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By definition, it is accepted worldwide that environmental risk assessment is a scientific practice that identifies and evaluates the likelihood of an activity or substance to threaten the environment in general, and/or living organisms, natural habitats and ecosystems, in particular. In other words, it is a term used to describe the overall process or method allowing (a) the identification of hazards and risk factors that have the potential to cause harm (hazard identification) and (b) the analysis and evaluation of the risk associated with that particular hazard (risk analysis and risk evaluation). It also suggests all the appropriate ways to eliminate or minimize the hazard, or control the risk when the hazard cannot be eliminated (risk control).

The growing impact of anthropogenic activities generates huge quantities of toxic residues that can have direct or indirect detrimental effects on the quality of our environment. The increase in their jeopardizing effects can exert short-, medium- and long-term consequences, affecting not only human health but also environmental health in general. Furthermore, these disturbances are usually capable, in most cases, of reducing the biodiversity of native flora and fauna, which will, in turn, encourage the resistance and emergence of new pests and diseases. This book, Bird and Reptile Species in Environmental Risk Assessment Strategies, intends to provide an outline, relevant examples and stimulate practical discussions on the use of these non-conventional groups of vertebrates, bird and reptile species, within the scientific challenges faced by the Ecotoxicology and Risk Assessment academic world. It includes relevant tools which may be of use in the analysis prior to the implementation of decisions leading to actions which will hopefully reduce environmental health risks against environmental factors. Our aim was to compile information from a diversity of sources into a single volume. The rationale is to give some real-life examples in order to widen the concept that the use of non-conventional animal models, far from being a scientific conundrum, may instead provide useful avenues to be explored and further developed as a way to provide help and real answers to some of the actual problems the whole planet is dealing with currently. Concomitantly, these concrete examples extend concepts of hazardous factors to living species worldwide that may stimulate new research ideas and trends in their respective important fields.

Imazethapyr is an herbicide that is used in a variety of crops worldwide, including soybean and corn. The aim of the present study was to evaluate the biomarkers responses of adult Leptodactylus latinasus exposed to the formulation Pivot® H (10.59% imazethapyr) in the laboratory at concentrations and under conditions that simulate two potential field exposure scenarios: an immersion in field runoff (Scenario 1: 10 mg/L) and a direct exposure to the droplets emitted by spray noozles (Scenario 2: 1000 mg/L). In both scenarios, the experimental procedure involved completely immersing the frogs over a period of 15 s. Different endpoints were evaluated at several ecotoxicological levels 48 and 96 h after the herbicide exposure. These included individual (biometric indices and behavior alterations), histological (liver pigments and lesions), biochemical (catalase, glutathione system and cholinesterase activities) and genotoxic effects (micronuclei induction and nuclear abnormalities). Forty-eight hours after imazethapyr exposure, frogs submitted to Scenario 1 presented an inhibition of liver glutathione-S-transferase activity, whereas histological alterations and increased hepatic cholinesterase levels were observed in frogs exposed under Scenario 2. Ninety-six hours after exposure to the imazethapyr formulation, frogs from the Scenario 1 treatment presented a decrease in liver melanin and hemosiderin, increased hepatic catalase activity and micronuclei induction. For their part, frogs exposed to Scenario 2 presented a decrease in the hepatosomatic index, an increase in liver alterations, melanin reduction and micronuclei induction. The multivariate analysis enables correlations to be made between biomarkers of different organizational level in exposed anurans. Our result indicates that real exposure to imazethapyr formulations under field conditions may pose a risk to Leptodactylus latinasus populations living in the agroecosystems., Facultad de Ciencias Naturales y Museo

Environmental risk assessment is a scientific practice that identifies and evaluates the likelihood of an activity or substance to threaten the environment in general, and/or living organisms, natural habitats and ecosystems, in particular. It is a term used to describe the overall process or method allowing (a) the identification of hazards and risk factors that have the potential to cause harm and (b) the analysis and evaluation of the risk associated with that particular hazard. It also suggests all the appropriate ways to eliminate or minimize the hazard, or control the risk when the hazard cannot be eliminated. This book covers many non-traditional marsupial and placental mammalian species employed to analyse the jeopardising effects of environmental pollutants, as well as examples of applications of models in integrated assessment solutions. We hope that this book will provide answers or at least provide some clues to the inherent potential risk of the presence of these xenobiotics in our environment. This book aims to shed some light on the matter, whilst offering relevant tools for evaluating risk and providing a framework of practical discussions. These will foster decisions and actions required to reduce environmental health risks against environmental factors. This book presents some real-life examples, extending concepts (of hazardous factors) to living species that may stimulate new research ideas and trends in the relevant fields.

