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.     

Crude oil effects on microsomal mixed-function oxidase system components in the striped mullet (Mugil cephalus)

The effects of exposure to two types of crude oil on microsomal mixed-function oxidase system components in livers of juvenile striped mullet ($\text{Mugil cephalus}$) were investigated. Mullet were exposed for 4 days to emulsified Empire Mix or Saudi Arabian crude oils at an initial concentration of 75 ppm and an average of 1 ppm in the water column. Liver size was increased by about 50% following exposure to both oils. Since neither total hepatic protein nor microsomal protein increased as rapidly as did liver size, the concentrations of both were reduced following oil exposures. The proportion of microsomal protein to total hepatic protein or wet weight was not altered following crude oil exposure. Both cytochromes P-450 and $\text{b}$₅ were induced following oil treatment. NADPH-dependent enzymes assayed with cytochrome $\text{c}$ and dichlorophenolindophenol as substrates showed increases in activity after exposure to Empire Mix crude oil but only the latter enzyme activity was increased on a microsomal protein basis following Saudi Arabian crude oil treatment. Activities of NADH cytochrome $\text{c}$ and NADH cytochrome $\text{b}$₅ reductases appeared to vary with the protein level. However, since liver size was increased, oil-treated mullet had more of all parameters measured than did control mullet. Although the acute toxicity of Saudi Arabian crude oil to mullet is greater than that of Empire Mix crude oil, Empire Mix crude oil had greater inductive effects on microsomal oxidase components.     

原油和消油剂对鱼类毒性的研究进展

原油对鱼类的毒性主要来自其水溶性成分,可导致鱼卵死亡或发育畸形,并造成鱼类麻痹、发炎、粘膜受损和死亡。消油剂的使用在分散了油膜的同时也带来二次污染,其活性剂对鱼类产生新的毒性影响。消油剂将原油分散为乳化颗粒并能进入鱼体内,加大了原油的毒性并延长其作用时间。综述了原油和消油剂对鱼类毒性的研究进展,展望了鱼类在相关毒性试验领域的应用及发展方向。     

The effect of several crude oils and some petroleum distillation fractions on intestinal absorption in ducklings (Anas platyhynchos).

Ducklings given hypertonic saline drinking water show significant increases in the rates of Na+ and water transfer across the intestinal mucosa. These increased rates of transfer are maintained as long as the birds are fed dypertonic saline. Oral administration of a single small dose of crude oil had no effect on the basal rate of mucosal transfer in freshwater-maintained ducklings but the adaptive response of the mucosa is suppressed in birds given hypertonic saline. When crude oils from eight different geographical locations were tested, the degree of inhibition varied between them; the greatest and smallest degrees of inhibition being observed following administration of Kuwait and North Slope, Alaska, crude oils respectively. The effects of distallation fractions derived from two chemically different crude oils were also examined. The volume of each distallation fraction administered corresponded to its relative abundance in the crude oil from which it was derived. The inhibitory effect was not associated exclusively with the same distallation fractions from each oil. A highly naphthenic crude oil from the San Joaquin Valley, California, showed the greatest inhibitory activity in the least abundant (2%), low boiling point (smaller than 245 degrees C) fraction and the least inhibitory activity in the highest boiling point (greater than 482 degrees C) most abundant (47%) fraction. In contrast, a highly paraffinic crude oil from Paradox Basin, Utah, showed the greatest inhibitory effect with the highest boiling point fraction and a minimal effect with the lowest boiling point fraction; the relative abundances of these two fractions in the crude oil represented 27 and 28% respectively. Water-soluble extracts of both crude oils also had inhibitory effects on mucosal transfer rates and these roughly proportionate to the inhibitory potency of the low boiling point fraction of each oil. Weathered samples of San Joaquin Valley, California, and the Paradox Basin, Utah, oils showed greater effects than corresponding samples of unweathered oils even though most of the low molecular weight material from both oils was either evaporated or solubilized in the underlying water during the 36-h weathering period.     

