric tons of hydrocarbons (>C9 saturated and aromatic fractions) were deposited in 56% of the studied area, containing 21± 10% (up to 47%) of the total amount of oil discharged and not recovered from the DWH spill. Examination of the spatial trends and chemical diagnostic ratios indicate large deposition of weathered DWH oil in coastal and deep-sea areas and negligible deposition on the continental shelf (behaving as a transition zone in the northern GoM). The large-scale analysis of deposited hydrocarbons following the DWH spill helps understanding the possible long-term fate of the released oil in 2010, including sedimentary transformation processes, redistribution of deposited hydrocarbons, and persistence in the environment as recycled petrocarbon.

Sediment Core Extrusion Method at Millimeter Resolution Using a Calibrated, Threaded-rod.

Aquatic sediment core subsampling is commonly performed at cm or half-cm resolution. Depending on the sedimentation rate and depositional environment, this resolution provides records at the annual to decadal scale, at best. An extrusion method, using a calibrated, threaded-rod is presented here, which allows for millimeter-scale subsampling of aquatic sediment cores of varying diameters. Millimeter scale subsampling allows for sub-annual to monthly analysis of the sedimentary record, an order of magnitude higher than typical sampling schemes. The extruder consists of a 2 m aluminum frame and base, two core tube clamps, a threaded-rod, and a 1 m piston. The sediment core is placed above the piston and clamped to the frame. An acrylic sampling collar is affixed to the upper 5 cm of the core tube and provides a platform from which to extract sub-samples. The piston is rotated around the threaded-rod at calibrated intervals and gently pushes the sediment out the top of the core tube. The sediment is then isolated into the sampling collar and placed into an appropriate sampling vessel (e.g., jar or bag). This method also preserves the unconsolidated samples (i.e., high pore water content) at the surface, providing a consistent sampling volume. This mm scale extrusion method was applied to cores collected in the northern Gulf of Mexico following the Deepwater Horizon submarine oil release. Evidence suggests that it is necessary to sample at the mm scale to fully characterize events that occur on the monthly time-scale for continental slope sediments.

Was the extreme and wide-spread marine oil-snow sedimentation and flocculent accumulation (MOSSFA) event during the Deepwater Horizon blow-out unique?

During the Deepwater Horizon blowout, thick layers of oiled material were deposited on the deep seafloor. This large scale benthic concentration of oil is suggested to have occurred via the process of Marine Oil Snow Sedimentation and Flocculent Accumulation (MOSSFA). This meta-analysis investigates whether MOSSFA occurred in other large oil spills and identifies the main drivers of oil sedimentation. MOSSFA was found to have occurred during the IXTOC I blowout and possibly during the Santa Barbara blowout. Unfortunately, benthic effects were not sufficiently studied for the 52 spills we reviewed. However, based on the current understanding of drivers involved, we conclude that MOSSFA and related benthic contamination may be widespread. We suggest to collect and analyze sediment cores at specific spill locations, as improved understanding of the MOSSFA process will allow better informed spill responses in the future, taking into account possible massive oil sedimentation and smothering of (deep) benthic ecosystems.

Non-lethal approach identifies variability of δ (15)N values in the fin rays of Atlantic Goliath Grouper, Epinephelus itajara.

The Atlantic Goliath Grouper, Epinephelus itajara, is critically endangered throughout its range but has begun to show initial signs of recovery in Florida state waters. As the population continues to rebound, researchers face a pressing need to fill the knowledge gaps about this iconic species. Here, we examined the δ (15)N isotopic records in fin rays collected from Atlantic Goliath Grouper, and related changes of isotopic ratios over time to life history characteristics. Fin-ray analysis was used as a non-lethal technique to sample individuals from two locations at similar latitudes from the west and east coasts of Florida, USA. δ (15)N data were acquired by mechanically separating the annuli of each fin ray and then analyzing the material in an Irradiance Elemental Analyzer Mass Spectrometer. The δ (15)N values were consistent among individuals within populations from each coast of Florida, and mirrored the expected changes over the lives of the fish. Overall, differences were found between δ (15)N values at juvenile life history phases versus adult phases, but the patterns associated with these differences were unique to each coastal group. We demonstrated, for the first time, that δ (15)N values from fin rays can be used to assess the life histories of Atlantic Goliath Grouper. The non-lethal strategies outlined here can be used to acquire information essential to the management of species of concern, such as those that are threatened or endangered.

