Kavezno izlaganje lubina (Dicentrarchus labrax) u procjeni genotoksičnog utjecaja onečišćenja

Genotoxic effects are often the earliest signs of pollution-related environmental disturbance. In this study, we used the comet assay and micronucleus test to assess DNA damage in the erythrocytes of the European sea bass (Dicentrarchus labrax) exposed to environmental pollution in situ. Fish were collected from a fi sh farm in the Trogir Bay and their cages placed at an unpolluted reference site Šolta (Nečujam Bay) and a polluted site Vranjic (Kaštela Bay) for four weeks. A group of fi sh which remained at the fi sh farm Trogir Bay were used as the second control group. Fish exposed at the Vranjic site showed a signifi cantly higher erythrocyte DNA damage, measured by the comet assay, than either control group. Micronucleus induction showed a similar gradient of DNA damage, but did not reach statistical signifi cance. Our results show that cage exposure of a marine fi sh D. labrax can be useful in environmental biomonitoring and confi rm the comet assay as a suitable tool for detecting pollution-related genotoxicity.Genotoksični učinak često je jedan od najranijih pokazatelja štetnog djelovanja onečišćenja okoliša. U ovom radu procijenjeno je oštećenje DNA u eritrocitima lubina (Dicentrarchus labrax) izloženima okolišnom onečišćenju s pomoću komet-testa i mikronukleus-testa. Lubini su prikupljeni na ribogojilištu i kavezno izloženi u periodu od četiri tjedna na dvije postaje različitog stupnja onečišćenja na jadranskoj obali: na kontrolnoj postaji Šolta (zaljev Nečujam) i na onečišćenoj postaji Vranjic (Kaštelanski zaljev). Zasebna skupina lubina skupljena na ribogojilištu poslužila je kao druga kontrola. Rezultati komet-testa pokazali su statistički značajan porast oštećenja DNA na postaji Vranjic u usporedbi s obje kontrolne postaje. Rezultati mikronukleus-testa pokazali su sličan gradijent onečišćenja, iako nisu dosegli statističku značajnost. Ovi rezultati upućuju na primjenjivost kaveznog izlaganja lubina D. labrax u biomonitoringu vodenog okoliša te potvrđuju korisnost komet-testa kao prikladne metode za detekciju genotoksičnog utjecaja onečišćenja

Role of non-enzymatic antioxidants on the bivalves' adaptation to environmental mercury: Organ-specificities and age effect in Scrobicularia plana inhabiting a contaminated lagoon

This study aimed to investigate the role of non-enzymatic antioxidants on adaptive skills over time in the bivalve Scrobicularia plana environmentally exposed to mercury. Inter-age (2+, 3+, 4+, 5+ year old) and organ-specific (gills, digestive gland) approaches were applied in bivalves collected from moderately and highly contaminated sites at Ria de Aveiro (Portugal). S. plana's adaptive skills were dependent on the contamination extent; under moderate contamination scenario, the intervention of the different antioxidants took place harmoniously, evidencing an adjustment capacity increasing with the age. Under higher contamination degree, S. plana failed to cope with mercury threat, showing an age-dependent deterioration of the defense abilities. In organ-specific approach, the differences were particularly evident for thiol-compounds, since only gills displayed the potential to respond to moderate levels by increasing non-protein thiols and total glutathione. Under high contamination degree, both organs were unable to increase thiol-compounds, which were compensated by the ascorbic acid elevation

Immunosuppression in the infaunal bivalve Scrobicularia plana environmentally exposed to mercury and association with its accumulation

This study aimed to test the hypothesis whether mercury (Hg) activates or suppresses inappropriately the immunity of the bivalve Scrobicularia plana inhabiting a Hg contaminated area (Laranjo basin, Ria de Aveiro, Portugal). Immunity endpoints, as well as lipid peroxidation (LPO) as a sign of damage, were evaluated in parallel with total Hg burden. Bivalves from both moderately (MO) and highly (HI) contaminated sites displayed higher haemolymph Hg load and reduced plasma agglutination. Increased haemocytes density and decreased phagocytosis were observed at HI, whereas increased oxidative burst activity (OBA) was observed at MO, pointing out that the immunotoxicity is a result of Hg direct contact involving no ROS intervention. OBA observed at MO was concomitantly associated to peroxidative damage as depicted by LPO increase in haemocytes and haemolymph plasma. Thus, S. plana can be suggested as a suitable bioindicator of metal pollution in coastal areas on the basis of Hg bioaccumulation and immunotoxicity responses

