Effects of synthetic and plant-based fungicides on biomarkers in the early stages of zebrafish development
Data
2018-12-14
Autores
Título da revista
ISSN da revista
Título do Volume
Editora
Resumo
A produção agrícola é frequentemente afetada pelas condições ambientais inerentes às mudanças climatéricas. Esta situação acarreta o aumento da utilização de pesticidas, como é o exemplo de fungicidas para controlar o míldio e oídio na vitivinicultura. Neste sentido, a utilização dos fungicidas têm sido associados ao aparecimento dos seus resíduos no solo e lençóis de água com possíveis efeitos tóxicos nos ecossistemas. Contudo, o estudo e o conhecimento toxicológico acerca dos resíduos e efeitos destes compostos são limitados ou inexistentes. Assim este trabalho teve como objetivo avaliar a toxicidade de fungicidas sintéticos (azoxistrobina, tebuconazole e mancozeb) e de compostos de origem natural (extrato de Mimosa tenuiflora, extrato de Equisetum arvense e timol) recorrendo à utilização de embriões de peixe-zebra como modelo. Numa primeira abordagem procedeu-se ao cálculo dos valores das concentrações médias letais para 96h de exposição (96h-LC50) iniciada em embriões na fase de blástula (~2 horas pós-fertilização). Com base no valor de 96h-LC50, os embriões foram expostos a concentrações que variaram entre 0.001 e 0.1 mgL-1 para a azoxistrobina (LC50 = 1.15 mgL-1 ), 0.0005 e 0.05 mgL-1 para o mancozebe (LC50 = 5.13 mgL-1 ) e 0.05 e 5 mgL-1 para o tebuconazole (LC50 = 7.25 mgL-1 ). Para os compostos de origem natural, os embriões foram expostos a concentrações que variaram entre 0.00625 e 0.625 mgL-1 para o extrato aquoso de Equisetum arvense (LC50 = 435.31 mgL-1 ), 0.008 e 0.8 mgL-1 para o extrato etanólico de Mimosa tenuiflora (LC50 = 123.87 mgL-1 ) e 0.01 e 1 mgL-1 para o timol (LC50 = 32.67 mgL-1 ). Durante a exposição foram avaliados diversos parâmetros letais (mortalidade, destacamento da cauda e da cabeça), sub-letais (desenvolvimento dos somitos, olhos, otólitos, edema, pigmentação, movimentos espontâneos, sistema circulatório e eclosão), teratogénicos (malformações), bem como a análise morfométrica das larvas. Outros parâmetros bioquímicos como enzimas ligadas ao stresse oxidativo (SOD, CAT, GPx e GR), níveis de ROS, níveis de glutationas (GSH e GSSG), enzimas relativas a degradação de compostos (GST e CarE), ligadas a neurotransmissão (AcHE) e respiração anaeróbia (LDH), foram também analisados. Os embriões expostos a compostos sintéticos apresentaram uma maior percentagem de efeitos letais, sub-letais e a nível enzimático. Os embriões expostos a mancozebe apresentam um decréscimo na sua taxa de eclosão muito acentuado para todas as concentrações avaliadas, e um grande número de malformações (edemas cardíacos e do saco vitelino, bem como torções na coluna) apresentavam uma maior prevalência na concentração mais elevada, sendo assim efeitos dose-dependentes. Já os expostos a azoxistrobina na concentração mais baixa apresentam um aumento dos ROS bem como um aumento da SOD, GST, CarE e AcHE, bem como uma diminuição da atividade da CAT e dos níveis de GSH e GSSG. Não se verificaram diferenças no desenvolvimento dos embriões expostos a tebuconazole, bem como a fungicidas à base dos compostos naturais. Conclui-se assim que a presença de compostos sintéticos no ambiente pode causar alterações significativas nos ecossistemas aquáticos, podendo estes serem substituídos pelos compostos naturais. Contudo, mais estudo são necessários para comprovar na totalidade a segurança e sensibilidade dos compostos testados.
