First baseline for bioenergetic biomarkers in Cnesterodon decemmaculatus as test organism in ecotoxicological studies.

Cnesterodon decemmaculatus is a Neotropical teleost fish frequently used in ecotoxicological evaluations, whose biology has been thoroughly studied. Although there is considerable information on its response to different toxicants, no range of reference values has been so far established for the different biological parameters proposed as biomarkers of effect or exposure. Moreover, no study has yet examined the possible influence of the metabolic status of the exposed animals on their response to toxic stress. Therefore, the aim of this work was to provide a first baseline for a set of bioenergetic biomarkers in C. decemmaculatus adults exposed to a control medium under previously standardized conditions, and to assess their possible intrinsic seasonal variability. The responses of the biomarkers obtained from the controls were contrasted with those from the reference toxicant (Cadmio-Cd) and receiving waters (surface waters of the Reconquista River RR, Buenos Aires Province, Argentina). We conducted four 12-day assays (one in each season) of exposure to control media, (reconstituted moderate hard water, MHW) and two assays of exposure to Cd in MHW and surface river water (RR) in both summer and autumn. The variables recorded were: Food intake (In), fecal production (F), specific assimilation (A) and cumulative mortality, oxygen extraction efficiency (OEE), specific metabolic rate (SMR), ammonia excretion (N), ammonia quotient (AQ) and scope for growth (SFG). The seasonal variation shown by some physiological parameters, points to the need for establishing a baseline obtained from standardized media, preferably on a seasonal basis. Moreover, SFG and A appeared as the most sensitive biomarkers, emphasizing the importance to consider the metabolic status of the test organisms for the appropriate interpretation of results from ecotoxicological studies performed under controlled experimental conditions. The obtained results provide useful information on C. decemmaculatus as model species in ecotoxicological bioassays involving biomarkers of early effect.

Assessment of cytotoxicity biomarkers on the microalga Chlamydomonas reinhardtii exposed to emerging and priority pollutants.

Contamination of aquatic ecosystems linked to anthropogenic activity is currently a major concern; therefore, ecotoxicological studies are needed to assess its effect on organisms. The main objective of this study was to investigate the effects of different pollutants on microalgae in search of sensitive biomarkers that can promote a common cytotoxic response regardless of the contaminant. Cultures of the freshwater microalga Chlamydomonas reinhardtii were exposed for 24 h to four chemicals, three emerging pollutants (benzophenone-3, bisphenol A and oxytetracycline) and one priority substance (atrazine). A cytometric panel was carried out to assess toxicity biomarkers including cellular growth, inherent cell properties, viability, vitality, cytoplasmic membrane potential and ROS levels. Lipid peroxidation, photosynthetic efficiency and transcriptional responses of photosynthesis- and oxidative stress-related genes using RT-qPCR were also studied. Some toxicity responses showed a similar pattern; a decrease in growth rate, vitality and photosynthetic efficiency and an increase in autofluorescence and in the number of cells with depolarised cytoplasmic membrane and were found for all chemicals tested. However, ATZ and OTC provoked a decrease in cell size, whereas BP-3 and BPA caused an increase in cell size, intracellular complexity and ROS levels and a decrease in cell viability. Assayed pollutants generally promoted an overexpression of genes related to cellular antioxidant defence system and a subexpression of photosynthesis-related genes. In addition to the traditional growth endpoint, cell vitality, autofluorescence and gene expression of catalase, glutathione peroxidase and Fe-superoxide dismutase were significantly affected for all chemicals tested, showing a common cytotoxic response. Among the tested substances, BP-3 provoked the strongest cytotoxic alterations on this microalga, pointing out that some emerging contaminants could be more harmful to organisms than priority pollutants.

Estimation of toluene exposure in air from BMA (S-benzylmercapturic acid) urinary measures using a reverse dosimetry approach based on physiologically pharmacokinetic modeling.

