An assessment of the genotoxic impact of the Sea Empress oil spill by the measurement of DNA adduct levels in selected invertebrate and vertebrate species

The grounding of the Sea Empress oil tanker resulted in the release of 72,000 tonnes of crude oil into Milford Haven, Wales, UK. Our initial studies indicated that this contamination resulted in elevated levels of DNA adducts in one of the area's native marine species Lipophrys pholis [B.P. Lyons, J.S. Harvey, J.M. Parry, An initial assessment of the genotoxic impact of the Sea Empress oil spill by the measurement of DNA adduct levels in the intertidal teleost Lipophrys pholis, Mutat. Res. 390 (1997) 263-268]. These original studies were extended and the genotoxic impact of the oil contamination was investigated in the invertebrates Halichondria panicea and Mytilus edulis, along with the vertebrate fish species L. pholis, Pleuronectes platessa and Limanda limanda. DNA adduct levels were assessed in these species over a period of 2-17 months after the incident. The studies indicate differences in the impact of acute oil contamination upon vertebrate and invertebrate species. The oil contamination did not induce any detectable elevations in adduct levels in the invertebrate species H. panicea and M. edulis. In contrast, the oil contamination did appear to induce adducts in the vertebrate teleost species L. pholis, P. platessa and Lim. limanda. Despite some difficulties in sampling, the data obtained 12-17 months after the spill suggested that the affected species recovered from the oil contamination. While the studies indicate that the genetic impact of the oil contamination was less severe than might have been expected, it remains possible that the DNA adducts detected in the teleosts could lead to genetic changes in these species in the future.     

Population trends and reproductive success of the European shag Phalacrocorax aristotelis on the Iberian Peninsula following the Prestige oil spill

In 2003, immediately following the Prestige oil spill in Galicia, Spain, we studied the population trends and reproductive performance of European shags (Phalacrocorax aristotelis) at oiled and unoiled colonies. This bird is an important member of the nearshore marine community, breeding in the area affected by the Prestige oil spill. The European shag feeds around the breeding colonies throughout the year, making it a useful indicator of environmental change. Before the oil spill, population trends were similar between oiled and unoiled colonies. Nevertheless, colonies located within the path of the oil suffered greater declines (ca. 10%) compared with pre-spill trends and with population trends at unoiled colonies. In 2003, the breeding success was 50% lower in oiled colonies compared with unoiled colonies. The data available from pre-spill years suggest that the annual reproductive success did not differ among colonies before the impact. European shags breeding at colonies affected by oil showed a negative initial impact from the Prestige oil spill. The reduction in reproductive success at oiled colonies may be due to sub-lethal effects of oil exposure or low food availability after the oil spill.     

DNA damage in eelpout (Zoarces viviparus) from Göteborg harbour

The relationship between DNA damage and the exposure of marine organisms to environmental contaminants was examined in the G?teborg harbour area. This research is part of a wider ecotoxicological study planned to evaluate the biological impact of chemical contamination in the River G?ta estuary, following a bunker oil (10-100 tonnes) spill occurred in June 2003. Here we present data on the DNA strand breaks derived using the comet assay and the presence of apoptotic cells using the diffusion assay in nucleated erythrocytes of the eelpout (Zoarces viviparus) from the study area and at a clean reference site. Polycyclic aromatic hydrocarbon metabolites were also analyzed in the bile of exposed fish. The results showed a high level of damaged DNA, paralleled by a peak in bile PAH metabolites, in fish from the most impacted site, 3 weeks after the oil spill. A significant recovery was observed in specimens from the spill site, 5 months later, but not in fish caught in the middle part of G?teborg harbour, which is chronically subjected to heavy chemical pollution. The levels of apoptic cells did not show any marked variations, but a significant recovery was observed in fish from the oil impacted site 5 months after the spill.     

