Estudos com oligoquetas aquáticos aplicados à ecotoxicologia são quase inexistentes no Brasil, embora diversas agências reguladoras já tenham demonstrado interesse na inclusão da espécie Branchiura sowerbyi (Oligochaeta, Naididae) em protocolos de monitoramento de ambientes aquáticos tropicais. Com o objetivo de ampliar o conhecimento sobre o potencial de utilização desta espécie em ensaios ecotoxicológicos, efetuou-se ampla revisão bibliográfica entre 1950 e 2012, verificando-se discrepâncias metodológicas, o uso quase exclusivo da matriz água e a inserção da turfa como fonte de matéria orgânica nos testes com sedimento artificial, recomendado como forma de reduzir a diferença entre os diversos tipos de sedimento avaliados. A partir desta revisão, bioensaios de toxicidade com sedimento natural e artificial (SAT, com a fibra de coco em substituição à turfa) foram realizados para avaliar a sobrevivência, crescimento e reprodução de B. sowerbyi. Posteriormente, testes de toxicidade aguda e crônica (incluindo a bioacumulação) foram realizados com B. sowerbyi eTubifex tubifex (espécie com uso padronizado) para avaliar os efeitos dos metais arsênio e zinco. Pelos resultados obtidos verifica-se que a reprodução e taxa de crescimento de B. sowerbyi, quando mantida em SAT (número de casulo: 2 ± 0,01; taxa de crescimento diário: - 0,44 ± 0,24%), foram inferiores (p < 0,05) em relação ao sedimento natural (número de casulos: 19,00 ± 6,24; taxa de crescimento diário: 0,35 ± 0,24%), demonstrando que o sedimento natural é mais favorável ao cultivo da espécie. Nos testes com metais, B. sowerbyi aparentou ser mais sensível ao arsênio e ao zinco, do que a espécie Tubifex tubifex, com valores de CL50 e CE50 inferiores aos observados para T. tubifex (CL50(96h): 297 ± 24,50 &#956;molAs·L-1 e 14,89 ± 1,01 &#956;molZn·L-1 para B. sowerbyi, contra > 1577,43 &#956;molAs·L-1 e 132,51 ± 12,95 &#956;molZn·L-1 para T. tubifex e CL50(14d): 4,28 ± 0,03 e 9,72 ± 0,38 &#956;molAs·g-1 para B. sowerbyi e T. tubifex, respectivamente). A partir da análise das metodologias utilizadas para as duas espécies e daquelas sugeridas em trabalhos publicados com B. sowerbyi, elaborou-se um protocolo para realização de bioensaios com a espécie B. sowerbyi em condições tropicais. / Aquatic oligochaetes studies applied to ecotoxicology are rare in Brazil, although several regulatory agencies had shown interest in the inclusion of the species B. sowerbyi (Oligochaeta, Naididae) in the tropical aquatic ecosystems monitoring. Aiming to increase the knowledge about the potential use of this species on ecotoxicological bioassays, an extensive literature review from 1950 to 2012 was performed, verifying methodological discrepancies; the majority use of the water sample and the insertion of peat as a source of organic matter on the bioassays with artificial sediment, which it is recommended to reduce the difference between the sediment samples that were collected from different places. From this review, toxicity bioassays with natural and artificial sediments (SAT, with coir instead of peat) were made to evaluate the survival, growth and reproduction of B. sowerbyi. After that, acute and chronic bioassays were made with B. sowerbyi and Tubifex tubifex (a standardized species) to evaluate the effects of arsenic and zinc. The results showed that the reproduction and growth rate of B. sowerbyi, kept in SAT (number of cocoons: 2 ± 0.01; daily growth rate: - 0.44 ± 0.24%), were smaller (p < 0.05) compared to the natural sediment (number of cocoons: 19.00 ± 6.24; diary growth rate: 0.35 ± 0.24%), showing that the natural sediment is more favorable to the cultivation of this species. In the bioassays with metals, B. sowerbyi appeared to be more sensible to arsenic and zinc than T. tubifex (CL50(96h): 297 ± 24.50 &#956;molAs·L-1 and 14.89 ± 1.01 &#956;molZn·L-1 to B. sowerbyi, against > 1577.43 &#956;molAs·L-1 and 132.51 ± 12.95 &#956;molZn·L-1 to T. tubifex and CL50(14d): 4.28 ± 0.03 and 9.72 ± 0.38 &#956;molAs·g-1 to B. sowerbyi and T. tubifex, respectively). From the analysis of the methodological approaches from the two species and from those suggested by other studies with B. sowerbyi, a bioassay protocol for the species B. sowerbyi was elaborated.