Mercury contaminated systems under recovery can represent an increased risk to seafood human consumers: A paradox depicted in bivalves' body burdens

The present study, carried out in a mercury (Hg) contaminated estuary, aimed to investigate: (i) the long-term evolution of the Hg bioavailability in the environmental matrices, in a period of 5 years (2003– 2008), without new anthropogenic inputs; (ii) the temporal evolution of Hg load (organic and inorganic forms) in the native bivalve Scrobicularia plana , inferring the progression of human health risk associated to its consumption and the dependence on the animals’ size. The area selected was Laranjo basin of Ria de Aveiro (Portugal), where a Hg gradient was identified because of past discharges from a chlor-alkali plant. Two sites termed M (moderately contaminated) and H (highly contaminated) were compared with an uncontaminated reference (R) site. Results displayed a persistence of Hg in the environment, though the levels in sediment decreased at site H, confirming the ecosystem recovery. The risk associated to clam consumption remained stable in 2008 considering their total Hg (T-Hg) load and the limits established by public health authorities, though T-Hg levels significantly decreased at H site for size classes C3 (2 year) and C4 (4 year). Organic Hg (O-Hg) accumulation increased from 2003 to 2008, reaching threatening lev- els and suggesting an increased bioavailability of this Hg form. This evolution towards an increase of O- Hg accumulation was particularly prominent under a moderate contamination scenario (site M). Overall, it was demonstrated that a period of 5 years of ecosystem recovery was not enough to eliminate the risk to human consumers, highlighting Hg contamination in estuaries as a long-lasting legacy. Paradoxically, it was pointed out that in a given step of the long-term restoration process, occurring naturally in aquatic systems, the risk associated to bivalves’ consumption can appear augmented due to O-Hg accumulation increments

Lipid peroxidation vs. antioxidant modulation in the bivalve Scrobicularia plana in response to environmental mercury-Organ specificities and age effect

This study aimed at the assessment of mercury burden and its association to damage vs. antioxidant protection in the bivalve Scrobicularia plana environmentally exposed to mercury. Inter-age and organ-specific approaches were applied by using different annual size classes (2+, 3+, 4+ and 5+ year old) and assessing specific organs (gills, digestive gland), respectively. Bivalves were collected from moderately and highly contaminated sites at Laranjo basin – Ria de Aveiro (Portugal), where a mercury gradient was identified, and compared with those from a reference site. Besides total and organic mercury accumulation, endpoints combining lipid peroxidation (LPO), as a damage sign, and antioxidant protection (catalase, CAT; glutathione peroxidase, GPX; glutathione S-transferase, GST, activities) were determined. The whole-body accumulation as total mercury changed according to the environmental gradient for all age classes, while in terms of organic mercury, only 5+-year-old animals showed increased accumulation with increased environmental mercury level. Mercury induced peroxidative damage, showing that antioxidative mechanisms were insufficient. The adaptive capacity to pro-oxidant challenge, expressed as antioxidant induction and lesser vulnerability to enzyme inhibition, increased with age. The specific analyses of gills and digestive gland revealed that both organs were able to mirror external levels of exposure in the accumulation of total and organic mercury. Nevertheless, gills displayed higher potential to accumulate organic forms. The organ specificity was evident for antioxidant response with a clear pattern of overall increase in gills and decrease in digestive gland. In addition, only gills displayed statistical correlations between oxidative stress responses and mercury accumulation. Besides the contribution to understand mercury toxicodynamics, specific organs approach is strongly recommended in order to avoid misinterpretations. The use of whole-body analyses can be particularly compromising when oxidative stress responses (rather than mercury accumulation) are addressed

Phagocytic cell responses to silica-coated dithiocarbamate-functionalized iron oxide nanoparticles and mercury co-exposures in Anguilla anguilla L.

