Advances in Chemical Analysis of Oil Spills Since the Deepwater Horizon Disaster.

Analytical techniques for chemical analysis of oil, oil photochemical and biological transformation products, and dispersants and their biodegradation products benefited significantly from research following the 2010 Deepwater Horizon (DWH) disaster. Crude oil and weathered-oil matrix reference materials were developed based on the Macondo well oil and characterized for polycyclic aromatic hydrocarbons, hopanes, and steranes for use to assure and improve the quality of analytical measurements in oil spill research. Advanced gas chromatography (GC) techniques such as comprehensive two-dimensional GC (GC × GC), pyrolysis GC with mass spectrometry (MS), and GC with tandem MS (GC-MS/MS) provide a greater understanding at the molecular level of composition and complexity of oil and weathering changes. The capabilities of high-resolution MS (HRMS) were utilized to extend the analytical characterization window beyond conventional GC-based methods to include polar and high molecular mass components (>400 Da) and to provide new opportunities for discovery, characterization, and investigation of photooxidation and biotransformation products. Novel separation approaches to reduce the complexity of the oil and weathered oil prior to high-resolution MS and advanced fluorescence spectrometry have increased the information available on spilled oil and transformation products. HRMS methods were developed to achieve the required precision and sensitivity for detection of dispersants and to provide molecular-level characterization of the complex surfactants. Overall, research funding following the DWH oil spill significantly advanced and expanded the use of analytical techniques for chemical analysis to support petroleum and dispersant characterization and investigations of fate and effects of not only the DWH oil spill but future spills.

Exposure methodologies for dissolved individual hydrocarbons, dissolved oil, water oil dispersions, water accommodated fraction and chemically enhanced water accommodated fraction of fresh and weathered oil.

Characterizing the nature and effects of oil released into the marine environment is very challenging. It is generally recognized that "environmentally relevant" conditions for exposure involve a range of temporal and spatial conditions, a range of exposure pathways (e.g., dissolved, emulsions, sorbed onto particulates matter), and a multitude of organisms, populations, and ecosystems. Various exposure methodologies have been used to study the effects of oil on aquatic organisms, and uniform protocols and exposure methods have been developed for the purposes of regulatory toxicological assessments. Ultimately, all exposure methods have drawbacks, it is impossible to totally mimic field conditions, and the choice of exposure methodology depends on the specific regulatory, toxicological, or other research questions to be addressed. The aim of this paper is to provide a concise review of the state of knowledge to identify gaps in that knowledge and summarize challenges for the future.

Occurrence, distribution, and associated pollutants of plastic pellets (nurdles) in coastal areas of South Texas.

Nurdles, also known as plastic resin pellets, are now a major source of plastic pollution on beaches globally, thus it is important to elucidate their weathering patterns and environmental fates as well as the associated pollutants. In this study we collected nurdles from 24 sites in the coastal bend region of south Texas, covering areas from the near shore railway stations to the adjacent bays and barrier islands. The morphologies of nurdles and associated pollutants including polycyclic aromatic hydrocarbons (PAHs), polychlorinated biphenyls (PCBs), and mercury, were investigated. The results showed that the nurdles varied greatly in color, shape, polymer composition, and oxidation degree. More than 80 % of the nurdles were made with polyethylene, and the rest with polypropylene, polyester, polystyrene, polyethylene-vinyl acetate, and polyvinyl chloride based on Fourier Transform Infrared Spectroscopy (FTIR) analysis. PCBs were not detected on nurdles. PAHs and mercury on nurdles were detected at 12 % and 20 % of the sampling sites. The total concentrations of detectable PAHs ranged from 92.59 to 1787.23 ng/g-nurdle, and the detectable mercury concentrations ranged from 1.23 to 22.25 ng/g-nurdle. Although the concentrations of these pollutants were not at the acute toxic effect level, the presence of PAHs and mercury suggested the potential risk of pollutant exposure to marine organisms in ecosystems, given the fact that nurdles are persistent in the environment.

