Early Biological Modulations Resulting from 1-Week Venlafaxine Exposure of Marine Mussels Mytilus galloprovincialis Determined by a Metabolomic Approach.

There is growing evidence of the presence of pharmaceuticals in natural waters and their accumulation in aquatic organisms. While their mode of action on non-target organisms is still not clearly understood, their effects warrant assessment. The present study assessed the metabolome of the Mediterranean mussel (Mytilus galloprovincialis) exposed to a 10 µg/L nominal concentration of the antidepressant venlafaxine (VLF) at 3 time-points (1, 3, and 7 days). Over the exposure period, we observed up- or down-modulations of 113 metabolites, belonging to several metabolisms, e.g., amino acids (phenylalanine, tyrosine, tryptophan, etc.), purine and pyrimidine metabolisms (adenosine, cyclic AMP, thymidine, etc.), and several other metabolites involved in diverse functions. Serotonin showed the same time-course modulation pattern in both male and female mussels, which was consistent with its mode of action in humans, i.e., after a slight decrease on the first day of exposure, its levels increased at day 7 in exposed mussels. We found that the modulation pattern of impacted metabolites was not constant over time and it was gender-specific, as male and female mussels responded differently to VLF exposure.

An integrated metabolomics and proteogenomics approach reveals molecular alterations following carbamazepine exposure in the male mussel Mytilus galloprovincialis.

Carbamazepine is one of the most abundant pharmaceutical active compounds detected in aquatic systems. Based on laboratory exposures, carbamazepine has been proven to adversely affect aquatic organisms. However, the underlying molecular events remain poorly understood. This study aims to investigate the molecular mechanisms potentially associated with toxicological effects of carbamazepine on the mussel Mytilus galloprovincialis exposed for 3 days at realistic concentrations encountered in coastal environments (80 ng/L and 8 µg/L). An integrated metabolomics and proteogenomics approach, including data fusion strategy, was applied to gain more insight in molecular events and cellular processes triggered by carbamazepine exposure. Consistent metabolic and protein signatures revealed a metabolic rewiring and cellular stress at both concentrations (e.g. intensification of protein synthesis, transport and catabolism processes, disruption of lipid and amino acid metabolisms). These highlighted molecular signatures point to the induction of autophagy, closely related with carbamazepine mechanism of action, as well as a destabilization of the lysosomal membranes and an enzymatic overactivity of the peroxisomes. Induction of programmed cell death was highlighted by the modulation of apoptotic cognate proteins. The proposed integrative omics data analysis was shown to be highly relevant to identify the modulations of the two molecular levels, i.e. metabolites and proteins. Multi-omics approach is able to explain the resulting complex biological system, and document stronger toxicological pieces of evidence on pharmaceutical active compounds at environmental concentrations in sentinel organisms.

Occurrence, distribution, and ecological risk assessment of emerging and legacy contaminants in the Kadicha river in Lebanon.

The Kadicha river basin in Northern Lebanon is an illustrative example of multiple pressures encountered in the Mediterranean region: it is a small coastal river affected by rapid urbanization, population growth (drastically impacted by the influx of Syrian refugees), and a chronic default of wastewater treatment. In this context, multiple classes of contaminants may attain the river accumulating in sediment. However, very little information is available in the literature on the contamination status in such stressed Mediterranean contexts. This study proposed a first contamination evaluation of a small Mediterranean river submitted to multiple pressures. Two sediment sampling campaigns along sites impacted by increasing urban gradient within the Kadicha river basin were performed to determine the occurrence and the environmental risks of both emerging and legacy contaminants. The results revealed the detection of the 41 studied compounds. The highest concentrations were attained by PAHs and polycyclic musks (up to 311.79, 94.22, and 81.13 ng/g of dry weight for PAH, cashmeran, and galaxolide, respectively). The discontinuous urbanized upstream area and the estuary were the most contaminated areas of the river. An environmental risk assessment showed a hazard quotient (HQ) higher than 1 for both legacy and emerging compounds (EHMC and 4-MBC), indicating a potential risk to benthic species. Monitoring campaigns and implementation of wastewater treatment plants should be encouraged as the anthropogenic pressure on small Mediterranean rivers will increase over the years.

Diurnal variations in personal care products in seawater and mussels at three Mediterranean coastal sites.

The presence of personal care products (PCPs) in the marine environment is of major concern. PCPs, UV filters, and musks can enter the marine environment indirectly through wastewater or directly via recreational activities. We conducted this study to document patterns in the occurrence of seven PCPs at three coastal sites impacted by recreational activities during 1 day. The study focused on diurnal variations in these seven PCPs in seawater and indigenous mussels. In seawater, UV filters showed diurnal variations that mirrored variations in recreational activities at the sites. Ethylhexyl methoxycinnamate (EHMC) and octocrylene (OC) water concentrations increased from under the limit of quantification in the morning to 106 and 369 ng/L, respectively, when recreational activities were the highest. In mussels, diurnal variations in OC were observed, with the lowest concentrations recorded in the morning and then increasing throughout the day. As Mytilus spp. are widely used as sentinels in coastal pollution monitoring programs (mussel watch), our findings on diurnal variations could enhance sampling recommendations for recreational sites impacted by PCPs.

Metabolomics assessment of the effects of diclofenac exposure on Mytilus galloprovincialis: Potential effects on osmoregulation and reproduction.