The growing impact of anthropogenic activities generates huge quantities of toxic residues that can have direct or indirect detrimental effects upon the quality of our environment. The increase in their jeopardizing effects can exert short-, medium- and long-term consequences, affecting not only human health but also environmental health in general. Furthermore, these disturbances are usually capable, in most cases, of reducing the biodiversity of native flora and fauna, which will, in turn, encourage the resistance and emergence of new pests and diseases. This book intends to provide an outline, relevant examples and stimulate practical discussions on the use of this non-conventional group of vertebrates. The book includes relevant tools which may be of use in the analysis prior to the implementation of decisions leading to actions which will hopefully reduce environmental health risks against environmental factors. Our aim was to compile information from a diversity of sources into a single volume. The rationale is to give some real-life examples in order to widen the concept that the use of non-conventional animal models, far from being a scientific conundrum, may instead provide useful avenues to be explored and further developed as a way to provide help and real answers to some of the actual problems the whole planet is dealing with currently. Concomitantly, these concrete examples extend concepts of hazardous factors to living species worldwide that may stimulate new research ideas and trends in their respective important fields.

Acute genotoxicity of commercial glyphosate (GLY) (Credit®)-, 2,4-D-acid (2,4-D) (Dedalo Elite)-, 2,4-D-amine (2,4-D DMA) (Weedar Full®)- and 2,4-D-ester (2,4-D BE) (Herbifen Super®)-based herbicide formulations alone and their combinations were analysed in Cnesterodon decemmaculatus. Mortality was evaluated as a lethal end-point and the single cell gel electrophoresis (SCGE) bioassay was used as a sublethal end-point. LC50₉₆ₕ values for Dedalo Elite was 0.46 mg/L and Herbifen Super® was 2.67 mg/L based on 2,4-D and 2,4-D BE, respectively. Results reveal a higher toxicity exerted on C. decemmaculatus after exposure to 2,4-D- rather than 2,4-D BE-based herbicide formulations. Overall, results demonstrated an enhancement in the genetic damage index committed to an enhancement of damaged erythrocytes of C. decemmaculatus when exposed to Credit®, Dedalo Elite, Weedar Full® and Herbifen Super® at 5% and 10% of LC50₉₆ₕ values alone as well as in their combinations. Overall, the combination of GLY plus 2,4-D or GLY plus 2,4-D DMA showed a synergistic pattern whereas the combination of GLY plus 2,4-D BE was antagonic. Furthermore, this research is pioneer in the assessment of lethality and genotoxicity induced by 2,4-D-, 2,4-D DMA- and 2,4-D BE-based formulations when combined with GLY-based formulated herbicides in fish after they are acutely exposed., Facultad de Ciencias Naturales y Museo

We analyzed the acute toxicity of the 48% glyphosate (GLY)-based Credit®, the 57.71% dicamba (DIC)-based Kamba®, and the 83.5% 2,4-dichlorophenoxyacetic acid (2,4-D)-based Weedar® Full, alone and as mixtures on the fish Cnesterodon decemmaculatus. Mortality revealed the LC50 96h values of 91.73 mg L−1 (range: 86.80-98.00 mg L−1), 1401.57 mg L−1 (range: 1243.78-1527.35) and 678.04 mg L−1 (range: 639.35-718.04 mg L−1) for GLY, DIC and 2,4-D, respectively. Mean values for the toxic unit (TU) that induced 50% mortality (TU50 96h) of fish exposed to equitoxic mixtures were 1.67 (range: 1.65-1.69) for Credit® and Kamba® and 1.28 (range: 1.20-1.36) for Credit® and Weedar® Full suggesting that both mixtures are antagonic. Non-equitoxic combinations demonstrated an antagonistic interaction of herbicides Credit® and Kamba®, whereas a synergistic effect was observed for Credit® and Weedar® Full formulations. GLY and DIC as a mixture demonstrated lower toxicity on non-target species compared to GLY and 2,4-D in combination, at least for C. decemmaculatus, leading to the conclusion that the former combination could be strongly recommended in further agricultural practices. Equitoxic and non-equitoxic mixtures of glyphosate with auxinic herbicides evidence opposite effects in C. decemmaculatus. Fil: Ruiz de Arcaute, Celeste. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata; Argentina. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. Cátedra de Citología; Argentina Fil: Soloneski, Sonia Maria Elsa. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata; Argentina. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. Cátedra de Citología; Argentina Fil: Larramendy, Marcelo Luis. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata; Argentina. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. Cátedra de Citología; Argentina