The impacts of weathering on concentration and bioaccessibility of organic pollutants associated with plastic pellets (nurdles) in coastal environments.

Nurdles, the pre-production plastic pellets, are a major source of plastic pollution in marine environments due to unregulated spills during production and transportation. We analyzed the types of plastics and associated organic pollutants on nurdles collected along the shoreline of Gulf of Mexico in Texas. Our results showed that the nurdles were made from polyethylene (81.9%) and polypropylene (18.1%). Polycyclic aromatic hydrocarbons (PAHs, 16 US EPA priority) and polychlorinated biphenyls (PCBs, 7 commercial congeners) sorbed to the nurdles were in concentration ranges of 1.6-14,700 ng/ g and 0-642 ng/ g, respectively. Heavily weathered nurdles tended to have higher concentrations of PAHs and PCBs than lightly weathered ones. The bioaccessibility of sorbed contaminants was evaluated using a simulated intestinal fluid. The results showed that the associated PAHs were more bioaccessible in lightly weathered nurdles (13.1 ± 2.3%) than heavily weathered one (5.3 ± 0.1%), and that no PCBs were bioaccessible. These findings are informative for toxicity evaluation and resource management of plastic debris in coastal environments.

Initial oil concentration affects hydrocarbon biodegradation rates and bacterial community composition in seawater.

During oil spills in the field or for laboratory incubation studies, different oil concentrations are often encountered or applied, yet how initial oil concentration affects biodegradation rates of hydrocarbons and the development of oil degraders remains unclear. We incubated seawater for 50 d with different oil concentrations (0, 50, 100, 200, 400 and 800 ppm). n-Alkanes and polycyclic aromatic hydrocarbons (PAHs), and the bacterial community were analyzed periodically. Results show that the biodegradation rates of alkanes, derived from first order kinetics, decreased with increasing oil concentration, but percent residual was ~50% regardless of the initial concentration. In contrast, the biodegradation rates of PAHs increased with concentration, and the percent residual increased with oil concentration. Increasing oil concentration resulted in increased abundances of Rhodobacterales, Altererythrobacter, and Neptuniibacter. However, Alcanivorax abundance was barely detected in 400 and 800 ppm. Overall, oil concentration critically affected the degradation of hydrocarbons and the bacterial community.

Potential effects of bacterial communities on the formation of blooms of the harmful dinoflagellate Prorocentrum after the 2014 Texas City "Y" oil spill (USA).

The association between phytoplankton blooms and oil spills is still controversial despite numerous studies. Surprisingly, to date, there have been no studies on the effect of bacterial communities (BCs) exposed to crude oil on phytoplankton growth, even though crude oil changes BCs, which can then affect phytoplankton growth and species composition. Co-culture with crude oil-exposed BCs significantly stimulated the growth of Prorocentrum texanum in the laboratory. To gain more direct evidence, oil-degrading bacteria from oil-contaminated sediment collected after the Texas City "Y" oil spill were isolated, and changes in dinoflagellate growth when co-cultured with single bacterial isolates was investigated. The oil-degrading bacterial isolates significantly stimulated the growth of dinoflagellates (axenic and xenic cultures) through releasing growth-promoting substances. This study provides new evidence for the potential role of oil-degrading bacteria in the formation of phytoplankton blooms after an oil spill.

Hydrocarbon degradation and response of seafloor sediment bacterial community in the northern Gulf of Mexico to light Louisiana sweet crude oil.