Hemocyte parameters of the Pacific oyster Crassostrea gigas a year after the Hebei Spirit oil spill off the west coast of Korea

In marine bivalves, hemocytes support various physiological functions, including immune defense, nutrient transport, shell repair, and homeostatic maintenance. Although the effects of marine contaminants on the immunological functions of bivalves have been extensively investigated, the impacts of oil spills are not well understood. Therefore, we investigated hemocyte parameters in the Pacific oyster Crassostrea gigas 13 months after the Hebei Spirit oil spill (December 2007) off the west coast of Korea. The parameters studied included hemocyte concentration and mortality, relative proportion of hemocyte populations, and immunological functions such as phagocytosis and oxidative activity using flow cytometry. These immune-related parameters in oysters damaged by the oil spill were also compared to control oysters that were collected from an area unaffected by the spill. The flow cytometry study indicated that granulocyte population, phagocytic capacity, and reactive oxygen species production in oysters exposed to crude oil 13 months prior were depressed compared to the unexposed control oysters. Our data suggest that immunocompetence in oysters affected by the oil spill had not fully recovered 1 year after the accident, although more detailed studies on the physiology and disease resistance should be performed.     

Home sweet home: evidence for nest‐fidelity in the rocky intertidal fish,the shanny Lipophrys pholis

The ability to be faithful to a particular area or site was analysed in the shanny Lipophrys pholis. Using passive integrated transponders, adults from a population of L. pholis at Cabo Raso, Portugal, were followed over a period of 3 years. The findings showed that site fidelity is a consistent behaviour during the breeding season with specific breeding males being found only in particular sectors within the area, and in specific nests throughout the years. The fact that, in general, L. pholis individuals were absent from the study area during the non‐breeding season and breeding males were recorded returning to the same nests and sectors for consecutive breeding seasons suggests that they have developed excellent homing abilities. Translocation data corroborate this idea showing that breeding males successfully returned to their nests after a displacement of >100 m. Altogether, these findings highlight the relevance of life‐history traits (e.g. nesting) in the conditioning of site fidelity and homing for this species of rocky intertidal fish, and more importantly, provide evidence for the need of a well‐developed navigational system.     

Ten Years after the Prestige Oil Spill: Seabird Trophic Ecology as Indicator of Long-Term Effects on the Coastal Marine Ecosystem

Major oil spills can have long-term impacts since oil pollution does not only result in acute mortality of marine organisms, but also affects productivity levels, predator-prey dynamics, and damages habitats that support marine communities. However, despite the conservation implications of oil accidents, the monitoring and assessment of its lasting impacts still remains a difficult and daunting task. Here, we used European shags to evaluate the overall, lasting effects of the Prestige oil spill (2002) on the affected marine ecosystem. Using δ ¹⁵N and Hg analysis, we trace temporal changes in feeding ecology potentially related to alterations of the food web due to the spill. Using climatic and oceanic data, we also investigate the influence of North Atlantic Oscillation (NAO) index, the sea surface temperature (SST) and the chlorophyll a (Chl a) on the observed changes. Analysis of δ ¹⁵N and Hg concentrations revealed that after the Prestige oil spill, shag chicks abruptly switched their trophic level from a diet based on a high percentage of demersal-benthic fish to a higher proportion of pelagic/semi-pelagic species. There was no evidence that Chl a, SST and NAO reflected any particular changes or severity in environmental conditions for any year or season that may explain the sudden change observed in trophic level. Thus, this study highlighted an impact on the marine food web for at least three years. Our results provide the best evidence to date of the long-term consequences of the Prestige oil spill. They also show how, regardless of wider oceanographic variability, lasting impacts on predator-prey dynamics can be assessed using biochemical markers. This is particularly useful if larger scale and longer term monitoring of all trophic levels is unfeasible due to limited funding or high ecosystem complexity.     

Long-term reproductive impairment in a seabird after the Prestige oil spill

Large oil spills are dramatic perturbations on marine ecosystems, and seabirds are one of the worst affected organisms in such events. It has been argued that oil spills may have important long-term consequences on marine organisms, but supporting evidence remains scarce. The European shag (Phalacrocorax aristotelis) was strongly impacted at population level by the Prestige oil spill, the biggest spillage in the eastern North Atlantic. In this paper, we report on the long-term consequences on reproduction of this coastal seabird, using temporal and spatial replicated data (before–after–control–impact design). Our study revealed long-term reproductive impairment during at least the first 10 years since the Prestige oil spill. Annual reproductive success did not differ before the impact, but after the impact it was reduced by 45% in oiled colonies compared with unoiled ones. This is a rare documentation of long-term effects after a major oil spill, highlighting the need for long-term monitoring in order to assess the real impact of this type of disturbance on marine organisms.     