Dicamba (DIC) and 2,4-dichlorophenoxyacetic acid (2,4-D) are two of the most applied auxinic herbicides worldwide, both individually and as part of a mixture. However, the toxicity and interactions achieved when applied as a mixture have not yet been characterised. The equitoxic and non-equitoxic acute toxicity exerted by binary mixtures of Banvel® (57.71% DIC) and DMA® (58.4% 2,4-D) on the Neotropical fish Cnesterodon decemmaculatus were evaluated. Results revealed mean values of 1.02 (range, 0.96–1.08) for the toxic unit (TU) that induced 50% mortality (TU50 96 h) to the fish exposed to binary equitoxic mixtures of the commercial formulations Banvel®–DMA®. These results suggest that the mixture is nearly concentration additive. Furthermore, results demonstrated the occurrence of synergistic interaction when non-equitoxic combinations of Banvel®-or DMA®-formulated herbicides were assayed. In this context and regardless of their concentrations, either Banvel®- or DMA®-induced toxicity were synergised by the presence of the counterpart within mixtures. The present study represents the first evidence of the lethality exerted by mixtures of two auxinic herbicides—namely, DIC and 2,4-D—reported to date for fish and other biotic matrices. When C. decemmaculatus is used as the target organism, a synergistic pattern is observed following exposure to a mixture of both herbicides. Synergic effects of equitoxic and non-equitoxic mixtures of auxinic herbicides in C. decemmaculatus. Fil: Ruiz de Arcaute, Celeste. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. Cátedra de Citología; Argentina Fil: Soloneski, Sonia Maria Elsa. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. Cátedra de Citología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Larramendy, Marcelo Luis. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. Cátedra de Citología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina

A combined approach employing alkaline single cell gel electrophoresis (SCGE) and cytokinesis-blocked micronucleus (MNs) cytome bioassays was adopted to assess the deleterious properties of the auxinic 2,4-dichlorophenoxyacetic acid (2,4-D) and its microparticulated low volatility product Dedalo Elite (30% a.i.) on Chinese hamster ovary (CHO-K1) cells. Cytotoxicity was estimated by neutral red uptake (NRU), succinic dehydrogenase activity (MTT) and apoptosis assessment. Both compounds were assayed at 0.1–10 μg/ml concentration range. Whereas exposed CHO-K1 cells revealed a statistically significant enhancement of MNs when 10 μg 2,4-D/ml was assayed, MNs were only achieved in cells treated with 2 μg Dedalo Elite/ml. A diminution in the nuclear division index was only achieved after exposure to Dedalo Elite within the 1–10 μg/ml concentration range. Whereas increased genetic damage index was achieved when 6 and 10 μg 2,4-D/ml were assayed, GDI induction was observed in treatments employing 4 μg Dedalo Elite/ml. Both compounds induced cytotoxicity by inhibition of both lysosomal and MTT activities by enhancing the frequencies of early and late apoptotic cells. Our results not only indicate the genotoxic and cytotoxic potential of 2,4-D and its microparticulated marketplace formulation, but also highlight the risk of these agrochemicals present towards the biota and human health.

Glyphosate (GLY)-dicamba (DIC) and GLY-flurochloridone (FLC) are herbicide mixtures which are widely used for treating fallow containing glyphosate resistant weeds. The aim of this study was to evaluate the acute toxic effects and the prevailing interactions on stage 36 tadpoles of the anuran species Rhinella arenarum when exposed to equitoxic and non-equitoxic combinations of these herbicide combinations. Experiments were realized using the following combinations of commercial formulations: 48% GLY-based Credit® + 57.71% DIC-based Banvel® and 48% GLY-based Credit® + 25% FLC-based Twin Pack Gold®. GLY-DIC and GLY-FLC equitoxic mixtures were assayed mixing each constituent with an equivalent individual toxicity able to induce the same lethality effect. After 96 h of exposure, GLY-DIC and GLY-FLC equitoxic mixtures presented toxic unit 50 values (TU50 96h) of 1.74 (confidence interval: 1.58–1.92) and 1.54 (confidence interval: 1.46–1.62) respectively, indicating the presence of a weak antagonistic interaction as TU values were greater than 1. For their part, most non-equitoxic combinations of GLY-DIC and GLY-FLC tested did not significantly differ from additivity, the only exception being when DIC and FLC were fixed at 0.33 TUs, where a weak antagonism was observed. Overall, results indicate that the toxicity of both GLY-DIC and GLY-FLC mixtures to R. arenarum tadpoles vary from additive to slightly antagonistic, depending on the proportion of constituting herbicide formulations present in the mixture.

