REMOVAL OF AMMONIACAL NITROGEN FROM MUNICIPAL LANDFILL LEACHATE WITH FLOATING Typha domingensis (TYPHACEAE) / Eliminación del nitrógeno amoniacal del lixiviado de vertedero municipal con Typha domingensis (TYPHACEAE) en flotación

ABSTRACT A promising method for the treatment of effluents is the use of floating macrophytes. Ammoniacal nitrogen is a typical compound present in urban landfill leachates and its removal is important due its toxicity to several organisms. Therefore, the study evaluated Typha domingensis survival and nitrification potential artificially floating in domestic solid waste leachate. Plants were exposed for 35 days to leachate (100, 75 and 50 %) and to rainwater with N:P:K (control). Dissolved oxygen (DO) of the treatments was periodically measured, and ammoniacal nitrogen, nitrite and nitrate were analyzed before and after exposure. At the end of the experiment, plant survival rate was calculated. After two weeks, DO increased twice in the control, three times in 50 % leachate, four times in 75 % leachate, and eight times in 100 % leachate. At the end of the experiment, ammoniacal nitrogen was no longer detected, there was a significant reduction of nitrite, and a significant increase of nitrate in the treatments containing leachate. Plant survival was higher in those individuals exposed to 100 % leachate and decreased at lower leachate concentrations: 98 %, 94 %, 92 %, and 86 %. The study demonstrated that the ammoniacal nitrogen concentration of the leachate was not toxic to T. domingensis, and that it was efficient in the removal of this compound from the effluent, indicating that the species may be used artificially floating for the removal of this contaminant from domestic solid waste landfill leachate when in low concentrations.

RESUMEN Un método prometedor para el tratamiento de efluentes es el uso de macrófitas flotantes. El nitrógeno amoniacal es un componente típico de lixiviados de vertederos urbanos y su remoción es importante debido a su toxicidad para diversos organismos. Por eso, el estudio evaluó la supervivencia y el potencial de nitrificación de Typha domingensis flotando artificialmente en lixiviados de residuos sólidos domésticos. Las plantas fueron expuestas por 35 días a lixiviado (100, 75 y 50 %) y al agua de lluvia con N:P:K (control). El oxígeno disuelto (OD) fue medido periódicamente; el nitrógeno amoniacal, el nitrito y el nitrato fueron analizados antes y después de la exposición; al final del experimento, la tasa de supervivencia de las plantas fue calculada. Después de dos semanas, el OD aumentó dos veces en el control, tres veces en el lixiviado 50 %, cuatro veces en el lixiviado 75 % y ocho veces en el lixiviado 100 %. Al final del experimento, el nitrógeno amoniacal no fue más detectado, hubo una reducción significativa de nitrito y un aumento significativo de el nitrato en los tratamientos que contenían lixiviado. La supervivencia de las plantas fue mayor en aquellos individuos expuestos al lixiviado 100 % y se redujo en las menores concentraciones: 98 %, 94 %, 92 % y 86 %. El estudio demostró que la concentración de nitrógeno ammoniacal del lixiviado no fue tóxica para T. domingensis y que ésta fue eficiente en la remoción del compuesto del efluente, indicando que la especie puede ser utilizada flotando artificialmente para la remoción del contaminante de lixiviados de residuos sólidos domésticos en bajas concentraciones.

Germination and gametophyte development of Cyathea corcovadensis (Raddi) Domin (Cyatheaceae) from spores stored at low temperatures / Germinação e desenvolvimento gametofítico de Cyathea corcovadensis (Raddi) Domin (Cyatheaceae) com esporos armazenados em baixas temperaturas