Effect of Crude Oil and Chemical Additives on Metabolic Activity of Mixed Microbial Populations in Fresh Marsh Soils

Abstract Hydrocarbons increase abundance of hydrocarbon-degrading microorganisms, but also decrease microbial diversity. This could disrupt ecosystem dynamics by altering soil organic matter mineralization and resultant nutrient remineralization rates. Crude oil, which is known to contain toxins and reduce microbial diversity, was hypothesized to reduce gross metabolic activity of mixed microbial populations in wetland soils. Soil respiration and Eh were compared, for 6 months, among microcosms containing marsh soils that differed in soil organic matter (Panicum hemitomon Shult. or Sagittaria lancifolia L. dominated marshes), crude oil (Arabian crude, Louisiana crude, or no oil), and additives (a cleaner, a dispersant, fertilizer, or no additive). No treatment slowed activity; instead, Louisiana plus fertilizer and all Arabian treatments temporarily accelerated activity. Additional C respired from oiled microcosms exceeded C added as crude oil by 1.4 to 3.5 times. Thus, much additional C originated from soil organic matter rather than crude oil. Crude oils temporarily lowered soil Eh, which is consistent with accelerated metabolism and demand for electron acceptors. The lack of inhibition observed at the community level does not necessarily indicate an absence of toxicity. Instead, tolerant species with metabolic versatility probably maintained activity. Stimulation probably resulted from removal of micronutrient limitation, rather than removal of grazing pressure or macronutrient limitation. Regardless, accelerated soil organic matter mineralization surely accelerated nutrient remineralization. This might explain some reports of crude oil stimulating plant growth. These results are not inconsistent with theoretical and experimental conclusions regarding effects of biodiversity on ecosystem stability and productivity, nor are they inconsistent with conclusions that crude oils contain components that are toxic to microbes, vegetation, and fauna. However, these data do indicate that crude oils also contain components that temporarily stimulate metabolic activity of surviving microbes. Received: 27 April 1998; Accepted: 15 July 1998     

Effects of visible and UV light on the characteristics and properties of crude oil-in-water (O/W) emulsions

The effects of visible and UV light on the characteristics and properties of Prudhoe Bay (PB) and South Louisiana (SL) emulsions were investigated to better understand the role of sunlight on the fate of spilled crude oils that form emulsions with a dispersant in the aquatic environment. Before irradiation, crude oil emulsions showed the presence of dispersed crude oil micelles in a continuous water phase and crude oil components floating on the surface. The crude oil micelles decreased in size with irradiation, but emulsions retained their high degree of polydispersity. UV irradiation reduced the stability of emulsions more effectively than visible light. The reduction of micelles size caused the viscosity of emulsions to increase and melting point to decrease. Further, irradiation increased acid concentrations and induced ion formation which lowered the pH and increased the conductivity of emulsions, respectively. Ni and Fe in PB emulsions were extracted from crude oil with UV irradiation, which may provide an efficient process for metal removal. The emulsions were stable toward freeze/thaw cycles and their melting temperatures generally decreased with irradiation. Evidence of ˙OH production existed when emulsions were exposed to UV but not to visible light. The presence of H(2)O(2) enhanced the photodegradation of crude oil. Overall, the changes in emulsion properties were attributed to direct photodegradation and photooxidation of crude oil components.     

Physiological response of 10 phytoplankton species exposed to macondo oil and the dispersant,Corexit

Culture experiments were conducted on ten phytoplankton species to examine their biological and physiological responses during exposure to oil and a combination of oil and dispersant. The species tested included a range of taxa typically found in the Gulf of Mexico such as cyanobacteria, chlorophytes, and diatoms. Cultures were exposed to Macondo surrogate oil using the water accommodated fraction (WAF), and dispersed oil using a chemically enhanced WAF (CEWAF) and diluted CEWAF, to replicate conditions following the Deepwater Horizon spill in the Gulf of Mexico. A range of responses were observed, that could broadly class the algae as either “robust” or “sensitive” to oil and/or dispersant exposure. Robust algae were identified as Synechococcus elongatus, Dunaliella tertiolecta, two pennate diatoms Phaeodactylum tricornutum and Navicula sp., and Skeletonema grethae CCMP775, and were largely unaffected by any of the treatments (no changes to growth rate or time spent in lag phase relative to controls). The rest of the phytoplankton, all centric diatoms, exhibited at least some combination of reduced growth rates or increased lag time in response to oil and/or dispersant exposure. Photophysiology did not have a strong treatment effect, with significant inhibition of photosynthetic efficiency (F_v/F_m) only observed in the CEWAF, if at all. We found that the effects of oil and dispersants on phytoplankton physiology were species‐dependent, and not always detrimental. This has significant implications on how oil spills might impact phytoplankton community structure and bloom dynamics in the Gulf of Mexico, which in turn impacts higher trophic levels.     