Hydrocarbons in Deep-Sea Sediments following the 2010 Deepwater Horizon Blowout in the Northeast Gulf of Mexico.

The Deepwater Horizon (DWH) spill released 4.9 million barrels of oil into the Gulf of Mexico (GoM) over 87 days. Sediment and water sampling efforts were concentrated SW of the DWH and in coastal areas. Here we present geochemistry data from sediment cores collected in the aftermath of the DWH event from 1000-1500 m water depth in the DeSoto Canyon, NE of the DWH wellhead. Cores were analyzed at high-resolution (at 2 mm and 5 mm intervals) in order to evaluate the concentration, composition and input of hydrocarbons to the seafloor. Specifically, we analyzed total organic carbon (TOC), aliphatic, polycyclic aromatic hydrocarbon (PAHs), and biomarker (hopanes, steranes, diasteranes) compounds to elucidate possible sources and transport pathways for deposition of hydrocarbons. Results showed higher hydrocarbon concentrations during 2010-2011 compared to years prior to 2010. Hydrocarbon inputs in 2010-2011 were composed of a mixture of sources including terrestrial, planktonic, and weathered oil. Our results suggest that after the DWH event, both soluble and highly insoluble hydrocarbons were deposited at enhanced rates in the deep-sea. We proposed two distinct transport pathways of hydrocarbon deposition: 1) sinking of oil-particle aggregates (hydrocarbon-contaminated marine snow and/or suspended particulate material), and 2) advective transport and direct contact of the deep plume with the continental slope surface sediments between 1000-1200 m. Our findings underline the complexity of the depositional event observed in the aftermath of the DWH event in terms of multiple sources, variable concentrations, and spatial (depth-related) variability in the DeSoto Canyon, NE of the DWH wellhead.

A decline in benthic foraminifera following the deepwater horizon event in the northeastern Gulf of Mexico.

Sediment cores were collected from three sites (1000-1200 m water depth) in the northeastern Gulf of Mexico from December 2010 to June 2011 to assess changes in benthic foraminiferal density related to the Deepwater Horizon (DWH) event (April-July 2010, 1500 m water depth). Short-lived radioisotope geochronologies (²¹°Pb, ²³4Th), organic geochemical assessments, and redox metal concentrations were determined to relate changes in sediment accumulation rate, contamination, and redox conditions with benthic foraminiferal density. Cores collected in December 2010 indicated a decline in density (80-93%). This decline was characterized by a decrease in benthic foraminiferal density and benthic foraminiferal accumulation rate (BFAR) in the surface 10 mm relative to the down-core mean in all benthic foraminifera, including the dominant genera (Bulimina spp., Uvigerina spp., and Cibicidoides spp.). Cores collected in February 2011 documented a site-specific response. There was evidence of a recovery in the benthic foraminiferal density and BFAR at the site closest to the wellhead (45 NM, NE). However, the site farther afield (60 NM, NE) recorded a continued decline in benthic foraminiferal density and BFAR down to near-zero values. This decline in benthic foraminiferal density occurred simultaneously with abrupt increases in sedimentary accumulation rates, polycyclic aromatic hydrocarbon (PAH) concentrations, and changes in redox conditions. Persistent reducing conditions (as many as 10 months after the event) in the surface of these core records were a possible cause of the decline. Another possible cause was the increase (2-3 times background) in PAH's, which are known to cause benthic foraminifera mortality and inhibit reproduction. Records of benthic foraminiferal density coupled with short-lived radionuclide geochronology and organic geochemistry were effective in quantifying the benthic response and will continue to be a valuable tool in determining the long-term effects of the DWH event on a larger spatial scale.

