Applicability of the Caenorhabditis elegans survival, growth and reproduction test to assess the effects of biosolids used in agriculture

Modern high productivity agriculture has led to an impoverishment of nutrients and organic carbon in field soils, and as a consequence plants nutrition complements like fertilisers or wastes have to be used to keep high yield. These complex materials can also be a source of toxicants, depending on their origin, so there is a necessity to assess their environmental effects before field use. In this study, biosolids used in agriculture were tested as a mix with standard soil at different field dose through ecotoxicity test on the nematode Caenorhabditis elegans, endpoints for this organism being survival, growth and reproduction. In order to run these tests we first followed the standardized protocol for sediment and soil testing with C. elegans (ISO 10872, 2010). These results showed that the mixtures could be significantly different according to the type of soil (e.g. two phases system with overlaying water). Indeed, soil moistening in the standardized protocol is based on soil dry weight which means that the same amount of water is added for every soil, regardless of the water holding capacity (WHC) of the samples. Moreover, some biosolids can have a high WHC what increase humidity's differences among soil and soil mixed with biosolids when moistened as recommended by this protocol. To prevent this, improvements of the standardized protocol were developed, in which water addition is based on soil WHC instead of soil dry weight. Moreover, food volume added to the samples was lowered in order to be able to include it in the volume for soil moistening. In this context and in order to validate this new method, comparisons between standardized and optimized protocols were carried out for endpoints responses in five soils with different textures. This optimized protocol was used to assess the effect of different biosolids (limed sludge, manure ...) mixed with standard soil at different field rates and an example for a limed sludge is given. Regarding these results, improvements of the standardized protocol are adapted to assess the effects of these materials on C. elegans

Effect of non-ageing and ageing ceria nanoparticles suspensions on fresh water micro-algae

When assessing the hazards properties of nanomaterials in the environment, the main research challenges are numerous. Firstly, determining if nanomaterials are more or less toxic than the bulk forms of the same materials and the extent to which toxicity is governed or influenced by the physico-chemicals properties of the nanoparticles. Secondly, it appears necessary to study the effect of nanomaterials and nanoparticles throughout their life cycle including both initial forms and physico-chemically modified form (i.e. aggregated or agglomerated forms) resulting from an ageing process. Our work focused on the effect of commercial ceria nanoparticle (nCeO2) suspensions, towards freshwater micro-algae assessing the effect nCeO2 suspensions with different agglomeration/aggregation state obtained by using an artificial ageing process. Both ageing and non-ageing nCeO2 suspensions were fully characterized using dynamic light scattering (ZetaSizer, Malvern Instruments) or laser diffraction (MasterSizer, Malvern Instruments) and transmission electron microscopy (TEM). In addition, the interaction between NPs and algae were investigated using flow-cytometry and environmental scanning electron microscope technique (E-SEM).The results obtained showed that the algae growth inhibition was similar after exposure to non-ageing or ageing nCeO2 suspensions. The results obtained from flow-cytometry and E-SEM proved that the ceria NPs are able to tightly entrap the algae cells, which could in part contribute to the effect recorded. Those results also support the fact that aggregation or agglomeration has a few influences when focusing on the standardized algae ecotoxicity test. Moreover by comparison to our previous studies performed with other ceria suspensions, it was shown that the primary particle size and consequently the particle surface area is a relevant parameter in assessing the ecotoxicity of nanoparticles

Investigating a transcriptomic approach on marine mussel hemocytes exposed to carbon nanofibers: An in vitro/in vivo comparison

Manufactured nanomaterials are an ideal test case of the precautionary principle due to their novelty and potential environmental release. In the context of regulation, it is difficult to implement for manufactured nanomaterials as current testing paradigms identify risk late into the production process, slowing down innovation and increasing costs. One proposed concept, namely safe(r)-by-design , is to incorporate risk and hazard assessment into the design process of novel manufactured nanomaterials by identifying risks early. When investigating the manufacturing process for nanomaterials, differences between products will be very similar along key physicochemical properties and biological endpoints at the individual level may not be sensitive enough to detect differences whereas lower levels of biological organization may be able to detect these variations. In this sense, the present study used a transcriptomic approach on Mytilus edulis hemocytes following an in vitro and in vivo exposure to three carbon nanofibers created using different production methods. Integrative modeling was used to identify if gene expression could be in linked to physicochemical features. The results suggested that gene expression was more strongly associated with the carbon structure of the nanofibers than chemical purity. With respect to the in vitro/in vivo relationship, results suggested an inverse relationship in how the physicochemical impact gene expression