This study aimed to use a reverse dosimetry PBPK modeling approach to estimate toluene atmospheric exposure from urinary measurements of S-benzylmercapturic acid (BMA) in a small group of individuals and to evaluate the uncertainty associated to urinary spot-sampling compared to 24-h collected urine samples. Each exposure assessment technique was developed namely to estimate toluene air exposure from BMA measurements in 24-h urine samples (24-h-BMA) and from distributions of daily urinary BMA spot measurements (DUBSM). Model physiological parameters were described based upon age, weight, size and sex. Monte Carlo simulations with the PBPK model allowed converting DUBSM distribution (and 24-h-BMA) into toluene air levels. For the approach relying on DUBSM distribution, the ratio between the 95% probability of predicted toluene concentration and its 50% probability in each individual varied between 1.2 and 1.4, while that based on 24-h-BMA varied between 1.0 and 1.1. This suggests more variability in estimated exposure from spot measurements. Thus, estimating toluene exposure based on DUBSM distribution generated about 20% more uncertainty. Toluene levels estimated (0.0078-0.0138 ppm) are well below Health Canada's maximum chronic air guidelines. PBPK modeling and reverse dosimetry may be combined to interpret urinary metabolites data of VOCs and assess related uncertainties.

Polyclonal antibody-based immunoassay of vitellogenin in Van fish (Alburnus tarichi).

Van Lake is the third largest closed lake in the world and the biggest lake in Turkey. An ELISA method has developed with the aim of determining the pollution caused by estrogens and estrogen-like chemicals that have come to the lake Van in recent years. First, the vitellogenin in estrogen-treated male fish plasma was purified by ion exchange chromatography, injected into rats, and the obtained polyclonal antibodies were tested for specificity by Western blot and immunohistochemical methods. Immunohistochemical labeling of the vitellogenin-synthesized liver resulted in the intense marking of the liver of the animals injected with estrogen, while no markings were observed in the control group. The limit of detection of the developed enzyme-linked immunosorbent assay was 4.6 µg L-1, and the working range was 7.8 to 2000 µg L-1. Intra- and inter-assay variations were 13.0 % and 13.3%. The highest level of vitellogenin in male fishes measured was 23.56 µg mL-1.

Critical knowledge gaps and relevant variables requiring consideration when performing aquatic ecotoxicity assays.

The increasing diversity and complexity of contaminants released in the environment continuously lead to new challenges when applying ecotoxicity assays. This paper comprises a review concerning exposure assessment and highlights important variables that should be taken into account when investigating aquatic media toxicity under both laboratory or field conditions. Thus, to reflect as much as possible what occurs in nature, ecotoxicity assays must carefully consider these variables in their experimental design. This includes contaminant properties, the selected bioindicators and biomarkers, the dose mode/regime, concentration vs. load, exposure to single vs. multiple contaminants and exposure of single vs. multiple species. Many of these, however, are not usually taken into account, leading to critical knowledge gaps in this area, discussed in detail herein.

How metals directly affect the antioxidant status in the liver and kidney of Oreochromis niloticus? An in vitro study.

BACKGROUND: Metals can disturb the integrity of physiological and biochemical mechanisms in fish. Thus components of defense as an antioxidant system are significant biomarkers due to their vital role in coping with metal stress. The aim of the current study is to investigate the direct effects of Cd, Cu, and Zn sublethal exposures (in vitro) on the antioxidant system parameters in the liver and kidney of Nile tilapia. METHODS: The antioxidant enzyme activities and GSH levels were analyzed after in vitro sublethal metal (200 and 400 µg/L Cd, Cu, and Zn) treatments of Oreochromis niloticus liver and kidney supernatants. RESULTS: Metals even at lower levels caused significant changes in the levels of antioxidant system parameters due to concentration, metal, and tissue type. GSH metabolism parameters were more responsive to the metal effect. TBARS levels and GPX activity were mostly increased while CAT, SOD, rGSH, and GSH/GSSG levels decreased. The kidney was more affected than the liver in vitro conditions. Cu was more effective in the liver whereas it was Zn for the kidney. Cd caused negative correlations among the antioxidant enzymes. Significant correlations were found between enzymes and GSH levels upon Zn and Cu exposures. CONCLUSIONS: Direct metal effects may trigger different response trends due to their nature and tissue differences. The current data provide a knowledge about which antioxidant biomarkers can define better the oxidative stress caused by direct metal effect for further studies including in vivo experiments.

Comparative acute toxicity of glyphosate-based herbicide (GBH) to Daphnia magna, Tisbe longicornis, and Emerita analoga.