The presence of pharmaceutically active compounds in aquatic environments has become a major concern over the past 20years. Elucidation of their mode of action and effects in non-target organisms is thus now a major ecotoxicological challenge. Diclofenac (DCF) is among the pharmaceutical compounds of interest based on its inclusion in the European Union Water Framework Directive Watch List. In this study, our goal was to investigate the potential of a metabolomic approach to acquire information without any a priori hypothesis about diclofenac effects on marine mussels. For this purpose, mussel's profiles were generated by liquid chromatography combined with high resolution mass spectrometry. Two main metabolic pathways were found to be impacted by diclofenac exposure. The tyrosine metabolism was mostly down-modulated and the tryptophan metabolism was mostly up-modulated following exposure. To our knowledge, such DCF effects on mussels have never been described despite being of concern for these organisms: catecholamines and serotonin may be involved in osmoregulation, and in gamete release in mollusks. Our results suggest potential impairment of mussel osmoregulation and reproduction following a DCF exposure.

Spatial and temporal trends in water quality in a Mediterranean temporary river impacted by sewage effluents.

This paper analyzes how changes in hydrological conditions can affect the water quality of a temporary river that receives direct inputs of sewage effluents. Data from 12 spatial surveys of the Vène river were examined. Physico-chemical parameters, major ion, and nutrient concentrations were measured. Analyses of variance (ANOVA) and multivariate analyses were performed. ANOVA revealed significant spatial differences for conductivity and major ion but no significant spatial differences for nutrient concentrations even if higher average concentrations were observed at stations located downstream from sewage effluent discharge points. Significant temporal differences were observed among all the parameters. Karstic springs had a marked dilution effect on the direct disposal of sewage effluents. During high-flow periods, nutrient concentrations were high to moderate whereas nutrient concentrations ranged from moderate to bad at stations located downstream from the direct inputs of sewage effluents during low-flow periods. Principal component analysis showed that water quality parameters that explained the water quality of the Vène river were highly dependent on hydrological conditions. Cluster analysis showed that when the karstic springs were flowing, water quality was homogeneous all along the river, whereas when karstic springs were dry, water quality at the monitoring stations was more fragmented. These results underline the importance of considering hydrological conditions when monitoring the water quality of temporary rivers. In view of the pollution observed in the Vène river, "good water chemical status" can probably only be achieved by improving the management of sewage effluents during low-flow periods.

In vitro biomonitoring of contamination by estrogenic compounds in coastal environments: comments on the use of M. galloprovincialis.

The use of mussel extracts in in vitro bioassays which express the estrogen receptor could provide valuable information on the bioavailability of endocrine disruptors in coastal environments. The aim of this study was to assess the temporal variability of the estrogenic responses in bioassays in Mytilus galloprovincialis. A 6-month in situ experiment was conducted in order to follow the estrogenic activity on MELN cell line during the reproduction stages of mussels. Estradiol equivalents (EEQ) determined in mussels using the MELN cell lines ranged from 0.79 to 3.72 ng/g dry weight (d.w.) in males, from 0.42 to 2.33 ng/g d.w. in females and from 3.41 to 4.2 d.w. in undifferentiated bivalves. We observed an increase in EEQ values during the spawning stage for males, not for female. The maximal EEQ values were observed at the indifferent stage. We discuss these results in regards to the actual knowledge on mussels' reproductive cycle and to the possible impact of xeno-estrogens. Variations of E2 levels in mussels must be taken into account for further studies on xeno-estrogens monitoring using hER reporter cell-lines bioassays.

Implication of two in-stream processes in the fate of nutrients discharged by sewage system into a temporary river.

The aim of this study was to better understand the fate of nutrients discharged by sewage treatment plants into an intermittent Mediterranean river, during a low-flow period. Many pollutants stored in the riverbed during the low-flow period can be transferred to the downstream environments during flood events. The study focused on two processes that affect the fate and the transport of nutrients, a physical process (retention in the riverbed sediments) and a biological process (denitrification). A spatial campaign was carried out during a low-flow period to characterize the nutrient contents of both water and sediments in the Vène River. The results showed high nutrient concentrations in the water column downstream of the treated wastewater disposal (up to 13,315 µg N/L for ammonium and 2,901 µg P/L for total phosphorus). Nutrient concentrations decreased rapidly downstream of the disposal whereas nutrient contents in the sediments increased (up to 1,898 and 784 µg/g for total phosphorus and Kjeldahl nitrogen, respectively). According to an in situ experiment using sediment boxes placed in the riverbed for 85 days, we estimated that the proportion of nutrients trapped in the sediments represents 25% (respectively 10%) of phosphorus (respectively nitrogen) loads lost from the water column. In parallel, laboratory tests indicated that denitrification occurred in the Vène River, and we estimated that denitrification likely coupled to nitrification processes during the 85 days of the experiment was significantly involved in the removal of nitrogen loads (up to 38%) from the water column and was greater than accumulation processes.

Monitoring organic contaminants in small French coastal lagoons: comparison of levels in mussel, passive sampler and sediment.

In this study, the distribution of organic contaminants was investigated in the particular context of three Mediterranean coastal lagoons, where pollution input was hypothesised to come mainly from sediments resuspension. Mussels and semi-permeable membrane devices (SPMDs) were exposed to the water column for one month and then their content in estrogen-, benzo[a]pyrene- and dioxin-like substances as well as polycyclic aromatic hydrocarbons (PAH), polychlorinated biphenyls and alkylphenols was determined with biological and chemical analyses. PAH concentration was high in sediments (up to 1028 ng g(-1) dry weight), however the aqueous PAH concentrations estimated from SPMD data could be considered below the levels inducing adverse effects according to the environmental quality standards proposed by the Water Framework Directive. Dioxin-like activity was observed in sediments but not in mussels and SPMDs. In the two sewage-impacted lagoons, nonylphenols could be quantified in sediments, SPMDs and mussels. Nonylphenol concentrations in mussels were among the highest found in the literature. However, since nonylphenols contributed only to a small part of the estrogenic activities observed, natural or synthetic steroids originating from wastewater discharges could be also implicated in these responses in sediments.