Adapting oecd aquatic toxicity tests for use with manufactured nanomaterials: key issues and consensus recommendations

The unique or enhanced properties of manufactured nanomaterials (MNs) suggest that their use in nanoenabled products will continue to increase. This will result in increased potential for human and environmental exposure to MNs during manufacturing, use, and disposal of nanoenabled products. Scientifically based risk assessment for MNs necessitates the development of reproducible, standardized hazard testing methods such as those provided by the Organisation of Economic Cooperation and Development (OECD). Currently, there is no comprehensive guidance on how best to address testing issues specific to MN particulate, fibrous, or colloidal properties. This paper summarizes the findings from an expert workshop convened to develop a guidance document that addresses the difficulties encountered when testing MNs using OECD aquatic and sediment test guidelines. Critical components were identified by workshop participants that require specific guidance for MN testing: preparation of dispersions, dose metrics, the importance and challenges associated with maintaining and monitoring exposure levels, and the need for reliable methods to quantify MNs in complex media. To facilitate a scientific advance in the consistency of nanoecotoxicology test results, we identify and discuss critical considerations where expert consensus recommendations were and were not achieved and provide specific research recommendations to resolve issues for which consensus was not reached. This process will enable the development of prescriptive testing guidance for MNs. Critically, we highlight the need to quantify and properly interpret and express exposure during the bioassays used to determine hazard values

Population Structure and Gene Flow of the Yellow Anaconda (Eunectes notaeus) in Northern Argentina

Yellow anacondas (Eunectes notaeus) are large, semiaquatic boid snakes found in wetland systems in South America. These snakes are commercially harvested under a sustainable management plan in Argentina, so information regarding population structuring can be helpful for determination of management units. We evaluated genetic structure and migration using partial sequences from the mitochondrial control region and mitochondrial genes cyt-b and ND4 for 183 samples collected within northern Argentina. A group of landscape features and environmental variables including several treatments of temperature and precipitation were explored as potential drivers of observed genetic patterns. We found significant population structure between most putative population comparisons and bidirectional but asymmetric migration in several cases. The configuration of rivers and wetlands was found to be significantly associated with yellow anaconda population structure (IBD), and important for gene flow, although genetic distances were not significantly correlated with the environmental variables used here. More in-depth analyses of environmental data may be needed to fully understand the importance of environmental conditions on population structure and migration. These analyses indicate that our putative populations are demographically distinct and should be treated as such in Argentina's management plan for the harvesting of yellow anacondas

Prediction of second neurological attack in patients with clinically isolated syndrome using support vector machines

The aim of this study is to predict the conversion from clinically isolated syndrome to clinically definite multiple sclerosis using support vector machines. The two groups of converters and non-converters are classified using features that were calculated from baseline data of 73 patients. The data consists of standard magnetic resonance images, binary lesion masks, and clinical and demographic information. 15 features were calculated and all combinations of them were iteratively tested for their predictive capacity using polynomial kernels and radial basis functions with leave-one-out cross-validation. The accuracy of this prediction is up to 86.4% with a sensitivity and specificity in the same range indicating that this is a feasible approach for the prediction of a second clinical attack in patients with clinically isolated syndromes, and that the chosen features are appropriate. The two features gender and location of onset lesions have been used in all feature combinations leading to a high accuracy suggesting that they are highly predictive. However, it is necessary to add supporting features to maximise the accuracy. © 2013 IEEE

Prise en compte de l’évolution de l’état d’agglomération dans l’étude de l’écotoxicité des nanoparticules