Pesticides might increase the production of reactive oxygen species (ROS). Dicamba (DIC) and 2,4-dichlorophenoxyacetic acid (2,4-D) are auxinic herbicides commonly applied in agroecosystems to control unwanted weeds. We analysed the oxidative damage exerted on the fish Cnesterodon decemmaculatus by an acute exposure to DIC- and 2,4-D-based herbicides formulations Banvel® and DMA®, respectively. The Endo III- and Fpg-modified alkaline comet assay was employed for detecting DNA damage caused by oxidative stress, whereas enzymatic and non-enzymatic biomarkers such as the activities of catalase (CAT), glutathione-S-transferase (GST), acetylcholinesterase (AChE), and glutathione content (GSH) were used to assess antioxidant response to these two herbicides. At the DNA level, results demonstrate that both auxinic herbicides induce oxidative damage at purines level. An increase on CAT and GST activities were detected in 48 h- and 96 h-treated specimens with both auxinics. GSH content decreased in fish exposed to DIC during 48 h and to 2,4-D after 96 h of exposure. Additionally, a diminished AChE activity in specimens treated with DIC and 2,4-D was observed only after 96 h. Total protein content decreased in fish exposed to both auxinics during 96 h. These results represent the first evaluation of oxidative damage related to DIC and 2,4-D exposure on a fish species as the Neotropical freshwater teleost C. decemmaculatus., Facultad de Ciencias Naturales y Museo

Glyphosate (GLY) and imazethapyr (IMZT) are two herbicides commonly used worldwide, either alone or in mixtures. They represent key pesticides in modern agricultural management. The toxicity that results when employed as mixtures has not been characterized so far. Acute toxicity of the 48% GLY-based herbicide (GBH) Credit® and the 10.59% IMZT-based herbicide (IBH) Pivot® H alone and their binary combinations was analyzed in Rhinella arenarum tadpoles exposed in a semi-static renewal test. Lethal effects were determined using mortality as the end-point, whereas sublethal effects were determined employing the single-cell gel electrophoresis (SCGE) bioassay. Based on mortality experiments, results revealed LC5096 h values of 78.18 mg/L GBH and 0.99 mg/L IBH for Credit® and Pivot® H, respectively. An increase in the genetic damage index (GDI) was found after exposure to Credit® or Pivot® H at 5 and 10% of LC5096 h values. The combinations of 5% Credit®-5% Pivot® H LC5096 h and 10% Credit®-10% Pivot® H LC5096 h concentrations significantly enhanced the GDI in comparison with tadpoles exposed only to Credit® or Pivot® H. Thus, the effect of interaction between GBH and IBH inducing DNA damage in R. arenarum blood cells can be considered to be synergistic., Facultad de Ciencias Naturales y Museo

Cytotoxic and genotoxic effects of flurochloridone (FLC) and its formulations Twin Pack Gold® and Rainbow® were evaluated in CHO-K1 cells. Using the alkaline single-cell gel electrophoresis (SCGE) assay, we observed that FLC (15 μg/ml), Twin Pack Gold® or Rainbow® induced primary DNA damage, increasing the frequency of damaged nucleoids. Vitamin E pretreatment did not modify the effect. Decreased cell viability was observed only in Twin Pack Gold®-treated cultures and was significantly ameliorated by vitamin E. Post-treatment of herbicide-damaged CHO-K1 cells with the enzymes Endo III or Fpg did not increase FLC-, Twin Pack Gold®-, or Rainbow®-induced DNA damage. These results demonstrate that neither FLC nor FLC-based formulations induce DNA damage through hydroxyl radical or lipid alkoxyl radical production, and that the induced DNA lesions were not related to oxidative damage at the purine/pyrimidine level. Our observations strongly suggest that the cytotoxic effects observed after Twin Pack Gold® exposure are due to the excipients contained within the technical formulation rather than FLC itself. Fil: Soloneski, Sonia Maria Elsa. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. Cátedra de Citología; Argentina Fil: Nikoloff, Noelia. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. Cátedra de Citología; Argentina Fil: Larramendy, Marcelo Luis. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. Cátedra de Citología; Argentina