The Deepwater Horizon (DWH) blowout resulted in the deposition to the seafloor of up to 4.9% of 200 million gallons of oil released into the Gulf of Mexico. The petroleum hydrocarbon concentrations near the wellhead were high immediately after the spill, but returned to background levels a few years after the spill. Microbial communities in the seafloor are thought to be responsible for the degradation of hydrocarbons, however, our knowledge is primarily based upon gene diversity surveys and hydrocarbon concentration in field sediment samples. Here, we investigated the oil degradation potential and changes in bacterial community by amending seafloor sediment collected near the DWH site with crude oil and both oil and Corexit dispersant. Polycyclic aromatic hydrocarbons were rapidly degraded during the first 30 days of incubation, while alkanes were degraded more slowly. With the degradation of hydrocarbons, the relative abundances of Colwelliaceae, Alteromonadaceae, Methylococales, Alcanivorax, Bacteriovorax, and Phaeobacter increased remarkably. However, the abundances of oil-degrading bacteria changed with oil chemistry. Colwelliaceae decreased with increasing oil degradation, whereas Alcanivorax and Methylococcales increased considerably. We assembled seven genomes from the metagenome, including ones belonging to Colwellia, Alteromonadaceae, Rhodobacteraceae, the newly reported genus Woeseia, and candidate phylum NC10, all of which possess a repertoire of genes for hydrocarbon degradation. Moreover, genes related to hydrocarbon degradation were highly enriched in the oiled treatment, suggesting that the hydrocarbons were biodegraded, and that the indigenous microflora have a remarkable potential for the natural attenuation of spilled oil in the deep-sea surface sediment.

Petroleum hydrocarbon persistence following the Deepwater Horizon oil spill as a function of shoreline energy.

An important aspect of oil spill science is understanding how the compounds within spilled oil, especially toxic components, change with weathering. In this study we follow the evolution of petroleum hydrocarbons, including n-alkanes, polycyclic aromatic hydrocarbons (PAHs) and alkylated PAHs, on a Louisiana beach and salt marsh for three years following the Deepwater Horizon spill. Relative to source oil, we report overall depletion of low molecular weight n-alkanes and PAHs in all locations with time. The magnitude of depletion, however, depends on the sampling location, whereby sites with highest wave energy have highest compound depletion. Oiled sediment from an enclosed bay shows high enrichment of high molecular weight PAHs relative to 17α(H),21ß(H)-hopane, suggesting the contribution from sources other than the Deepwater Horizon spill, such as fossil fuel burning. This insight into hydrocarbon persistence as a function of hydrography and hydrocarbon source can inform policy and response for future spills.

Can gelatinous zooplankton influence the fate of crude oil in marine environments?

Gelatinous zooplankton are known for their capacity to excrete copious amounts of mucus that can be utilized by other organisms. The release of mucus is exacerbated by stressful conditions. Despite the recognized importance of cnidarian mucus to production and material flux in marine ecosystems, the role of gelatinous zooplankton in influencing the fate of oil spills is unknown. In this study we used laboratory experiments to observe the influence of mucus from the moon jellyfish (Aurelia aurita) on the aggregation and degradation of crude oil. The results show that jellyfish swimming in a dispersed solution of oil droplets produced copious amounts of mucus and the mucus aggregates that were shed by the animals contained 26 times more oil than the surrounding water. Incubation experiments showed that hydrocarbon degrading bacteria cell densities more than doubled in the presence of mucus and after 14days, resulted in a significant increase in oil degradation. These results suggest that jellyfish can aggregate dispersed oil droplets and embed them within a matrix that favors hydrocarbon degrading bacteria. While this study lends support to the hypothesis that the presence of gelatinous zooplankton can impact oil spills large scale mesocosm studies will be needed to fully quantify the influence on a natural system.

Potential Environmental Factors Affecting Oil-Degrading Bacterial Populations in Deep and Surface Waters of the Northern Gulf of Mexico.

