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10 Pág. Departamento de Medio Ambiente y Agronomía​, Rodents are considered one of the animal pests with the greatest impact on agricultural production and public health. Anticoagulant rodenticides (ARs), used as one of the most effective ways to control rodent populations worldwide, inhibit the vitamin K 2,3-epoxide reductase (VKORC1) enzyme involved in blood coagulation. Resistances to ARs are mainly associated with mutations or single nucleotide polymorphisms (SNPs) in the vkorc1 gene. Since the information on this subject is scarce in Spain, we monitored and discovered rodent SNPs that could favour genetic resistance in its populations. For that, more than 200 samples of stools and tails from brown rat (Rattus norvegicus), black rat (Rattus rattus) and mouse (Mus musculus) were collected from 12 Spanish regions previously identified with low AR efficacy in coordination with the National Association of Environmental Sanitation Companies (ANECPLA) and the managing entities of four locations. We then sequenced their vkorc1 exon 3 corresponding genomic DNA. We identified genotypic vkorc1 variations corresponding to amino acid changes at the VKORC1 protein at the S149I - S149T and the E155K - E155Q mutations, depending on the rodent species. Computational analysis of binding predictions found out that the brown rat S149I mutation predicted a high reduction of the binding affinity of chlorophacinone and brodifacoum ARs while, the black rat S149T, E155K and E155Q mutations slightly reduced bromadiolone AR binding. These results suggest that these mutations may be one of the causes of the increased resistance to those ARs., This research was funded by a charge of the Ministry for Ecological Transition and Demographic Challenge to Instituto Nacional de Investigación y Tecnología Agraria y Alimentaria (INIA), charge number EG17-017 and the project PID2019-108053RJ-I00/AEI/10.13039/501,100,011,033 funded by the Agencia Estatal de Investigación (AEI)

Background This study was conducted to provide much needed information on the potential of low solubility, highly biodurable nanoparticles (NPs) (TiO2 and CeO2 NPs), to bioaccumulate in fish and to investigate the relationship between an engineered nanomaterials surface coating, uptake and biokinetics in vivo. Rainbow trout (Oncorhynchus mykiss) were fed diets spiked with uncoated or surface coated (polyethylene glycol (PEG) or citrate (CIT)) TiO2 and CeO2 NPs (100 mg NPs/kg feed) for 10 days and thereafter fish were allowed to depurate for 42 days. Special care was taken to measure the real dispersed and actual administered concentrations, taking into consideration any potential losses from leaching, and to characterise the form (size, aggregation state, charge) of the NPs to which the fish were exposed. Results The coatings had an influence on levels of uptake and distributions. Most notably a higher uptake of PEG and CIT coated TiO2 and CeO2 NPs compared to fish exposed to uncoated materials was observed. The elimination of any Ti from tissues was rapid during the first day of depuration, whereas Ce levels remained in fish tissues (stomach, intestine and liver) with differences in depuration and redistribution of the three types of CeO2 NPs. However, no bioaccumulation potential for both tested metal oxide NPs in fish irrespective of coatings is expected according to BMF values Conclusions Distinct uptake, distribution and depuration kinetics in rainbow trout have been evidenced for different metal oxide NPs (TiO2 and CeO2 NPs). Coatings influenced uptake and in some cases led to slower depuration and distinct distributions, but do not make the uncoated NPs studied bioaccumulative (BMF > 1). Graphical Abstract

The Acacias River in Colombia receives large volumes of industrial effluents mostly derived from the oil industry. To contribute to the study of the possible effects of industrial wastewaters on the aquatic environment and particularly on fish populations, a native neotropical fish, Aequidens metae was used as a sentinel species. Wild specimens of A. metae were caught at three different places of the Acacias River taking as reference the point of discharge of an oil industry effluent; upstream, downstream, and at the vicinity of the discharge pipe. A fourth sampling site was chosen as a reference site away from urban settlements. Samplings were performed twice, during the rainy and dry seasons. After anesthesia animals were weighted and measured, and humanely sacrificed. Livers were extracted, frozen on site and transported to the laboratory. Condition indices were calculated. Total protein content and the detoxification 7-ethoxyresorufin-O-deethylase (EROD) enzyme activity were estimated. Histopathological alterations were also evaluated. Water quality was estimated through the measurement of several variables. Results obtained evidenced that the highest induction in EROD activity and the strongest histological alterations in liver of the monitored fish appeared during the dry seasons at the discharge site and downstream to this point., The authors wish to thank the students and research assistants of the Research Group BioTox for their valuable participation during the sampling procedures. This study was funded by Colciencias, ANH and Universidad de los Llanos Project number 112272151869, contrato 162-2016, grant 721–2015 “Convocatoria para la formación de recurso humano en Colombia en el área de hidrocarburos, a través de proyectos de investigación”).