The organophosphorus herbicide glyphosate (GLY) and the organophosphate insecticide chlorpyriphos (CPF) are key pesticides in modern management cultures worldwide. Sublethal toxicity of the commercial herbicide formulation Roundup® and the insecticide formulation Terfos® were evaluated on Eisenia fetida coelomocytes exposed under in vivo and ex vivo laboratory conditions. Induction of DNA single-strand breaks evaluated by the single cell gel electrophoresis assay and coelomocyte viability as well as alterations in coelomocyte trophic indexes were employed as endpoints for genotoxicity and cytotoxicity, respectively. Specimens were exposed at concentrations corresponding to recommended pesticide field application rate, and endpoints were evaluated after 7 and 14 days of treatment (in vivo exposure). In addition, coelomocytes were exposed to aqueous leachate of pesticide-contaminated soils during 1 h (ex vivo exposure). Earthworms exposed to Roundup® and Terfos® showed an increased frequency of DNA damage. Also, a decrease of coelomocyte viability and decrease of trophic indexes were observed in all treatments. The results demonstrate that either GLY- and CPF-based formulations exerted genotoxic as well as cytotoxic effects in coelomocytes of E. fetida exposed in vivo and ex vivo. Fil: Curieses, Silvana Patricia. Universidad Nacional de Luján. Departamento de Ciencias Básicas. Programa de Investigación en Ecotoxicología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Sáenz, María Elena. Universidad Nacional de Luján. Departamento de Ciencias Básicas. Programa de Investigación en Ecotoxicología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Alberdi, José Luis. Universidad Nacional de Luján. Departamento de Ciencias Básicas. Programa de Investigación en Ecotoxicología; Argentina Fil: Martinez, Ricardo Santiago. Universidad Nacional de Luján. Departamento de Ciencias Básicas. Programa de Investigación en Ecotoxicología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Larramendy, Marcelo Luis. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata; Argentina Fil: Di Marzio, Walter Dario. Universidad Nacional de Luján. Departamento de Ciencias Básicas. Programa de Investigación en Ecotoxicología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina

Long-term genotoxic effects of two auxinic herbicide formulations, namely, the 58.4% 2,4-dichlorophenoxyacetic acid (2,4-D)-based DMA® and the 57.7% dicamba (DIC)-based Banvel® were evaluated on Cnesterodon decemmaculatus. Primary DNA lesions were analyzed by the single-cell gel electrophoresis methodology. Two sublethal concentrations were tested for each herbicide corresponding to 2.5% and 5% of the LC5096h values. Accordingly, fish were exposed to 25.2 and 50.4 mg/L or 41 and 82 mg/L for 2,4-D and DIC, respectively. Fish were continuously exposed for 28 days with replacement of test solutions every 3 days. Genotoxicity was evaluated in ten individuals from each experimental point at the beginning of the exposure period (0 day) and at 7, 14, 21 and 28 days thereafter. Results demonstrated for first time that 2,4-D-based formulation DMA® induced primary DNA strand breaks after 7–28 days exposure on C. decemmaculatus regardless its concentration. On the other hand, DIC-based formulation Banvel® exerted its genotoxic effect after exposure during 7–14 days and 7 days of 2.5 and 5% LC5096h, respectively. The present study represents the first evidence of primary DNA lesions induced by two widely employed auxinic herbicides on C. decemmaculatus, namely 2,4-D and DIC, following long-term exposure. 2,4-D and dicamba exert primary DNA lesions in blood cells of C. decemmaculatus after long-term exposure. Fil: Ruiz de Arcaute, Celeste. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Facultad de Ciencias Naturales y Museo, Universidad Nacional de la Plata; Argentina Fil: Larramendy, Marcelo Luis. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Facultad de Ciencias Naturales y Museo, Universidad Nacional de la Plata; Argentina Fil: Soloneski, Sonia Maria Elsa. Facultad de Ciencias Naturales y Museo, Universidad Nacional de la Plata; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina

