New protocol to affix sample for environmental and high vacuum electron microscopy.

Electron microscopy has become increasingly essential as an investigative research resource in various fields of science. Despite the evolution of the equipment and preparation techniques, little has been published on techniques for affixation and removal of samples on the stubs after analysis. This work aims to illustrate a quick and low-cost procedure for fixing and removing a sample from the support for analysis by electronic microscopy, applied to crustacean and insect species, as well as other invertebrates. It thus makes it feasible to use specimens or anatomical portions of specimens represented by a few individuals, or designated as paratypes or holotypes in a collection. RESEARCH HIGHLIGHTS: This study presents a quick and low-cost procedure for fixing and removing samples from support for electron microscopy analysis. This technique makes it possible to use individual specimens or anatomical portions represented in a collection. Fluidization of the adhesive and its complete removal allow easy recovery and repositioning of samples.

Three new records of Cladocera (Crustacea: Branchiopoda) for the State of Maranhão, Northeastern Brazil.

Brazilian biodiversity is still poorly studied, especially in the northeast region of Brazil. In the state of Maranhão, the rich fauna is still under-explored in relation to the Cladocera, and this geographic area covers four biomes, which makes it an important field of research on the Cladocera community. In this work, the record of three species is reported: Dadaya macrops Sars, 1901 first record of the genus for the state, Simocephalus serrulatus (Koch, 1841) and Simocephalus latirostris (Stingelin, 1906). The work presents brief descriptions and illustrations of the morphology of the studied species.

Monogamy in the Burrowing Shrimp Axianassa australis Rodrigues & Shimizu, 1992 (Decapoda, Gebiidea, Axianassidae).

Our knowledge of the mating systems in burrowing shrimps (infraorders Axiidea and Gebiidea) is still rather limited. Here we describe the burrow use pattern, sex ratio, and sexual dimorphism of the burrowing shrimp Axianassa australis to test for monogamy, considering that monogamous species live in heterosexual pairs and exhibit a low degree of sexual dimorphism. To this end, a total of 226 individuals of A. australis were collected from the northeast region of Brazil. Our results showed that A. australis inhabited its burrows mainly as pairs, most of which were male-female pairs. In agreement with the expectations, specimens of A. australis were found dwelling as heterosexual pairs more frequently than expected by chance alone. The presence of ovigerous females was associated with the burrow occupation; that is, brooding females were more frequently observed in male-female combinations than solitarily. Also supporting theoretical considerations, we did not observe sexual dimorphism in body size between males and females of the population and the different categories of the burrow occupation. Conversely, sexual dimorphism in cheliped size was evident in the population, with larger chelipeds in males than in females. This observation agrees with that reported for most burrowing shrimps in which male-male competition is the main evolutionary force of sexual selection. The observations above favor the hypothesis that A. australis is primarily monogamous, with a small fraction of the males moderately promiscuous.

Richness and composition of the Cladocera community (Crustacea: Branchiopoda) from the Maranho State, Northeast Brazil.

The state of Maranho, northeast Brazil, has four different biomes. The water bodies in Maranho belongs to three hydrographic regions. Thus, the state potentially contributes to a high biodiversity. Despite that, in the state, the study of cladocerans can still be considered incipient. This work aims to summarize the list of species of cladocerans in the state of Maranho, including the results of new fauna surveys. We report ten new species records for the state, two species represent the new record of the genus Pseudosida Herrick, 1884. The results found point to the occurrence of 82 valid taxa in the state of Maranho.

A predictive multimetric index based on macroinvetebrates for Atlantic Forest wadeable streams assessment / Índice Multimétrico Preditivo baseado em macroinvertebrados para avaliação de riachos vadeáveis da Mata Atlântica

Abstract: Multimetric Indices (MMIs) have been widely applied for ecological assessment in freshwater ecosystems. Most MMIs face difficulties when scaling up from small spatial scales because larger scales usually encompass great environmental variability. Covariance of anthropogenic pressures with natural environmental gradients can be a confounding factor in assessing biologic responses to anthropogenic pressures. This study presents the development and validation of a predictive multimetric index to assess the ecological condition of Atlantic Forest wadeable streams using benthic macroinvertebrates. To do so, we sampled 158 sites for the index development. We adjusted each biological metric to natural variation through multiple regression analyses (stepwise-forward) and considered that the residual distribution describes the metric variation in the absence of natural environmental influence. For metric selection we considered normal distribution, variation explained by the models, redundancy between metrics and sensitivity to differentiate reference from impaired sites. We selected five metrics to the final index: total richness, %MOLD, %Coleoptera, EPT richness and Chironomidae abundance. The residuals were transformed into probabilities and the final index was obtained through the mean of these probabilities. This index performed well in discriminating the impairment gradient and it showed a high correlation (r = 0.85, p <0.001) with a specific index developed for a particular basin indicating a similar sensitivity. This index can be used to assess wadeable streams ecological condition in Atlantic Forest biome, so we believe that this type of approach represents an important step towards the application of biomonitoring tools in Brazil.

