Stress Response, Immunity, and Organ Mass in Toads (Rhinella diptycha) Living in Metal-Contaminated Areas.

Mining is one of the main activities that drive the economy of Brazil. Mining activity is associated with risk of contamination of environment and local fauna by metals. Amphibians have a life cycle that requires a transition between aquatic and terrestrial environments, increasing their vulnerability to metal contamination in the water and substrate. Metals are ubiquitous, with high bioaccumulative and biomagnifying potential, and may lead to immune and endocrine disruption. In this study, we analyzed two different components of the innate immune response, bacterial killing ability (BKA) and phytohemagglutinin edema (PHA), and two stress biomarkers, corticosterone plasma levels (CORT) and the neutrophil to lymphocyte ratio (N:L), of toads (Rhinella diptycha) living in places contaminated by metals. Blood samples were collected pre- and post-restraint (1h), followed by an immune challenge with PHA and tissue collection (liver, spleen, and kidneys). Toads liver metal bioaccumulation did not correlate with the immune response or stress biomarkers. Post-restraint, animals had increased CORT and reduced BKA, independently of the collection site, and these variables were not correlated with liver metal bioaccumulation. Interestingly, toads with the larger spleen (immune organ) showed increased N:L post-restraint and greater edema after the PHA challenge. Our results indicate that toads living in metal-contaminated environments responded to acute stressor, activating the hypothalamic-pituitary-interrenal axis and the immune response. Keep tracking the physiological variables of these animals and the presence of metals in the environment and tissues should provide valuable health status indicators for the population, which is vital for proposing amphibian conservation strategies in these areas.

Predicting zebrafish spatial avoidance triggered by discharges of dairy wastewater: An experimental approach based on self-purification in a model river.

Wastewater discharges from dairy industries can cause a range of harmful effects in aquatic ecosystems, including a decline in biodiversity due to species evasion. Therefore, it is important to know the purification potential of rivers for the removal of pollutants released in dairy wastewater (DWW). The hypothesis adopted in this work was that the release of DWW into stretches of the Ribeirão dos Pombos River (São Paulo State, Brazil) might trigger an avoidance response, resulting in fish migrating to other regions, with the response being greater when the self-cleaning potential of the river is smaller. Therefore, the goals of the present study were to: (i) investigate how land use and seasonality of the rainfall regime influence the quality of the water in different areas of the river (P1: river source; P2: urban region; P3: rural region); (ii) assess the potential of the river to purify DWW; and (iii) evaluate the potential toxicity and repellency of DWW to the freshwater fish Danio rerio, using acute toxicity (mortality) and non-forced avoidance tests, respectively. P1 was shown to be the most preserved area. The chemical composition of the river varied seasonally, with higher concentrations of Cl- and SO42- at P3 during the rainy period. The river purification potential for DWW was higher at P2, due to greater microbiological activity (associated with higher BOD). The DWW was more acutely toxic in water from P2. The avoidance response was strongly determined by the concentration of DWW, especially for water from P2. The high capacity for self-cleaning at P2 did not seem sufficient to maintain the stability of the ecosystem. Finally, the non-forced exposure system proved to be a suitable approach that can assist in predicting how contaminants may affect the spatial distributions of organisms.

Taxonomic and non-taxonomic responses of benthic macroinvertebrates to metal toxicity in tropical reservoirs. The case of Cantareira Complex, São Paulo, Brazil.

Benthic macroinvertebrates are organisms that are recognized as water quality bio-indicators. A wide variety of indices and metrics have been shown to respond to a variety of anthropogenic impacts, usually under a general condition of environmental impairment. The absence of a clear distinction in the relations between specific pollutants and biotic variables is very common and can lead to biased interpretation of biomonitoring. The aims of this research were to test taxonomic and non-taxonomic responses to specific environmental conditions instead to general conditions. For this purpose, we estimated the theoretical toxicity by comparing toxicity values published by EPA with metal concentrations in water and sediments. Then we tested the responses of biological variables to toxicity and other environmental conditions using the linear mixed effects models approach. We generated 32 models considering 24 different biological metrics and indices that were grouped in five levels. Taxonomic and abundance metrics were best predictor than functional or tolerance-based indexes. The strongest model was that which considered subfamily taxonomic resolution responding to Al_w and Cr_s.

Factors that control the spatial and temporal distributions of phosphorus, nitrogen, and carbon in the sediments of a tropical reservoir.