Demographic trends of Brown Pelicans in Louisiana before and after the Deepwater Horizon oil spill

Marine oil spills may have extensive and deleterious effects on coastal waterbirds, but pre‐spill data sets are often not available for making comparisons of demographics to the period following a spill. The 2010 Deepwater Horizon oil spill allowed us to compare Brown Pelican (Pelecanus occidentalis) demographics during pre‐ and post‐spill years. We banded 1114 pelicans on Louisiana barrier islands from 2007 to 2009, tracked their distribution via band re‐sighting surveys from 2008 to 2011, and conducted age‐structure surveys. Across Louisiana coastal islands in 2011, we detected 7% of pelicans that had been oiled during the 2010 spill and released following rehabilitation. Similarly, 6% of pelicans (not oiled) banded at the same release site in 2007 were observed across coastal islands 1 yr after banding. We observed variation in proportions of pelicans that were 1, 2, and 3 or more years old among years (2008–2011) and across islands, but little variation could readily be assigned to spill‐related mortality. These Brown Pelican demographic trends one year following the Deepwater Horizon oil spill are contrary to other assessments of the impacts of oil contamination on marine birds. However, additional research is required to evaluate potential long‐term population trends.     

Further Studies on Haemogregarina bigemina Laveran & Mesnil,the Marine Fish Blennius pholis L., and the Isopod Gnathia maxillaris Montagu1

Haemogregarina bigemina is redescribed from the blood of the marine fish Blennius pholis, and stages presumed to be those of the haemogregarine are recorded from the hematophagous praniza larva of the isopod Gnathia maxillaris. At College Rocks, Aberystwyth, Wales, the main study area, a high incidence of infection occurred in B. pholis. No exoerythrocytic stages were observed in these fish, nor was sexual dimorphism of the gametocyte evident. As in an earlier study, ecological evidence favored transmission by G. maxillaris rather than by leeches. Gametocytes, syzygy, oocysts, sporoblasts, and sporozoites were identified in the anterior hindgut of the isopod. The stages observed in G. maxillaris are compared with those of other haemogregarines described from the digestive tract of leeches. Mention is made of an intraleucocytic haemogregarine of another fish, Crenilabrus melops.     

Oiling rates and condition indices of shorebirds on the northern Gulf of Mexico following the Deepwater Horizon oil spill

The coastline of the Gulf of Mexico in the United States is an important wintering and stopover region for migratory shorebirds. The Deepwater Horizon oil spill (April–August 2010) impacted more than 1700 km of this coastline and could potentially affect shorebirds through long‐term exposure to toxins, degraded habitats, and altered food chains. We investigated the exposure to Deepwater Horizon oil of seven species of shorebirds that winter or stopover along the northern Gulf of Mexico. From October 2010 to May 2012, we captured and banded 691 shorebirds at six sites that experienced varying levels of oil contamination. Of birds sampled, 22 were lightly oiled, with species that forage on the coast having higher rates of oiling than those that forage in more estuarine habitats. Although only 8.6% of birds captured from October 2010 to May 2011 and 0.6% of the birds captured from August 2011 to June 2012 showed signs of oiling, an unknown, but potentially larger, number of shorebirds were likely exposed to indirect effects of the spill, such as decreased foraging time due to oiling of sites or disturbance from cleanup activities. Fuel stores and fattening rates of Dunlins (Calidris alpina) during spring migration, as measured using plasma metabolites, were not influenced by site oiling level. However, the level of disturbance at study sites was a significant predictor of both fuel stores and glycerol levels, suggesting that Dunlins stopping over during spring migration may have had difficulty reaching necessary fuel stores in spring 2011 due to disturbance from cleanup activity on oiled beaches. These effects from disturbance were only observed at sites with high cleanup activity, suggesting that the impact of oil‐spill cleanup on shorebirds may be minimized by limiting cleanup activities to specific areas and times of day.     

