Water temperature modulates mercury accumulation and oxidative stress status of common goby (Pomatoschistus microps).

Mercury (Hg) is a widespread pollutant across estuarine and coastal areas, raising concern on its potential impact on aquatic organisms. Hg may origin from natural and anthropogenic sources, being persistent and potentially toxic to biota, ultimately representing a serious risk to human health. Hg accumulation and toxicity may also induce reactive oxygen species (ROS) production in marine organisms, responsible for cell and tissue damage. Additionally, the temperature is undoubtedly an important environmental factor to consider regarding accumulation, due to its marked influence on the physiology and ecology of aquatic organisms. This study aimed to investigate the effect of different temperature scenarios (15, 20 and 25 °C) on the Hg accumulation in Pomatoschistus microps (Krøyer, 1838) liver and muscle, as well as on oxidative stress responses and energy metabolism, after short-term exposure to a naturally contaminated sediment with an environmentally relevant [Hg] (1.2 µg g-1). The results showed that Hg accumulation tends to increase along the temperature gradient with higher values of Hg accumulated in liver than in muscle tissue. The action of antioxidant enzymes and stress proteins seems to be effective in combating oxidative stress in the liver. Despite the action of antioxidant defences in the muscle, oxidative damage was observed at the protein level concomitantly with a decrease in aerobic energy production after exposure to Hg at higher temperatures. These findings are ecologically relevant and highlight the importance of further investigation of combined effects of Hg and other stressors, especially in a scenario of a changing climate where events leading to rapid alterations on water parameters are more frequent.

Essential oils from Ocimum basilicum cultivars: analysis of their composition and determination of the effect of the major compounds on Haemonchus contortus eggs.

The continuous use of synthetic anthelmintics against gastrointestinal nematodes (GINs) has resulted in the increased resistance, which is why alternative methods are being sought, such as the use of natural products. Plant essential oils (EOs) have been considered as potential products for the control of GINs. However, the chemical composition and, consequently, the biological activity of EOs vary in different plant cultivars. The aim of this study was to evaluate the anthelmintic activity of EOs from cultivars of Ocimum basilicum L. and that of their major constituents against Haemonchus contortus. The EOs from 16 cultivars as well the pure compound linalool, methyl chavicol, citral and eugenol were used in the assessment of the inhibition of H. contortus egg hatch. In addition, the composition of three cultivars was simulated using a combination of the two major compounds from each. The EOs from different cultivars showed mean Inhibition Concentration (IC50) varying from 0.56 to 2.22 mg/mL. The cultivar with the highest egg-hatch inhibition, Napoletano, is constituted mainly of linalool and methyl chavicol. Among the individual compounds tested, citral was the most effective (IC50 0.30 mg/mL). The best combination of compounds was obtained with 11% eugenol plus 64% linalool (IC50 0.44 mg/mL), simulating the Italian Large Leaf (Richters) cultivar. We conclude that different cultivars of O. basilicum show different anthelmintic potential, with cultivars containing linalool and methyl chavicol being the most promising; and that citral or methyl chavicol isolated should also be considered for the development of new anthelmintic formulations.

Dietary intake from complementary feeding is associated with intestinal barrier function and environmental enteropathy in Brazilian children from the MAL-ED cohort study.

A child's diet contains nutrients and other substances that influence intestinal health. The present study aimed to evaluate the relations between complementary feeding, intestinal barrier function and environmental enteropathy (EE) in infants. Data from 233 children were obtained from the Brazilian site of the Etiology, Risk Factors, and Interactions of Enteric Infections and Malnutrition and the Consequences for Child Health and Development Project cohort study. Habitual dietary intake from complementary feeding was estimated using seven 24-h dietary recalls, from 9 to 15 months of age. Intestinal barrier function was assessed using the lactulose-mannitol test (L-M), and EE was determined as a composite measure using faecal biomarkers concentrations - α-1-antitrypsin, myeloperoxidase (MPO) and neopterin (NEO) at 15 months of age. The nutrient adequacies explored the associations between dietary intake and the intestinal biomarkers. Children showed adequate nutrient intakes (with the exception of fibre), impaired intestinal barrier function and intestinal inflammation. There was a negative correlation between energy adequacy and L-M (ρ = -0·19, P < 0·05) and between folate adequacy and NEO concentrations (ρ = -0·21, P < 0·01). In addition, there was a positive correlation between thiamine adequacy and MPO concentration (ρ = 0·22, P < 0·01) and between Ca adequacy and NEO concentration (ρ = 0·23; P < 0·01). Multiple linear regression models showed that energy intakes were inversely associated with intestinal barrier function (ß = -0·19, P = 0·02), and fibre intake was inversely associated with the EE scores (ß = -0·20, P = 0·04). Findings suggest that dietary intake from complementary feeding is associated with decreased intestinal barrier function and EE in children.