In the present study, the damage recovery capabilities of Boana pulchella tadpoles after acute exposure (96 h) to 0.39 mg/L concentration of the imazethapyr (IMZT)-based herbicide formulation Pivot® H (25% IMZT LC50 value) were assessed during a period of 7 to −21 days. To appraise the recovery capabilities, frequency of micronuclei (MNs), other nuclear abnormalities and DNA single-strand breaks evaluated by single cell gel electrophoresis assay on circulating blood cells were employed as endpoints for genotoxicity. Growth, development, body mass, and morphological abnormalities were also employed as individual endpoints in the recovery assay. Results demonstrated that IMZT induced sublethal effects at both the individual (i.e., loss of keratodonts) and cytogenetic levels (e.g., increase of MN frequency, other nuclear abnormalities and DNA single-strand breaks). At 11 days of the exposure phase, tadpoles recovered their basal levels of frequency of MNs, other nuclear abnormalities, and comets. However, loss of keratodonts, observed at the end of the exposure period, was present up to 21 days thereafter. Finally, axial abnormalities and delay in development stage were observed only during the postexposure phase in IMZT-exposed tadpoles at 18 and 25 days, respectively and were observed until the end of the experiment. This is the first evidence of use the comet assay as cytogenetic biomarker of genotoxicity in evaluating the recovery capabilities of amphibians in general and also those of B. pulchella after exposure to IMZT., Facultad de Ciencias Naturales y Museo, Facultad de Ciencias Exactas, Centro de Investigaciones del Medioambiente

Acute lethal and sublethal toxicity of the pirimicarb-based commercial formulation Aficida® were evaluated on Boana pulchella tadpoles. Whereas mortality was used as end point for lethality, frequency of micronuclei and other nuclear abnormalities as well as alterations in the frequency of erythroblasts in circulating blood as biomarkers for genotoxicity and cytotoxicity, respectively. Swimming, growth, developmental and morphological abnormalities were also employed as sublethal end points. Results show that the species is within the 13th percentile of the distribution of acute sensitivity of species to pirimicarb for aquatic vertebrates. Results revealed values of 23.78 and 101.45 mg/L pirimicarb as LC5096 h for GS25 and GS36 tadpoles, respectively. The most evident effects were related with the swimming activity with NOEC and LOEC values within the 0.005–0.39 mg/L pirimicarb concentration range. Aficida® induced DNA damage at the chromosomal level by increasing micronuclei frequency and other nuclear abnormalities, i.e., lobbed and notched nuclei and binucleated cells. Cellular cytotoxicity was found after Aficida® treatment. The presence of abdominal oedemas in exposed organisms and thus flotation response of organisms could be proposed as a new sensitive exposure parameter. The multiple end point assessment approach used allowed a complete understanding the multi level of effects occurring by exposure to pirimicarb, at least in B. pulchella. Fil: Natale, Guillermo Sebastian. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata; Argentina. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Departamento de Química. Centro de Investigaciones del Medio Ambiente; Argentina Fil: Vera Candioti, Josefina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata; Argentina. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. Cátedra de Citología; Argentina Fil: Ruiz de Arcaute, Celeste. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata; Argentina. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. Cátedra de Citología; Argentina Fil: Soloneski, Sonia Maria Elsa. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata; Argentina. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. Cátedra de Citología; Argentina Fil: Larramendy, Marcelo Luis. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata; Argentina. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. Cátedra de Citología; Argentina Fil: Ronco, Alicia Estela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata; Argentina. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Departamento de Química. Centro de Investigaciones del Medio Ambiente; Argentina

In the present study, the damage recovery capabilities of Boana pulchella tadpoles after acute exposure (96h) to 0.39mg/L concentration of the imazethapyr (IMZT)-based herbicide formulation Pivot

We evaluated the role of oxidative stress in the genotoxic damage induced by imazethapyr (IMZT) and its formulation Pivot® in mammalian CHO-K1 cell line. Using the alkaline comet assay, we observed that a concentration of 0.1 μg/mL of IMZT or Pivot® was able to induce DNA damage by increasing the frequency of damaged nucleoids. To test whether the DNA lesions were caused by oxidative stress, the DNA repair enzymes endonuclease III (Endo III) and formamidopyrimidine-DNA glycosylase (Fpg), which convert base damage to strand breaks, were used. Our results demonstrate that after treatment of CHO-K1 cells with the pure active ingredient as well as the commercial formulation Pivot®, an increase in DNA strand breaks was observed after incubation of both Endo III and Fpg enzymes, indicating that both compounds induce DNA damage involving both pyrimidine and purine-based oxidations, at least in CHO-K1 cells. Our findings confirm the genotoxic potential of IMZT and suggest that this herbicide formulation must be employed with great caution, especially not only for exposed occupational workers but also for other living species., Facultad de Ciencias Naturales y Museo