The tree fern Cyathea corcovadensis (Raddi) Domin is an endangered species in the state of Rio Grande do Sul, Brazil. It currently occurs only in the northern segment of the coastal region. Spore storage would help in conservation programs since it maintains genetic variability and provides material for in vitro cultures. Current study evaluates the effect of low temperatures combined to different spore storage times on the germination and initial gametophyte development of C. corcovadensis. Spores were divided into two groups: spores of the first group were sowed immediately in Meyer culture medium with nystatin, at pH 4.0, while spores of the second group were stored at 7, -20 and -196°C during 60, 120, 180, and 365 days and then sowed in the same medium. Spore storage at 7 and -196°C for 365 days not only provided higher germination percentages than those reported for recently-collected spores but also stimulated gametophytic development. The latter was demonstrated by the higher percentages of laminar gametophytes in these treatments. The possibility of storing spores provides material for in vitro experiments, which is of special interest for C. corcovadensis due to its ornamental potential and conservation status.

A samambaia arborescente Cyathea corcovadensis (Raddi) Domin está em perigo de extinção no Estado do Rio Grande do Sul, Brasil, ocorrendo somente no norte da região costeira. O armazenamento de esporos pode auxiliar programas de conservação, uma vez que oferece a possibilidade de manutenção da variabilidade genética e fornece material para culturas in vitro. O objetivo deste estudo foi avaliar o efeito de temperaturas baixas combinadas com diferentes tempos de armazenamento de esporos sobre a germinação e o desenvolvimento inicial de gametófitos de C. corcovadensis. Os esporos foram divididos em dois grupos: os do primeiro grupo foram semeados imediatamente em meio de cultura Meyer com nistatina, com pH 4,0 e os do segundo grupo foram armazenados a 7, -20 e -196°C durante 60, 120, 180 e 365 dias e então semeados no meio supra mencionado. O armazenamento de esporos a 7 e -196°C durante 365 dias propiciou porcentagens de germinação superiores àquelas observadas para esporos recém coletados e também estimulou o desenvolvimento gametofítico, o que foi demonstrado pelos elevados valores de porcentagem de gametófitos laminares observados nestes tratamentos. A possibilidade de armazenar esporos oferece material para experimentos in vitro, o que é de especial interesse para C. corcovadensis, considerando seu potencial ornamental e estado de conservação.

In vitro propagation of Cyathea atrovirens (Cyatheaceae): spore storage and sterilization conditions / Propagación in vitro de Cyathea atrovirens (Cyatheaceae): almacenamiento de esporas y condiciones de esterilización

Cyathea atrovirens occurs in a wide range of habitats in Brazil, Paraguay, Uruguay and Argentina. In the Brazilian State of Rio Grande do Sul, this commonly found species is a target of intense exploitation, because of its ornamental characteristics. The in vitro cultura is an important tool for propagation which may contribute toward the reduction of extractivism. However, exogenous contamination of spores is an obstacle for the success of aseptic long-term cultures. This study evaluated the influence of different sterilization methods combined with storage conditions on the contamination of the in vitro cultures and the gametophytic development of C. atrovirens, in order to establish an efficient propagation protocol. Spores were obtained from plants collected in Novo Hamburgo, State of Rio Grande do Sul, Brazil. In the first experiment, spores stored at 7oC were surface sterilized with 0.5, 0.8 and 2% of sodium hypochlorite (NaClO) for 15 minutes and sown in Meyer’s culture medium. The cultures were maintained in a growth room at 26±1ºC for a 12-h photoperiod and photon flux density of 100μmol/m²/s provided by cool white fluorescent light. Contamination was assessed at 60 days, and gametophytic development was scored at 30, 60, 120 and 130 days of in vitro culture, analyzing 300 individuals for each treatment. There was no significant difference in culture contamination among the different sodium hypochlorite concentrations tested, and all treatments allowed for the development of cordiform gametophytes at 130 days of culture. In the second experiment, spores stored at 7 and -20°C were divided into two groups. Half of the spores were surface sterilized with 2% of NaClO for 15 minutes and the other half was not sterilized. All spores were sown in Meyer’s medium supplemented with one of the following antibiotics: nystatin, Micostatin® and actidione. The culture conditions and the procedures used for evaluating contamination and gametophytic development were the same described for the first experiment. No contamination was observed in spores stored at -20°C and treated with NaClO and actidione. In all treatments, cordiform gametophytes presenting antheridia were observed at 120 days. The percentages of these gametophytes increased from 120 to 130 days and no significant differences were observed among treatments. Archegonia were observed on cordiform gametophytes at 130 days. The findings provide data relevant to in vitro propagation procedures of this species, which may increase the availability of plants for ornamental purposes, therefore contributing to the reduction of the exploitation of endangered tree ferns species. Rev. Biol. Trop. 62 (1): 299-308. Epub 2014 March 01.

