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In order to assess the effects of tannery effluents (TE) in organism health, juveniles of Prochilodus lineatus were submitted to in situ tests at four different river locations: site A – upstream of the tannery; site B – next to the tannery; and sites C and D – downstream of the tannery. After 96 h exposure in the river, samples of fish tissue, river water and sediment were collected in order to quantify chromium (Cr) concentrations. Tissue samples were used to assess the activity of ethoxyresorufin-O-deethylase (EROD) and glutathione S-transferase (GST), the content of glutathione (GSH) and metallothionein (MT) and the occurrence of lipid peroxidation (LPO) and DNA damage. Higher Cr concentrations were detected in the water and sediments from site B and in the liver of fish confined at site B, compared to the other sites. Fish caged at site B demonstrated higher levels of liver MT and hepatic EROD activity in relation to fish caged at the other sites. Moreover, fish from site B presented increased liver and branchial GST activities, as well as more GSH in the liver, than fish from site A. There were no significant variations in the occurrence of LPO and DNA damage among fish caged at the different sites. Thus, TE increased Cr levels in the water, sediments, and fish livers and stimulated the synthesis of MT and GSH and the activities of EROD and GST. In conclusion, TE affect the quality of the river and promote changes in biochemical biomarkers and Cr accumulation in P. lineatus.

Manganese (Mn) might stimulate the valve closure reflex in the freshwater bivalve Anodontites trapesialis, leading to metabolic suppression, whereas zinc (Zn) is not able to modify this behavior. To investigate this particular response, we exposed A. trapesialis specimens to Mn (0.5 mg L-1 ) and Zn (1.0 mg L-1 ) alone, and to their mixture, to measure further endpoints in different clam tissues: glycogen level in gills, and calcium (Ca2+ ), sodium (Na+ ), and chloride (Cl- ) concentrations in the hemolymph. Furthermore, we used cutting-edge technology, proteomics, to evaluate modifications in protein patterns under the 3 exposure tests. The main results highlighted that only Mn caused a clear drop in glycogen levels in gills, an increase in Ca2+ and Na+ , and a simultaneous decrease in Cl- concentration in the hemolymph. The proteomic analysis confirmed that Mn promoted more effects in A. trapesialis than the other tested conditions, because the number of proteins modulated was higher than the results obtained after exposure to Zn and the mixture. Moreover, 11 of the 12 modulated proteins were down-expressed. These results consolidate the hypothesis that Mn might suppress gill metabolic rate in A. trapesialis. Environ Toxicol Chem 2019;38:2480-2485. © 2019 SETAC.

As one of the most impactful industries, coal mining can promote several alterations at surrounding environment. In surface water, elevated concentrations of metals like Mn, Zn, Fe and Al are often observed. Thus, the aim of this study was to investigate the bioaccumulation and the sub-lethal effects of these metals on various organs of the Neotropical bivalve Anodontites trapesialis confined along a stream located near a coal mine, in order to assess a set of biomarkers that could be used for effectively monitoring coal mining areas. Clams were caged, for 96 h, at two sites located upstream (Up1 and Up2) and two sites downstream (Dw1 and Dw2) from the mine. Metals bioaccumulation was determined in gills, mantle, digestive gland, muscle and hemolymph and the following biomarkers were measured in A. trapesialis tissues: total antioxidant capacity against peroxyl radicals, metallothionein content, lipid peroxidation (LPO), proteins carbonylation, glutathione S-transferase activity, superoxide dismutase activity and acetylcholinesterase (AChE) activity. The results showed that Al and Fe bioaccumulation in the gills and hemolymph, Al bioaccumulation in the mantle and muscle, increased LPO in the gills (Dw1 and Dw2) and mantle (Dw1), as well as reduced AChE activity in the muscle (Dw1 and Dw2) should be considered effective biomarkers for monitoring coal mining areas. A. trapesialis proved to be an efficient biological model, considering that biomarkers responses were observed in the clams after only 96 h of confinement at Dw sites, accordingly this species could be a good candidate for monitoring Neotropical freshwaters.