The impacts of anthropic activities have had profound effects on the nitrogen (N) and phosphorus (P) cycles in many aquatic ecosystems. We investigated the spatial and temporal distributions of carbon (C), N, and P in the sediments of a tropical Paiva Castro Reservoir (São Paulo, Brazil), as well as their release and retention in the system. In 2010, surface sediments were collected at nine sites in the reservoir, and a core was obtained in the limnetic zone, in 2010. The core was dated using the 210Pb technique. The organic C content was estimated from organic matter concentration, which was measured by the loss-on-ignition method, and the concentrations of P and N were determined by spectrophotometry. Marked spatial heterogeneity in the Paiva Castro sediments associated with both natural variations in the water body and variations induced by human impacts was observed. Heterogeneity was evidenced by a decrease in the allochthonous contribution of organic matter (C/N) in the upstream-downstream direction and increases of N and P, mainly associated with water flows in the different compartments of the reservoir. In the core, C and N concentrations display significant positive correlations with increases in population and agricultural activities in the drainage basin through time. The C/P molar ratios in surface sediments are indicative of human impacts in the region, as C:P ratios in the sediment are low (7.8:1) compared to the Redfield ratio (C:P = 108:1). Predominance of oxic conditions at the sediment surface and particles sizes < 63 µm provided favorable conditions for P retention in the sediments, which helps prevent eutrophication. Approaches used in this research should be extended to other locations, especially in mesotrophic and oligotrophic reservoirs, to provide information on historical impacts in such aquatic ecosystems.

Influence of interspecific interactions on avoidance response to contamination.

An increasing number of studies have shown the ability of organisms to escape from toxic effects due to contamination, by moving spatially towards less contaminated habitats. However, this issue has been investigated in monospecific scenarios, without considering possible interactions between species during the contamination avoidance process. It is widely known that the spatial distribution of one species can be affected by another one, in different ways. Therefore, the main question addressed in the present study was as follows: Might interspecific interaction between the freshwater fish Danio rerio (zebrafish) and Poecilia reticulata (guppy) change their behavior patterns in terms of avoidance in the presence of a copper gradient? Zebrafish and guppies exposed to a copper gradient were tested for avoidance responses in a free-choice, non-forced, static, multi-compartmented exposure system, using two distinct approaches: (1) monospecific tests, in which only one species was exposed to the copper gradient, at two different population densities; and (2) multispecific tests, in which both species were tested simultaneously. In the control (with no copper) monospecific tests, both species were randomly distributed; however, in the control multispecific test, P. reticulata tended to aggregate. In the monospecific tests with a copper gradient, both species avoided copper in a similar way, with AC50 (concentration triggering avoidance in 50% of the exposed population) values between 15 and 18 µg·L-1, irrespective of the population density. However, in the multispecific tests, P. reticulata displaced D. rerio to previously avoided copper levels, consequently increasing the AC50 of D. rerio to 75 µg·L-1. This study shows the importance of understanding the interactions among species in contaminated areas, and the way that one species can prevent the avoidance behavior of another.

Bisphenol risk in fish exposed to a contamination gradient: Triggering of spatial avoidance.

Bisphenol A (BPA) is an emerging contaminant widely used in various industrial products. Sublethal toxicity of BPA on aquatic organisms is expected to occur at a concentration of around 500 µg L-1, which is much higher than environmentally realistic concentrations found in water bodies (up to 0.41 µg L-1). However, there is no information concerning how a BPA contamination gradient could affect the spatial displacement of organisms. We hypothesized that fish might be able to detect an environmentally realistic BPA contamination gradient and avoid potential toxic effects due to continuous exposure. Therefore, the objectives of this work were: (i) to determine if BPA could trigger an avoidance response in the freshwater fish Poecilia reticulata; (ii) to assess whether BPA-driven avoidance occurs at environmentally relevant concentrations; and (iii) to estimate the population immediate decline (PID) at the local scale, considering avoidance and mortality as endpoints. Avoidance experiments were performed in a seven-compartment non-forced exposure system, in which a BPA contamination gradient was simulated. The results indicated that BPA triggered avoidance in P. reticulata. In a traditional forced acute toxicity test, lethal effects in 50% of the population occurred at a BPA concentration of 1660 µg L-1, while in the non-forced system with a BPA concentration gradient, avoidance of 50% of the population occurred at a concentration four orders of magnitude lower (0.20 µg L-1). At environmentally relevant BPA concentrations, PID was mainly determined by the avoidance response. Avoidance in P. reticulata populations is expected to occur at BPA concentrations below those that cause sublethal effects on fish and are considered safe by international agencies (≤1 µg L-1). The approach used in the present study represents a valuable tool for use in environmental risk assessment strategies, providing a novel and ecologically relevant response that is complementary to traditional ecotoxicological tests.

Habitat fragmentation caused by contaminants: Atrazine as a chemical barrier isolating fish populations.