Mercury bioaccumulation in the long-fin squid Loligo forbesi near the Mid-Atlantic Ridge: Implications to human exposure.

Due to growing commercial interest as a fishing resource and its intermediate position in the marine trophic chains as both prey and predator, cephalopods can play an important role in mercury (Hg) transfer along the marine food webs, since they can bioaccumulate this metal in its tissues. Our study aims to analyze Hg accumulation in Loligo forbesi caught in the Azores Archipelago (Portugal) quantifying Hg in different tissues (mantle and stomach), as well in the squid stomach content, in order to evaluate the efficiency of Hg transfer from prey to predator. Hg data from the tissues was used to estimate the weekly tolerable Hg intake due to squid consumption. Overall data indicate that Hg measured in the stomach tissue (0.1 ± 0.01 µg g-1) was significantly higher than Hg levels found in the mantle (0.04 ± 0.001 µg g-1) and stomach contents (0.01 ± 0.001 µg g-1). BMF (bioaccumulation factor) was >1 for all the samples, indicating a biomagnification process from prey to predator. Hg concentration in the mantle tissue was correlated with mantle size; although females present higher Hg levels than males, the difference was found to be not related to gender but rather to the fact that females had larger bodies. Finally, considering the Hg concentration found in the mantle and the permitted Hg levels, it is advisable to consume up to 1050-1890g of squid per week, according to the regulatory agencies. Thus, our results indicate that, since these doses are respected, consumption of squids from the Azorean waters do not pose a risk to humans.

Effects of diclofenac and salicylic acid exposure on Lemna minor: Is time a factor?

The global occurrence of pharmaceuticals in the aquatic environment has been considered a particularly concerning problem with unknown consequences. Non-steroidal anti-inflammatory drugs (NSAIDs) including diclofenac (DCF) and salicylic acid (SA), are among the most frequently prescribed drugs in the world, being consequently commonly found in the aquatic environment. Prolonged experiments (with duration of exposure that surpass those recommended by already established testing guidelines) are important to obtain ecologically relevant data to address the issue of NSAIDs ecotoxicity, because by being more realistically (namely in terms of levels and durations of exposure), such tests may indicate realistic challenges posed to aquatic organisms. Among the most common test species that are used for assessing environmental quality, plants play a leading role. Lemna species are among the most important plants used for ecotoxicity testing. Therefore, the aim of this study was to evaluate the temporal effect of a prolonged exposure of DCF and SA on Lemna minor. To attain this purpose, L. minor plants were chronically exposed to 0, 4, 20, and 100 µg/L of both pharmaceuticals, and samplings were performed at 6, 10 and 14 days of exposure. The analyzed endpoints were: levels of chlorophyll a, b and total, carotenoids; and enzymatic biomarkers, such as catalase, ascorbate peroxidase and glutathione-S-transferases. Diclofenac was responsible for alterations in all analyzed parameters in different intervals of exposure. Salicylic acid exposure was not capable of causing alterations on pigment contents of L. minor, however, enzymatic biomarkers were altered at all sampling intervals. Thus, it is possible to conclude that both pharmaceuticals can cause damage on the tested macrophyte species, biochemical parameters being more sensitive than physiological ones. Additional prolonged experiments are required to understand the chronic effects of different pharmaceuticals in the aquatic environment, especially in plants.

Multi-generational exposure to Pb in two monophyletic Daphnia species: Individual, functional and population related endpoints.