The aim of this work was to know the differential composition of the dissolved fraction of a glyphosate-based herbicide (GBH), commercialized as GLIFOPAC, when reaches different aquatic environments and its ecotoxicological effects on crustaceans species living in them. Daphnia magna, Tisbe longicornis, and Emerita analoga were exposed to glyphosate herbicide called GLIFOPAC (480 g L-1 of active ingredient or a.i.) at concentrations between 0.5 and 4.8 g a.i. L-1. Acute toxicity in D. magna (48 h-LC50), E. analoga (48 h-LC50), and T. longicornis (96 h-LC50) was studied. Chromatographic analysis of the GBH composition used and water (freshwater/sea water) polluted with GLIFOPAC were evaluated. Results reported acute toxicity (48-96 h-LC50) values for D. magna, E. analoga and T. longicornis of 27.4 mg L-1, 806.4 mg L-1, and 19.4 mg L-1, respectively. Chromatographic evaluation described around 45 substances of the GLIFOPAC composition, such as from the surfactant structures (aliphatic chain with esther/ether group), metabolites (AMPA), and other substances (glucofuranose, glucopyranoside, galactopyranose). This study evidenced differences in the GLIFOPAC composition in freshwater and marine water, which may differentiate the toxic response at the crustacean-level in each aquatic environment.

Cytoprotective and anti-apoptotic action of HSP70 stress protein in Oreochromis niloticus exposed to residual dilutions of insecticides with fipronil and ethiprole.

The objective of this research was to investigate the potential damage caused by the residual concentrations of the insecticides Regent® WS 800 and Curbix® SC 200, containing fipronil and ethiprole, respectively as active ingredients, on the liver of Oreochromis niloticus. The analyses of HSP70 shock protein labelling and cell death process by TUNEL method were performed in order to measure the effects of the exposure of cell repair system of fish to both insecticides. Statistical analyses showed no significant molecular damage to the hepatic tissue of animals. Nevertheless, variations in HSP70 and DNA fragmentation levels, endpoint of cell repair system response and cellular death, respectively, were observed in several groups. These results indicate that the cell repair machinery was efficient when in contact with residual concentrations of insecticides. However, the DNA fragmentation detected by the TUNEL method suggests that even in face of the cytoprotective action of the HSP70 protein, there are damages that become irreparable. To finish, it is worth mentioning that given the results obtained from residual concentrations, use in the field should be with caution.

Biochemical and genotoxic biomarkers and cell cycle assessment in the zebrafish liver (ZF-L) cell line exposed to the novel metal-insecticide magnesium-hespiridin complex.

The flavonoid metal-insecticide magnesium-hesperidin complex (MgHP) has recently been considered as a novel insecticide to replace some persistent pesticides. However, it is important to evaluate its action on non-target species, mainly those living in an aquatic environment, as these ecosystems are the final receptors of most chemicals. Reactive oxygen species, antioxidant and oxidative stress biomarkers, genotoxicity as well as cell cycle was evaluated in the liver cell line from zebrafish (Danio rerio; ZF-L) exposed to 0, 0.1, 1, 10, 100 and 1000 ng mL-1 MgHP. MgHP affected cell stability by increasing reactive oxygen species (ROS) in both exposure times (24 and 96 h) at high concentrations. Catalase (CAT) activity decreased after 24 h exposure, and glutathione and metallothionein values increased, avoiding the lipid peroxidation. Genotoxicity increased as MgHP concentration increased, after 24 h exposure, exhibiting nuclear abnormalities; it was recovered after 96 h exposure, evidencing possible stimulation of DNA repair mechanisms. The alteration in the cell cycle (increasing in the Sub-G1 phase and decreasing in the S-phase) was associated with chromosomal instability. In conclusion, the responses of ROS and the antioxidant defense system depended on MgHP concentration and time exposure, while DNA exhibited some instability after 24 h exposure, which was recovered after 96 h.

Correct Fish Muscle AChE Extraction for Use in Environmental Risk Assessment and Ecotoxicological Studies Has Been Neglected.

Several articles published in the field of ecotoxicology and environmental risk assessment have failed to adequately use fish muscle cholinesterases as biomarkers. This letter seeks to underline the importance of developing and applying correct protocols for the extraction of these enzymes from the tissues of these animals.

Bioaccumulation and oxidative damage of polycyclic aromatic hydrocarbon mixtures in Manila clam Ruditapes philippinarum.