When assessing the hazardous properties of nanomaterials for the environment, there are numerous significant research challenges. Among them, it appears necessary to study the effect of nanomaterials and nanoparticles throughout their life cycle including both initial forms and physico-chemically modified forms (i. e. aggregated or agglomerated forms) that may result from an ageing process. We addressed this question by focusing on the effect of commercial ceria nanoparticle (nCeO2) suspensions on freshwater micro-algae. We assessed the effect of nCeO2 suspensions with different agglomeration/aggregation states obtained by using an artificial ageing process. It was shown that the algae growth inhibition was similar after exposure to non-degraded or degraded nCeO2 suspensions. Those results support the fact that aggregation or agglomeration has little influence when focusing on the growth of algal cells. Moreover by comparison to our previous studies performed with other ceria particle suspensions, it was shown that the primary particle size and consequently the particle surface area is a relevant parameter in assessing the ecotoxicity of nanoparticles.L’utilisation grandissante de produits contenant des nanoparticules peut entraîner une exposition des différents compartiments environnementaux. Par conséquent, l’évaluation des effets pour l’environnement de ces nanoparticules apparaît indispensable. La spécificité et la réactivité importante de ce type de composés conduit également à s’interroger sur leur comportement lors d’un relargage éventuel dans l’environnement. Cet aspect, bien que rarement renseigné, se révèle fondamental pour mieux appréhender l’écotoxicité de ces nanoparticules au cours de leur cycle de vie, incluant les formes altérées, potentiellement agrégées et/ou agglomérées. Ces questions ont été récemment abordées . Dans ce cadre, l’étude d’une suspension de nanoparticules de dioxyde de cérium (CeO2 ; taille primaire = 8 nm) possédant un coating organique a été réalisée. Cette suspension est commercialisée en tant qu’agent protecteur (anti-rayure anti-UV) à destination des peintures extérieures. Le comportement dans l’eau de cette suspension de nanoparticules avant et après un processus artificiel de vieillissement ainsi que les effets sur les producteurs primaires ont été étudiés

Devenir et impact des contaminants dans l'environnement : application à la gestion des sédiments et des boues d'épuration. Contribution à l'élaboration d'une méthodologie de caractérisation de l'écotoxicité des sédiments dulçaquicoles contaminés : intérêt de nouveaux outils écotoxicologiques. Rapport final

Au cours de ces dernières années, différentes méthodologies d’évaluation de l’écotoxicité des sédiments ont été proposées.Depuis, de nouveaux outils de caractérisation des dangers des sédiments ont été normalisés à l’ISO. C’est le cas des essais sur :- Nématodes : détermination de l'effet toxique d'échantillons de sédiment et de sol sur la croissance, la fertilité et la reproduction de Caenorhabditis elegans (NF ISO 10872 :2010) ;- Ostracodes : détermination de la toxicité des sédiments d'eau douce envers Heterocypris incongruens (NF ISO 14371:2012) ;- Bactéries : détermination de l'inhibition de l'activité déshydrogénase d'Arthrobacter globiformis - essai de contact sur solide avec le colorant redox résazurine (ISO/FDIS 18187) ;- Macrophytes : détermination de l'effet toxique des sédiments sur la croissance de Myriophyllum aquaticum (NF ISO 16191:2014).Bien que récemment standardisés, peu de travaux renseignant sur l’utilisation de ces nouveaux protocoles d’essai dans un contexte de caractérisation de l’écotoxicité des sédiments sont disponibles (Höss et al. 2010 ; Feiler et al. 2013). Le présent projet avait pour objectif d’évaluer l’applicabilité effective de ces nouveaux protocoles d’essai dans ce contexte et de juger de la pertinence et de la sensibilité de ces organismes par rapport aux essais plus classiquement réalisés (essais sur larves de chironomes notamment). La sensibilité des organismes vis-à-vis de substances chimiques de référence, la question des substrats proposés en tant que témoins ont été discutés précédemment. Les résultats de ces travaux ont fait l’objet des deux premiers rapports d’avancement de 2013 et 2014 (DRC13-136875-13811A et DRC-15-136875-04705A).Au cours de l’année 2015, les travaux portant sur l’utilisation de ces bioessais pour la caractérisation des sédiments ont été finalisés. Une dernière campagne de prélèvement de sédiments a été effectuée. L’ensemble des sédiments prélevés au cours des campagnes de 2014 et 2015 a été testé à l’aide des essais sur Caenorhabditis elegans (nématode), Heterocypris incongruens (ostracode), Myriophyllum aquaticum (macrophyte) et Arthrobacter globiformis (bactérie), mais aussi à l’aide de l’essai sur Chironomus riparius renseignant sur la mortalité, l’inhibition de la croissance et l’inhibition de l’émergence des larves de diptères. Les sédiments des campagnes de 2014 et 2015 ont également été analysés afin de les caractériser sur un plan physico-chimique (granulométrie, paramètres géochimiques, oligo-éléments) ainsi que sur leur niveau de contamination en polluants principaux (métaux et HAP, essentiellement)

Nanoparticules : influence des conditions d’essais et des paramètres physico-chimiques sur l’écotoxicité pour le milieu aquatique