To better evaluate chemical damage in chronically contaminated habitats, a nine-generational exposure to Lead (Pb) was done with two monophyletic Daphnia species, from temperate (Daphnia magna) and tropical (Daphnia similis) environments. The multi-generational test consisted generally of a continuous Pb exposed set of organisms, plus an extra control set running simultaneously. To assess daphnids recovery after Pb exposure, some organisms from the sixth generation were transferred to clean media for three extra generations (recovery period; F6 to F9), while others were keep Pb exposed. All setups (control, Pb exposure and recovery period) were submitted to two different dietary regimes, the standard (3 × 105 cells/mL) and restricted food (1.5 × 105 cells/mL) regimes. To evaluate the effects of generational Pb exposure and food regimes, individual, functional and population related endpoints were assessed (number of offspring, body length and rate of population increase (r) and feeding rate (FR)). The tests were conducted on the first (F0) and last generations (F9). No differences were shown on number of offspring and feeding among F9 control and continuous Pb exposed D. magna, although a higher r was shown for F9 Pb exposed organisms. F9 Pb exposed D. similis also presented a higher r than F9 control, however, lethality was induced at high Pb exposure levels. At food restriction the patterns were opposite and D. magna died at high Pb exposure while Pb exposed D. similis was the only setup (compared to control and recovery period) to survive at high Pb exposure levels. Regarding the recovery period, D. magna (standard food) did not cope well with the Pb re-exposure and lethality was induced, while D. similis indicate a decreased Pb sensitivity (only setup that survived high Pb exposure levels). Under food restriction, both species presented a decreased Pb sensitivity and consequent failed recovery (possibly due to epigenetic changes). Both species presented similar patterns regarding generations. Organisms from F0 presented enhanced reproductive outputs in comparison to F9 and the contrary occurred to the FR (even in control organisms). Data show an acclimation under a generational Pb exposure, which could increase the population of adapted organisms in natural habitats. And, since there was not a full recovery after three generations in clean media, an indication of epigenetic changes for both species may also be considered.

Toxicity of lead and mancozeb differs in two monophyletic Daphnia species.

Lead and mancozeb are two important chemicals used for different human purposes and activities worldwide. Hazard assessment in different areas of the world is carried out with different but phylogenetically similar species, adapted to different climatic conditions, in order to increase relevance. This study evaluated the sensitivity of two monophyletic species, the tropical species Daphnia similis and the temperate species Daphnia magna, to the two chemicals lead and mancozeb. Standard acute and chronic ecotoxicological tests (reproduction and growth), as well as other sublethal measurements such as the intrinsic rate of population increase (r), feeding rate (FR) and O2 consumption, were recorded along with the analysis of the AChE activity to determine the neurotoxicity of both contaminants. Albeit their similar evolutionary status, D. magna generally presented a lower sensitivity to Pb in comparison to D. similis. Despite the differences in sensitivity, both species presented similar patterns of response under Pb exposure, with diminished reproductive outputs, feeding impairment, reduced O2 consumption and no effect on AChE activity. Mancozeb decreased the reproduction, rate of population increase and feeding rate, increased the AChE activity in both species and increased O2 consumption only in D. magna. While D. magna increased O2 consumption under mancozeb exposure, no effects were observed for D. similis. Thus, species may present different responses and sensitivities to different pollutants, regardless of their phylogeny. Therefore, the use of ecotoxicological assays with native species is crucial for a better ecological risk assessment in contaminated areas.

Influence of biochar particle size on biota responses.

Despite the increasing interest for biochar as a soil amendment, a knowledge gap remains on its impacts on non-target soil and aquatic species. We hypothesised that biochar particle size and application rate can play a role in the toxicity to biota. Pine woodchip biochar was incorporated in a clean soil at three particle size classes: small (<0.5 mm), medium (1-2 mm), and large (<4 mm), and at two concentrations: 1% and 6% w/w. A laboratory screening with earthworm Eisenia andrei avoidance behaviour bioassay was carried out to test the most adequate application rates, particle sizes and soil-biochar pre-incubation period. Thereafter, a 28-day greenhouse microcosm experiment was conducted as an ecologically more representative approach. Survival, vertical distribution and weight changes of E. andrei, and bait-lamina consumption were recorded. Soil leachates from the microcosms were collected to evaluate their impact on Daphnia magna immobilisation and Vibrio fischeri (Microtox®) bioluminescence. A feeding experiment with E. andrei was also performed to address earthworm weight changes and to conduct a screening of PAH-type metabolites in their tissue. The 6% <0.5 mm treatment pre-incubated for 96 h induced significant avoidance of the earthworms. Significantly lower bait-lamina consumption was observed in microcosms for the 6% <0.5 mm treatment. Moreover, particle size was a statistically significant factor regarding the loss of weight in the feeding experiment and higher concentration of naphthalene-type metabolites detected in E. andrei tissue, when exposed to <0.5 mm biochar particles. The leachates had no adverse effects on the aquatic species. The results suggest that particles <0.5 mm of pine woodchip biochar can pose sub-lethal effects on soil biota.