The aim of this study was to investigate the bioaccumulation and oxidative damage of Manila clam, Ruditapes philippinarum, exposed to four selected mixtures of polycyclic aromatic hydrocarbons (PAHs; benzo (a) pyrene (BaP), benzo (a) anthracene (BaA), benzo (b) fluoranthene (BbF), and chrysene (Chr) in equal proportion. For this purpose, clams were exposed to PAHs (BaP:BbF:BaA:Chr = 1:1:1:1) at different concentrations (0.05, 0.5, and 5 µg/L) for 21 days, followed by a 15-day depuration period. All four PAHs accumulated in the gill, digestive gland, adductor muscle, and soft tissue of Manila clams, and all PAH treatment groups showed clear time and dose dependence. The decreasing order of bioaccumulation for the four PAHs in the exposure experiment was Chr > BaA > BaP > BbF. Moreover, the order of PAH bioaccumulation for the four tissues during the whole experiment was digestive gland > gill > soft tissues > adductor muscles. Although the initial concentrations of the four PAHs were the same, the final accumulated contents were different. Therefore, we also determined the detoxification processes of the four PAH mixtures in gills and digestive glands. The bioaccumulation of Chr was higher than the other three PAHs, probably because clams have a lower metabolic capacity for Chr than for BaP, BbF, and BaA. Exposure to PAH mixtures can result in oxidative damage, as indicated by the fact that DNA strand breaks, lipid peroxidation (LPO), and protein carbonyl (PC) were induced significantly (P < 0.05), except in the low-dose groups of PAHs, and different trends were detected with time of exposure. According to the correlation analysis, aryl hydrocarbon hydroxylase, glutathione s-transferase, superoxide dismutase, DNA strand break, PC, and LPO in both the gill and digestive gland are potential early indicators of PAH mixtures. We investigated the accumulation rules of R. philippinarum exposed to the selected PAHs and screened the potential biomarkers. The results of our study provide important scientific information for the purpose of monitoring marine pollution.

Effects of Microplastics Associated with Triclosan on the Oyster Crassostrea brasiliana: An Integrated Biomarker Approach.

Urban waste is a complex mixture of different substances, including microplastics and pharmaceuticals and personal care products. Microplastics have a high affinity for hydrophobic substances. One of these substances is triclosan, a bactericide used in a variety of hygiene products. Therefore, microplastics (MPs) may serve as a vector between triclosan and aquatic organisms. The current study sought to evaluate the effects of the interaction between microplastics and triclosan based on a mechanistic approach in which the oyster Crassostrea brasiliana was used as a model. The organisms were exposed to three conditions: the control, microplastic (MP), and microplastic contaminated with triclosan (MPT). The organisms were exposed for 3 or 7 days. After the exposure time, hemolymph was sampled for performing the neutral red retention time assay and, subsequently, the gills, digestive glands, and adductor muscles were dissected for measuring biomarkers responses (EROD, DBF, GST, GPx, GSH, lipid peroxidation, DNA strand breaks, and AChE). Our results demonstrate combined effects of MPs associated with triclosan on oyster physiology and biochemistry, as well as on lysosomal membrane stability. These results contribute to understanding the effects of contaminants of emerging concern and microplastics on aquatic organisms.

Effects of a glyphosate-based herbicide on the development and biochemical biomarkers of the freshwater copepod Notodiaptomus carteri (Lowndes, 1934).

In this work we analyzed the effects of Sulfosato Touchdown®, a glyphosate-based herbicide, on the ontogenic development and biochemical markers of the freshwater copepod Notodiaptomus carteri. A 30-days life-cycle experiment was carried out with three different glyphosate concentrations (0, 0.38, and 0.81 mg L-1) to analyze the developmental time from nauplii to adult copepods and their individual growth. An additional 10-days experiment with the same glyphosate concentrations was designed to evaluate the energy reserves (glycogen, proteins and lipids) and the activity of three antioxidant enzymes, superoxide dismutase (SOD), catalase, and glutathione-S-transferase (GST) in adult copepods, separately for females and males. We found that the lowest glyphosate concentration increased the nauplii and total development time. The highest glyphosate concentration prevented copepods from reaching the adult stage, inhibited the growth of the first copepodite stage and increased the GST and SOD activity in adult females. According to our results, the presence of this herbicide in freshwater systems could impose a risk in the ecological role of copepods in nature. This study will contribute to propose the Notodiaptomus genus as model specie for monitoring purposes in the Neotropical aquatic systems.