Cyathea atrovirens (Langsd. & Fisch.) Domin (Cyatheaceae) se presenta en una amplia gama de hábitats en Brasil, Paraguay, Uruguay y Argentina. Debido a sus características ornamentales, la especie es objeto de intensa explotación. El cultivo in vitro es una herramienta importante para la propagación lo que puede contribuir a la reducción del impacto de las actividades extractivas. Sin embargo, la contaminación exógena de esporas es un obstáculo para el éxito de cultivos asépticos a largo plazo. Este estudio evaluó la influencia de diferentes métodos de esterilización en combinación con las condiciones de almacenamiento sobre la contaminación de los cultivos in vitro y el desarrollo gametofítico de C. atrovirens. En el primer experimento, las esporas almacenadas a 7°C se esterilizaron superficialmente con 0.5, 0.8 y 2% de hipoclorito de sodio (NaClO) durante 15 minutos y se sembraron en medio de cultivo de Meyer. Aunque no hubo diferencia en la contaminación de lós cultivos entre las concentraciones de hipoclorito de sodio de las diferentes pruebas, en el tratamiento con 2% NaClO se observó un mayor porcentaje de gametofitos cordiformes a los 130 días. En el segundo experimento, las esporas almacenadas a 7 y -20°C fueron divididas en dos grupos. La mitad de las esporas se esterilizaron con 2% de NaClO durante 15 minutos y la otra mitad no fue esterilizada. Todas las esporas se sembraron en medio de Meyer suplementado con uno de los siguientes antibióticos: nistatina, Micostatin® o actidiona. No se observó contaminación de las esporas almacenadas a -20°C y tratadas con NaClO y actidiona. En todos los tratamientos, se observaron gametofitos cordiformes con anteridios y arquegonios. Los resultados proporcionan datos relevantes para la propagación in vitro de C. atrovirens, que pueden aumentar la disponibilidad de las plantas para fines ornamentales, contribuyendo así a la reducción de la exploración de las especies de helechos arborescentes en peligro de extinción.

Leaf dimorphism of Microgramma squamulosa (Polypodiaceae): a qualitative and quantitative analysis focusing on adaptations to epiphytism

The epiphytic fern Microgramma squamulosa occurs in the Neotropics and shows dimorphic sterile and fertile leaves. The present study aimed to describe and compare qualitatively and quantitatively macroscopic and microscopic structural characteristics of the dimorphic leaves of M. squamulosa, to point more precisely those characteristics which may contribute to epiphytic adaptations. In June 2009, six isolated host trees covered by M. squamulosa were selected close to the edge of a semi-deciduous seasonal forest fragment in the municipality of Novo Hamburgo, State of Rio Grande do Sul, Brazil. Macroscopic and microscopic analyzes were performed from 192 samples for each leaf type, and permanent and semi-permanent slides were prepared. Sections were observed under light microscopy using image capture software to produce illustrations and scales, as well as to perform quantitative analyses. Fertile and sterile leaves had no qualitative structural differences, being hypostomatous and presenting uniseriate epidermis, homogeneous chlorenchyma, amphicribal vascular bundle, and hypodermis. The presence of hypodermal tissue and the occurrence of stomata at the abaxial face are typical characteristics of xeromorphic leaves. Sterile leaves showed significantly larger areas (14.80cm²), higher sclerophylly index (0.13g/cm²) and higher stomatal density (27.75stomata/mm²) than fertile leaves. The higher sclerophylly index and the higher stomatal density observed in sterile leaves are features that make these leaves more xeromorphic, enhancing their efficiency to deal with limited water availability in the epiphytic environment, compared to fertile leaves.