Infant feeding practices and determinant variables for early complementary feeding in the first 8 months of life: results from the Brazilian MAL-ED cohort site.

OBJECTIVE: The present study aimed to describe breast-feeding, complementary feeding and determining factors for early complementary feeding from birth to 8 months of age in a typical Brazilian low-income urban community. DESIGN: A birth cohort was conducted (n 233), with data collection twice weekly, allowing close observation of breast-feeding, complementary feeding introduction and description of the WHO core indicators on infant and young child feeding. Infant feeding practices were related to socio-economic status (SES), assessed by Water/sanitation, wealth measured by a set of eight Assets, Maternal education and monthly household Income (WAMI index). Two logistic regression models were constructed to evaluate risk factors associated with early complementary feeding. RESULTS: Based on twice weekly follow-up, 65 % of the children received exclusive breast-feeding in the first month of life and 5 % in the sixth month. Complementary feeding was offered in the first month: 29 % of the children received water, 15 % infant formulas, 13 % other milks and 9·4 % grain-derived foods. At 6 months, dietary diversity and minimum acceptable diet were both 47 % and these increased to 69 % at 8 months. No breast-feeding within the first hour of birth was a risk factor for the early introduction of water (adjusted OR=4·68; 95 % CI 1·33, 16·47) and low WAMI index a risk factor for the early introduction of other milks (adjusted OR=0·00; 95 % CI 0·00, 0·02). CONCLUSIONS: Data suggest local policies should promote: (i) early breast-feeding initiation; (ii) SES, considering maternal education, income and household conditions; (iii) timely introduction of complementary feeding; and (iv) dietary diversity.

Participation of TLR2 and TLR4 in Cytokines Production by Patients with Symptomatic and Asymptomatic Chronic Chagas Disease.

Chagas disease (CD), caused by the protozoan Trypanosoma cruzi, is a serious public health issue. Its evolution involves an acute stage, characterized by no specific symptoms, and the chronic stage during most individuals are asymptomatic, but about 30-40% of them become symptomatic presenting the cardiac or digestive disease. Host immune response mechanisms involved in symptomatic or asymptomatic chronic disease are not fully understood. The pro-inflammatory cytokines are crucial in host resistance. However, a fine control of this inflammatory process, by action of anti-inflammatory cytokines, is necessary to avoid tissue injury. This control was found to be responsible for no clinical manifestations in asymptomatic individuals. Toll-like receptors (TLRs) are extremely important in defining the cytokine profile released in response to a micro-organism. We found that patients with the cardiac form predominantly released the pro-inflammatory cytokines: IFN-γ, TNF-α and IL-17 with the involvement of both, TLR2 and TLR4. In contrast, patients with asymptomatic disease release predominantly the anti-inflammatory cytokines IL-10 and TGF-ß, but also with TLR2 and TLR4 participation. The mechanisms by which stimulation of the same TLRs results in release of different pattern of cytokines, depending on the patients group that is being evaluated, are discussed.

Fungal Planet description sheets: 558-624.