Understanding bacterial community dynamics as a result of an oil spill is important for predicting the fate of oil released to the environment and developing bioremediation strategies in the Gulf of Mexico. In this study, we aimed to elucidate the roles of temperature, water chemistry (nutrients), and initial bacterial community in selecting oil degraders through a series of incubation experiments. Surface (2 m) and bottom (1537 m) waters, collected near the Deepwater Horizon site, were amended with 200 ppm light Louisiana sweet crude oil and bacterial inoculums from surface or bottom water, and incubated at 4 or 24°C for 50 days. Bacterial community and residual oil were analyzed by pyrosequencing and gas chromatography-mass spectrometry (GC-MS), respectively. The results showed that temperature played a key role in selecting oil-degrading bacteria. Incubation at 4°C favored the development of Cycloclasticus, Pseudoalteromonas, Sulfitobacter, and Reinekea, while 24°C incubations enhanced Oleibacter, Thalassobius, Phaeobacter, and Roseobacter. Water chemistry and the initial community also had potential roles in the development of hydrocarbon-degrading bacterial communities. Pseudoalteromonas, Oleibacter, and Winogradskyella developed well in the nutrient-enriched bottom water, while Reinekea and Thalassobius were favored by low-nutrient surface water. We revealed that the combination of 4°C, crude oil and bottom inoculum was a key factor for the growth of Cycloclasticus, while the combination of surface inoculum and bottom water chemistry was important for the growth of Pseudoalteromonas. Moreover, regardless of the source of inoculum, bottom water at 24°C was a favorable condition for Oleibacter. Redundancy analysis further showed that temperature and initial community explained 57 and 19% of the variation observed, while oil and water chemistry contributed 14 and 10%, respectively. Overall, this study revealed the relative roles of temperature, water chemistry, and initial bacterial community in selecting oil degraders and regulating their evolution in the northern Gulf of Mexico.

High Phosphate Concentrations Accelerate Bacterial Peptide Decomposition in Hypoxic Bottom Waters of the Northern Gulf of Mexico.

Despite extensive studies of the development and dynamics of hypoxia in coastal oceans, factors controlling the decomposition rates and pathways of labile organic matter (OM) in hypoxic waters are not well understood. Here we investigate peptide decomposition in a stratified water column in the hypoxic region of the northern Gulf of Mexico by conducting on-deck incubation experiments amended with tetrapeptide ala-val-phe-ala (AVFA), a fragment of RuBisCO. Our results show that decomposition efficiency of AVFA was limited by the availability of soluble reactive phosphorus (Pi) in the surface water (<0.3 µM), as it was greatly enhanced after Pi addition to the incubation water. In contrast, peptide decomposition rate in the subsurface water, enriched with Pi (0.4-1.2 µM), was twice as high as that in the surface water, concomitant with the development of fast-growing bacteria during the incubation. Consistent with the Growth Rate Hypothesis, these results indicate that a high level of Pi is crucial in stimulating the growth of bacterial strains with high RNA contents and thus faster OM decomposition in marine environments. This high decomposition potential of OM in subsurface hypoxic waters presents a positive feedback on hypoxia formation in Pi-enriched coastal subsurface waters, as a higher OM decomposition rate leads to rapid consumption of dissolved oxygen (DO).

Natural Sunlight Shapes Crude Oil-Degrading Bacterial Communities in Northern Gulf of Mexico Surface Waters.

Following the Deepwater Horizon (DWH) spill in 2010, an enormous amount of oil was observed in the deep and surface waters of the northern Gulf of Mexico. Surface waters are characterized by intense sunlight and high temperature during summer. While the oil-degrading bacterial communities in the deep-sea plume have been widely investigated, the effect of natural sunlight on those in oil polluted surface waters remains unexplored to date. In this study, we incubated surface water from the DWH site with amendments of crude oil, Corexit dispersant, or both for 36 days under natural sunlight in the northern Gulf of Mexico. The bacterial community was analyzed over time for total abundance, density of alkane and polycyclic aromatic hydrocarbon degraders, and community composition via pyrosequencing. Our results showed that, for treatments with oil and/or Corexit, sunlight significantly reduced bacterial diversity and evenness and was a key driver of shifts in bacterial community structure. In samples containing oil or dispersant, sunlight greatly reduced abundance of the Cyanobacterium Synechococcus but increased the relative abundances of Alteromonas, Marinobacter, Labrenzia, Sandarakinotalea, Bartonella, and Halomonas. Dark samples with oil were represented by members of Thalassobius, Winogradskyella, Alcanivorax, Formosa, Pseudomonas, Eubacterium, Erythrobacter, Natronocella, and Coxiella. Both oil and Corexit inhibited the Candidatus Pelagibacter with or without sunlight exposure. For the first time, we demonstrated the effects of light in structuring microbial communities in water with oil and/or Corexit. Overall, our findings improve understanding of oil pollution in surface water, and provide unequivocal evidence that sunlight is a key factor in determining bacterial community composition and dynamics in oil polluted marine waters.

