The effect of PFOs on the uptake and translocation of emerging contaminants by crops cultivated under soil and soilless conditions.

The Mediterranean is a region of substantial agriculture production that faces concurrent environmental stresses and freshwater pollution given the occurrence of emerging contaminants (ECs). Among these pollutants, the surface-active substances have been suggested to enhance the bioavailability of other ECs. This research evaluates a comparative uptake and translocation assessment of irrigation exposure to atenolol (ATN, 60 µg/L), carbamazepine (CBZ, 60 µg/L) and triclosan (TCS, 30 µg/L) alone vs. these combined with perfluorooctanesulfonic acid (PFOS, 10 µg/L) under semifield (i.e., soil experimental set) vs. hydroponics (i.e., soilless experimental set) growing conditions with lettuce, radish and tomato plants. Both experimental sets revealed efficient root uptake and translocation for the three ECs regardless of their co-existence with PFOS. The overall results of the uptake and translocation of the ECs in the lettuce and tomato plants suggested a simultaneous treatment-plant organ interaction, which was not affected by PFOS being present in both experimental sets. PFOS in irrigation water did not increase cellular perviousness to the other three ECs. These observations support the hypothesis of factors other than PFOS being responsible for the differential bioaccumulation and translocation potentials seen in both experimental sets. However, the radish plants co-irrigated with PFOS brought about increased movement of ECs from roots to aerial parts, more specifically ATN and CBZ in the soil experimental set, and ATN and TCS in the soilless set. These results support the notion that factors inherent to the physiological characteristics of this root vegetable contributed to ECs' increased tendency to move from roots to aerial parts. Despite the three ECs efficiently accumulating, the risk to humans from eating the edible parts of these plants grown under soil or soilless conditions was low.

Uptake of atenolol, carbamazepine and triclosan by crops irrigated with reclaimed water in a Mediterranean scenario.

Water scarcity is a natural condition in the Mediterranean rim countries. In this region, reuse of reclaimed water (RW) from wastewater treatment plants (WWTPs) is becoming a potential source for highly water-demanding activities such as agriculture. However, the removal capacity of contaminants in regular WWTPs has been found to be limited. Considering a Mediterranean scenario, this research investigated the plant uptake and translocation of three representative pharmaceuticals and personal care products (PPCPs) typically present in RW samples from a WWTP located in an urban area in Spain, and assessed the potential risk to humans from plant consumption. The RW samples were collected and analyzed for three representative PPCPs (atenolol -ATN-, carbamazepine -CBZ- and triclosan -TCS-). The target contaminants were also spiked at two levels in the RW samples to consider two worst-case scenarios. Three plant models (lettuce, maize and radish) were grown outdoors and irrigated with four treatments: tap water; RW samples, and the two spiked RW samples. Generally speaking, results revealed an efficient root uptake for the three PPCPs regardless of plant species and fortification level, and suggested an interaction effect of treatment and plant organ. Different bioaccumulation and translocation potentials of the three PPCPs were seen into the aerial organs of the plants. Overall, these observations support the idea that factors including the physico-chemical properties of the PPCPs and physiological plant variables, could be responsible for the differential accumulation and translocation potentials observed. These variables could be critical for crops irrigated with RW in regions with extended dry seasons, high solar incidence and low annual rainfall such as those in the Mediterranean rim where plants are subjected to high transpiration rates. However, the results obtained from this experimental approach suggested a negligible risk to humans from consumption of edible plants irrigated with RW samples with presence of PPCPs, despite the fact that the three representative PPCPs under study accumulated efficiently in the plants.

Alterations in gene expression levels provide early indicators of chemical stress during Xenopus laevis embryo development: A case study with perfluorooctane sulfonate (PFOS).