El helecho epífito Microgramma squamulosa se encuentra en el Neotrópico y tiene hojas estériles y fértiles dimorfas. El objetivo de este estudio fue describir y comparar cuantitativa y cualitativamente la organización estructural de las hojas de la M. squamulosa, investigando las características morfológicas y anatómicas, y señalando los factores que contribuyen a la adaptación al ambiente epífito. Los análisis macroscópicos y microscópicos se realizaron a partir de 192 muestras de hojas de cada tipo. Láminas permanentes y semi-permanentes fueron preparadas y las secciones fueron observadas en el microscopio de luz. Hojas fértiles y estériles no mostraron diferencias estructurales cualitativas, son hipostomáticas, presentan epidermis uniseriada, haz vascular anficrival e hipodermis. La presencia de hipodermis y aparatos estomáticos en la nsuperficie abaxial de la epidermis son típicas de hojas xeromórficas. Hojas estériles presentaran áreas significativamente mayores, tasa más alta de esclerofilia y mayor densidad de estomas que hojas fértiles. La mayor tasa de esclerofilia y mayor densidad de estomas son características que hacen las hojas estériles más xeromórficas, aumentando su eficiencia para hacer frente a la disponibilidad de agua limitada en el ambiente epífito, en comparación con las hojas fértiles.

Genotoxicity biomonitoring of sewage in two municipal wastewater treatment plants using the Tradescantia pallida var. purpurea bioassay

The genotoxicity of untreated and treated sewage from two municipal wastewater treatment plants (WTP BN and WTP SJN) in the municipality of Porto Alegre, in the southern Brazilian state of Rio Grande do Sul, was evaluated over a one-year period using the Tradescantia pallida var. purpurea (Trad-MCN) bioassay. Inflorescences of T. pallida var. purpurea were exposed to sewage samples in February (summer), April (autumn), July (winter) and October (spring) 2009, and the micronuclei (MCN) frequencies were estimated in each period. The high genotoxicity of untreated sewage from WTP BN in February and April was not observed in treated sewage, indicating the efficiency of treatment at this WTP. However, untreated and treated sewage samples from WTP SJN had high MCN frequencies, except in October, when rainfall may have been responsible for reducing these frequencies at both WTPs. Physicochemical analyses of sewage from both WTPs indicated elevated concentrations of organic matter that were higher at WTP SJN than at WTP BN. Chromium was detected in untreated and treated sewage from WTP SJN, but not in treated sewage from WTP BN. Lead was found in all untreated sewage samples from WTP SJN, but only in the summer and autumn at WTP BN. These results indicate that the short-term Trad-MCN genotoxicity assay may be useful for regular monitoring of municipal WTPs.

In vitro culture of Vriesea gigantea and Vriesea philippocoburgii: two vulnerable bromeliads native to Southern Brazil

A importância das bromélias como plantas ornamentais e o fato de várias espécies nativas do Rio Grande do Sul estarem ameaçadas de extinção leva ao interesse pela sua micropropagação. Uma cultura in vitro de Vriesea gigantea e Vriesea philippocoburgii foi estabelecida a partir de sementes. A taxa de germinação de V. gigantea foi superior à taxa de germinação de V. philippocoburgii. Plântulas de V. philippocoburgii deram origem a vários brotos adventíceos quando cultivadas em meio basal de Knudson. Por outro lado, para V. gigantea, foi necessária uma maior concentração de sais, sendo que o número de brotos foi aumentado com o meio Murashige and Skoog. A adição de carvão ativado e ácido naftalenoacético ao meio de regeneração permitiu o desenvolvimento de brotos e a formação raízes, confirmando o sucesso da cultura in vitro. A expressão diferencial dos genótipos justifica a necessidade de continuadas pesquisas para que se estabeleçam as condições ideais para os tecidos específicos.