Agriculture is affected by the environmental conditions inherent in climate change. This phenomenon is associated with increased use of pesticides in order to control fungi and pests (eg, mildew and powdery mildew) that attack agricultural, especially wine production. In this sense, the use of fungicides have been associated with the appearance of their residues in the soil and water sheets with possible toxic effects on ecosystems However, the study and toxicological knowledge about the residues and effects of these compounds are limited or non-existent. The objective of this work was to evaluate the toxicity of synthetic fungicides (azoxystrobin, tebuconazole and mancozeb) and compounds of natural origin (extract of Mimosa tenuiflora, extract of Equisetum arvense and thymol) using zebrafish embryos as a toxicological model. In a first approach, we calculated the mean lethal concentration values for 96h exposure (96h-LC50) in embryos in the blastula phase (~ 2 hours post-fertilization). Based on the 96h-LC50 value, the embryos were exposed to concentrations ranging from 0.001 to 0.1 mgL-1 for azoxystrobin (LC50 = 1.15 mgL-1 ), 0.0005 and 0.05 mgL-1 for mancozeb = 5.13 mgL-1 ) and 0.05 and 5 mgL-1 for tebuconazole (LC50 = 7.25 mgL-1 ). For the compounds of natural origin, the embryos were exposed to concentrations ranging from 0.00625 to 0.625 mgL-1 for the aqueous extract of Equisetum arvense (LC50 = 435.31 mgL-1 ), 0.008 and 0.8 mgL-1 for the Mimosa tenuiflora extract (LC50 = 123.87 mgL-1 ) and 0.01 and 1 mgL-1 for thymol (LC50 = 32.67 mgL-1 ). During the exhibition, several lethal parameters (mortality, tail and head detachment), sub-lethal (development of somites, eyes, otoliths, edema, pigmentation, spontaneous movements, circulatory system and hatching), teratogenic (malformations) such as the morphometric analysis of the larvae. Other biochemical parameters such as oxidative stress-linked enzymes (SOD, CAT, GPx and GR), ROS levels, glutathione levels (GSH and GSSG), compounds degradation enzymes (GST and CarE), neurotransmission and anaerobic respiration (LDH), were also analyzed. Embryos exposed to synthetic compounds showed a higher percentage of lethal, sublethal and enzymatic effects. Embryos exposed to mancozeb showed a marked decrease in their hatching rate for all concentrations evaluated, and a large number of malformations (cardiac and yolk sac edema as well as spinal torsions) had a higher prevalence at the highest concentration, thus being dose-dependent effects. On the other hand, those exposed to azoxystrobin at the lowest concentration present an increase in ROS as well as an increase in SOD, GST, CarE and AcHE, as well as a decrease in CAT activity and GSH and GSSG levels. There were no differences in the development of embryos exposed to tebuconazole, as well as fungicides based on natural compounds. It is thus concluded that the presence of synthetic compounds in the environment can cause significant changes in aquatic ecosystems, and these can be replaced by natural compounds. However, further study is needed to fully demonstrate the safety and sensitivity of the compounds tested.
Agriculture is affected by the environmental conditions inherent in climate change. This phenomenon is associated with increased use of pesticides in order to control fungi and pests (eg, mildew and powdery mildew) that attack agricultural, especially wine production. In this sense, the use of fungicides have been associated with the appearance of their residues in the soil and water sheets with possible toxic effects on ecosystems However, the study and toxicological knowledge about the residues and effects of these compounds are limited or non-existent. The objective of this work was to evaluate the toxicity of synthetic fungicides (azoxystrobin, tebuconazole and mancozeb) and compounds of natural origin (extract of Mimosa tenuiflora, extract of Equisetum arvense and thymol) using zebrafish embryos as a toxicological model. In a first approach, we calculated the mean lethal concentration values for 96h exposure (96h-LC50) in embryos in the blastula phase (~ 2 hours post-fertilization). Based on the 96h-LC50 value, the embryos were exposed to concentrations ranging from 0.001 to 0.1 mgL-1 for azoxystrobin (LC50 = 1.15 mgL-1 ), 0.0005 and 0.05 mgL-1 for mancozeb = 5.13 mgL-1 ) and 0.05 and 5 mgL-1 for tebuconazole (LC50 = 7.25 mgL-1 ). For the compounds of natural origin, the embryos were exposed to concentrations ranging from 0.00625 to 0.625 mgL-1 for the aqueous extract of Equisetum arvense (LC50 = 435.31 mgL-1 ), 0.008 and 0.8 mgL-1 for the Mimosa tenuiflora extract (LC50 = 123.87 mgL-1 ) and 0.01 and 1 mgL-1 for thymol (LC50 = 32.67 mgL-1 ). During the exhibition, several lethal parameters (mortality, tail and head detachment), sub-lethal (development of somites, eyes, otoliths, edema, pigmentation, spontaneous movements, circulatory system and hatching), teratogenic (malformations) such as the morphometric analysis of the larvae. Other biochemical parameters such as oxidative stress-linked enzymes (SOD, CAT, GPx and GR), ROS levels, glutathione levels (GSH and GSSG), compounds degradation enzymes (GST and CarE), neurotransmission and anaerobic respiration (LDH), were also analyzed. Embryos exposed to synthetic compounds showed a higher percentage of lethal, sublethal and enzymatic effects. Embryos exposed to mancozeb showed a marked decrease in their hatching rate for all concentrations evaluated, and a large number of malformations (cardiac and yolk sac edema as well as spinal torsions) had a higher prevalence at the highest concentration, thus being dose-dependent effects. On the other hand, those exposed to azoxystrobin at the lowest concentration present an increase in ROS as well as an increase in SOD, GST, CarE and AcHE, as well as a decrease in CAT activity and GSH and GSSG levels. There were no differences in the development of embryos exposed to tebuconazole, as well as fungicides based on natural compounds. It is thus concluded that the presence of synthetic compounds in the environment can cause significant changes in aquatic ecosystems, and these can be replaced by natural compounds. However, further study is needed to fully demonstrate the safety and sensitivity of the compounds tested.
Descrição
Original Dissertation for the purpose of obtaining a Master's Degree in Biochemistry at the University of Trás-os-Montes and Alto Douro
Palavras-chave
Toxicology , Fungicides , Zebrafish , Embryonic development , Oxidative Stress