The Broad-Scale Analysis of Metals, Trace Elements, Organochlorine Pesticides and Polycyclic Aromatic Hydrocarbons in Wetlands Along an Urban Gradient, and the Use of a High Trophic Snake as a Bioindicator.

Wetlands and their biodiversity are constantly threatened by contaminant pollution from urbanisation. Despite evidence suggesting that snakes are good bioindicators of environmental health, the bioaccumulation of contaminants in reptiles is poorly researched in Australia. We conducted the first broad-scale analysis of 17 metals and trace elements, 21 organochlorine pesticides, and 14 polycyclic aromatic hydrocarbons in the sediments (4 samples per site, December 2018) from four wetlands along an urban gradient in Perth, Western Australia, and from the livers (5 livers per site, February-April 2019) of western tiger snakes Notechis scutatus occidentalis captured at those sites. All 17 metals and trace elements were detected in the sediments of wetlands as well as 16 in the livers of tiger snakes. Arsenic, Cu, Hg, Pb, Se, and Zn were at concentrations exceeding government trigger values in at least one sediment sample. Two organochlorine pesticides and six of seven polycyclic aromatic hydrocarbons were detected in the sediments of a single wetland, all exceeding government trigger values, but were not detected in tiger snakes. Metals and trace elements were generally in higher concentration in sediments and snake livers from more heavily urbanised wetlands. The least urbanised site had some higher concentrations of metals and trace elements, possibly due to agriculture contaminated groundwater. Concentrations of nine metals and trace elements in snake livers were statistically different between sites. Arsenic, Cd, Co, Hg, Mo, Sb, and Se near paralleled the pattern of contamination measured in the wetland sediments; this supports the use of high trophic wetland snakes, such as tiger snakes, as bioindicators of wetland contamination. Contamination sources and impacts on these wetland ecosystems and tiger snakes are discussed herein.

Environmental behavior of and gastropod biomarker response to trace metals from a backwater area of Xian'nv lake.

Combined with sediment pollutant analysis, the gastropod Cipangopaludina cahayensis was chosen as an indicator organism to evaluate the environmental behavior of trace metals and the aquatic ecological risk that they present in a backwater area of Xian'nv Lake. Based on hydrological characteristics, 24 sampling sites representing the main stream (MS), tributaries (TR), lake area (LA) and lake tributaries (LT) were collected. The results revealed that cadmium (Cd) was the main pollutant and that it significantly accumulated in sediments of the research area. Based on the pollutant concentrations, the degree of Cd pollution was ranked in the following order: LA > MS > TR > LT. Several intersections between the rivers and Xian'nv Lake, including LA1, LA7 and LA 10, were observed to have higher Cd deposition. There was a significant difference in the spatial distribution of pollutants, which resulted in a higher accumulation of trace metals in the backwater area and its tributary. The Cd content in the visceral sac of C. cahayensis was positively correlated with the concentration of heavy metals in the sediment. The response of multiple antioxidant biomarkers, including superoxide dismutase (SOD), catalase (CAT), and glutathione S-transferase (GST), as well as the glutathione (GSH) content and the level of by-products of lipid peroxidation (TBARS), in C. cahayensis revealed a potential relationship to the environmental behavior of the pollutants. By combining the different biomarkers responses, the integrated biomarker response index (IBR) corresponded well with the pollution distribution characteristics in different areas.

Risk assessment of heavy metal toxicity by sensitive biomarker δ-aminolevulinic acid dehydratase (ALA-D) for onion plants cultivated in polluted areas in Kosovo.

Biomarkers allow an integrated risk assessment of heavy metal pollution effects in living organisms. In this study, the biochemical effects of Cd, Cr, Ni, Pb and Zn pollution in agricultural soil and their accumulation in Alium cepa L. (onion) were evaluated with ALA-D enzyme response as a biomarker, along with δ-aminolevulinic acid (ALA) and total chlorophyll contents in leaves of this plant. Soil samples were randomly selected from agricultural areas in two regions, Mitrovica and Obiliqi, which are considered the most industrially polluted regions in Kosovo. Results show that Pb and Zn concentrations in soil samples from Mitrovica (1953-2576 mg kg -1) and Obiliqi regions (138-179 mg kg -1) and their bioaccumulation levels in onion were significantly higher in comparison with the control group. There was an adverse negative correlation between Pb or Zn concentration and ALA-D activity and total chlorophyll content, and a positive correlation with ALA content. This study indicates that ALA-D activity can be used as a very sensitive biomarker for evaluation of heavy metal pollution. The bioaccumulation of heavy metals from soil polluted areas poses a threat for food contamination and public health.