In the present study, Xenopus laevis embryos were exposed to a range of perfluorooctane sulfonate (PFOS) concentrations (0, 0.5, 6, 12, 24, 48 and 96mg/L) for 96h in laboratorial conditions to establish toxicity along with possible gene expression changes. Mortality and deformities were monitored daily and head-tail length was measured at the end of the assay as an indicator of growth. At 24 and 96h post-exposure (hpe), the mRNA expression levels of the genetic markers involved in general stress responses (hsp70, hsp47, crh-a and ucn1), oxidative stress (cat.2 and sod), lipid metabolism (ppard) and apoptosis (tp53 and bax) were analyzed by RT-qPCR. Malformations were significantly higher in the embryos exposed to the highest PFOS concentration (41.8% to 56.4%) compared to controls (5.5%) at 48, 72 and 96hpe. Growth inhibition was observed in the embryos exposed to PFOS concentrations≥48mg/L. At 24 hpe, a statistically significant up-regulation of genes hsp70, hsp47, ppard, tp53 and bax in relation to controls was found. Similar responses were found for genes hsp70, hsp47, crh-a, ucn1, sod and ppard at 96 hpe. Alterations in the mRNA expression levels indicated both a stress response to PFOS exposure during X. laevis embryo development, and alterations in the regulation of oxidative stress, apoptosis, and differentiation. These molecular alterations were detected at an earlier exposure time or at lower concentrations than those producing developmental toxicity. Therefore, these sensitive warning signals could be used together with other biomarkers to supplement alternative methods (i.e. the frog embryo test) for developmental toxicity safety evaluations, and as tools in amphibian risk assessments for PFOS and its potential substitutes.

Effects of life cycle exposure to dietary 2,2', 4,4'-tetrabromodiphenyl ether (BDE-47) on medaka fish (Oryzias latipes).

Previous studies conducted in our laboratory, which resorted to 40-day oral exposures to BDE-47 in specific developmental windows of medaka (Oryzias latipes) did not evidence effects on growing or breeding periods. In this new study, full life cycle (i.e. 140-day) dietary exposure to 1000 ng of BDE-47/g was performed with medaka to evaluate effects on growth and reproduction (i.e. fecundity, fertility, hatchability), and to analyze the bioacumulated BDE-47 in and transferred to offspring. No significant effects were observed for the biometric analyses during the growth and maturation periods and no biased sex ratios were found. Reproductive capacity was not affected by the presence of BDE-47 in diet. There was no evidence for apparent effects from parental exposure during embryo and eleutheroembryo development. The analytical results revealed steady BDE-47 bioaccumulation during the growing period, which remained in the reproductive phase in males, and a decreasing tendency was noted in females. These lowering BDE-47 levels in females coincided with the detected BDE-47 levels offloaded in embryos. In the 10-day-old post-hatch larvae, the BDE-47 concentrations dropped to comparatively lower values than the concentrations detected in parents. This finding suggests an efficient metabolic process in the eleutheroembryonic and post-eleutheroembryonic phases. Our 140-day dietary approach found no BDE-47 effects on medaka growth and reproduction, or in early progeny stages despite effective bioaccumulation and maternal transfer.

Effects of life cycle exposure to polystyrene microplastics on medaka fish (Oryzias latipes).

The number of published studies evaluating the effects of microplastics (MPs) in fish has increased in the last decade. However, of the available studies, few have explored the long-term effects of MPs on fish growth and reproduction and have resorted to MPs in the form of µm-sized beads/microspheres. In this study, 6-10 day-old post-hatch medaka (Oryzias latipes) fish were exposed to 50 (i.e. 1X) and 500 (i.e. 10X) µg of heterogeneously sized and irregularly shaped virgin polystyrene (PS) MP particles (200-µm range)/L for 150 days. These concentrations corresponded to respective daily mean values of 247 and 3087 particles/L administered through the diet. The PS MPs dietary exposure resulted in body burdens of 114 and 440 particles/g fish on day 50, and of 78 and 173 particles/g fish on day 100 since the respective exposures to the 1X and the 10X treatments started. The biometric analyses found no incidence of PS MPs ingestion on overall fish growth and development. The histological survey in the 10X group did not reveal alterations in gills or in the digestive tract. Mild alterations in other organs were seen and included increased fluid material in the peritoneal cavity, glomerular and tubular alterations in kidneys, and differences in the diameter of the thyroid follicles and thickness of the follicular epithelial cells. The initial days of the reproductive phase revealed MP-related differences in the number of gravid females, fecundity, and fertilization rates. Overall, these values reverted to normal rates throughout the succeeding days. No significant effects of PS MPs exposure were evidenced on offspring success. The 150-day PS MPs dietary exposure used in this study provided clues of histological effects and a reproduction delay. However, it did not seem to compromise overall growth/thriving and the ongoing reproduction.