Information on how atrazine can affect the spatial distribution of organisms is non-existent. As this effect has been observed for some other contaminants, we hypothesized that atrazine-containing leachates/discharges could trigger spatial avoidance by the fish Poecilia reticulata and form a chemical barrier isolating upstream and downstream populations. Firstly, guppies were exposed to an atrazine gradient in a non-forced exposure system, in which organisms moved freely among the concentrations, to assess their ability to avoid atrazine. Secondly, a chemical barrier formed by atrazine, separating two clean habitats (extremities of the non-forced system), was simulated to assess whether the presence of the contaminant could prevent guppies from migrating to the other side of the system. Fish were able to avoid atrazine contamination at environmentally relevant concentrations (0.02 µg L-1), below those described to cause sub-lethal effects. The AC50 (atrazine concentration causing avoidance to 50% of the population) was 0.065 µg L-1. The chemical barrier formed by atrazine at 150 µg L-1 (concentration that should produce an avoidance around 82%) caused a reduction in the migratory potential of the fish by 47%; while the chemical barrier at 1058 µg L-1 (concentration that produces torpidity) caused a reduction in the migratory potential of the fish by 91%. Contamination by atrazine, besides driving the spatial distribution of fish populations, has potential to act as a chemical barrier by isolating fish populations. This study includes a novel approach to be integrated in environmental risk assessment schemes to assess high-tier contamination effects such as habitat fragmentation and population displacement and isolation.

Potential effects of triclosan on spatial displacement and local population decline of the fish Poecilia reticulata using a non-forced system.

Triclosan (TCS) is an emerging contaminant of concern in environmental studies due to its potential adverse effects on fish behavior. Since avoidance has been shown to be a relevant behavioral endpoint, our aims were: (i) to determine if TCS is able to trigger an avoidance response in Poecilia reticulata; (ii) to predict the population immediate decline (PID) caused by TCS exposure, by integrating lethality and avoidance responses; and (iii) to verify the overestimation of risk when mortality is assessed under forced exposure. Fish were exposed to TCS in a forced exposure system, to assess mortality, and to a TCS gradient in a non-forced exposure (NFE) system. Two NFE scenarios were simulated: (#1) a spatially permanent gradient, including low and high concentrations; and (#2) a scenario with high concentrations, simulating a local discharge. The fish avoided TCS concentrations as low as 0.2 µg L-1 (avoidance of 22%). The AC50 obtained from scenario #1 (8.04 µg L-1) was about 15 times more sensitive than that from scenario #2 (118.4 µg L-1). In general, up to the highest concentration tested (2000 µg L-1), the PID was determined by the avoidance. Mortality from the forced exposure was overestimated (48 h-LC50 of 1650 mg L-1), relative to the NFE. The reduced mortality in a non-forced environment does not imply a lower effect, because part of the population is expected to disappear by moving towards favorable environments. TCS is a potential environmental disturber, since at environmentally relevant concentrations (<2 µg L-1) it could cause a decline in the fish population.

Fatty Acid Composition of Tropical Fish Depends on Reservoir Trophic Status and Fish Feeding Habit.

Eutrophication results in a deficiency of n-3 LC-PUFA (long-chain polyunsaturated fatty acids) in aquatic food chains, affecting fish nutrition and physiology. The trophic transfer of FA (fatty acids) to fish species of different feeding habits was investigated in two reservoirs in southeast Brazil-the mesotrophic Ponte Nova Reservoir (PN) and the hypereutrophic Billings Reservoir (Bil). Total FA profile of stomach contents and adipose tissue, triacylglycerols (TAG), and phospholipids (PL) from liver and muscle of the omnivorous Astyanax fasciatus and the carnivorous Hoplias malabaricus were analyzed by gas chromatography. A prevalence of n-6PUFA, as 18:2n-6 (linoleic acid) and 20:4n-6 (arachidonic acid, ARA) was observed in the stomach contents and in the tissues of A. fasciatus from the PN reservoir. In contrast, n-3 LC-PUFA, as 20:5n-3 (eicosapentaenoic acid, EPA) was accumulated in fish tissues from Bil, resulting in higher n3/n6 and EPA/ARA ratios, compared to fish from PN. This differential FA accumulation was also observed for H. malabaricus, but differences were slightly minor, and no changes were observed in the EPA/ARA ratios between fish from both reservoirs. Regardless reservoir, FA profiles of TAG resembled that of their diet, whereas FA profiles of PL were more conservative and mainly comprised by LC-PUFA. We conclude that reservoir trophic status affected the FA composition of food resources available to these fish species, resulting in differential allocation of n-3 and n-6 FA. As expected, FA profile of the investigated fish species also reflected their feeding habit and physiological demands.