Effects of preservation methods of muscle tissue from upper-trophic level reef fishes on stable isotope values (δ (13)C and δ (15)N).

Research that uses stable isotope analysis often involves a delay between sample collection in the field and laboratory processing, therefore requiring preservation to prevent or reduce tissue degradation and associated isotopic compositions. Although there is a growing literature describing the effects of various preservation techniques, the results are often contextual, unpredictable and vary among taxa, suggesting the need to treat each species individually. We conducted a controlled experiment to test the effects of four preservation methods of muscle tissue from four species of upper trophic-level reef fish collected from the eastern Gulf of Mexico (Red Grouper Epinephelus morio, Gag Mycteroperca microlepis, Scamp Mycteroperca phenax, and Red Snapper Lutjanus campechanus). We used a paired design to measure the effects on isotopic values for carbon and nitrogen after storage using ice, 95% ethanol, and sodium chloride (table salt), against that in a liquid nitrogen control. Mean offsets for both δ (13)C and δ (15)N values from controls were lowest for samples preserved on ice, intermediate for those preserved with salt, and highest with ethanol. Within species, both salt and ethanol significantly enriched the δ (15)N values in nearly all comparisons. Ethanol also had strong effects on the δ (13)C values in all three groupers. Conversely, for samples preserved on ice, we did not detect a significant offset in either isotopic ratio for any of the focal species. Previous studies have addressed preservation-induced offsets in isotope values using a mass balance correction that accounts for changes in the isotope value to that in the C/N ratio. We tested the application of standard mass balance corrections for isotope values that were significantly affected by the preservation methods and found generally poor agreement between corrected and control values. The poor performance by the correction may have been due to preferential loss of lighter isotopes and corresponding low levels of mass loss with a substantial change in the isotope value of the sample. Regardless of mechanism, it was evident that accounting for offsets caused by different preservation methods was not possible using the standard correction. Caution is warranted when interpreting the results from specimens stored in either ethanol or salt, especially when using those from multiple preservation techniques. We suggest the use of ice as the preferred preservation technique for muscle tissue when conducting stable isotope analysis as it is widely available, inexpensive, easy to transport and did not impart a significant offset in measured isotopic values. Our results provide additional evidence that preservation effects on stable isotope analysis can be highly contextual, thus requiring their effects to be measured and understood for each species and isotopic ratio of interest before addressing research questions.

Stable isotopes in fish eye lenses as potential recorders of trophic and geographic history.

We evaluated eye lenses as potential recorders of stable isotope histories in fish because they consist of metabolically inert optical proteins that are deposited in successive, concentric circles (laminae) much like otolith circuli and tree rings. We conducted four different tests on lenses from red snapper, red grouper, gag, and white grunt. The first test was a low-resolution screening of multiple individuals (4-5 radial groups of laminae per lens, all species except white grunt). Along the radial axis, all individuals exhibited substantial isotopic variability. Red snapper individuals separated into two groups based on δ15N and gag separated into two groups based on δ13C. Two gag with the greatest variation were chosen for high-resolution temporal analysis using individual laminae from their second eye lenses. The first-order patterns from the high-resolution analysis generally mimicked patterns from the low-resolution screening of grouped laminae, yet the high-resolution plots revealed early-life details that were not apparent in the low-resolution screenings. For the third test, left- versus right-eye variation was compared using high-resolution methods. White grunt left- and right-eye radial isotopic patterns were almost identical for both δ13C and δ15N, suggesting the variations observed among individual fish were not artifacts. The final test evaluated intra-laminar variation; multiple samples were analyzed from different parts of the same lamina. Seven laminae from three individuals of two species were analyzed in this manner; variations among laminae were found to be much higher than variations within laminae. However, nominal intra-laminar variations were comparable to nominal differences between left and right lenses, suggesting intra-laminar variation established measurement precision. Eye lens isotopes appear to be useful for reconstructing the isotopic histories of individual fish; these histories can be compared with spatially-derived isoscapes to reconstruct individual histories for site fidelity, movement and trophic position.