Osmo-ionic regulation and carbonic anhydrase, Na+/K+-ATPase and V-H+-ATPase activities in gills of the ancient freshwater crustacean Aegla schmitti (Anomura) exposed to high salinities.

Aeglidae anomuran crabs originated in the sea, but invaded and diversified in southern South American freshwater (FW) streams. We here aimed at examining their tolerance of increased salinity, after a long time of evolution in FW (~33 million years). Aegla schmitti were exposed to FW and dilute seawater of salinities 15, 20, and 25‰ for 1, 5 and 10 days. Mortality in 35‰ was also assessed. Hemolymph osmolality, Na+, K+, Cl-, and Mg2+ ions, and hydration levels of the abdominal muscle were assayed. The activities of the Carbonic Anhydrase (CA), Na+/K+-ATPase (NKA) and V-H+-ATPase (VHA) were also assayed in the gills. A. schmitti preserves osmoregulatory mechanisms of its marine ancestors. It is able to survive in high salinities (25‰) for at least 10 days. Mortality in 35‰ was of 56% after 1 day, and of 100% after 7 days. In 25‰, NaCl is apparently hyporegulated at all times, while hemolymph osmolality rises after 5 days. CA and NKA activities remained unchanged in all experimental conditions, while VHA activity decreased after 10 days in 25‰. Hemolymph NaCl data was compatible with either hyporegulation and/or putative influx of NaCl into cells for regulatory volume increase (RVI). Further studies should deepen the understanding of the roles of low permeabilities and saturation of high affinity uptake systems in truly FW decapods, in their responses to high salinities. Moreover, the fate of extracellular NaCl as secretion in true hypo-regulation and/or influx into cells for RVI should also be investigated.

Integrated biomarker response index to assess toxic effects of environmentally relevant concentrations of paracetamol in a neotropical catfish (Rhamdia quelen).

The nonsteroidal anti-inflammatory drugs (NSAIDs) are amongst the most commonly detected classes of pharmaceuticals in freshwater environments, with paracetamol being the most abundant. The aim of this study was to evaluate the possible toxic effects of environmentally relevant concentrations (0.25, 2.5 and 25 µg.L-1) of paracetamol in Rhamdia quelen fish exposed for 14 days using different biomarkers. The total count of leukocytes and thrombocytes was reduced at the highest concentration. In the gills, all concentrations of paracetamol reduced the glutathione S-transferase (GST) activity and the reduced glutathione (GSH) levels compared to the control group. The activity of catalase (CAT) was not altered and glutathione peroxidase (GPx) activity increased at the highest concentrations. The superoxide dismutase (SOD) activity decreased at 25 µg.L-1 and the LPO levels increased at 2.5 µg.L-1 when compared to the control group. The concentration of ROS was not different among the groups. In the posterior kidney the activities of GST (2.5 µg.L-1), CAT (2.5 µg.L-1 and at 25 µg. L-1) and GPx and GSH levels increased at all concentrations when compared to the control group. The SOD activity and LPO levels did not change. Paracetamol caused genotoxicity in the blood and gills at concentrations of 2.5 µg.L-1 and in the posterior kidney at 2.5 and 25 µg.L-1. An osmoregulatory imbalance in plasma ions and a reduction in the carbonic anhydrase activity in the gills at 0.25 µg.L-1 were observed. Histopathological alterations occurred in the gills of fish exposed to 25 µg.L-1 and in the posterior kidney at 0.25 and 25 µg.L-1 of paracetamol. The integrated biomarker index showed that the stress caused by the concentration of 25 µg.L-1 was the highest one. These results demonstrated toxic effects of paracetamol on the gills and posterior kidneys of fish, compromising their physiological functions and evidencing the need for monitoring the residues of pharmaceuticals released into aquatic environment.

A systematic evaluation on the relationship between hypo-osmoregulation and hyper-osmoregulation in decapods of different habitats.