Effect assessment of reclaimed waters and carbamazepine exposure on the thyroid axis of Xenopus laevis: Gene expression modifications.

Reclaimed water (RW) obtained from wastewater treatment plants (WWTP) is used for irrigation, groundwater recharge, among other potential uses. Although most pollutants are removed, traces of them are frequently found, which can affect organisms and alter the environment. The presence of a myriad of contaminants in RW makes it a complex mixture with very diverse effects and interactions. A previous study, in which tadpoles were exposed to RW and RW spiked with Carbamazepine (CBZ), presented slight thyroid gland stimulation, as suggested by the development acceleration of tadpoles and histological findings in the gland provoked by RW, regardless of the CBZ concentration. To complement this study, the present work analysed the putative molecular working mechanism by selecting six genes coding for the thyroid-stimulating hormone (TSHß), thyroid hormone metabolising enzymes (DIO2, DIO3), thyroid receptors (THRA, THRB), and a thyroid hormone-induced DNA binding protein (Kfl9). Transcriptional activity was studied by Real-Time PCR (RT-PCR) in brains, hind limbs, and tails on exposure days 1, 7, and 21. No significant differences were observed between treatments for each time point, but slight alterations were noted when the time response was analysed. The obtained results indicate that the effects of RW or RW spiked with CBZ are negligible for the genes analysed during the selected exposure periods.

The role of PFOS on triclosan toxicity to two model freshwater organisms.

Surface-active substances may enhance the bioavailability of certain pollutants by modifying the permeability of cell membranes. However, they could also interact in a positive manner by increasing toxicity to aquatic organisms. A comparative effects assessment of waterborne exposure to triclosan (TCS) alone vs. combined with perfluorooctanesulfonic acid (PFOS) was herein investigated in daphnids (Daphnia magna) and medaka (Oryzias latipes) early life stages (ELS) using a battery of non-invasive behavioral, physiological and anatomical endpoints. Additionally, TCS bioaccumulation was evaluated in medaka embryos to help discern differences in effects caused by either, changes in TCS permeability or by a positive interaction with PFOS. The TCS analytical measurements in the medaka ELS exposure media revealed fast dissipation with half-lives < 12 h. The D. magna immobilization and feeding inhibition assays suggested an increased response when TCS (≥200 and 37.50 µg/L, respectively) was co-exposed with PFOS. Concentrations <800 µg TCS/L did not affect medaka ELS. However, exposures to ≥400 µg TCS/L + PFOS had effects on the embryo and eleutheroembryo viability. The morphometric analysis of the embryonic gallbladder area and the oxidative stress, determined in vivo by the manifestation of reactive oxygen species (ROS), revealed a hormetic response in both experimental sets. The TCS + PFOS experimental set generally resulted in increased gallbladder areas and ROS activity levels compared to those quantified in the corresponding TCS set. The bioaccumulation studies in the medaka embryos revealed comparable TCS levels regardless of PFOS presence. Without disregarding any TCS's enhanced bioavailability caused by the surface-active substance, overall results primarily indicate increased biological effects of TCS due to a potentiation action of PFOS as a binary mixture with TCS.

Use of a novel battery of bioassays for the biological characterisation of hazardous wastes.

Four toxicity bioassays were used for the biological characterisation of nine hazardous wastes and extracts. This evaluation included conventional and novel bioassays, and combined in vivo and in vitro tests in order to facilitate the effect characterisation. This test battery incorporated different relevant taxonomic groups for the aquatic compartment and covered several ecotoxicological endpoints. The toxicity bioassays used for this characterisation were the acute immobilisation daphnia test, an acute toxicity test with larvae of Xenopus laevis, an in vitro test with the fish cell line RTG-2 comprising endpoints for cellular defence and viability, and finally the DR-CALUX assay to detect dioxin-like compounds. The aim of this study is to contribute to the development of a cost-effective battery of toxicity tests for the acute screening of hazardous and toxic wastes for the aquatic compartment. For this objective, the correlations between toxicity data derived from all bioassay were studied using a multivariate analysis, including the Principal Component Analysis. The results showed that Daphnia and Xenopus were effective assays to detect toxicity and they could be incorporated to a screening test battery. On the other hand, the toxicity results with the in vitro test RTG-2 showed that this test could be a good alternative to in vivo tests, demonstrating an acceptable sensitivity for toxicity detection and contributing other advantages as reducing assays cost and animal testing. Finally DR-CALUX test implemented the tests-batteries in the screening of hazardous wastes when there is a suspicious that dioxin-like compounds are presented in the samples.

