REMEDIATION OF TETRACYCLINE FROM WATER SOURCES USING VETIVER GRASS (Chrysopogon zizanioides L. Nash) AND TETRACYCLINE-TOLERANT ROOT-ASSOCIATED BACTERIA

Antibiotics are emerging contaminants worldwide. Due to insufficient policy regulations, public awareness, and the constant exposure of the environment to antibiotic sources has created a major environmental concern. Wastewater treatment plants (WWTP) are not equipped to filter-out these compounds before the discharge of the disinfected effluent into water sources (e.g., lakes and streams) and current available technologies are not equipped to remediate these compounds from environmental sources. Hence, the challenge remains to establish a biological system to remove these antibiotics from wastewater. An invitro hydroponic remediation system was developed using vetiver grass (Chrysopogon zizanioides L. Nash) to remediate tetracycline (TC) from water. Comparative metabolomics studies were conducted to investigate the metabolites/pathways associated with tetracycline metabolism in plants and TC-degrading bacteria. The results show that vetiver plants effectively uptake tetracycline from water sources. Vetiver root-associated bacteria recovered during the hydroponic remediation trial were highly tolerant to TC (as high as 600 ppm) and could use TC as a sole carbon and energy source. Growth conditions (pH, temperature, and oxygen requirement) for TC-tolerant bacteria were optimized for higher TC remediation capability from water sources. The plant (roots and shoots) and bacterial species were further characterized for the metabolites produced during the TC degradation process using GC-MS to identify the possible biochemical mechanism involved. Also, the plant root zone was screened for metabolites/enzymes that were secreted during antibiotic degradation and could potentially enhance the degradation process. The root zone was selected for this analysis because this region of the plant has shown a greater capacity for antibiotic degradation compared to the shoot zone. The role of antioxidant enzymes in TC degradation process revealed glutathione-S-transferase (GSTs) as an important group of enzymes in both plant and bacteria potentially involved in TC degradation process. Metabolomics results also suggest potential GST activity in the TC remediation/ transformation process used by plants. This information could be useful in gaining insights for the application of biological remediation systems for the mitigation of antibiotics from waste-water.

Avaliação do desenvolvimento e a eficiência do capim vetiver (chrysopogon zizanioides) em sistemas híbridos de alagados construídos

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

O uso de fosfatos associado à fitorremediação em solos de Santo Amaro (BA) contaminados por metais tóxicos

A contaminação do solo no município de Santo Amaro (BA) por metais tóxicos provocada pelas atividades da empresa Plumbum Mineração tem gerado impactos sobre a saúde ambiental e humana. O objetivo da presente pesquisa foi avaliar a viabilidade da remediação deste solo contaminado por chumbo e cádmio, utilizando diferentes fontes de fosfatos e fitorremediação com o capim vetiver [Vetiveria zizanioides (L.)]. O estudo foi realizado em colunas de PVC onde amostras de solo foram colocadas com a aplicação de fosfato dihidrogênio de potássio (KH2PO4) (T1); fertilizante fosfato natural reativo (FNR) (T2) e; uma mistura do KH2PO4 e de fertilizante FNR (T3). Amostras de solo contaminado sem tratamento (T0) foram utilizadas como controle. Após 60, 120 e 180 dias, alíquotas do solo foram retiradas das colunas para análises. Ao final de cada período, mudas de capim vetiver [(Vetiveria zizanioides (L.)] foram plantadas em vasos com as amostras de solo: T0, T1, T2 e T3 em triplicata. Para a determinação das concentrações de chumbo e cádmio no solo e tecidos vegetais foi utilizado o ICP-OES. A partir das análises física e química constatou-se que o solo possui textura argilosa e capacidade de troca catiônica (CTC) elevadas. As extrações com solução de ácido dietilenotriaminopentaacético (DTPA) e Toxicity Characteristic Leaching Procedure (TCLP) mostraram que o tratamento T1 seguido do T3 foram os mais eficientes na imobilização de Pb e Cd. Entretanto, todos os tratamentos resultaram em concentrações de metais ainda disponíveis no solo que excediam os limites estabelecidos pela USEPA, sendo o solo, portanto, considerado tóxico mesmo após o tratamento. Com base nas concentrações de metais extraídos através da extração sequencial pelo método BCR após a remediação e a fitorremediação do solo, foi verificado que todos os três tratamentos com fosfatos foram eficientes em imobilizar o Pb e Cd nas formas menos solúveis, porém, o Cd permaneceu mais solúvel e com maior mobilidade do que o Pb. Os ensaios de letalidade utilizando minhoca Eisenia andrei mostraram que a mortalidade observada no solo após 60 dias de tratamento foi significativamente reduzida após 120 e 180 dias de tratamento. A perda de biomassa pelas minhocas também foi reduzida de acordo com o tempo de tratamento. O teste de germinação com alfaces (Lactuca sativa L.) indicou que as amostras de solo tratadas continuam bastante tóxicas, apesar da disponibilidade reduzida do Pb e do Cd como visto nos resultados da extração por TCLP e por BCR. A avaliação de risco ecológico potencial indicou que os tratamentos do solo com fosfatos associado à fitorremediação reduziram a mobilidade do Pb, principalmente nos tratamentos T1 e T3. Para o Cd o risco ecológico potencial aumenta consideravelmente quando comparado com o Pb demonstrando que esse elemento, apesar dos tratamentos com fosfatos mais a fitorremediação continua móvel. O tratamento com KH2PO4 (T1) foi o mais eficiente na redução da mobilidade, disponibilidade e da toxicidade dos metais, seguido pelo T3 e T2 para o Pb e o T3 seguido pelo T1 e T2 para o Cd.