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.     

Changes in mutagenicity during crude oil degradation by fungi

Two fungal strains, Cunninghamella elegans and Penicillium zonatum, that grow with crude oil as a sole carbon source were exposed to three crude oils that exhibit a range of mutagenic activity. At regular time intervals following fungal incubation with the various crude oils, extracts were tested for the presence of mutagenic activity using the spiral Salmonella assay. When the most mutagenic of the oils, Pennsylvania crude oil, was degraded by C. elegans or by P. zonatum, its mutagenicity was significantly reduced; corresponding uninoculated (weathered) controls of Pennsylvania crude remained mutagenic. West Texas Sour crude oil, a moderately mutagenic oil, exhibited little change in mutagenicity when incubated with either C. elegans or P. zonatum. Swanson River Field crude oil from Cook Inlet, Alaska is a slightly mutagenic oil that became more mutagenic when incubated with C. elegans; weathered controls of this oil showed little change in mutagenicity. Mycelial mat weights measured during growth on crude oils increased corresponding to the biodegradation of about 25% of the crude oil.     

Effect of water-soluble fraction of Cook Inlet crude oil on swimming performance and plasma cortisol in juvenile coho salmon (Oncorhynchus kisutch)

Swimming performance of juvenile coho salmon decreased and plasma cortisol increased, following 48-hr exposure to the water-soluble fraction (WSF) of Cook Inlet crude oil at 75% of the LC50. Exposure to 25 and 50% of the LC50 did not significantly reduce swimming performance. Plasma cortisol concentrations were highest in fish exposed to both the combined stress of WSF exposure and of forced swimming in a stamina tunnel.     

Application of biosurfactant produced from peanut oil cake by Lactobacillus delbrueckii in biodegradation of crude oil

Lactobacillus delbrueckii cultured with peanut oil cake as the carbon source yielded 5.35 mg ml(-1) of biosurfactant production. Five sets of microcosm biodegradation experiments were carried out with crude oil as follows: set 1 - bacterial cells+crude oil, set 2 - bacterial cells+crude oil+fertilizer, set 3 - bacterial cells+crude oil+biosurfactant, set 4 - bacterial cells+crude oil+biosurfactant+fertilizer, set 5 - with no bacterial cells, fertilizer and biosurfactant (control). Maximum degradation of crude oil was observed in set 4 (75%). Interestingly, when biosurfactant and bacterial cells were used (set 3), significant oil biodegradation activity occurred and the difference between this treatment and that in set 4 was 7% higher degradation level in microcosm experiments. It is evident from the results that biosurfactants alone is capable of promoting biodegradation to a large extent without added fertilizers.     

Effect of acute exposure to crude petroleum on some reproductive hormones in salmon and flounder

1. The concentration of total (free + conjugated) androgens in plasma of sexually mature male salmon and flounder was generally lower in oil-exposed fish. 2. Exposure to crude oil inhibited some testicular development of salmon during the final stages of maturation. 3. Oil exposure had no effect on levels of total plasmatic androgens or estradiol in male and female flounder during gonadal recrudescence. 4. Aryl hydrocarbon hydroxylase activity increased in liver and kidney of both species. 5. Sperm, collected from salmon exposed to oil, fertilized eggs from non-exposed females to produce normal alevins.     

Unexpected Interaction with Dispersed Crude Oil Droplets Drives Severe Toxicity in Atlantic Haddock Embryos