Acute toxicity and genotoxicity of the dicamba-based commercial herbicide formulation Banvel® were evaluated on Cnesterodon decemmaculatus (Pisces, Poeciliidae) exposed under laboratory conditions. A lethal effect was used as the end point for mortality, whereas frequency of micronuclei (MNs) and DNA single-strand breaks evaluated by the single cell gel electrophoresis assay were employed as end points for genotoxicity. Mortality studies revealed an LC50 96 h value of 1639 mg/L (range, 1471–1808) of dicamba. Furthermore, behavioral changes, e.g., gathering at the bottom of the aquarium, slowness in motion, abnormal swimming, and slow reaction, were observed. Whereas increased frequency of MNs was observed when 1229 mg/L dicamba was assayed for 48 h, no induction of MNs was observed in fish exposed to the herbicide for 96 h, regardless of the concentration of dicamba. Furthermore, other nuclear abnormalities, i.e., binucleated cells and lobed and notched nuclei, were induced in fish exposed for 48 h but not 96 h. Increase in the genetic damage index was observed in those treatments (lasting for both 48 and 96 h) within the 410–1229 mg/L dicamba concentration-range. This study represents the first evidence of acute lethal and sublethal effects exerted by dicamba on a piscine species native to Argentina. The results could indicate that dicamba-based formulation Banvel® is the less toxic emerging pollutant reported so far for C. decemmaculatus. Finally, our findings highlight the properties of this herbicide that jeopardize nontarget living species exposed to this agrochemical. Fil: Ruiz de Arcaute, Celeste. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. Cátedra de Citología; Argentina Fil: Soloneski, Sonia Maria Elsa. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. Cátedra de Citología; Argentina Fil: Larramendy, Marcelo Luis. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. Cátedra de Citología; Argentina

The in vitro effects of S-metolachlor and its formulation Twin Pack Gold® (96% a.i.) were evaluated in human hepatoma (HepG2) cells. Cytokinesis-blocked micronucleus cytome (CBMN-cyt) and MTT assays as well as Neutral Red uptake were employed for genotoxicity and cytotoxicity evaluation. Activities were tested within the concentration range of 0.25–15 μg/ml S-metolachlor for 24 h of exposure. Both compounds rendered a minor reduction in the NDI although not reaching statistical significance. Results demonstrated that the S-metolachlor was not able to induce MNs. On the other hand, 0.5–6 μg/ml Twin Pack Gold® increased the frequency of MNs. When cytotoxicity was estimated, S-metolachlor was not able to induce either a reduction of lysosomal or mitochondrial activity. Contrarily, whereas 1–15 μg/ml Twin Pack Gold® induced a significant reduction of mitochondrial activity, all tested concentrations of the formulated product induced a significant decrease of lysosomal performance as a function of the concentration of the S-metolachlor-based formulation titrated into cultures. Genotoxicity and cytotoxicity differences obtained with pure S-metolachlor and the commercial S-metolachlor-based formulation indicate that the latter may contain additional unsafe xenobiotics and support the concept of the importance of evaluating not only the active principle but also the commercial formulation when estimating the real hazard from agrochemicals. Fil: Nikoloff, Noelia. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. Cátedra de Citología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Escobar, Luciana Ines. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. Cátedra de Citología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Soloneski, Sonia Maria Elsa. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. Cátedra de Citología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Larramendy, Marcelo Luis. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. Cátedra de Citología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina

The ability of two 48 percent chlorpyrifos-based insecticides (Lorsban* 48E® and CPF Zamba®), two 50 percent pirimicarb-based insecticides (Aficida® and Patton Flow®), and two 48 percent glyphosate-based herbicides (Panzer® and Credit®) to induce DNA single-strand breaks in peripheral blood erythrocytes of Cnesterodon decemmaculatus (Jenyns, 1842) (Pisces, Poeciliidae) exposed under laboratory conditions was evaluated by the single-cell gel electrophoresis (SCGE) assay. In those fish exposed to Lorsban* 48E®, CPF Zamba®, Aficida®, Patton Flow®, Credit®, and Panzer®, a significant increase of the genetic damage was observed for all formulations regardless of the harvesting time. This genotoxic effect was achieved by an enhancement of Type II-IV comets and a concomitant decrease of Type 0-I comets over control values. A regression analysis revealed that the damage varied as a negative function of the exposure time in those Lorsban* 48E®- and Aficida®-treated fish. On the other hand, a positive correlation between damage increase and exposure time was achieved after Patton Flow® and Credit® treatment. Finally, no correlation was observed between increase in the genetic damage and exposure time after treatment with CPF Zamba® or Panzer®. These results highlight that all agrochemicals inflict primary genotoxic damage at the DNA level at sublethal concentrations, regardless of the exposure time of the aquatic organisms under study, at least within a period of 96 h of treatment. Fil: Vera Candioti, Josefina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Santa Fe; Argentina. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. Catedra de Citologia; Argentina Fil: Soloneski, Sonia Maria Elsa. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Cientifico Tecnológico la Plata; Argentina. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. Catedra de Citologia; Argentina Fil: Larramendy, Marcelo Luis. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Cientifico Tecnológico la Plata; Argentina. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. Catedra de Citologia; Argentina