Trace Elements in Crustaceans, Mollusks and Fish in the Kenyan Part of Lake Victoria: Bioaccumulation, Bioindication and Health Risk Analysis.

This is the first comprehensive report on the accumulation of Cr, Ni, As, and Ag in the fish species Nile tilapia Oreochromis niloticus and Nile perch Lates niloticus from Lake Victoria, complemented with recent data on Cu, Zn, Cd, and Pb. This also is the first report on Cr, Ni, As, and Ag levels in invertebrates: the shrimp Caridina nilotica, gastropod Pila ovata, and bivalve Mutela bourguignati. The study was conducted at five sites in the Kenyan part of Lake Victoria: four sites in Winam Gulf influenced by various anthropogenic pressures, including a site near Kisumu City, and one in the main lake, with lesser direct anthropogenic influence. Apart from Cu and Ag, which were highest in O. niloticus liver, the invertebrates had higher levels of trace elements than fish. Contamination of the gulf with trace elements was best mirrored by the invertebrates, whose mobility is limited; they accumulated Cr, Cd, Ag, and Pb corresponding to the levels in the surface sediment. The accumulation of trace elements in fish species and their bioindicative potential corresponded to their habitats and feeding behaviour. The tissue contents of most trace elements were higher in the inshore-dwelling, omnivorous O. niloticus compared to the pelagic, piscivorous L. niloticus. Cu (465 ± 689 mg/kg dw) and Ag (3.45 ± 1.49 mg/kg dw) in the liver of O. niloticus were up to 10 and 119 times higher than in L. niloticus, respectively. Oreochromis niloticus therefore has bioindicative potential for Cu and Ag contamination. Both the invertebrates and fish showed positive correlations between Cu and Ag concentrations, indicating similar source and/or uptake route. The target hazard quotients (THQ) show that there is no human health risk associated with the consumption of these fish. However, the levels of Zn, Cd, and Pb in P. ovata surpassed maximum food safety limits and are hence potentially unsafe for human consumption.

Oxidative Stress and Damage Biomarkers in Clam Ruditapes decussatus Exposed to a Polluted Site: The Reliable Biomonitoring Tools in Hot and Cold Seasons.

In the present study, a multi-biomarker approach was used to assess the biological effects of metal pollution in the southern lagoon of Tunis, on clam Ruditapes decussatus both in "hot" (in summer) and "cold" (in winter) seasons. Clams were collected in August 2015 and February 2016 from three sites of the lagoon and from Louza considered a reference site. The concentrations of five trace metals (cadmium, copper, iron, lead, and zinc) in the soft tissues of R. decussatus were evaluated at the sampling sites. A core of biomarkers indicative of (a) neurotoxicity (acetylcholinesterase, AChE); (b) biotransformation (glutathione S-transferase, GST); (c) oxidative stress (catalase, CAT; total glutathione peroxidase, T-GPx; total glutathione peroxidase, T-GPx; selenium-dependent glutathione peroxidase, Se-GPx; glutathione reductase, GR; superoxide dismutase, SOD) (d) lipid peroxidation (malondialdhyde, MDA level), and (e) apoptotic process (caspase 3-like, CSP3) was selected for measurements of environmental effects on the populations of clams collected from the different sampling sites. The results of metal bioaccumulation in soft tissues of Ruditapes decussatus revealed a high pollution in the South Lagoon of Tunis with spatial variation and relatively high levels at the navigation channel. Anthropogenic pollutants in the lagoon led to the activation of antioxidant defense and biotransformation enzymes to oxidative damage of the membrane and activation of apoptosis, and revealed neurotoxicity. Among this core of biomarkers, the antioxidants enzymes (CAT, SOD, GR, and GPx) were very sensitive, allowing the discrimination among sites and pointing to the navigation channel as the most impacted site in the southern lagoon of Tunis. Moreover, a significant effect of season was recorded on biomarkers responses (e.g., CAT, GR, SOD, AChE, and CSP3 activities and MDA levels) with higher levels in winter than in summer, probably influenced by the reproductive stage and food availability. Finally, the measurement of the selected core of biomarkers in the whole soft tissues of clams was considered as an integrated indicator of environmental stress. Moreover, R. decussatus proved to be a remarkable sentinel species capable to establish a reliable diagnosis of the health status of the marine environment in different areas of the southern lagoon of Tunis, both in "hot" and "cold" seasons.