Avaliação de um sistema descentralizado de tratamento de esgotos domésticos em escala real composto por tanque séptico e wetland construída híbrida

A concentração demográfica e de sistemas coletivos de esgotamento sanitário dentro do perímetro urbano da maioria dos municípios brasileiros, dificulta a viabilização do fornecimento de serviços de tratamento de esgotos domésticos à habitações e núcleos habitacionais isolados situados em áreas periurbanas e rurais, intensificando os danos provocados pela poluição de origem antrópica ao meio ambiente e à preservação da saúde pública. Para contribuir no equacionamento deste problema, o presente estudo teve por finalidade avaliar uma wetland construída híbrida em escala real, composta por uma unidade com fluxo contínuo subsuperficial vertical seguida por uma unidade de fluxo contínuo subsuperficial horizontal, cultivadas com capim Vetiver, para o tratamento de efluente proveniente de tanque séptico. A presente configuração experimental busca uma solução de baixo custo e simplificada para o tratamento descentralizado de esgotos domésticos. A estação experimental de tratamento de esgotos, parte integrante e um dos produtos da Rede Nacional de Tratamento de Esgotos Descentralizados RENTED, foi construída no Centro Tecnológico de Hidráulica CTH / Escola Politécnica EPUSP, campus Butantã da USP, em São Paulo. O esgoto bruto foi proveniente do Conjunto Residencial da USP e do restaurante central da Cidade Universitária. As vazões média e máxima de esgoto bruto, respectivamente, de 640L.d-1 e 1600L.d-1, foram aplicadas à entrada do TS de 5.000L e deste escoaram por gravidade à entrada da wetland construída híbrida, com TDH total de 2,8d e 1,1d, respectivamente, sob aplicação das vazões média e máxima de projeto. O monitoramento do experimento em campo, incluindo o período inicial de partida, durou 6 meses consecutivos. Foram avaliados os parâmetros físico-químicos e microbiológicos do esgoto bruto e do efluente do tanque séptico e das câmaras da wetland construída híbrida durante 97 dias consecutivos. Os resultados indicaram que tanto as mudas jovens quanto as adultas de capim Vetiver adaptaram-se bem às condições ambientais. As eficiências médias de remoção no efluente tratado final quanto à matéria orgânica carbonácea foram de 96 por cento para DBO5,20 e 90 por cento para DQO, 40 por cento para N-total, 23 por cento para N-amoniacal total, 60 por cento para P-total, 52 por cento para P-PO4, 74 por cento para SST, 96 por cento para SSV, 75 por cento para sólidos sedimentáveis, 44 por cento para SDV, 88 por cento para sulfeto total, e 97 por cento para óleos e graxas totais, variando entre 73 por cento a 100 por cento . Cerca de 80 por cento da fração orgânica da matéria nitrogenada presente no esgoto bruto foi removida. A remoção de coliformes termotolerantes foi, em média, de 2 e 3 unidades log, e de Escherichia Coli, média de 1 e 3 unidades log, respectivamente, sob aplicação das vazões máxima e média, Giardia sp, média de 99,995 por cento , Cryptosporidium sp, média de 98,7 por cento , Enterovírus, média de 99,6 por cento , e Ascaris sp, média de 0,10 ovo/L. A remoção de sulfetos propiciou a geração de efluente tratado sem odores desagradáveis. A diminuição da vazão aplicada e a elevação do TDH influenciaram positivamente no desempenho do sistema com relação às remoções dos parâmetros físico-químicos e microbiológicos. Com base no presente estudo, ficou evidente o benefício da associação de wetlands construídas com fluxo vertical e horizontal no tratamento de efluente de tanque séptico. O capim Vetiver apresentou bom potencial de utilização em wetlands 8 construídas para tratamento de esgotos domésticos. O sistema experimental de tratamento apresentou flexibilidade operacional, mantendo bom desempenho inclusive nos períodos de sobrecarga. A qualidade do efluente tratado final obtido no presente estudo atende às exigências de lançamento e aos padrões de emissão de efluentes líquidos em corpos dágua e em sistemas públicos de esgotamento sanitário definidos na legislação ambiental federal e do Estado de São Paulo. A tecnologia de wetlands construídas híbridas associada ao tratamento de efluentes de tanque séptico apresentou bom potencial para o tratamento descentralizado de esgotos domésticos, inclusive em regiões com pouca disponibilidade de área livre.