Estudos com oligoquetas aquáticos aplicados à ecotoxicologia são quase inexistentes no Brasil, embora diversas agências reguladoras já tenham demonstrado interesse na inclusão da espécie Branchiura sowerbyi (Oligochaeta, Naididae) em protocolos de monitoramento de ambientes aquáticos tropicais. Com o objetivo de ampliar o conhecimento sobre o potencial de utilização desta espécie em ensaios ecotoxicológicos, efetuou-se ampla revisão bibliográfica entre 1950 e 2012, verificando-se discrepâncias metodológicas, o uso quase exclusivo da matriz água e a inserção da turfa como fonte de matéria orgânica nos testes com sedimento artificial, recomendado como forma de reduzir a diferença entre os diversos tipos de sedimento avaliados. A partir desta revisão, bioensaios de toxicidade com sedimento natural e artificial (SAT, com a fibra de coco em substituição à turfa) foram realizados para avaliar a sobrevivência, crescimento e reprodução de B. sowerbyi. Posteriormente, testes de toxicidade aguda e crônica (incluindo a bioacumulação) foram realizados com B. sowerbyi eTubifex tubifex (espécie com uso padronizado) para avaliar os efeitos dos metais arsênio e zinco. Pelos resultados obtidos verifica-se que a reprodução e taxa de crescimento de B. sowerbyi, quando mantida em SAT (número de casulo: 2 ± 0,01; taxa de crescimento diário: - 0,44 ± 0,24%), foram inferiores (p < 0,05) em relação ao sedimento natural (número de casulos: 19,00 ± 6,24; taxa de crescimento diário: 0,35 ± 0,24%), demonstrando que o sedimento natural é mais favorável ao cultivo da espécie. Nos testes com metais, B. sowerbyi aparentou ser mais sensível ao arsênio e ao zinco, do que a espécie Tubifex tubifex, com valores de CL50 e CE50 inferiores aos observados para T. tubifex (CL50(96h): 297 ± 24,50 &#956;molAs·L-1 e 14,89 ± 1,01 &#956;molZn·L-1 para B. sowerbyi, contra > 1577,43 &#956;molAs·L-1 e 132,51 ± 12,95 &#956;molZn·L-1 para T. tubifex e CL50(14d): 4,28 ± 0,03 e 9,72 ± 0,38 &#956;molAs·g-1 para B. sowerbyi e T. tubifex, respectivamente). A partir da análise das metodologias utilizadas para as duas espécies e daquelas sugeridas em trabalhos publicados com B. sowerbyi, elaborou-se um protocolo para realização de bioensaios com a espécie B. sowerbyi em condições tropicais. / Aquatic oligochaetes studies applied to ecotoxicology are rare in Brazil, although several regulatory agencies had shown interest in the inclusion of the species B. sowerbyi (Oligochaeta, Naididae) in the tropical aquatic ecosystems monitoring. Aiming to increase the knowledge about the potential use of this species on ecotoxicological bioassays, an extensive literature review from 1950 to 2012 was performed, verifying methodological discrepancies; the majority use of the water sample and the insertion of peat as a source of organic matter on the bioassays with artificial sediment, which it is recommended to reduce the difference between the sediment samples that were collected from different places. From this review, toxicity bioassays with natural and artificial sediments (SAT, with coir instead of peat) were made to evaluate the survival, growth and reproduction of B. sowerbyi. After that, acute and chronic bioassays were made with B. sowerbyi and Tubifex tubifex (a standardized species) to evaluate the effects of arsenic and zinc. The results showed that the reproduction and growth rate of B. sowerbyi, kept in SAT (number of cocoons: 2 ± 0.01; daily growth rate: - 0.44 ± 0.24%), were smaller (p < 0.05) compared to the natural sediment (number of cocoons: 19.00 ± 6.24; diary growth rate: 0.35 ± 0.24%), showing that the natural sediment is more favorable to the cultivation of this species. In the bioassays with metals, B. sowerbyi appeared to be more sensible to arsenic and zinc than T. tubifex (CL50(96h): 297 ± 24.50 &#956;molAs·L-1 and 14.89 ± 1.01 &#956;molZn·L-1 to B. sowerbyi, against > 1577.43 &#956;molAs·L-1 and 132.51 ± 12.95 &#956;molZn·L-1 to T. tubifex and CL50(14d): 4.28 ± 0.03 and 9.72 ± 0.38 &#956;molAs·g-1 to B. sowerbyi and T. tubifex, respectively). From the analysis of the methodological approaches from the two species and from those suggested by other studies with B. sowerbyi, a bioassay protocol for the species B. sowerbyi was elaborated.