Stage-dependent effects of chlorpyrifos on medaka (Oryzias latipes) swimming behavior using a miniaturized swim flume.

By considering chlorpyrifos (CPF), an organophosphorus pesticide with known mechanisms of action that affect neurobehavioral development, we assessed the validity and sensitivity of a miniaturized swim flume by investigating the effects of the insecticide on swimming behavior in medaka (Oryzias latipes) fish growing stages. Medaka in three developmental periods, namely 0, 20 and 40 day-old post-hatch (i.e. time points 0, 20 and 40, respectively), were exposed to CPF (12.5, 25, 50 and 100 µg/L) for 48 h under semi-static conditions. The CPF half-lives during exposures were evaluated and the swimming patterns in a flume section (arena) were presented on two-dimensional gradient maps of forced movement of fish against water current. A comparative numerical analysis of fish residence times between each time point control and the corresponding CPF groups was performed by dividing arenas into 15 proportional areas. The time point 0 control group gradient map showed a noticeably different swim pattern from those of the ≥12.5 µg CPF/L groups, which was statistically supported by the differences for residence times seen in ≥12 corresponding areas. The control group gradient maps for time points 20 and 40 differed from those of the respective ≥12.5 µg CPF/L groups. The comparative analysis of the residence times in the corresponding 15 areas revealed differences in ≥5 areas for time point 20 and in ≥3 areas for time point 40. The integrative analysis of the gradient maps and the numerical statistics revealed stage-specific effects and a concentration-response relationship between CPF and alterations on forced medaka swimming despite the dissipation of CPF from the water column. These results indicate the validity of the miniaturized swim flume toward a more environmentally realistic scenario for the evaluation of neurodevelopmental and behavioral toxicity in small fish models.

Effects of dietary exposure of polycyclic musk HHCB on the metamorphosis of Xenopus laevis.

The compound 1,3,4,6,7,8-hexahydro-4,6,6,7,8,8-hexamethylcyclopenta-[γ]-2-benzopyrane (HHCB; galaxolide, Chemical Abstracts Service number 1222-05-5) is a synthetic musk used extensively as a fragrance in many consumer products and classified as an emerging pollutant. The ecotoxicological information available for HHCB addresses exposure via water, but this compound is frequently adsorbed into particulate matter. The goal of the present study was to assess the effects of dietary exposure to several environmentally relevant HHCB concentrations adsorbed in food during Xenopus laevis metamorphosis. The authors sought to determine if such exposure to this synthetic musk resulted in histological changes in the thyroid gland in conjunction with changes in development (staging, timing to metamorphosis), body weight, and length. Developmental acceleration on day 14, together with hypertrophy of the thyroid follicular epithelium in tadpoles, suggested a possible agonistic effect of HHCB, which would have been compensated after metamorphosis by regulatory mechanisms to maintain homeostasis. Further research into the potential thyroid-related mechanisms of action of HHCB should be conducted. Environ Toxicol Chem 2016;35:1428-1435. © 2015 SETAC.

Acute and chronic toxicity of emerging contaminants, alone or in combination, in Chlorella vulgaris and Daphnia magna.

This work presents the toxicity results of different compounds classified as emerging contaminants on primary producers and primary consumers in the aquatic compartment. The objectives were to (1) obtain acute and chronic toxicity results for algae and Daphnia magna using standardised or currently used tests, (2) study the relationship between the effects on the impaired feeding rate for daphnia and the effects of reproduction and (3) examine the responses on daphnia and algae after binary combinations of environmentally relevant compounds and perfluorooctane sulfonate (PFOS). Toxicity data on personal care products (PCPs), not reported in the scientific literature up to now, are presented. The results confirmed that the Daphnia feeding bioassay can be a sensitive, ecologically relevant endpoint to detect sublethal effects and could complement the information obtained with the reproduction test on Daphnia. The results also suggested that the concomitant occurrence of PFOS and other emerging contaminants in the aquatic compartment could affect the toxicity of some compounds according to their lipophilicity.