Risk posed by the invasive defoliator Uraba lugens to New Zealand native flora

• 1 The risk posed to New Zealand native flora by the recently‐established pest of Australian origin Uraba lugens Walker (Lepidoptera: Nolidae) (gum leaf skeletonizer) was assessed. Weed biological control host range testing methods were applied to identify those New Zealand plant species potentially at risk. Native plants tested were primarily in the Myrtaceae, the family that contains all the Australian hosts of U. lugens.
• 2 Experimental methods included no‐choice larval feeding assays and field cage and laboratory oviposition trials. Difficulty in ascertaining reliable oviposition preference data from cage oviposition trials limited the confidence with which the invader's field host range could be predicted.
• 3 Field surveys of plants attacked by U. lugens in the infested area supported the initial predictions but only some of the at‐risk native Myrtaceae were present in the area.
• 4 The risk to native New Zealand plants is presented in terms of two mechanisms: development of self‐sustaining populations and temporary spill‐over of solitary larvae. Development of self‐sustaining populations of the pest within native forests is deemed to be highly improbable.
• 5 Temporary spill‐over impacts are most likely in urban areas within mixed species plantings or boundaries between native/exotic forests and coinciding with the mobile larval life stages. Spill‐over impacts from U. lugens have only been recorded to date on Metrosideros excelsa.

     

Comparison of the Acute Toxicity of Corexit 9500 and Household Cleaning Products

Surfactant formulations used in chemical dispersing agents are derived from the same functional components used in numerous household products such as dishwashing soaps and laundry detergents. During the Deep Water Horizon (DWH) oil spill response, a significant volume of chemical dispersant was deployed, causing members of the public and the media to question the role of chemical dispersant (Corexit 9500) usage in mitigating oil spill effects. Consequently, laboratory tests were conducted by regulatory agencies to further evaluate and substantiate the existing aquatic toxicity of Corexit dispersants. To help put dispersant toxicity in context, two independent accredited labs were commissioned to conduct parallel studies that compared the acute toxicity of Corexit 9500 to common household cleaning agents. The results indicate that the acute toxicity of Corexit 9500 to marine aquatic organism is either within the median range or less toxic than the household cleaning agents tested. The median LC50 value for Corexit 9500 exposures to Americamysis bahia was 42.5 mg/L (four products were less toxic and four products were more toxic); whereas, the median LC50 value for Corexit 9500 exposures to Menidia beryllina was 73.1 mg/L (one product was less toxic and seven products were more toxic).     

A permanent fish cell line (EPC) for genotoxicity testing of marine sediments with the comet assay

Genotoxicity and cytotoxicity were evaluated in an in vitro system with a permanent cell line Epithelioma papulosum cyprini (EPC) derived from a skin tumour of carp (Cyprinus carpio L.). EPC cells were exposed to different concentrations of organic sediment extracts from the North Sea for 24 h. After incubation the cells were analysed for viability and DNA strand breaks with the comet assay or single cell gel electrophoresis (SCGE). The results confirm the sensitivity of this assay. Out of 10 marine sediment samples from the North Sea, 9 showed a dose-dependent genotoxic effect. The EC₅₀ of sediment extracts ranged from 7 to 307 mg sediment dry weight/ml assay volume. Hepatic microsomal enzymes from dab (Limanda limanda L.) was proposed for enzymatic activation of benzo[a]pyrene (BAP) or sediment extracts, respectively. The suitability of this in vitro test system for assessing genotoxic and cytotoxic effects of marine sediment extracts on EPC cells could be demonstrated.     