Fil: Vera Candioti, Josefina. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. Catedra de Citologia; Argentina. Consejo Nacional de Investigaciones Cientificas y Tecnicas; Argentina

Mortality, genotoxicity, and cytotoxicity of the 48% glyphosate-based formulations Panzer and Credit ® were evaluated on Cnesterodon decemmaculatus (Jenyns, 1842) (Pisces, Poeciliidae) under laboratory conditions. Induction of micronuclei (MN) and alterations in the erythrocytes:erythroblasts ratio were employed as end points for genotoxicity and cytotoxicity, respectively. For Panzer ® , mean values of 16.70 and 15.68 mg/L were determined for LC 50 at 24 and 96 h, respectively, and these concentrations reached mean values of 98.50 and 91.73 mg/L for Credit ® . LC 50 values decreased as a negative linear function of Panzer ® exposure time within the 0–96 h period, but not for Credit ® . LC 50 values indicated that the fish were more sensitive to Panzer ® than to Credit ® . Both 3.9 and 7.8 mg/L of Panzer ® increased MN frequency at 48 and 96 h of treatment. When fish were exposed to Credit ® , an increased frequency of MN over control values was found after 96 h for all concentrations assayed, but not after 48 h. No cellular cytotoxicity was found after Panzer ® and Credit ® treatment, regardless of both the concentration and the sampling time. Furthermore, our results demonstrated that Panzer ® and Credit ® should be considered as glyphosate-based commercial formulations with genotoxic but not cytotoxic effect properties.

Ecotoxicology and Genotoxicology: Non-traditional Aquatic Models intends to stimulate practical discussions on the use of non-conventional biotic matrices to address scientific challenges faced in these fields. It provides real-life examples to convey that the use of a non-conventional animal model, far from being a scientific conundrum, may instead provide real answers to some of our planet's problems. In this epilogue, we summarise the book and we hope that the importance of the status of the environment using autochthonous, non-target species, which are truly exposed to locally used xenobiotic agents, is realised.

Imazethapyr (IMZT) is a selective postemergent herbicide with residual action. Available data analyzing its effects in aquatic vertebrates are scarce. In previous studies, we demonstrated that IMZT induces lesions into the DNA of Hypsiboas pulchellus tadpoles using the single-cell gel electrophoresis (SCGE) assay as a biomarker for genotoxicity. Currently, this assay can be modified by including incubation with lesion-specific endonucleases, e.g., endonuclease III (Endo III) and formamidopyrimidine-DNA glycosylase (Fpg), which detect oxidized pyrimidine and purine bases, respectively. The aim of this study was to evaluate the role of oxidative stress in the genotoxic damage in circulating blood cells of H. pulchellus tadpoles exposed to the IMZT-based Pivot H® formulation (10.59% IMZT) at a concentration equivalent to 25% of the LC50 (96 h) value (0.39 mg/L IMZT) during 48 and 96 h. Our results demonstrate that the herbicide induces oxidative DNA damage on H. pulchellus tadpoles at purines bases but not at pyrimidines. Our findings represent the first evidence of oxidative damage caused by IMZT on anuran DNA using the alkaline restriction enzyme-modified SCGE assay., Centro de Investigaciones del Medioambiente, Facultad de Ciencias Naturales y Museo

Fil: Larramendy, Marcelo Luis. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. Catedra de Citologia; Argentina. Consejo Nacional de Investigaciones Cientificas y Tecnicas. Centro Cientifico Tecnologico Conicet - La Plata; Argentina

Amphibians as animal models have long been employed as indicators of environmental quality and to detect toxicity risks of environmental pollutants. The increased attention to the use of non-traditional species of amphibians in ecotoxicology and genotoxicology research lies in their ability not only to reveal the toxic and genotoxic effects of many potential environmental xenobiotics, but also to help researchers understand the behaviour of real ecosystems. This chapter presents an overview of selected research that has led to the use of the common tree frog Hypsiboas pulchellus as a reliable and valid model in in vivo and in situ studies of aquatic pollution.