Novel species of fungi described in this study include those from various countries as follows: Australia: Banksiophoma australiensis (incl. Banksiophoma gen. nov.) on Banksia coccinea, Davidiellomycesaustraliensis (incl. Davidiellomyces gen. nov.) on Cyperaceae, Didymocyrtis banksiae on Banksia sessilis var. cygnorum, Disculoides calophyllae on Corymbia calophylla, Harknessia banksiae on Banksia sessilis, Harknessia banksiae-repens on Banksia repens, Harknessia banksiigena on Banksia sessilis var. cygnorum, Harknessia communis on Podocarpus sp., Harknessia platyphyllae on Eucalyptus platyphylla, Myrtacremonium eucalypti (incl. Myrtacremonium gen. nov.) on Eucalyptus globulus, Myrtapenidiella balenae on Eucalyptus sp., Myrtapenidiella eucalyptigena on Eucalyptus sp., Myrtapenidiella pleurocarpae on Eucalyptuspleurocarpa, Paraconiothyrium hakeae on Hakea sp., Paraphaeosphaeria xanthorrhoeae on Xanthorrhoea sp., Parateratosphaeria stirlingiae on Stirlingia sp., Perthomyces podocarpi (incl. Perthomyces gen. nov.) on Podocarpus sp., Readeriella ellipsoidea on Eucalyptus sp., Rosellinia australiensis on Banksia grandis, Tiarosporella corymbiae on Corymbia calophylla, Verrucoconiothyriumeucalyptigenum on Eucalyptus sp., Zasmidium commune on Xanthorrhoea sp., and Zasmidium podocarpi on Podocarpus sp. Brazil: Cyathus aurantogriseocarpus on decaying wood, Perenniporia brasiliensis on decayed wood, Perenniporia paraguyanensis on decayed wood, and Pseudocercospora leandrae-fragilis on Leandrafragilis.Chile: Phialocephala cladophialophoroides on human toe nail. Costa Rica: Psathyrella striatoannulata from soil. Czech Republic: Myotisia cremea (incl. Myotisia gen. nov.) on bat droppings. Ecuador: Humidicutis dictiocephala from soil, Hygrocybe macrosiparia from soil, Hygrocybe sangayensis from soil, and Polycephalomyces onorei on stem of Etlingera sp. France: Westerdykella centenaria from soil. Hungary: Tuber magentipunctatum from soil. India: Ganoderma mizoramense on decaying wood, Hodophilus indicus from soil, Keratinophyton turgidum in soil, and Russula arunii on Pterigota alata.Italy: Rhodocybe matesina from soil. Malaysia: Apoharknessia eucalyptorum, Harknessia malayensis, Harknessia pellitae, and Peyronellaea eucalypti on Eucalyptus pellita, Lectera capsici on Capsicum annuum, and Wallrothiella gmelinae on Gmelina arborea.Morocco: Neocordana musigena on Musa sp. New Zealand: Candida rongomai-pounamu on agaric mushroom surface, Candida vespimorsuum on cup fungus surface, Cylindrocladiella vitis on Vitis vinifera, Foliocryphia eucalyptorum on Eucalyptus sp., Ramularia vacciniicola on Vaccinium sp., and Rhodotorula ngohengohe on bird feather surface. Poland: Tolypocladium fumosum on a caterpillar case of unidentified Lepidoptera.Russia: Pholiotina longistipitata among moss. Spain: Coprinopsis pseudomarcescibilis from soil, Eremiomyces innocentii from soil, Gyroporus pseudocyanescens in humus, Inocybe parvicystis in humus, and Penicillium parvofructum from soil. Unknown origin: Paraphoma rhaphiolepidis on Rhaphiolepsis indica.USA: Acidiella americana from wall of a cooling tower, Neodactylaria obpyriformis (incl. Neodactylaria gen. nov.) from human bronchoalveolar lavage, and Saksenaea loutrophoriformis from human eye. Vietnam: Phytophthora mekongensis from Citrus grandis, and Phytophthora prodigiosa from Citrus grandis. Morphological and culture characteristics along with DNA barcodes are provided.

Effects of ivermectin on Danio rerio: a multiple endpoint approach: behaviour, weight and subcellular markers.