Generalist hydrocarbon‐degrading bacterial communities in the oil‐polluted water column of the North Sea

The aim of this work was to determine the effect of light crude oil on bacterial communities during an experimental oil spill in the North Sea and in mesocosms (simulating a heavy, enclosed oil spill), and to isolate and characterize hydrocarbon‐degrading bacteria from the water column. No oil‐induced changes in bacterial community (3 m below the sea surface) were observed 32 h after the experimental spill at sea. In contrast, there was a decrease in the dominant SAR11 phylotype and an increase in Pseudoalteromonas spp. in the oiled mesocosms (investigated by 16S rRNA gene analysis using denaturing gradient gel electrophoresis), as a consequence of the longer incubation, closer proximity of the samples to oil, and the lack of replenishment with seawater. A total of 216 strains were isolated from hydrocarbon enrichment cultures, predominantly belonging to the genus Pseudoaltero monas; most strains grew on PAHs, branched and straight‐chain alkanes, as well as many other carbon sources. No obligate hydrocarbonoclastic bacteria were isolated or detected, highlighting the potential importance of cosmopolitan marine generalists like Pseudoalteromonas spp. in degrading hydrocarbons in the water column beneath an oil slick, and revealing the susceptibility to oil pollution of SAR11, the most abundant bacterial clade in the surface ocean.     

Assessment of the genotoxicity in toad Bufo raddei exposed to petrochemical contaminants in Lanzhou Region, China

Single cell gel electrophoresis or comet assay, micronucleus (MN) test and global DNA methylation detection were used to assess the genotoxicity in toad Bufo raddei exposed to the petrochemical (mainly oil and phenol) polluted area in Lanzhou Region (LZR) comparing with a relatively unpolluted area in Liujiaxia Region (LJXR). The results from the present study indicated that DNA damage and MN frequency in toad from LZR were significantly higher than those from LJXR at the same sampling month, whereas the degree of global DNA methylation was lower, which implies that the petrochemical contaminants at environmental level in LZR were genotoxic to B. raddei. The degree of genotoxic damage was obviously related with the extent of pollution among the three sampling months in LZR. The significantly positive correlations between DNA damage and concentrations of oil and/or phenol existed in liver cells but erythrocytes, implying that liver is more suitable as a sentinel tissue for the assessment of genotoxic impact of low-level contamination. The results from both comet assay and global DNA methylation detection on liver cells showed that the genotoxicity varied significantly with oil and/or phenol concentrations, suggesting that these two methods are relatively sensitive and suitable for monitoring the genotoxicity of petrochemical pollutants on amphibians.     

Spatial and temporal impacts of a diesel fuel spill on stream invertebrates

1. We assessed the effects of a 26 500 L diesel fuel spill on the macroinvertebrate fauna of a small trout stream in central New York, U.S.A. To determine the spatial extent of the spill we sampled three locations (0.7, 5.0 and 11.8 km downstream of the spill), each containing a reference site (unaffected tributary) and an impact site (downstream of spill). Sampling was repeated four times over a 15‐month period to assess temporal recovery. 2. Immediately after the spill, invertebrate density at all three locations below the spill was significantly lower than reference density. Three months after the spill, density up to 5 km below the spill was still far lower (<100 individuals per sample) than reference density (800–1200 individuals per sample). A year after the spill, density was similar between reference and impact sites, suggesting that invertebrates had recovered numerically. 3. Taxonomic richness up to 5.0 km below the spill was less than half the reference taxonomic richness and this difference persisted for at least 3 months. Some significant differences between reference and impact sites were observed after 15 months, but these differences could not be attributed to the oil spill. 4. For at least 3 months following the spill, the site immediately downstream of the spill was dominated by Optioservus, a petrochemical‐tolerant riffle beetle. Twelve to 15 months after the spill, both the reference and impact sites near the spill were dominated numerically by the mayfly Ephemerella, but the degree of dominance was twice as large at the impact site. 5. We concluded that the diesel fuel spill significantly reduced the density of invertebrates (by 90%) and taxonomic richness (by 50%) at least 5.0 km downstream, but density recovered within a year. Throughout the study, however, the fauna immediately below the spill was species poor and significantly over‐represented by a single dominant taxon, suggesting that 15 months was not sufficient for full community recovery from the oil spill.     