Les travaux décrits dans ce mémoire ont été conduits dans le but de tester l'utilisation potentielle de l'autotomie d'un invertébré d'eau douce Tubifex tubifex (Annélide, Oligochète) comme biomarqueur de la qualité sanitaire et environnementale des milieux aquatiques et apporter notre contribution à l'étude de la toxicité létale et sublétale de divers contaminants (le paraquat sous forme de Gramoxone® ; le fludioxonil sous forme Géoxe® ; l'isoproturon et le fenhexamide) couramment utilisés dans le vignoble champardennais. Nous avons réalisé des expérimentations au laboratoire et au niveau d'un site comportant plusieurs bassins de recueil des eaux de ruissellement du vignoble et présentant des taux de contamination différents et contrastés. Nous avons constaté une forte induction de l'autotomie pendant l'exposition aux contaminants. L'isoproturon et le Géoxe® s'accumulent de façon contrastée selon les régions morphologiques du ver. Des perturbations histopathologiques et biochimiques ont été observées chez les vers exposé aux contaminants. La sévérité des atteintes histopathologiques a été moins importante chez les vers autotomisés que les vers entiers. Les systèmes de défenses antioxydants et le métabolisme de phase II (catalase, glutathion-S-transférase) montrent des perturbations variables selon le contaminant et la région du ver sans qu'un schéma général de réponse n'apparaisse. L'approche expérimentale montre que les eaux des bassins provoquent rapidement une autotomie et un stress chez les vers. Le taux d'autotomie est corrélé avec la durée d'exposition et la concentration en toxique. De même une bonne corrélation existe « in situ » entre ce taux d'autotomie et la qualité sanitaire des bassins. Cette réponse morphologique originale constitue donc un biomarqueur potentiel de contamination et peut être compris comme une réponse adaptative du vers à la présence d'un ou plusieurs contaminants. / The work described in this thesis was conducted in order to test the potential use of autotomy of a freshwater invertebrate Tubifex tubifex (Annelida, Oligochaeta) as a biomarker of the sanitary and environnemental quality of aquatic environments and make our contribution to the study of lethal and sublethal toxicity of various contaminants (paraquat as Gramoxone® ; fludioxonil form Géoxe®; isoproturon and fenhexamid) commonly used in the vineyard Champagne-Ardenne. We conducted experiments in the laboratory and at a site with several ponds collecting vineyard runoff and presenting contrasted contamination rates. We found a significant induction of autotomy during exposure to contaminants. The isoproturon and Géoxe® accumulate in the worm with contrasted pattern according to morphological worm regions. Histopathological and biochemical disturbances were observed in worms exposed to contaminants. Rates of histopathology alterations were less important in worms autotomized than in entire worms. The antioxidant defense systems and phase II metabolism (catalase, glutathione-S-transferase) varied in different way depending on the parameter tested but not general pattern of the biochemical responses occured. The experimental approach showed that the pond water rapidly induced autotomy and stressed worms. The autotomy rate was correlated with the exposure time and toxicant concentration. Similarly a good correlation existed "in situ" between autotomy rates and the sanitary quality of ponds. Thus the autotomy is an original morphological response that might be a potential biomarker of contamination and might be understood as an adaptive response to the presence of contamination.

Le but de cette thèse était de déterminer l’impact de deux types de nanoparticules métalliques (sulfure de cadmium – CdS - et or - Au) sur deux organismes aquatiques modèles (vers tubifex et poisson zèbre Danio rerio). L’approche a consisté à doper le sédiment de la Garonne avec les deux types de nanoparticules et caractériser l’effet de cette contamination sur les deux organismes en utilisant plusieurs marqueurs à différents niveaux d'organisation biologique (réponses biochimiques, activité de neurotransmission, génotoxicité, expression génétique et activité de bioturbation). Les effets ont été évalués après exposition aux sédiments contaminés dans des systèmes expérimentaux de types microcosmes durant une vingtaine de jours. Dans un certain nombre de cas, ce travail a révélé un effet des nanoparticules qui n’était pas dû au métal mais aux propriétés nanoparticulaires. En effet, dans le cas des nanoparticules de CdS, nous avons observé des altérations du génome ainsi que la modification de l’activité de bioturbation des vers tubifex en réponse à la contamination aux nanoparticules mais pas à celle de la forme ionique. Dans ce travail nous avons pu observer le relargage des nanoparticules d’or depuis le sédiment vers la colonne d’eau, et sa biodisponibilité potentielle pour les poissons. Ces nanoparticules provoquaient des altérations de l’ADN chez les deux organismes, la modification de l’expression de certains gènes et l’augmentation de l’activité de l’acétylcholinestérase chez le poisson zèbre. Les résultats de bioturbation n’ont pas montré d’effets des nanoparticules d’or sur le mouvement des vers. / The aim of this thesis was to determine the impact of two types of metal nanoparticles (cadmium sulphide - CdS - and gold - Au) on two aquatic models (tubifex worms and zebrafish Danio rerio). The approach has been to dope the Garonne sediment with the two types of nanoparticles and characterize the effect of this contamination on the two animals using multiple markers at different levels of biological organization (biochemical responses, neurotransmission activity, genotoxicity, gene expression and bioturbation activity). The effects were evaluated after exposure to contaminated sediment in experimental microcosms systems for twenty days. In a number of cases this work revealed an effect of the nanoparticles that was not due to metal but to the nanoparticulate properties. Indeed, in the case of CdS nanoparticles, we observed alterations in the genome as well as the modification of the bioturbation activity of worms tubifex in response to the contamination of nanoparticles but not to that of the ionic form. In this work we observed the release of gold nanoparticles from the sediment into the water column, and its potential bioavailability to fish. These nanoparticles were causing DNA damage in both organisms, modifying the expression of certain genes and increased acetylcholinesterase activity in zebrafish. Bioturbation results showed no effects of gold nanoparticles on the movement of worms.