Ivermectin (IVM) is a broad acting antihelmintic used in various veterinary pharmaceuticals. It has been shown that IVM enters the aquatic compartment and adversely affects organisms including fish. This study is based on the hypothesis that long term exposure to IVM affects fish and thus, the main objective was to assess the chronic effects of 0.25 and 25 µg IVM/L to zebrafish using multiple endpoints representative of several levels of biological organization: weight, behaviour (swimming and feeding) and subcellular markers including biomarkers for oestrogenicity (vitellogenin-VTG), oxidative stress (catalase-CAT and glutathione-S-transferase-GST) and neurotransmission (cholinesterase-ChE). Concentrations as low as 0.25 µg IVM/L disrupted the swimming behaviour, causing fish to spend more time at the bottom of aquaria. Such reduction of the swimming performance affected the feeding ability which is likely responsible for the weight loss. The effects on weight were gender differentiated, being more pronounced in males (0.25 µg IVM/L) than in females (25 µg IVM/L). Fish exposed to 25 µg/L exhibited darker coloration and mild curvature of the spine. No effects on VTG and AChE were observed, but a reduction on CAT and GST levels was observed in fish exposed to 25 µg IVM/L, although these alterations probably only reflect the general condition of the fish which was significantly compromised at this concentration. Despite that predicted environmental concentrations of IVM are below 0.25 µg/L, the behavioural effects may be translated into important ecological impacts, e.g. at predator-prey interactions where fish competitive advantage can be decreased. Future work should address the link between behaviour disruption and population fitness. The current study was based on a one experiment and multiple endpoint (anchored) approach, allowing the results to be integrated and linked towards a mechanistic understanding.

Effects of the Pyrethroid Esfenvalerate on the Oligochaete, Lumbriculus variegatus.

Esfenvalerate is a neurotoxic pyrethroid insecticide widely used for agricultural and residential purposes and is considered toxic to nontarget organisms such as fish and aquatic invertebrates. In this study, we evaluated the toxicity of esfenvalerate on the aquatic oligochaete Lumbriculus variegatus. In the acute test, organisms showed visible signs of stress but no LC50 value could be determined. In the 28-day chronic test, a significant decrease in reproduction was observed with a NOEC value of 0.25 µg/kg and a LOEC value of 2.34 µg/kg. As for biomass per worm, a significant decrease was also observed with a NOEC value of 2.34 µg/kg and a LOEC value of 36.36 µg/kg. Reproductive impairment and reductions in biomass of L. variegatus exposed to environmentally realistic concentrations of esfenvalerate observed in laboratory tests suggests potential deleterious effects of this pyrethroid on oligochaete natural populations.

A protein extract and a cysteine protease inhibitor enriched fraction from Jatropha curcas seed cake have in vitro anti-Toxoplasma gondii activity.

Toxoplasma gondii is a parasite of great medical and veterinary importance that has worldwide distribution and causes toxoplasmosis. There are few treatments available for toxoplasmosis and the search for plant extracts and compounds with anti-Toxoplasma activity is of utmost importance for the discovery of new active drugs. The objective of this study was to investigate the action of a protein extract and a protease inhibitor enriched fraction from J. curcas seed cake on developing tachyzoites of T. gondii-infected Vero cells. The protein extract (JcCE) was obtained after solubilization of the J. curcas seed cake with 100 mM sodium borate buffer, pH 10, centrifugation and dialysis of the resulting supernatant with the extracting buffer. JcCE was used for the in vitro assays of anti-Toxoplasma activity at 0.01, 0.1, 0.5, 1.5, 3.0 and 5.0 mg/ml concentration for 24 h. The results showed that JcCE reduced the percentage of infection and the number of intracellular parasites, but had no effect on the morphology of Vero cells up to 3.0 mg/mL. The cysteine protease inhibitor enriched fraction, which was obtained after chromatography of JcCE on Sephadex G-75 and presented a unique protein band following SDS-PAGE, reduced both the number of T. gondii infected cells and intracellular parasites. These results suggest that both JcCE and the cysteine protease inhibitor enriched fraction interfere with the intracellular growth of T. gondii.

Cholinesterase activity on Echinogammarus meridionalis (Pinkster) and Atyaephyra desmarestii (Millet): characterisation and in vivo effects of copper and zinc.

