Cold sediment microbial community shifts in response to crude oil water-accommodated fraction with or without dispersant: a microcosm study.

In cold environments, the low temperature slows down microbial metabolisms, such as the biodegradation processes of hydrocarbons, which are often stimulated by the addition of dispersants in oil spill disasters. In this study, we investigated the effects of hydrocarbon water-accommodated fraction (WAF) prepared with and without dispersant on benthic microbial communities in a microcosm experiment in which hydrocarbon removal was observed. Both WAFs contained similar polycyclic aromatic hydrocarbon (PAH) content. The microcosm experiment, set up with either pristine or contaminated sediments, was conducted for 21 days at 4 °C under WAF and WAF + dispersant conditions. The behavior of bacterial communities in response to WAF and WAF + dispersant was examined at both DNA and RNA levels, revealing the effect of WAF and WAF + dispersant on the resident and active communities respectively. The contaminated sediment showed less taxa responsive to the addition of both WAF and WAF + dispersant than the pristine sediment, indicating the legacy effect by the presence hydrocarbon-degrading and dispersant-resistant taxa inhabiting the contaminated sediment.

Legacy and dispersant influence microbial community dynamics in cold seawater contaminated by crude oil water accommodated fractions.

Dispersants, used for combating oil spills, increase hydrocarbon bioavailability promoting their biodegradation. Oil weathering process introduces harmful soluble hydrocarbons, such as polycyclic aromatic hydrocarbons (PAHs), into the water column, resulting in water-accommodated fraction (WAF). The presence of dispersants can influence the weathering process by increasing PAHs solubility, toxicity and biodegradability. However, little is known on how dispersants affect microbial communities and their degradation capacities, especially in cold environment where low temperature decreases microbial activity and thus hydrocarbon degradation. Here, we investigated the microbial community dynamics in cold water contaminated by WAF prepared from crude oil with or without a commercial dispersant (Finasol OSR52). The WAFs, prepared with Naphthenic North Atlantic crude oil, were used to contaminate seawater from Norwegian cold sites, one oil-contaminated and the other pristine. The WAF-contaminated seawaters were maintained in microcosms at 4 °C for 21 days. The content of PAHs and microbial compositions (16S rRNA gene sequencing) were determined at days 0, 7, 14 and 21. In addition, the 96 h toxicity assay with adult Acartia tonsa revealed WAFs toxicity at days 0 and 21. The toxicity of WAF mixtures, with and without dispersant, against Acartia tonsa was reduced during the experiment, but PAHs removal was not increased. The water from the oil-contaminated site showed the highest PAHs removal revealing legacy effect (presence of microorganisms adapted to PAHs). Additionally, our results reveal: i) microbial community plasticity allowing the adaptation to the presence of PAHs and dispersant, ii) specific bacteria taxa probably involved in PAHs degradation, and iii) dispersants shape the microbial communities dynamics by stimulating potential dispersant-degrading taxa, such as Fusibacter. Thus, our results provide valuable insights on the role of microbial community in determining the fate of water-solubilized hydrocarbon in cold environment while questioning the role of dispersant used for fighting oil spill.

Application of a biological multilevel response approach in the copepod Acartia tonsa for toxicity testing of three oil Water Accommodated Fractions.

Copepods play a critical role in the marine food webs, being a food source for marine organisms. In this study, we investigated the toxic effects of Water Accommodated Fractions (WAFs) from three types of oil: Naphthenic North Sea crude oil (NNS), Intermediate Fuel Oil (IFO 180) and a commercial Marine Gas Oil (MGO). The WAFs were prepared at 10 °C and 30 PSU (practical salinity unit), and tested on the marine copepod Acartia tonsa at different endpoints and at different levels of biological organization. We determined the median lethal concentrations after 96 h (LC50) and reproduction capabilities were calculated in adult females following seven days of exposure to sublethal WAF doses. The total lipid content was measured in reproductive females using Nile red lipophilic dye after 96 h of WAF exposure. We also measured the transcription levels of genes involved in antioxidant response and xenobiotic biotransformation after short exposure for 48 h. High doses (7% WAF) of MGO affected survival, percentage of fecund females, egg hatching success, and total lipid content. The IFO 180 WAF affected, at medium (20%) and high (40%) doses, the number of fecund females, mortality and produced significant effects on gene expression levels. In conclusion, toxicity assays showed that the WAFs prepared from refined oils were more toxic than crude oil WAF to Acartia tonsa.