Mercury is a pervasive environmental contaminant which is globally distributed in freshwater ecosystems. In order to assess the risk that mercury and methylmercury pose to public health through consumption and trophic level transfer, it is first necessary to understand the interactions and uptake that occurs between benthic organisms and mercury impacted sediments. Delineation of these interactions currently rely on correlating measurements of bulk sediment concentrations with bioaccumulation of either total mercury or methylmercury. However, it has been proposed that porewater concentrations, rather than sediment concentrations, should be used to predict uptake and bioavailability. Diffusive gradient in thin films (DGTs) have been proposed as a viable technique for porewater measurements to assess the bioavailable fractions of mercury. DGTs were compared to traditional bulk solid sampling to assess their capabilities for the prediction of total and methylmercury bioaccumulation in benthic oligochaetes, T. tubifex. DGTs performed similarly to the bulk solids sampling approach in respect to their correlation with mercury bioaccumulation in the sediment matrix studied. Bioturbation was shown to impact redox profiles in the sediment which led to a decrease in porewater methylmercury concentrations in the uppermost surficial sediment depths. These results indicate that monitoring tools such as DGTs are necessary to better understand the fate of mercury at field scale contaminated sites. / text

TNT (2,4,6-trinitrotoluene) is a persistent contaminant at many military installations and poses a threat to aquatic ecosystems. Data from environmental fate and toxicity studies with TNT revealed that sediment toxicity test procedures required modification to accurately assess sediment TNT toxicity. Key modifications included aging TNT-spiked sediments 8-14 d, basing lethal dose on measured sediment concentrations of the molar sum of TNT and its main nitroaromatic (NA) transformation products (SNA), basing sublethal dose on average sediment SNA concentrations obtained from integration of sediment SNA transformation models, avoiding overlying water exchanges, and minimizing toxicity test durations. Solid phase microextraction fibers (SPMEs) were investigated as a biomimetic chemical measure of toxicity and bioavailability. Both organism and SPME concentrations provided measures of lethal dose independent of exposure scenario (TNT-spiked sediment or TNT-spiked water) for Tubifex tubifex. Among all benthic organisms tested (Chironomus tentans, Ceriodaphnia dubia, T. tubifex) and matrixes, median lethal dose (LC50) estimates based on SPME and organism concentrations ranged from 12.6 to 55.3 mmol SNA/ml polyacrylate and 83.4 to 172.3 nmol SNA/g tissue, ww, respectively. For Tubifex, LC50s (95% CI) based on SNA concentrations in sediment and SPMEs were 223 (209-238) nmol SNA/g, dw and 27.8 (26.0-29.8) mmol SNA/ml, respectively. Reproductive effects occurred at slightly lower exposures. Median effective dose (EC50) estimates (95% CI) for Tubifex cocoon production, based on sediment and SPME concentrations, were 118 (114-122) nmol SNA/g, dw and 21.8 (21.2-22.4) mmol SNA/ml, respectively. Bioconcentration experiments with Tubifex revealed that compound hydrophobicity predicted the toxicokinetics and bioconcentration of these compounds from water, however, there was a large discrepancy between the toxicokinetics of absorbed versus metabolically-generated aminodinitrotoluenes. A large portion of bioconcentrated, radiolabeled TNT transformation products could not be identified. In addition to their ability to provide matrix-independent measures of dose, SPME concentrations were more accurate indicators of bioavailable NAs than were sediment concentrations.