Resumo: Índices Multimétricos (MMIs) são ferramentas amplamente aplicadas ao monitoramento ecológico de ecossistemas aquáticos continentais. A maior dificuldade na utilização dos MMIs em amplas escalas espaciais consiste no aumento da variabilidade ambiental associado ao aumento da escala. A covariância entre os impactos antropogênicos e o gradiente ambiental natural pode atuar como fator de confusão, dificultando a avaliação das respostas biológicas às pressões antropogênicas. Este estudo consiste no desenvolvimento e validação de um índice multimétrico preditivo para avaliar a condição ecológica de riachos vadeáveis da Mata Atlântica, utilizando macroinvertebrados bentônicos. Todas as métricas biológicas foram ajustadas a variação natural através da análise de regressão múltipla ("stepwise-forward") e a distribuição dos resíduos foi utilizada para descrever variação de cada métrica na ausência de gradiente ambiental natural. Para a seleção das métricas foram considerados critérios de normalidade da distribuição, variação explicada pelos modelos, redundância entre métricas e sensibilidade para diferenciar gradiente de impacto. Foram selecionadas cinco métricas para compor o índice final: riqueza total, %MOLD (Mollusca+Diptera), %Coleoptera, riqueza de EPT (Ephemeroptera+Plecoptera+Trichoptera) e abundância de Chironomidae. Os resíduos das métricas selecionadas foram transformados em probabilidades e o valor final do índice foi obtido através da média dessas probabilidades. O índice teve ótimo desempenho em discriminar o gradiente de impacto e mostrou alta correlação (r = 0.85, p <0.001) com um índice desenvolvido especificamente para uma das bacias, indicando sensibilidade similar. O índice desenvolvido nesse trabalho pode ser utilizado em todo o estado do Rio de Janeiro e acreditamos que este tipo de abordagem constitui uma contribuição importante para o desenvolvimento e implementação de um programa de monitoramento biológico no Brasil.

New records of Anthalona acuta Van Damme, Sinev & Dumont 2011 and Anthalona brandorffi (Sinev & Hollwedel, 2002) in Brazil, with description of a new species of the simplex-branch (Crustacea: Cladocera: Chydoridae).

The range of geographical distribution of Anthalona acuta Van Damme, Sinev & Dumont 2011 and Anthalona brandorffi (Sinev & Hollwedel, 2002) in Brazil has increased by almost 2000 km to the south. New records of Anthalona verrucosa verrucosa (Sars, 1901) were also added. Populations of Anthalona brandorffi from Central Brazil showed a peculiar morphological variation, with some individuals having only a single denticle on the labral keel. A new species of the simplex-branch, Anthalona neotropica sp. nov., was described based on Brazilian material, and this is the first taxon of this branch registered in the Neotropics. It differs from Anthalona simplex Van Damme, Sinev & Dumont 2011, a Central African species, in the morphology of underneath sack of the lateral head pores, length of IDL setae and armature of first flaming-torch seta of limb IV. It could be distinguished from Anthalona sanoamuangae Sinev & Kotov, 2012 (distributed through the South- East Asia) by the morphology of the main head pores, length of IDL setae and armature of the pecten of postabdominal claw. Anthalona neotropica sp. nov. seems to have a benthic/hyporheic habit. All studied species have a wide geographical distribution and could be confused with Anthalona verrucosa Sars, 1901, thus at least some if not all previous records of this species on the continent must be revised.

A Survey of Escherichia coli and Salmonella in the Hyporheic Zone of a Subtropical Stream: Their Bacteriological, Physicochemical and Environmental Relationships.