Biomonitoring of aromatic hydrocarbons in clam Meretrix meretrix from an emerging urbanization area, and implications for human health.

Pollution with total petroleum hydrocarbons (TPHs) is a global concern and particularly in coastal environments. Polycyclic aromatic hydrocarbons (PAHs) are regarded as the most toxic components of TPHs and they can also be derived from other sources. Fangcheng Port is considered as a representative emerging coastal city in China, but the status, sources, and hazards to organisms and humans with respect to contamination with PAHs and TPHs are unknown in the coastal regions of this area. Therefore, in this study, we cloned cytochrome P450 family genes (CYP1A1, CYP3A, and CYP4) and heat shock protein 70 gene (HSP70) in the clam Meretrix meretrix as well as optimizing the method for measuring the 7-ethoxyresorufin O-deethylase activity. These molecular indicators and four specific physiological indexes were found to be appropriate biomarkers for indicating the harmful effects of PAHs and TPHs on clams after exposure to the crude oil water-soluble fraction. In field monitoring surveys, we found that the 2- and 3-ring PAHs were dominant in the clams whereas the 4- to 6-ring PAHs were dominant in the sediments at each site. The PAH levels (3.63-12.77 ng/g wet weight) in wild clams were lower, whereas the TPH levels (13.25-70.50 µg/g wet weight) were higher compared with those determined previous in China and elsewhere. The concentrations of PAHs and TPHs in the sediments (19.20-4215.76 ng/g and 3.65-866.40 µg/g dry weight) were moderate compared with those in other global regions. Diagnostic ratio analysis demonstrated that the PAHs were derived mainly from pyrogenic sources. The TPHs may have come primarily from industrial effluents, land and maritime transportation, or fishing activities. The Integrated Biomarker Response version 2 indexes indicated that the clams collected from site S5 exhibited the most harmful effects due to contamination by PAHs and TPHs. Human health risk assessments demonstrated that the risks due to PAHs and TPHs following the consumption of clams can be considered acceptable. Our results suggest that continuous monitoring of contamination by PAHs and TPHs is recommended in this emerging coastal city as well as assessing their human health risks.

Metal-Associated Biomarker Responses in Crabs from a Marine Protected Area in Southeastern Brazil.

The environmental quality of a Ramsar wetland site located at the Cananéia-Iguape-Peruíbe Protected Area (CIP-PA), in São Paulo, Brazil, was assessed by geochemical analyses and biomarker assessments (GPx, GST, GSH, GST, MT, LPO, DNA damage) performed in swimming crab Callinectes danae Smith, 1869 organs (posterior and anterior gills and hepatopancreas) to estimate sediment contaminant bioavailability. The results indicated that two sampling stations, PT and PM, exhibited the worst environmental conditions, as sediments collected at both points contained metal contamination, while crabs exhibited significant responses for GPx, GST, and LPO (mostly during winter). Sediment contamination tended to be associated to fine sediments (both seasons) and organic matter (winter). During the summer survey, Pb concentrations in sediments of station PT exceeded the Brazilian Sediment Quality Guidelines (SQGs) and the Canadian Interim Marine Sediment Quality Guidelines. Metal concentrations in sediments sampled in winter were higher compared with summer, with Co, Ni, and Pb exceeding SQGs levels at PT, whereas Co, Ni, Hg, Zn, and Pb exceeded SQGs at PM. Biomarker induction during summer appeared to be caused by natural variables (water salinity and temperature, and molting cycle), whereas oxidative stress and tissue damage during winter appeared to be more clearly linked to metal contamination. Anterior gills presented the clearest signs of seasonal variability, being more responsive to sediment contamination. The results suggest that metals originated from the upper Ribeira de Iguape River are transported toward the estuarine system, causing effects on C. danae individuals. Additionally, seasonality is a strong factor concerning CIP-PA toxicity, since the rainfall regime significantly modifies the freshwater flow and, consequently, estuarine water salinity, suspended particle and metal inputs, as well as the location of depositional areas. Thus, efforts to mitigate CIP-PA contamination should be based on the control of upstream pollution sources.