Important concentrations of Zn, Mn and Fe were detected in a stream near a coal mining area and promoted, in field, biomarkers alterations in the bivalve Anodontites trapesialis. In order to understand the isolated and mixed effects of these metals on these Neotropical bivalves, we run short-term experiments under laboratory controlled conditions. After 96 h-exposure, tissues (gills, mantle, digestive gland, muscle, hemolymph) were removed for metal bioaccumulation analysis, oxidative stress biomarkers (reactive oxygen species (ROS), total antioxidant capacity, lipoperoxidation (LPO), proteins carbonylation (PC), metallothionein (MT), activity of superoxide dismutase and glutathione S-transferase and hemocytes DNA damage) and cholinesterase (ChE versus ASCh activity) activity evaluation. We run three independent tests. In Zn test, clams were exposed to three concentrations of Zn (0.18 mg L−1, 1.0 mg L−1, 5.0 mg L−1); in Mn test, clams were exposed to three concentrations of Mn (0.1 mg L−1, 0.5 mg L−1, 5.0 mg L−1) and in Mix test, clams were exposed to the mixture Zn (1 mg L−1) + Mn (0.5 mg L−1), with and without Fe (5.0 mg L−1). After single exposure to 5.0 mg L−1, Zn bioaccumulated in all tissues, but only in mantle and hemolymph after exposure to 1.0 mg L−1. The increased MT in gills of A. trapesialis exposed to Zn appears to be sufficient to avoid damage, since LPO occurred only in digestive glands from animals exposed to 5.0 mg L−1. We suggested that A. trapesialis had a metabolic suppression in consequence of Mn presence, based on the following results: the decrease of ROS in gills, the decrease of the Zn and Mn concentrations in tissues and the decrease of ChE versus ASCh activity in muscle. Despite this, animals exposed to Mn suffer oxidative damages (LPO and PC) in the mantle and digestive gland and MT increased in the mantle. These results showed A. trapesialis responded differently to each metal and Mn caused more damage. When exposed to Fe, gills level of ROS was increased, despite no changes in metal accumulation occurred. On the other hand, after exposure to the mixtures, tissues bioaccumulated Zn and previously observed damages caused by Mn and Fe disappeared. Consequently, biomarkers were less affected under mixture treatments, demonstrating mixtures effects or responses were not simply a combination of single exposures to Zn, Mn and Fe, but depend on metals toxicokinetics.

O Zinco (Zn) é um metal essencial para os organismos vivos em quantidades traço, mas pode ser tóxico quando presente em concentrações mais elevadas. Assim, o objetivo deste estudo foi avaliar a bioacumulação e a ocorrência de alterações hematológicas, metabólicas e osmorregulatórias no peixe neotropical Prochilodus lineatus, após exposição ao Zn. Para isto, juvenis foram expostos à água contendo 0,2 (Zn 0,2) e 2,0 (Zn 2,0) mg L-1 de Zn, ou apenas à água (CTR). Após 24, 96 e 168 h de exposição os peixes foram amostrados para a retirada de sangue, brânquias, fígado e rim posterior, nos quais foi feita a determinação do acúmulo de Zn. O sangue também foi utilizado para as análises de hematócrito, hemoglobina e número de eritrócitos (RBC). No plasma foram medidas as concentrações de Na+, K+, Cl-, Ca2+, Mg2+ e glicose. O fígado foi utilizado para quantificação do glicogênio. E nas brânquias e rim posterior foram determinadas as atividades das enzimas: anidrase carbônica (AC), Na+/K+-ATPase (NKA), H+-ATPase (HATP), Ca2+-ATPase (CaATP) e Mg2+-ATPase (MgATP). Após 24 e 96 h de exposição, sangue e brânquias aumentaram significativamente a concentração de Zn em relação aos respectivos CTR. O rim posterior e o fígado aumentaram significativamente a concentração de Zn no grupo Zn 2,0, em relação aos seus grupos CTR, após 96 e 168 h de exposição, respectivamente. O RBC dos peixes dos grupos Zn 0,2 e Zn 2,0 foi significativamente menor do que o grupo CTR após 96 h. O glicogênio hepático diminuiu significativamente no grupo Zn 0,2, em relação ao CTR, após 168 h. A atividade da HATP branquial e renal