Immune system responses in fish are considered as suitable and sensitive biomarkers for monitoring aquatic pollution. However, a clear knowledge gap persists in the literture on the immunotoxic potential of engineered nanoparticles toward aquatic organisms such as fish. Employing major enzymatic- (glutathione reductase, GR; glutathione peroxidase, GPX; glutathione sulfo-transferase, GST; catalase, CAT) and thiol- (non-protein thiols, NP-SH; total glutathione, TGSH)-based defense biomarkers, this study assessed the response of phagocytes isolated from peritoneum (P-phagocytes), gill (G-phagocytes), head kidney (HK-phagocytes), and spleen (S-phagocytes) of European eel (Anguilla anguilla L.) to silica-coated magnetite particles (Fe3O4@SiO2/SiDTC, hereafter called IONP; size range: 82 +/- 21 to 100 +/- 30 nm; 2.5 mg L-1) alone and IONP and mercury (Hg; 50 mu g L-1) concomitant exposures. Responses of previous biomarkers were studied in P-phagocytes, G-phagocytes, HK-phagocytes, and S-phagocytes collected during 0, 2, 4, 8, 16, 24, 48, and 72 h of exposures. Contingent to hour of exposure to IONP, Hg, and IONP + Hg GST, GPX, CAT, NP-SH, and TGSH exhibited their differential responses in all the phagocytic cells considered. In particular, under IONP exposure, the potential occurrence of the GSH-independent antioxidant defense was indicated by the observed herein inhibition in the enzymatic- and thiol-based defense in A. anguilla phagocytes. In contrast, the response of P-, G-, HK-, and S-phagocytes to the increasing Hg exposure period reflected an increased detoxification activity. Notably, the occurrence of an antagonism between IONP and Hg was depicted during late hours (72 h) under IONP + Hg concomitant exposure, where elevations in the defense biomarkers were depicted. Overall, the P-, G-, HK-, and S-phagocytic cells exhibited a differential induction in the studied enzymes and thiols to counteract impacts of IONP, Hg, and IONP + Hg concomitant exposures. Future studies on the fish immunotoxicity responses to IONP exposure in multi-pollution conditions can be benefited with the major outcomes of the present study

Rescheduling the process of nanoparticle removal used for water mercury remediation can increase the risk to aquatic organism: evidence of innate immune functions modulation in European eel (Anguilla anguilla L.)

This study aimed to assess the mechanisms of innate immune function responses to silica-coated iron oxide nanoparticle functionalized with dithiocarbamate groups (IONP) exposure alone and its associated mercury (Hg) in European eel (Anguilla anguilla L.) phagocytes isolated from peritoneum (P-phagocytes), gill (G-phagocytes), head kidney (HK-phagocytes) and spleen (S-phagocytes). The study evaluated viability, phagocytosis, oxidative burst activity (OBA) and lipid peroxidation (LPO). Four groups were made: (1) 2 x 10(6) phagocytes + RPMI-1640 (control), (2) 2 x 10(6) phagocytes + IONP (2.5 mg L-1), (3) 2 x 10(6) phagocytes + Hg (50 mu g L-1) and (4) 2 x 10(6) phagocytes + IONP + Hg. Samplings were performed at 0, 2, 4, 8, 16, 24, 48 and 72 h of exposure. A. anguilla P-, G-, HK- and S-phagocytes in vitro exposure to IONP alone revealed either increased (except HK-phagocytes at 16 h) or no change in viability, suggesting that the cells are metabolically active and resistant to IONP exposure alone. In terms of phagocytes overactivation and reactive oxygen species (ROS) production as an indirect mechanism of immunotoxicity, the phagocytes responded in the following manner: P->S->HK-=G-phagocytes for IONP exposure alone, S->HK->P-=G-phagocytes for Hg exposure alone and HK->G-=S->P-phagocytes for concomitant exposure. Overall, considering Hg as a surrogate for metals and its association with IONP, as well as the likelihood that it could pose a serious threat to aquatic organisms by modulating their immune defense mechanisms if accidentally discharged into the aquatic environment, current results suggest that the step of IONP-metal complex removal must not be underrated and should be processed without any more ado