The toxicity resulting from exposure to oil droplets in marine fish embryos and larvae is still subject for debate. The most detailed studies have investigated the effects of water-dissolved components of crude oil in water accommodated fractions (WAFs) that lack bulk oil droplets. Although exposure to dissolved petroleum compounds alone is sufficient to cause the characteristic developmental toxicity of crude oil, few studies have addressed whether physical interaction with oil micro-droplets are a relevant exposure pathway for open water marine speices. Here we used controlled delivery of mechanically dispersed crude oil to expose pelagic embryos and larvae of a marine teleost, the Atlantic haddock (Melanogrammus aeglefinus). Haddock embryos were exposed continuously to two different concentrations of dispersed crude oil, high and low, or in pulses. By 24 hours of exposure, micro-droplets of oil were observed adhering and accumulating on the chorion, accompanied by highly elevated levels of cyp1a, a biomarker for exposure to aromatic hydrocarbons. Embryos from all treatment groups showed abnormalities representative of crude oil cardiotoxicity at hatch (5 days of exposure), such as pericardial and yolk sac edema. Compared to other species, the frequency and severity of toxic effects was higher than expected for the waterborne PAH concentrations (e.g., 100% of larvae had edema at the low treatment). These findings suggest an enhanced tissue uptake of PAHs and/or other petroleum compounds from attached oil droplets. These studies highlight a novel property of haddock embryos that leads to greater than expected impact from dispersed crude oil. Given the very limited number of marine species tested in similar exposures, the likelihood of other species with similar properties could be high. This unanticipated result therefore has implications for assessing the ecological impacts of oil spills and the use of methods for dispersing oil in the open sea.     

Dispersed oil disrupts microbial pathways in pelagic food webs

Most of the studies of microbial processes in response to the Deepwater Horizon oil spill focused on the deep water plume, and not on the surface communities. The effects of the crude oil and the application of dispersants on the coastal microbial food web in the northern Gulf of Mexico have not been well characterized even though these regions support much of the fisheries production in the Gulf. A mesocosm experiment was carried out to determine how the microbial community off the coast of Alabama may have responded to the influx of surface oil and dispersants. While the addition of glucose or oil alone resulted in an increase in the biomass of ciliates, suggesting transfer of carbon to higher trophic levels was likely; a different effect was seen in the presence of dispersant. The addition of dispersant or dispersed oil resulted in an increase in the biomass of heterotrophic prokaryotes, but a significant inhibition of ciliates, suggesting a reduction in grazing and decrease in transfer of carbon to higher trophic levels. Similar patterns were observed in two separate experiments with different starting nutrient regimes and microbial communities suggesting that the addition of dispersant and dispersed oil to the northern Gulf of Mexico waters in 2010 may have reduced the flow of carbon to higher trophic levels, leading to a decrease in the production of zooplankton and fish on the Alabama shelf.     

Aminotransferases in the bivalve mollusc Mytilus edulis L. and short term effects of crude oil in brackish water

Transaminases are among the crucial enzymes in amino acid metabolism, which in aquatic organisms is known to be affected by exposure to oil hydrocarbons. The transamination reactions in Mytilus edulis L. were studied to estimate their adequacy to indicate short term oil exposure in mussels. The transamination reactions were measured using paper chromatography and spectrophotometry. A high degree of transamination was observed between 2 oxoglutarate and alanine, aspartate and ornithine. A slight degree of transamination was shown with methionine, leucine, isoleucine, phenylalanine, serine, tryptophan, threonine, tyrosine and valine. No transamination was observed between 2 oxoglutarate and glycine, arginine, histidine, lysine, proline, citrulline and alanine. The effect of the water accommodated fraction WAF of crude oil on selected transaminase reactions was measured. The highest changes during the WAF exposure were mostly observed in the gills and mantle. Alanine aminotransferase EC 2.6.1.2 activity in the mantle was, at its highest, 55 over the control. Aspartate aminotransferase EC 2.6.1.1 activity increased in the gills by 52 . For ornithine transamination, in the gills the highest increase was by 75 and in the mantle by 50 . The metabolic pathways involved in the alterations of aminotransferase activities are discussed. It is concluded that ornithine transamination in gills is a potential indicator for short term crude oil exposure in Mytilus edulis. More studies are needed to evaluate the effects of other organic pollutants on ornithine transamination.     