Decapods occupy all aquatic, and terrestrial and semi-terrestrial environments. According to their osmoregulatory capacity, they can be osmoconformers or osmoregulators (hypo or hyperegulators). The goal of this study is to gather data available in the literature for aquatic decapods and verify if the rare hyporegulatory capacity of decapods is associated with hyper-regulatory capacity. The metric used to quantify osmoregulation was the osmotic capacity (OC), the gradient between external and internal (hemolymph) osmolalities. We employ phylogenetic comparative methods using 83 species of decapods to test the correlation between hyper OC and hypo OC, beyond the ancestral state for osmolality habitat, which was used to reconstruct the colonization route. Our analysis showed a phylogenetic signal for habitat osmolality, hyper OC and hypo OC, suggesting that hyper-hyporegulators decapods occupy similar habitats and show similar hyper and hyporegulatory capacities. Our findings reveal that all hyper-hyporegulators decapods (mainly shrimps and crabs) originated in estuarine waters. Hyper OC and hypo OC are correlated in decapods, suggesting correlated evolution. The analysis showed that species which inhabit environments with intense salinity variation such as estuaries, supratidal and mangrove habitats, all undergo selective pressure to acquire efficient hyper-hyporegulatory mechanisms, aided by low permeabilities. Therefore, hyporegulation can be observed in any colonization route that passes through environments with extreme variations in salinity, such as estuaries or brackish water.

Multiple biomarkers response in a Neotropical fish exposed to paralytic shellfish toxins (PSTs).

The Alagados Reservoir (Southern Brazil) is used as water supply, and since 2002 there have been reports with a presence of cyanobacterial blooms and cyanotoxins. In order to assess the water quality and the ecological integrity of the reservoir, we evaluated biochemical, genotoxic and osmoregulatory biomarkers in the freshwater cichlid fish (Geophagus brasiliensis) that were exposed to PSTs. The fish were sampled in the Alagados Reservoir in February 2016 (Summer) and were divided in three groups: 1) Reservoir group (RES): fish were collected immediately after sampling; 2) Depuration group (DEP): fish were submitted to the depuration experiment for 90 days in the laboratory; and 3) Reproduction group (REP): fish were kept in the laboratory until the fertilization and the chemical analyses were performed on the offspring (F1 generation). In the RES and DEP the blood, brain, muscle, liver and gills were collected for biochemical, genotoxic and osmoregulatory biomarkers analysis. Our results showed that the fish from the Alagados Reservoir (RES) presented oxidative stress and DNA damage; and after 90 days (DEP), the antioxidant system and DNA damage were recovered. Although PSTs were considered a risk to the ecological integrity of this water body; PSTs concentrations were not found in the tissues of the F1 generation. In addition, the biomarkers used were useful tools to evaluate the effects of environment contamination. Therefore, it is necessary to develop new technologies and monitoring programs in order to reduce cyanobaterial blooms, cyanotoxins and human activities that cause the contamination in aquatic environments.

Depuration time and sublethal effects of microcystins in a freshwater fish from water supply reservoir.

Microcystins (MCs) are hepatotoxins that have been considered to be a worldwide problem due the effects that they can cause to environmental and human health systems. The Iraí Reservoir, located in the South of Brazil, is used as a water supply and MCs concentrations have been reported in this ecosystem. This study aimed to determine the MCs concentrations in the Iraí Reservoir and to evaluate the MCs depuration time and the health of Geophagus brasiliensis using biomarkers. Water and fish samples were collected in the Iraí Reservoir from August 2015 to May 2016. Phytoplankton and chemical analyses were conducted using water samples and the fish were divided into two groups; the Immediate Group (IMM) and the Depuration Group (DEP). In the IMM group, the blood, liver, muscle, brain and gills were collected, in order to evaluate the genotoxic, biochemical and chemical biomarkers. The DEP group was used in the depuration experiment for 90 days, and after this period the fish were submitted to the same procedure as the IMM group. Our results suggested that fish accumulated MCs and it may have caused oxidative stress, neurotoxicity and molecular damage. Furthermore, MCs concentrations increased during the depuration time and it resulted in molecular damage over the first 30 days. After 90 days, the recovery of the antioxidant system occurred. The depuration started on the 15th day, however, the toxins were still present in the samples. Therefore, the effects and the persistence of MCs are a risk to environmental systems and human health.