Differentiating the roles of photooxidation and biodegradation in the weathering of Light Louisiana Sweet crude oil in surface water from the Deepwater Horizon site.

We determined the contributions of photooxidation and biodegradation to the weathering of Light Louisiana Sweet crude oil by incubating surface water from the Deepwater Horizon site under natural sunlight and temperature conditions. N-alkane biodegradation rate constants were ca. ten-fold higher than the photooxidation rate constants. For the 2-3 ring and 4-5 ring polycyclic aromatic hydrocarbons (PAHs), photooxidation rate constants were 0.08-0.98day(-1) and 0.01-0.07day(-1), respectively. The dispersant Corexit enhanced degradation of n-alkanes but not of PAHs. Compared to biodegradation, photooxidation increased transformation of 4-5 ring PAHs by 70% and 3-4 ring alkylated PAHs by 36%. For the first time we observed that sunlight inhibited biodegradation of pristane and phytane, possibly due to inhibition of the bacteria that can degrade branched-alkanes. This study provides quantitative measures of oil degradation under relevant field conditions crucial for understanding and modeling the fate of spilled oil in the northern Gulf of Mexico.

Effects of crude oil exposure on bioaccumulation of polycyclic aromatic hydrocarbons and survival of adult and larval stages of gelatinous zooplankton.

Gelatinous zooplankton play an important role in marine food webs both as major consumers of metazooplankton and as prey of apex predators (e.g., tuna, sunfish, sea turtles). However, little is known about the effects of crude oil spills on these important components of planktonic communities. We determined the effects of Louisiana light sweet crude oil exposure on survival and bioaccumulation of polycyclic aromatic hydrocarbons (PAHs) in adult stages of the scyphozoans Pelagia noctiluca and Aurelia aurita and the ctenophore Mnemiopsis leidyi, and on survival of ephyra larvae of A. aurita and cydippid larvae of M. leidyi, in the laboratory. Adult P. noctiluca showed 100% mortality at oil concentration ≥20 µL L(-1) after 16 h. In contrast, low or non-lethal effects were observed on adult stages of A. aurita and M. leidyi exposed at oil concentration ≤25 µL L(-1) after 6 days. Survival of ephyra and cydippid larva decreased with increasing crude oil concentration and exposition time. The median lethal concentration (LC50) for ephyra larvae ranged from 14.41 to 0.15 µL L(-1) after 1 and 3 days, respectively. LC50 for cydippid larvae ranged from 14.52 to 8.94 µL L(-1) after 3 and 6 days, respectively. We observed selective bioaccumulation of chrysene, phenanthrene and pyrene in A. aurita and chrysene, pyrene, benzo[a]pyrene, benzo[b]fluoranthene, benzo[k]fluoranthene, and benzo[a]anthracene in M. leidyi. Overall, our results indicate that (1) A. aurita and M. leidyi adults had a high tolerance to crude oil exposure compared to other zooplankton, whereas P. noctiluca was highly sensitive to crude oil, (2) larval stages of gelatinous zooplankton were more sensitive to crude oil than adult stages, and (3) some of the most toxic PAHs of crude oil can be bioaccumulated in gelatinous zooplankton and potentially be transferred up the food web and contaminate apex predators.

Interactions between zooplankton and crude oil: toxic effects and bioaccumulation of polycyclic aromatic hydrocarbons.