Metals are released into freshwater ecosystems from natural and anthropogenic sources, compromising their structural and functional equilibrium. As early warning tools, cholinesterases (ChEs) are usually used to assess the effects of organophosphate and carbamate pesticides, but are also known to be inhibited by metals. The objectives of this work were to characterise the activity of ChE present in the amphipod Echinogammarus meridionalis and the shrimp Atyaephyra desmarestii and to evaluate the in vivo effects of the metals copper and zinc in their ChE activity. To achieve this, firstly the activity of ChE forms were characterised using different in vitro assays with substrates and selective inhibitors. Then, the in vivo effects of 48 h exposures to increasing concentrations of copper and zinc on ChE activity were determined. The ChE form present in both species was acetylcholinesterase (AChE) since both revealed preference for the acetylthiocholine iodide substrate, total inhibition with eserine, the inhibitor of ChEs, and with 1,5-bis(4-allyldimethylammoniumphenyl)-pentan-3-one dibromide, the specific inhibitor of AChE, and presented insensitivity to iso-OMPA, a specific inhibitor of butyrylcholinesterase. The activity of ChEs was inhibited by zinc exposures in the amphipod species, but was not affected by copper. Exposure to copper and zinc did not affect ChEs activity in the shrimp at the concentrations tested. This work is a relevant contribution as foundation for the use of AChE in freshwater crustaceans in further studies including biomonitoring campaigns in different contamination scenarios.

Potential risk of biochar-amended soil to aquatic systems: an evaluation based on aquatic bioassays.

It is vital to address potential risks to aquatic ecosystems exposed to runoff and leachates from biochar-amended soils, before large scale applications can be considered. So far, there are no established approaches for such an assessment. This study used a battery of bioassays and representative aquatic organisms for assessing the acute toxicity of water-extractable fractions of biochar-amended soil, at reported application rates (80 t ha(-1)). Biochar-amended aqueous soil extracts contained cadmium (Cd), cobalt (Co), chromium (Cr), copper (Cu), manganese (Mn), zinc (Zn), nickel (Ni), lead (Pb), arsenic (As) and mercury (Hg) (Σmetals 96.3 µg l(-1)) as well as the 16 priority PAHs defined by the U.S. Environmental Protection Agency (Σ16PAHs 106 ng l(-1)) at contents in the range of current EU regulations for surface waters. Nevertheless, acute exposure to soil-biochar (SB) extracts resulted in species-specific effects and dose-response patterns. While the bioluminescent marine bacterium Vibrio fischeri was the most sensitive organism to aqueous SB extracts, there were no effects on the growth of the microalgae Pseudokirchneriella subcapitata. In contrast, up to 20 and 25% mobility impairment was obtained for the invertebrate Daphnia magna upon exposure to 50 and 100% SB extract concentrations (respectively). Results suggest that a battery of rapid and cost-effective aquatic bioassays that account for ecological representation can complement analytical characterization of biochar-amended soils and risk assessment approaches for surface and groundwater protection.

Water-extractable priority contaminants in LUFA 2.2 soil: back to basics, contextualisation and implications for use as natural standard soil.

The natural LUFA 2.2 standard soil has been extensively used in hazard assessment of soil contaminants, combining representation with ecological relevance for accurate risk evaluation. This study revisited the water-extractable fraction of LUFA 2.2 soil, through consecutive soil wet-dry cycles and discusses implications of use as standard substrate in derivation of ecotoxicological data and toxicity thresholds. Potentially bioavailable contents of metals (177.9-888.7 µg/l) and the 16 polycyclic aromatic hydrocarbons (PAHs; 0.064-0.073 µg/l) were dependent on the number of soil wetting-drying cycles applied. Such contents were screened based on current EU guidelines for surface waters and reported toxicological benchmarks for aquatic organisms. Aqueous concentrations generally fit within recommended Environmental Quality Standards (EQS), except for Hg (0.13-0.22 µg/l; >Maximum Allowable Concentration-MAC-of 0.07 µg/l) and for the sum of benzo(g,h,i)perylene and indeno(1,2,3-cd)pyrene (0.005 µg/l; >double the Annual Average of 0.002 µg/l). Further, aqueous As, Zn, Cd, Ni and Cr concentrations exceeded 'lower benchmark' values for aquatic organisms, possibly reflecting an inadequate derivation for ecotoxicological data. In turn, PAHs in LUFA 2.2 soil aqueous extracts, whilst individually, are not likely to constitute a hazard to test biota exposed to its aqueous fractions. This study urges for potentially bioavailable fractions of reference and standard natural soils to be adequately characterized and addressed as part of the research aim, experimental approach and design, as well as the expected scope of the outcomes.

Pesticide residue analyses and biomarker responses of native Costa Rican fish of the Poeciliidae and Cichlidae families to assess environmental impacts of pesticides in Palo Verde National Park.