aumentou significativamente nos grupos Zn 0,2 e 2,0, após 96 h. A atividade da NKA branquial diminuiu significativamente nos grupos Zn 0,2 e 2,0 após 168 h, com consequente aumento na concentração do K+. A atividade da CaATP branquial foi significativamente maior nos peixes expostos ao Zn 0,2 por 24 h, com redução significativa do Ca2+ neste tempo. Após 168 h, a atividade branquial da CaATP do grupo Zn 2,0 e da MgATP dos grupos Zn 0,2 e 2,0 foram significativamente menores em relação aos grupos CTR, com redução significativa do Mg2+ plasmático nestes mesmos grupos. No rim posterior, a atividade da CaATP e da MgATP dos grupos Zn 0,2 e 2,0 diminuiu significativamente após 96 h. Assim, a exposição de P. lineatus ao Zn, em concentrações dentro dos limites permitidos pela legislação brasileira, promoveu acúmulo do metal ao longo das 168 h de exposição, de acordo com o seguinte padrão temporal: 24 e 96 h sangue e brânquias, 96 h rim posterior, 96 e 168 h fígado. Além de reduzir o número de eritrócitos, importante no transporte de gases, o Zn promoveu alterações na atividade branquial e renal de enzimas essenciais para manutenção do equilíbrio iônico e ácido-base, resultando em desajustes nas concentrações plasmáticas principalmente de K+, Ca2+ e Mg2+. Zinc (Zn) is an essential metal for living organisms in trace amounts but it can be toxic when present in higher concentrations. Thus, the aim of this study was to evaluate the bioaccumulation and the occurrence of hematological, metabolic and osmoregulatory changes on the Neotropical fish Prochilodus lineatus, after exposure to Zn. For that, juveniles were exposed to 0.2 (Zn 0.2) and 2.0 (Zn 2.0) mg L-1 Zn, or only to water. After 24, 96 and 168 h of exposure fish were sampled for the withdraw of blood, gills, liver and posterior kidney, where Zn accumulation was determined. In addition, the blood was used for the determination of hematocrit, hemoglobin, and erythrocyte count (RBC). Plasma concentrations of Na+, K+, Cl-, Ca2+ and Mg2+ and glucose were measured. The liver was used for quantification of hepatic glycogen. In the gills and posterior kidney the activities of carbonic anhydrase (AC), Na+/K+-ATPase (NKA), H+-ATPase (HATP), Ca2+-ATPase (CaATP), Mg2+-ATPase (MgATP) were determined. After 24 and 96 h exposure, blood and gills significantly increased the concentration of Zn in relation to the respective CTR. The posterior kidney and the liver showed a significant increase in Zn concentration in fish from Zn 2.0 group, relative to CTR groups, after 96 and 168 h of exposure, respectively. RBC of fish from Zn 0.2 and Zn 2.0 groups were significantly lower than the CTR group after 96 h. Hepatic glycogen decreased significantly in Zn 0.2 group, in relation to the CTR, after 168 h. Gill and renal HATP activity increased significantly in Zn 0.2 and 2.0 groups after 96 h. Gill NKA activity decreased significantly in 0.2 and 2.0 Zn groups after 168 h, with a consequent increase in K+ concentration. Gill CaATP activity was significantly higher in fish exposed to Zn 0.2 for 24 h, when a significant decrease on plasma Ca2+ was observed. Conversely, after 168 h, branchial CaATP in fish from Zn 2.0 group and MgATP from Zn 0.2 and 2.0 groups were significantly lower in relation to the CTR groups, with a significant reduction of plasma Mg2+ in these same groups. In the posterior kidney, CaATP and MgATP in fish from Zn 0.2 and 2.0 groups decreased significantly after 96 h. Thus, the exposure of P. lineatus to Zn, in concentrations within the limits allowed by the Brazilian legislation, promoted accumulation of the metal during the 168 h of exposure, according to the following temporal pattern: 24 and 96h blood and gills, 96 h posterior kidney, 96 and 168 h liver. In addition to reducing the erythrocyte count, important in the transport of gases, Zn promoted changes in the gill and renal activity of essential enzymes for the maintenance of the ionic and acid-base balance, resulting in imbalances in plasma concentrations mainly of K+, Ca2+ and Mg2+.