Alteration in molecular markers of oocyte development and intersex condition in mullets impacted by wastewater treatment plant effluents.

Wastewater Treatment Plant (WWTP) discharges are an important source of endocrine disrupting chemicals (EDCs) into the aquatic environment. Fish populations inhabiting downstream of WWTP effluents show alterations in gonad and gamete development such as intersex condition, together with xenoestrogenic effects such as vitellogenin up-regulation. However, the molecular mechanisms participating in the development of intersex condition in fish are not elucidated. The aim of this study was to assess the impact of two WWTPs effluents (Gernika and Bilbao-Galindo situated in the South East Bay of Biscay) with different contaminant loads, in thicklip grey mullet (Chelon labrosus) populations inhabiting downstream, examining the presence and severity of intersex condition, during two seasons. Molecular markers of xenoestrogenicity and oocyte differentiation and development (vtgAa, cyp19a1a, cyp19a1b, cyp11b, foxl2, dmrt1 and gtf3a) were also studied. Intersex mullets were identified downstream of both WWTPs and vtgAa was upregulated in intersex and non intersex males. Sex dependent differential transcription levels of target genes were detected in mullets from Galindo. However, no such pattern was observed in mullets from Gernika, suggesting an attenuating effect over studied genes caused by a higher presence of EDCs in this site, as indicated by the elevated prevalence of intersex mullets in this population. In conclusion, no direct association between xenoestrogenic responses and intersex condition was established. Mullets from Gernika showed signs of severe EDC exposure compared to those from Galindo, as demonstrated by the higher prevalence of intersex males and the reduction in transcription profile differences between sexes of gametogenic gene markers.

Environmental estrogen-induced alterations of male aggression and dominance hierarchies in fish: a mechanistic analysis.

Environmental estrogens have been shown to affect aspects of fish behavior that could potentially impact on wild populations, but the physiological mechanisms underpinning these effects are unknown. Using small colonies of zebrafish (Danio rerio), we evaluated the impacts of estrogen exposure on the aggression of dominant males, the associated implications for their social status and reproductive success, and their signaling mechanisms. The aggression of dominant males exposed to 17α-ethinylestradiol (EE(2); 10 ng/L nominal) was reduced significantly, and half of these fish subsequently lost their dominance, behavioral changes that were reflected in their reproductive success. Plasma androgen and the expression of genes involved in sex steroid production/signaling (cyp19a1b, cyp17, hsd11b2, hsd17b3, ar) and aggression (avplrv1b, tph1b, htr1a, sst1, sstr1, th, slc6a3, ar) were higher in control dominant versus subordinate males, but suppressed by EE(2) exposure, such that the differences between the social ranks were not retained. The expression levels of avpl (brain), which promotes aggression and dominance, and ar and cyp17 (gonad) were elevated in nonexposed males paired with EE(2)-exposed males. Our findings illustrate that disruptions of behaviors affecting social hierarchy, and in turn breeding outcome, as a consequence of exposure to an environmental estrogen are signaled through complex interconnecting gonadal and neurological control mechanisms that generally conform with those established in mammalian models. The extensive molecular, genetic, physiological, and behavioral toolbox now available for the zebrafish makes this species an attractive model for integrated analyses of chemical effects spanning behavior to molecular effect mechanisms.

Effects of chronic exposure to dispersed oil on selected reproductive processes in adult blue mussels (Mytilus edulis) and the consequences for the early life stages of their larvae.

Mussels (Mytilus edulis) were continuously exposed to dispersed crude oil (0.015-0.25mg/l) for 7 months covering the whole gamete development cycle. After 1 month exposure to 0.25 mg oil/l, the level of alkali-labile phosphates (ALP) and the volume density of atretic oocytes in female gonads were higher than those in the gonads of control females, indicating that oil affected the level of vitellogenin-like proteins and gamete development. Spawning of mussels was induced after 7 months oil exposure. Parental oil exposure did not affect subsequent fertilization success in clean seawater but this was reduced in 0.25 mg oil/l. Parental exposure to 0.25 mg oil/l caused both slow development and a higher percentage of abnormalities in D-shell larvae 2 days post-fertilization; reduced growth 7 days post-fertilization. These effects were greatly enhanced when larval stages were maintained at 0.25 mg oil/l. Similar studies are warranted for risk assessment prognosis.

Application of a battery of biomarkers in mussel digestive gland to assess long-term effects of the Prestige oil spill in Galicia and Bay of Biscay: correlation and multivariate analysis.