Enhanced crude oil biodegradative potential of natural phytoplankton‐associated hydrocarbonoclastic bacteria

Phytoplankton have been shown to harbour a diversity of hydrocarbonoclastic bacteria (HCB), yet it is not understood how these phytoplankton‐associated HCB would respond in the event of an oil spill at sea. Here, we assess the diversity and dynamics of the bacterial community associated with a natural population of marine phytoplankton under oil spill‐simulated conditions, and compare it to that of the free‐living (non phytoplankton‐associated) bacterial community. While the crude oil severely impacted the phytoplankton population and was likely conducive to marine oil snow formation, analysis of the MiSeq‐derived 16S rRNA data revealed dramatic and differential shifts in the oil‐amended communities that included blooms of recognized HCB (e.g., Thalassospira, Cycloclasticus), including putative novel phyla, as well as other groups with previously unqualified oil‐degrading potential (Olleya, Winogradskyella, and members of the inconspicuous BD7‐3 phylum). Notably, the oil biodegradation potential of the phytoplankton‐associated community exceeded that of the free‐living community, and it showed a preference to degrade substituted and non‐substituted polycyclic aromatic hydrocarbons. Our study provides evidence of compartmentalization of hydrocarbon‐degrading capacity in the marine water column, wherein HCB associated with phytoplankton are better tuned to degrading crude oil hydrocarbons than that by the community of planktonic free‐living bacteria.     

Toxicological Aspects of the Essential Oil from Cinnamodendron dinisii

The objective of this study was to determine cytotoxic activity, hemolytic activity, and to evaluate the ability of the essential oil from Cinnamodendron dinisii to induce DNA fragmentation of human lymphocytes. The essential oil was obtained by hydrodistillation. Cytotoxic activity was determined by the MTT method. Hemolytic activity was evaluated by spectrophotometric quantification of hemoglobin released by erythrocytes. Damage to lymphocyte DNA molecules was assessed by the Comet assay. The essential oil under study showed high cytotoxic activity on Vero cells (CC₅₀ = 35.72 μg/mL) and induced hemolysis in both hematocrits, besides leading to the oxidation of hemoglobin released. The genotoxic activity of C. dinisii essential oil was also observed, which induced concentration‐dependent DNA fragmentation of human lymphocytes and, at 50 μL/mL, it was more active than the positive control. The essential oil from C. dinisii has a toxic action, suggesting a special attention in the application of this oil to health‐promoting activities; however, among its components, there are molecules with potential for future application in anticancer therapies.     

Genotoxic damage in polychaetes: A study of species and cell-type sensitivities

The marine environment is becoming increasingly contaminated by environmental pollutants with the potential to damage DNA, with marine sediments acting as a sink for many of these contaminants. Understanding genotoxic responses in sediment-dwelling marine organisms, such as polychaetes, is therefore of increasing importance. This study is an exploration of species-specific and cell-specific differences in cell sensitivities to DNA-damaging agents in polychaete worms, aimed at increasing fundamental knowledge of their responses to genotoxic damage. The sensitivities of coelomocytes from three polychaetes species of high ecological relevance, i.e. the lugworm Arenicola marina, the harbour ragworm Nereis diversicolor and the king ragworm Nereis virens to genotoxic damage are compared, and differences in sensitivities of their different coelomic cell types determined by use of the comet assay. A. marina was found to be the most sensitive to genotoxic damage induced by the direct-acting mutagen methyl methanesulfonate (MMS), and showed dose-dependent responses to MMS and the polycyclic aromatic hydrocarbon benzo(a)pyrene. Significant differences in sensitivity were also measured for the different types of coelomocyte. Eleocytes were more sensitive to induction of DNA damage than amoebocytes in both N. virens and N. diversicolor. Spermatozoa from A. marina showed significant DNA damage following in vitro exposure to MMS, but were less sensitive to DNA damage than coelomocytes. This investigation has clearly demonstrated that different cell types within the same species and different species within the polychaetes show significantly different responses to genotoxic insult. These findings are discussed in terms of the relationship between cell function and sensitivity and their implications for the use of polychaetes in environmental genotoxicity studies.