In order to identify new sustainable sources for producing cellulose nanofibers (CNFs), fast-growing poplar (Populus alba L.) wood was evaluated herein. For that purpose, bleached poplar kraft pulp was produced and submitted to TEMPO (2,2,6,6-tetramethylpiperidine-1-oxyl radical) mediated oxidation (TEMPO-ox) chemical pretreatment followed by microfluidization. The resulting CNFs were thoroughly characterized, including a rheological study at different pH values. Poplar CNFs showed properties comparable to eucalypt CNFs (reference material for CNFs production), showing high carboxylate content (1048 ± 128 µmol g−1 ), fibrillation yield (87.3% ± 8.1%), optical transmittance (83% at 700 nm) and thermal stability (up to more than 200 ◦C). Regarding the rheological study, whereas pH from 4 to 10 did not produce significant changes in rheological behavior, a reduction of pH down to 1 led to an order-of-magnitude increase on the viscoelastic functions. Therefore, poplar CNF shows potential in the pH-sensitive hydrogels application field. Finally, the possible ecotoxicity of poplar CNF was assessed. The decrease in cell viability was very low so that only concentrations causing a 10% cytotoxicity could be calculated for the assay detecting alterations in cell metabolism (10 µg mL−1 ) and plasma membrane integrity (60 µg mL−1), This research was funded by Comunidad de Madrid and MCIU/AEI/FEDER, EU via Projects SUSTEC-CM S2018/EMT-4348 and RTI2018-096080-B-C22, respectively. Silviculture and Forest Management Department (Forest Research Center, INIA, CSIC) is acknowledged for fast-growing poplar materials

Nanoenabled products (NEPs) have numerous outdoor uses in construction, transportation or consumer scenarios, and there is evidence that their fragments are released in the environment at low rates. We hypothesized that the lower surface availability of NEPs fragment reduced their environmental effects with respect to pristine nanomaterials. This hypothesis was explored by testing fragments generated by intentional micronisation ("the SUN approach"; Nowack et al. Meeting the Needs for Released Nanomaterials Required for Further Testing: The SUN Approach. Environmental Science & Technology, 2016 (50), 2747). The NEPs were composed of four matrices (epoxy, polyolefin, polyoxymethylene, and cement) with up to 5% content of three nanomaterials (carbon nanotubes, iron oxide, and organic pigment). Regardless of the type of nanomaterial or matrix used, it was observed that nanomaterials were only partially exposed at the NEP fragment surface, indicating that mostly the intrinsic and extrinsic properties of the matrix drove the NEP fragment toxicity. Ecotoxicity in multiple assays was done covering relevant media from terrestrial to aquatic, including sewage treatment plant (biological activity), soil worms (Enchytraeus crypticus), and fish (zebrafish embryo and larvae and trout cell lines). We designed the studies to explore the possible modulation of ecotoxicity by nanomaterial additives in plastics/polymer/cement, finding none. The results support NEPs grouping by the matrix material regarding ecotoxicological effect during the use phase. Furthermore, control results on nanomaterial-free polymer fragments representing microplastic had no significant adverse effects up to the highest concentration tested.

Aflatoxin B1 (AFB1) and fumonisin B1 (FB1) are mycotoxins widely found as cereal contaminants and their co-occurrence in corn has been associated with a high incidence of liver cancer. Both toxins are immunotoxic, with AFB1 being a procarcinogen, and its bioactivation through specific cytochrome P450 (Cyp) enzymes, such as Cyp1A, being a requirement for hepatocarcinogenic and toxic activities. This study evaluated the effects of these mycotoxins, alone or combined, on activation and expression of Cyp1A and its transcription factor aryl hydrocarbon receptor (Ahr) in hepatoma cell line H4IIE and spleen mononuclear cells of rats. The results demonstrate that in H4IIE cells, AFB1 induced an increase in Cyp1A activity and cyp1A transcription, associated with an enhanced Ahr activity, which suggests that this toxin can act as an Ahr agonist. Moreover, FB1 caused a small rise in Cyp1A activity and cyp1A expression. Similarly in spleen cells, AFB1 and FB1 induced overexpression of cyp1A and ahr genes. This work shows that the response potency was significantly higher for the mixture, indicating the existence of an interaction between both toxins. This study proposes the Ahr pathway activation as a toxicity mechanism of AFB1 and FB1, and highlights that FB1 may increase AFB1 bioactivation.