According to published reports, selected cell and tissue-level biomarkers revealed alterations in the health status of mussels collected after the Prestige oil spill (POS) in Galicia and the Bay of Biscay (2003-2006). Presently, univariate and multivariate analysis provides a comprehensive view of the whole data set. Correlation and principal component analysis (PCA) were performed using biomarkers corresponding to or derived from those data, say: induction, AOX(exp), and inhibition, AOX(eff), of Acyl-CoA oxidase enzyme activity; membrane labilisation period, LP, volume density, Vv(LYS), and surface-to-volume ratio, S/V(LYS), of digestive cell lysosomes; volume density of neutral lipids, Vv(NL); volume density of basophilic cells, Vv(BAS), mean luminal radius to mean epithelial thickness in digestive gland epithelium, MLR/MET; gonad index, GI; cumulative intensity of inflammatory responses, CI(IR) and parasitic infestation, CI(PI). The PCA results were used to classify sampling times and localities by means of hierarchical cluster analysis. Biomarkers were more frequently correlated with naphthalene than with total PAH concentration in mussel tissues. Many biological parameters were correlated with each other in agreement with their involvement in toxic processes or their role in the mussels' response against environmental insult. Naphthalene appears to be the most effective toxic herein, which supports that the reported effects may be mainly due to POS. According to the PCA, 4 groups of biomarkers explained 67.37% variability and the factors that most strongly contributed were S/V(LYS), CI(IR), Vv(BAS) and MLR/MET. These factors were used in the hierarchical cluster analyses carried out. The cluster analysis performed to classify sampling times discriminated 3 significantly different periods after POS: (a) a high affection period (until April-04); (b) the beginning of the recovery; and (c) an advanced recovery status in which geographical variability increased (from April-05 onwards). As regards the clustering of localities, major geographical differences were not detected but three outliers were identified: (a) early and highly impacted localities in Galicia and Bay of Biscay; (b) Aguiño that was characterised by extremely high Vv(BAS) during 2003; and (c) localities that were subjected to persistent chronic pollution.

Effects of the fuel oil spilled by the Prestige tanker on reproduction parameters of wild mussel populations.

The aim of this work was to assess possible effects of the Prestige oil spill on reproduction parameters of mussels along the Galician and Bay of Biscay coast. Studied endpoints included sex ratio, gonad histology and vitellogenin-like proteins using the alkali-labile phosphate (ALP) method. A high prevalence of haemocytic infiltration of follicles and severe oocyte atresia was found in most localities in April 2003. Spawning gonads were observed in most impacted populations in the same sampling. In April 2004 mature small sized follicles were observed. No histopathological changes were observed in April 2005 and 2006, except a high prevalence of necrotic gametes in 6 out of 22 localities in April 2006. Female ALP levels showed high interindividual variability in April 2004, which was reduced in April 2005 and 2006. No xenoestrogenic effects were observed in male mussels. Overall, gamete alterations were detected during 2003-2004 and a recovery trend was observed afterwards.

Specificity of the peroxisome proliferation response in mussels exposed to environmental pollutants.

Peroxisome proliferation has been proposed as novel biomarker of exposure to organic pollutants in aquatic organisms. Peroxisome proliferator compounds comprise a heterogeneous group of substances known for their ability to cause massive proliferation of peroxisomes and liver carcinogenesis in sensitive species such as rodents. Recently, several marine organisms (mussels and fish) have been shown as target species of peroxisome proliferators. In the present work, we aimed to investigate the specificity of the peroxisome proliferation response in mussels. For this purpose, mussels (Mytilus edulis) were exposed for three weeks to North Sea crude oil (NSO), a mixture of NSO, alkylphenols and extra PAHs (MIX), diallylphthalate (DAP), bisphenol-A (BPA) and tetrabromodiphenylether (TBDE), or transplanted for three weeks to four stations showing different copper concentrations in a copper mine. Peroxisome proliferation was assessed by measuring the activity of the peroxisomal beta-oxidation enzyme acyl-CoA oxidase (AOX) and the volume density occupied by peroxisomes (V(VP)) in the digestive gland. Mussels exposed to NSO and MIX showed significantly increased AOX activities and V(VP) compared to control animals. Significantly higher V(VP) was also found in DAP and TBDE exposed mussels. V(VP) did not vary in mussels transplanted into a copper concentration gradient. Our results confirm the usefulness and specificity of peroxisome proliferation as a suitable biomarker of exposure to organic contaminants such as oil derived hydrocarbons, phthalate plasticizers and polybrominated flame retardants in mussels.