Aminotransferases in the bivalve mollusc Mytilus edulis L. and short term effects of crude oil in brackish water

Transaminases are among the crucial enzymes in amino acid metabolism, which in aquatic organisms is known to be affected by exposure to oil hydrocarbons. The transamination reactions in Mytilus edulis L. were studied to estimate their adequacy to indicate short term oil exposure in mussels. The transamination reactions were measured using paper chromatography and spectrophotometry. A high degree of transamination was observed between 2 oxoglutarate and alanine, aspartate and ornithine. A slight degree of transamination was shown with methionine, leucine, isoleucine, phenylalanine, serine, tryptophan, threonine, tyrosine and valine. No transamination was observed between 2 oxoglutarate and glycine, arginine, histidine, lysine, proline, citrulline and alanine. The effect of the water accommodated fraction WAF of crude oil on selected transaminase reactions was measured. The highest changes during the WAF exposure were mostly observed in the gills and mantle. Alanine aminotransferase EC 2.6.1.2 activity in the mantle was, at its highest, 55 over the control. Aspartate aminotransferase EC 2.6.1.1 activity increased in the gills by 52 . For ornithine transamination, in the gills the highest increase was by 75 and in the mantle by 50 . The metabolic pathways involved in the alterations of aminotransferase activities are discussed. It is concluded that ornithine transamination in gills is a potential indicator for short term crude oil exposure in Mytilus edulis. More studies are needed to evaluate the effects of other organic pollutants on ornithine transamination.     

Hydrocarbon metabolism and cortisol balance in oil-exposed ringed seals, Phoca hispida

1. Ringed seals were exposed experimentally to oil contamination, by feeding of a [14C]naphthalene marked crude oil in fish for up to 4 days at a rate of 5 ml/day. 2. Mixed function oxygenase (MFO) activity, measured as aryl hydrocarbon hydroxylase in liver and kidney, was found to be induced, in particular in kidney tissue where the activity increased 3-fold. 3. MFO induction correlated with a high degree of conversion of crude oil hydrocarbons to water-soluble metabolites. Most of the radioactivity was found in the polar fraction of plasma and urine. 4. Plasma cortisol levels were somewhat elevated by captive holding, and increased markedly after oil-exposure. Cortisol half-life decreased after oil exposure from 1 3/4 to 1 hr.     

Corticosteroid and thyroid responses of larval and juvenile turbot exposed to the water-soluble fraction of crude oil

Turbot larvae (24–590 degree C days; 2–32 days post-hatch) and juveniles (1345 degree C days; 98 days post-hatch), were exposed for 6 h to 25, 33 and 50% water-soluble fraction (WSF) of crude oil in either static or flow-through test systems. Larvae showed generalized primary endocrine responses, identified by elevated whole body cortisol content from as early as 2 days post-hatch. In older larvae and juveniles, the response was related to the WSF concentration. This dose-response relationship was not apparent in younger and yolk-sac larvae. Whole body thyroxine content of turbot larvae exposed to the WSF of crude oil was increased, but triiodothyronine content remained stable. Aromatic hydrocarbon concentrations [benzene, toluene, ethylbenzene and xylene (BTEX) and naphthalenes] remained constant during flow-through tests, but 65% of the initial level of BTEX hydrocarbons and 40% of the naphthalenes were lost during static exposures. Larval mortalities increased with exposure to an increasing concentration of crude oil WSF. Larval activity was significantly reduced even at the lowest WSF concentration.     

The role of indigenous bacterial and fungal soil populations in the biodegradation of crude oil in a desert soil

The biodegradation capacity of indigenous microbial populations was examined in a desert soil contaminated with crude oil. To evaluate biodegradation, soil samples supplemented with 5, 10 or 20% (w/w) of crude oil were incubated for 90 days at 30 °C. The effect of augmentation of the soil with vermiculite (50% v/v) as a bulking agent providing increased surface/volume ratio and improved soil aeration was also tested. Maximal biodegradation (91%) was obtained in soil containing the highest concentration of crude oil (20%) and supplemented with vermiculite; only 74% of the oil was degraded in samples containing the same level of crude oil but lacking vermiculite. Gas chromatograms of distilled fractions of crude oil extracted from the soil before and after incubation demonstrated that most of the light and part of the intermediate weight fractions initially present in the oil extracts could not be detected after incubation. Monitoring of microbial population densities revealed an initial decline in bacterial viable counts after exposure to oil, presumably as a result of the crude oil’s toxicity. This decline was followed by a steep recovery in microbial population density, then by a moderate increase that persisted until the end of incubation. By contrast, the inhibitory effect of crude oil on the fungal population was minimal. Furthermore, the overall increased growth response of the fungal population, at all three levels of contamination, was about one order of magnitude higher than that of the bacterial population.