The Hyporheic Zone is among the most important interstitial freshwater habitats, but the relationship between biotic and abiotic factors in this zone remains under-explored. Enterobacteria were expected to be present, but no specific studies had ever confirmed this prediction. The aim of this study was, therefore, to evaluate the total coliforms, Escherichia coli and Salmonella spp. in hyporheic water and to determine the relationship of the physical, chemical and environmental factors at different depths in a rainforest stream. To this end, thirty-six water samples were collected at three depths in sites located in the first, second and third orders in diverse substrates. The total coliforms, Escherichia coli and Salmonella sp. were evaluated in terms of their CFU/ml. In the interstitial samples, coliforms were detected in 100% of the samples. The total coliform counts had higher values at intermediate depths, while E. coli and Salmonella spp. instead had higher values at intermediate and large depths, often reaching or exceeding the values of the surface samples. Our results revealed that Salmonella spp. and the coliforms have different microhabitat preferences. Salmonella spp. and coliform species prefer deposition areas, such as lateral sides of pools, curves and bars, but they have a tendency to distribute into different depths, likely due to temperature differences. Salmonella spp. prefer compact substrata, with fewer fluids passing through and with upwelling areas with lower oxygen inflow. The coliform species showed the opposite preference. Our results suggest that bacterial variation is related to environmental factors and physical-chemical parameters within the HZ and may play a key role in the microbial diversity and distribution in these ecosystems.

Assessing the ecological condition of streams in a southeastern Brazilian basin using a probabilistic monitoring design.

Prompt assessment and management actions are required if we are to reduce the current rapid loss of habitat and biodiversity worldwide. Statistically valid quantification of the biota and habitat condition in water bodies are prerequisites for rigorous assessment of aquatic biodiversity and habitat. We assessed the ecological condition of streams in a southeastern Brazilian basin. We quantified the percentage of stream length in good, fair, and poor ecological condition according to benthic macroinvertebrate assemblage. We assessed the risk of finding degraded ecological condition associated with degraded aquatic riparian physical habitat condition, watershed condition, and water quality. We describe field sampling and implementation issues encountered in our survey and discuss design options to remedy them. Survey sample sites were selected using a spatially balanced, stratified random design, which enabled us to put confidence bounds on the ecological condition estimates derived from the stream survey. The benthic condition index indicated that 62 % of stream length in the basin was in poor ecological condition, and 13 % of stream length was in fair condition. The risk of finding degraded biological condition when the riparian vegetation and forests in upstream catchments were degraded was 2.5 and 4 times higher, compared to streams rated as good for the same stressors. We demonstrated that the GRTS statistical sampling method can be used routinely in Brazilian rain forests and other South American regions with similar conditions. This survey establishes an initial baseline for monitoring the condition and trends of streams in the region.

Determining subsampling effort for the development of a rapid bioassessment protocol using benthic macroinvertebrates in streams of Southeastern Brazil.

The purpose of this study was to establish a subsampling procedure for benthic macroinvertebrates to aid in the development of a multimetric index to assess the biological condition of streams. Data from six streams that are considered minimally impaired were used. Subsampling was done using a device divided into 24 quadrats. The sediment from each quadrat was sorted, and all organisms were removed and identified. Richness metrics were the most affected by subsample size. Relative-abundance metrics were the most stable, proving that the sample was well distributed throughout the tray and abundance proportions were maintained. The results showed that the macroinvertebrate assemblage present in the six quadrats was similar to that present in the full sample. These analyses indicated that six quadrats, randomly designated, with a minimum of 200 collected specimens could be used in place of the full sample. In routine water management, managing time and costs are a major challenge; therefore, this type of simplification is absolutely necessary to ensure that a biomonitoring tool is useful for practical applications.

Biomonitoramento das águas: estratégias para prática de ensino / Biomonitoring of water: practical strategies for teaching