There is limited data on the sub-lethal oxidative stress effects of titanium dioxide nanoparticle aggregates (NM-TiO2) and its modulation by simulated solar radiation (SSR) to aquatic organisms. This study aimed to examine sublethal oxidative stress effects of aqueous exposure to three different types of NM-TiO2 differing in their coating or crystal structure but of similar primary size (20nm) plus a micron-sized bulk material to zebrafish embryos without and with SSR. Oxidative stress responses of known model prooxidant (tert-Butyl hydroperoxide) and photoprooxidant (fluoranthene) compounds were also studied. Results evidenced a low bio-availability of NM-TiO2 to embryos with detrimental effects on growth at 1mgml-1. Phototoxicity increased moderately, by 3 and 1.5 fold, under co-exposures to fluoranthene (100μgl-1) and to the NM-TiO2 P25 (1mgml-1), respectively, being unchanged in the other TiO2 aggregates. In vitro exposures under SSR confirmed that the NM-TiO2 P25 had the highest potential to generate reactive oxygen species (ROS). Antioxidant enzyme activities of superoxide dismutase increased shortly after exposure to the studied materials, whereas the levels of glutathione tend to be altered after longer exposures. All compounds were able to produce oxidative stress enhancing the senescence-associated β galactosidase pigment (SA-β-gal). Under SSR radiation the NM-TiO2 P25 affected antioxidant and oxidative stress responses as the phototoxic compound fluoranthene. These results indicated that despite the low bio-availability of NM-TiO2 to zebrafish embryos, P25 was phototoxic due to the production of reactive oxygen species. Nevertheless, overall our results indicated that fish development may not be at high risk in the face of NM-TiO2, even when combined with prooxidant conditions. © 2013 Elsevier B.V.

Graphene and its derivatives are an emerging class of carbon nanomaterial with great potential for a broad range of industrial and consumer applications. However, their increasing production and use is expected to result in release of nano-sized graphene platelets into the environment, where they may interact with chemical pollutants modifying their fate and toxic potential. The objective of this study was to assess whether graphene nanoplatelets can act as vector for aromatic environmental pollutants increasing their cellular uptake and associated hazardous effects in vitro. For this purpose, cell cultures of the topminnow fish (Poeciliopsis lucida) hepatoma cell line PLHC-1 were simultaneously (and successively) exposed to graphene nanoplatelets (graphene oxide (GO) or carboxyl graphene (CXYG)) and an aryl hydrocarbon receptor (AhR) agonist (β-naphthoflavone (β-NF), benzo(k)fluoranthene (BkF) or 3,3',4,4',5,5'-hexachlorobiphenyl (PCB169)). Following exposure cytochrome P450 1A (Cyp1A) induction was assessed by measuring cyp1A mRNA expression levels using reverse transcriptase-quantitative polymerase chain reaction (RT-qPCR) and Cyp1A-dependent ethoxyresorufin-O-deethylase (EROD) activity. It was observed that pre- and co-exposure of cells to GO and CXYG nanoplatelets had a potentiating effect on β-NF, BkF, and PCB169-dependent Cyp1A induction suggesting that graphene nanoplatelets increase the effective concentration of AhR agonists by facilitating their passive diffusion into the cells by damaging the cells' plasma membrane and/or by transporting them over the plasma membrane via a Trojan horse-like mechanism. The results demonstrate the existence of combination effects between nanomaterials and environmental pollutants and stress the importance of considering these effects when evaluating their respective hazard.

Carbaryl (1-naphthyl-N-methylcarbamate), a widely used carbamate insecticide, induces cytochrome P450 1A gene expression in mammalian cells. This activity is usually mediated by the interaction of the compound with the aryl hydrocarbon receptor. However, it has been proposed that this mechanism does not apply to carbaryl because its structure differs from that of typical aryl hydrocarbon receptor ligands. We show here that carbaryl promotes activation of target genes in a yeast-based bioassay expressing both aryl hydrocarbon receptor and aryl hydrocarbon receptor nuclear translocator. By contrast, carbaryl acted as a competitive inhibitor, rather than as an agonist, in a simplified yeast system, in which aryl hydrocarbon receptor nuclear translocator function is bypassed by fusing aryl hydrocarbon receptor to a heterologous DNA binding domain. This dual action of carbaryl, agonist and partial antagonist, was also observed by comparing carbaryl response in two vertebrate cell lines. A yeast two-hybrid assay showed that the mammalian coactivator cAMP response element-binding protein readily interacts with aryl hydrocarbon receptor bound to its canonical ligand beta-naphthoflavone, but not with the carbaryl-aryl hydrocarbon receptor complex. We propose that carbaryl interacts with aryl hydrocarbon receptor, but that its peculiar structure imposes a substandard configuration on the aryl hydrocarbon receptor ligand-binding domain that prevents interaction with key coactivators and activates transcription without the need for aryl hydrocarbon receptor nuclear translocator. This effect may be relevant in explaining its physiological effects in exposed animals, and may help to predict its effects, and that of similar compounds, in humans. Our data also identify the aryl hydrocarbon receptor/cAMP response element-binding protein interaction as a molecular target for the identification and development of new aryl hydrocarbon receptor antagonists.