Nanoparticles (NP) have been extensively applied in commercial products. It is consequently unavoidable that a large amount of NP will be discharged into environment, underlining the need to evaluate the ecotoxicological effect of NP towards aquatic organisms. However, relevant experts identified that “a large number of standardized test guidelines […] are unlikely to be applicable in their present state to engineered nanomaterials” (OECD). Indeed, the potential hazard of NP may be related to unusual physicochemical properties such as their particles size, agglomerate state in suspension and/or surface chemistry. Moreover, the method performed to disperse NP and achieve the testing suspensions can be seen as a drawback since it introduces an additional variable into the test design. Our work aims at characterizing the ecotoxicity of ceria NP toward aquatics micro-algae taking into account the influence of the dispersive methods and the physico-chemical properties of NP. The results obtained clearly show that ceria NP are ecotoxic toward micro algae compare to bulk materials. The toxic effect was principally attributing to the initial particles size and, to a lesser extent, to the agglomeration state. This suggest that exposure to NP could be expressed in surface area complementary to a mass basis. In addition, we have demonstrated that ceria NP were able to entrap and wrap the algae cells, which may contribute to the nanotoxicity.La singularité des propriétés physicochimiques des nanoparticules (NP) et le fait qu’un grand nombre de produits dits « nanostructurés » sont d’ores et déjà mis sur le marché en font un des sujets majeurs d’étude de ces dernières années. La dissémination de ces NP ou des résidus nanoparticulaires dans les différents compartiments de l’environnement est ainsi devenue une préoccupation grandissante, à l’instar de l’impact potentiel qu’ils peuvent avoir sur les organismes vivants. De nombreuses questions se posent, en effet, quant à l’applicabilité des outils actuels pour évaluer l’écotoxicité de ces produits émergents, notamment dans un cadre réglementaire. Les méthodes de préparation et les propriétés physico-chimiques des milieux d’essai (pH, force ionique) ont notamment été identifiées comme des éléments clés dans l’évaluation de l’écotoxicité des NP. Ils vont en effet conditionner certains paramètres physiques de la suspension tels que l’état d’agglomération des NP et leur stabilité, lesquels peuvent influencer leur écotoxicité. Cependant, peu de travaux concernant l’écotoxicité des NP intègrent de façon systématique l’influence des conditions d’essai, de même que l’importance des propriétés physicochimiques des NP. Ainsi, les objectifs de nos travaux sont multiples et visent à étudier l’influence de différentes méthodes de préparation des suspensions d’essai sur l’état d’agrégation de NP, et à identifier les paramètres physicochimiques (état d’agglomération, chimie de surface, taille initiale de la particule) potentiellement responsable de l’écotoxicité pour les organismes aquatiques. Nous avons pour cela entrepris l’étude de différentes NP d’oxydes de cérium en tant que particules modèles. Ces NP sont utilisées comme catalyseur dans certains moteurs diesels ou comme agent protecteur (anti-rayure, anti-UV) dans les peintures extérieures. Ils peuvent de ce fait se retrouver facilement dans l’environnement. Les effets pour le compartiment aquatique de ces NP ont été appréhendés à travers l’étude de leur écotoxicité vis-à-vis des micro-algues d’eau douce (P. subcapitata). Ces organismes, à la base des chaînes trophiques aquatiques, sont classiquement utilisés en tant qu’organismes modèles pour la détermination de l’écotoxicité des substance

What could be a relevant and cost & time effective tools-set for ecosystem risk assessment strategies ?

Socio-economic development and sustainable development politic programs are nowadays link to a sharp increase in performing ecosystems risk assessment (ERA) studies in UE countries. Some relevant methodologies have previously been proposed in order to give consensual guidances and directives in this field. Each states that, depending on the studies context, those assessments should be focused on the biological targets of one or more of the three inland environmental compartments (aquatic, terrestrial, air). This leads to usually describe ERA studies as an expensive and time consuming work, and in several cases one could question the relevancy of such extensive investigations. Many biological assessment tools from individuals bioassays to in situ studies are used in this field but few works were aimed to suggest a generic, scientifically and environmentally relevant and cost-time effective tools-set for ERA. Based on previous studies and our experience in the field of environmental risk assessment, we propose a decision tree taking into account the balance between time, cost and relevancy of various biological tools, including both ecotoxicological and ecological point of view. We also suggest that biological tools and environmental monitoring cannot be dissociated in an ERA strategies. Taking into account of this complementarity, we discuss how these tools could be selected and we finally propose an optimal base-set assessment package for ERA on the model of required data set for new existing substances in the European regulation. We think that this work is the next challenge in the way to further define consensual ERA strategies and methodologies