Pesticide chemical residues in water samples and biomarker responses in transplanted fish were used to monitor environmental hazards of pesticides in Palo Verde National Park (Costa Rica). The Costarican fish, Parachromis dovii (Ciclhidae) and Poecilia gillii (Poecillidae), were selected as sentinel species. Contaminant analyses detected up to 15 different pesticide residues in water with hexachlobenzene (2261 ng l(-1)), phorate (473 ng l(-1)), epoxiconazole (314) and bromacil (117 ng l(-1)) being the compounds found in higher concentrations. Biomarker responses evidenced impacts on cholinesterase activities in transplanted fish at Barbudal site probably due to the presence of organophosphate insecticides such as phorate. High enzyme activities of glutathione S-transferase and catalase and elevated levels of lipid peroxides were also observed at a site impacted by rice fields (Cabuyo); those effects could be associated with the presence of hexachloro benzene and triazole fungicides. In general, P. dovii biomarkers were affected to a greater extent than those of P. gillii in fish transplanted to sites associated with agriculture, which suggests the former species is a good candidate for future surveys.

Action of two phospholipases A2 purified from Bothrops alternatus snake venom on macrophages.

The in vitro effects of BaltTX-I, a catalytically inactive Lys49 variant of phospholipase A2 (PLA2), and BaltTX-II, an Asp49 catalytically active PLA2 isolated from Bothrops alternatus snake venom, on thioglycollate-elicited macrophages (TG-macrophages) were investigated. At non-cytotoxic concentrations, the secretory PLA2 BaltTX-I but not BaltTX-II stimulated complement receptor-mediated phagocytosis. Pharmacological treatment of TG-macrophages with staurosporine, a protein kinase C (PKC) inhibitor, showed that this kinase is involved in the increase of serum-opsonized zymosan phagocytosis induced by BaltTX-I but not BaltTX-II secretory PLA2, suggesting that PKC may be involved in the stimulatory effect of this toxin in serum-opsonized zymosan phagocytosis. Moreover, BaltTX-I and -II induced superoxide production by TG-macrophages. This superoxide production stimulated by both PLA2s was abolished after treatment of cells with staurosporine, indicating that PKC is an important signaling pathway for the production of this radical. Our experiments showed that, at non-cytotoxic concentrations, BaltTX-I may upregulate phagocytosis via complement receptors, and that both toxins upregulated the respiratory burst in TG-macrophages.

Soil habitat function after innovative nanoagriproducts application: Effect of ageing on the avoidance behaviour of the soil invertebrates Enchytraeus crypticus and Folsomia candida.

Research on nanotechnology with applications in agriculture has been gathering attention because it may achieve a good balance between agricultural production and environmental integrity. Among the vast nanomaterials, layered double hydroxides (LDHs) are a promising solution for supplying crops with macro- and/or micronutrients. Still, little is known about their safety implications for non-target organisms, such as soil invertebrates. The habitat function of soils might be impacted by potential stressors, which can be assessed through avoidance behaviour tests. This study aimed to assess the effect of two innovative agriproducts, Zn-Al-NO3 LDH and Mg-Al-NO3 LDH, on the avoidance behaviour of the enchytraeid Enchytraeus crypticus and the collembolan Folsomia candida, over time. Simultaneously, Zn and Mg potential release from LDHs to soil was evaluated. Overall, the behaviour of soil invertebrates differed between species, with enchytraeids being more sensitive to LDHs-treated soils than collembolans, possibly explained by their different physiological traits. The behaviour of soil organisms also depended on the LDH structural composition and was time-variable. Soil treated with Zn-Al-NO3 LDH was perceived as less favourable compared to Mg-Al-NO3 LDH, which was preferred to clean soil at most tested concentrations. LDHs toxicity was partly, but not exclusively, related to Zn and Mg release. Cations release over time was demonstrated in the chemical assessment. Still, LDHs toxicity to soil invertebrates decreased as increasing AC50 values were derived over time. Slower dissolution over time might explain the decrease in toxicity. Our study demonstrates that both soil invertebrates could sense LDHs in soil and eventually adapt their behaviour by avoiding or preferring, according to the type and level of LDH present.