O estudo, realizado no período 2007 a 2010, teve como objetivo desenvolver e avaliar recursos didáticos para atividades de ensino voltadas a formação de recursos humanos na área de biomonitoramento das águas para estudantes de cursos de graduação em biologia. Três foram os objetivos específicos: elaborar e desenvolver um livro técnico específico para a realidade brasileira, constituído por um manual de identificação de macroinvertebrados aquáticos do estado do Rio de Janeiro a partir do estudo de material didático existente e utilizado em outras localidades geográficas; idealizar e desenvolver um sistema de conservação para a realização de uma coleção didática específica, a partir de estudo sobre técnicas de conservação e problemas específicos de utilização; avaliar a aplicabilidade do livro técnico produzido em atividade de ensino. A investigação, de tipo teórico empírica, justificou-se pela ausência de material didático específico para área de ensino do biomonitoramento no Brasil e pela quase inexistente bibliografia relativa a sistemas de avaliação deste tipo de recurso didático. Para realização do livro técnico, utilizamos a transposição didática para a mediação de diferentes contextos culturais. Para a avaliação do material didático produzido utilizamos de maneira conjunta a metodologia qualitativa e a quantitativa. Do ponto de vista dos usuários buscamos entender dificuldades, exigências e desejos de todos os possíveis atores. Para a análise qualitativa utilizamos a análise do conteúdo. Para a análise quantitativa usamos o teste t de Student e a análise da variância (ANOVA) que foram considerados significativos por p-valor menor de 0,05. O processo de avaliação, realizado com estudantes de biologia, pesquisadores e profissionais da área de ensino evidenciou um alto grau de satisfação, mostrou que atende necessidades e desejos dos usuários, proporcionou detectar melhorias a serem aportadas e indicou que o material didático é adequado para os futuros programas de formação a ser implantados no Brasil. Os processos utilizados no presente estudo utilizados para realizar e avaliar o material didático serviram, no futuro, como base para o desenvolvimento de novo e mais adequado material técnico e didático uma vez que no Brasil sejam decididas as estratégias de aplicação do biomonitoramento das águas.

[Use of bioindicators for assessing and monitoring pesticides contamination in streams and rivers]. / Utilização de bioindicadores na avaliação de impacto e no monitoramento da contaminação de rios e córregos por agrotóxicos.

The objective of this article is to present an analysis of the main bioindicators that are currently used to assess the environmental impact of pollution in water resources. The simple quantification of chemicals in the environment is not enough to reveal the real effects of contamination on ecosystems, making necessary the assessment of the biological effects that pollution causes at different hierarchical levels. The bioindicators used in this article on two case studies comprehend different hierarchical levels: in case study 1, three organization levels were utilized: individual, cellular and molecular, to detect the early effects of exposition to environmental pollutants in three hydrographic basins. By observing the inhibition of AChE activity in fish it was possible to assess the effects of organophosphorate and carbamate pesticides, showing the effects of agricultural activities. In case study 2, we present an assessment at the macroinvertebrate community level using the Extended Biotic Index. We discuss the advantages and limitations in the production of reliable data that could be used in the implementation of adequate actions to protect and/or recover ecosystems.

Utilização de bioindicadores na avaliação de impacto e no monitoramento da contaminação de rios e córregos por agrotóxicos / Use of bioindicators for assessing and monitoring pesticides contamination in streams and rivers

Este artigo tem como objetivo apresentar uma análise dos principais bioindicadores utilizados para avaliação do impacto ambiental em recursos hídricos, já que a simples mensuração dos níveis de substâncias químicas presentes no ambiente não é suficiente para revelar os reais efeitos adversos da contaminação, tornando-se necessário a avaliação dos efeitos biológicos da contaminação em diversos níveis hierárquicos. Os bioindicadores foram tratados neste artigo através de dois estudos de casos que abrangem diferentes níveis; no caso 1, foram utilizados três níveis de organização: individual, celular e molecular, para detecção precoce dos efeitos reais da exposição de peixes aos poluentes ambientais em três bacias hidrográficas. A inibição da atividade da AchE em peixes possibilitou a avaliação dos efeitos dos agrotóxicos organofosforados e carbamatos, evidenciando o efeito dos cultivos agrícolas; no caso 2, apresentamos uma avaliação no nível da comunidade de macroinvertebrados, empregando o índice Biótico Estendido. Discutimos, as vantagens e limitações na produção de dados confiáveis que possibilitem a implementação de medidas adequadas para o diagnóstico em diferentes escalas, visando a proteção e/ou recuperação dos ecossistemas.

The objective of this article is to present an analysis of the main bioindicators that are currently used to assess the environmental impact of pollution in water resources. The simple quantification of chemicals in the environment is not enough to reveal the real effects of contamination on ecosystems, making necessary the assessment of the biological effects that pollution causes at different hierarchical levels. The bioindicators used in this article on two case studies comprehend different hierarchical levels: in case study 1, three organization levels were utilized: individual, cellular and molecular, to detect the early effects of exposition to environmental pollutants in three hydrographic basins. By observing the inhibition of AChE activity in fish it was possible to assess the effects of organophosphorate and carbamate pesticides, showing the effects of agricultural activities. In case study 2, we present an assessment at the macroinvertebrate community level using the Extended Biotic Index. We discuss the advantages and limitations in the production of reliable data that could be used in the implementation of adequate actions to protect and/or recover ecosystems.