We conducted ship-, shore- and laboratory-based crude oil exposure experiments to investigate (1) the effects of crude oil (Louisiana light sweet oil) on survival and bioaccumulation of polycyclic aromatic hydrocarbons (PAHs) in mesozooplankton communities, (2) the lethal effects of dispersant (Corexit 9500A) and dispersant-treated oil on mesozooplankton, (3) the influence of UVB radiation/sunlight exposure on the toxicity of dispersed crude oil to mesozooplankton, and (4) the role of marine protozoans on the sublethal effects of crude oil and in the bioaccumulation of PAHs in the copepod Acartia tonsa. Mortality of mesozooplankton increased with increasing oil concentration following a sigmoid model with a median lethal concentration of 32.4 µl L(-1) in 16 h. At the ratio of dispersant to oil commonly used in the treatment of oil spills (i.e. 1∶20), dispersant (0.25 µl L(-1)) and dispersant-treated oil were 2.3 and 3.4 times more toxic, respectively, than crude oil alone (5 µl L(-1)) to mesozooplankton. UVB radiation increased the lethal effects of dispersed crude oil in mesozooplankton communities by 35%. We observed selective bioaccumulation of five PAHs, fluoranthene, phenanthrene, pyrene, chrysene and benzo[b]fluoranthene in both mesozooplankton communities and in the copepod A. tonsa. The presence of the protozoan Oxyrrhis marina reduced sublethal effects of oil on A. tonsa and was related to lower accumulations of PAHs in tissues and fecal pellets, suggesting that protozoa may be important in mitigating the harmful effects of crude oil exposure in copepods and the transfer of PAHs to higher trophic levels. Overall, our results indicate that the negative impact of oil spills on mesozooplankton may be increased by the use of chemical dispersant and UV radiation, but attenuated by crude oil-microbial food webs interactions, and that both mesozooplankton and protozoans may play an important role in fate of PAHs in marine environments.

Evaluating bacterial community structures in oil collected from the sea surface and sediment in the northern Gulf of Mexico after the Deepwater Horizon oil spill.

Bacterial community structures were evaluated in oil samples using culture-independent pyrosequencing, including oil mousses collected on sea surface and salt marshes during the Deepwater Horizon oil spill, and oil deposited in sediments adjacent to the wellhead 1 year after the spill. Phylogenetic analysis suggested that Erythrobacter, Rhodovulum, Stappia, and Thalassospira of Alphaproteobacteria were the prevailing groups in the oil mousses, which may relate to high temperatures and strong irradiance in surface Gulf waters. In the mousse collected from the leaves of Spartina alterniflora, Vibrio of Gammaproteobacteria represented 57% of the total operational taxonomic units, suggesting that this indigenous genus is particularly responsive to the oil contamination in salt marshes. The bacterial communities in oil-contaminated sediments were highly diversified. The relatively high abundance of the Methylococcus, Methylobacter, Actinobacteria, Firmicutes, and Chlorofexi bacteria resembles those found in certain cold-seep sediments with gas hydrates. Bacterial communities in the overlying water of the oil-contaminated sediment were dominated by Ralstonia of Betaproteobacteria, which can degrade small aromatics, and Saccharophagus degradans of Gammaproteobacteria, a cellulose degrader, suggesting that overlying water was affected by the oil-contaminated sediments, possibly due to the dissolution of small aromatics and biosurfactants produced during biodegradation. Overall, these results provided key information needed to evaluate oil degradation in the region and develop future bioremediation strategies.

Chemical evolution of Macondo crude oil during laboratory degradation as characterized by fluorescence EEMs and hydrocarbon composition.

The fluorescence EEM technique, PARAFAC modeling, and hydrocarbon composition were used to characterize oil components and to examine the chemical evolution and degradation pathways of Macondo crude oil under controlled laboratory conditions. Three major fluorescent oil components were identified, with Ex/Em maxima at 226/328, 262/315, and 244/366 nm, respectively. An average degradation half-life of ∼20 d was determined for the oil components based on fluorescence EEM and hydrocarbon composition measurements, showing a dynamic chemical evolution and transformation of the oil during degradation. Dispersants appeared to change the chemical characteristics of oil, to shift the fluorescence EEM spectra, and to enhance the degradation of low-molecular-weight hydrocarbons. Photochemical degradation played a dominant role in the transformation of oil components, likely an effective degradation pathway of oil in the water column. Results from laboratory experiments should facilitate the interpretation of field-data and provide insights for understanding the fate and transport of oil components in the Gulf of Mexico.