Designing for participation in local social ridesharing networks : grass roots prototyping of IT systems

This abstract explores the possibility of a grass roots approach to engaging people in community change initiatives by designing simple interactive exploratory prototypes for use by communities over time that support shared action. The prototype is gradually evolved in response to community use, fragments of data gathered through the prototype, and participant feedback with the goal of building participation in community change initiatives. A case study of a system to support ridesharing is discussed. The approach is compared and contrasted to a traditional IT systems procurement approach.

Plastic traits of an exotic grass contribute to its abundance but are not always favourable

In herbaceous ecosystems worldwide, biodiversity has been negatively impacted by changed grazing regimes and nutrient enrichment. Altered disturbance regimes are thought to favour invasive species that have a high phenotypic plasticity, although most studies measure plasticity under controlled conditions in the greenhouse and then assume plasticity is an advantage in the field. Here, we compare trait plasticity between three co-occurring, C 4 perennial grass species, an invader Eragrostis curvula, and natives Eragrostis sororia and Aristida personata to grazing and fertilizer in a three-year field trial. We measured abundances and several leaf traits known to correlate with strategies used by plants to fix carbon and acquire resources, i.e. specific leaf area (SLA), leaf dry matter content (LDMC), leaf nutrient concentrations (N, C:N, P), assimilation rates (Amax) and photosynthetic nitrogen use efficiency (PNUE). In the control treatment (grazed only), trait values for SLA, leaf C:N ratios, Amax and PNUE differed significantly between the three grass species. When trait values were compared across treatments, E. curvula showed higher trait plasticity than the native grasses, and this correlated with an increase in abundance across all but the grazed/fertilized treatment. The native grasses showed little trait plasticity in response to the treatments. Aristida personata decreased significantly in the treatments where E. curvula increased, and E. sororia abundance increased possibly due to increased rainfall and not in response to treatments or invader abundance. Overall, we found that plasticity did not favour an increase in abundance of E. curvula under the grazed/fertilized treatment likely because leaf nutrient contents increased and subsequently its' palatability to consumers. E. curvula also displayed a higher resource use efficiency than the native grasses. These findings suggest resource conditions and disturbance regimes can be manipulated to disadvantage the success of even plastic exotic species.

An invasive grass shows colonization advantages over native grasses under conditions of low resource availability

Increased or fluctuating resources may facilitate opportunities for invasive exotic plants to dominate. This hypothesis does not, however, explain how invasive species succeed in regions characterized by low resource conditions or how these species persist in the lulls between high resource periods. We compare the growth of three co-occurring C4 perennial bunchgrasses under low resource conditions: an exotic grass, Eragrostis curvula (African lovegrass) and two native grasses, Themeda triandra and Eragrostis sororia. We grew each species over 12 weeks under low nutrients and three low water regimes differentiated by timing: continuous, pulsed, and mixed treatments (switched from continuous to pulsed and back to continuous). Over time, we measured germination rates, time to germination (first and second generations), height, root biomass, vegetative biomass, and reproductive biomass. Contrary to our expectations that the pulsed watering regime would favor the invader, water-supply treatments had little significant effect on plant growth. We did find inherent advantages in a suite of early colonization traits that likely favor African lovegrass over the natives including faster germination speed, earlier flowering times, faster growth rates and from 2 weeks onward it was taller. African lovegrass also showed similar growth allocation strategies to the native grasses in terms of biomass levels belowground, but produced more vegetative biomass than kangaroo grass. Overall our results suggest that even under low resource conditions invasive plant species like African lovegrass can grow similarly to native grasses, and for some key colonization traits, like germination rate, perform better than natives.

Unravelling the mechanisms behind the invasion of an introduced and now undesirable grass species

The introduction of Eragrostis curvula (African Lovegrass, herafter Lovegrass) for pasture improvement across Australia has not been successful. Instead Lovegrass, a C4 perennial grass originating from Southern African, has proven unpalatable to stock and to have low nutritional value if stocks do eat it. It has spread prolifically along roadsides, stream banks, conservation areas and pastures. Because control efforts have not been effective, our aim was to determine the putative mechanisms responsible for the dominance of Lovegrass, specifically disturbance (selective grazing) and competition.

Studies of assisted reproduction in the spotted grass frog Limnodynastes tasmaniensis: ovulation, early development and microinjection (ICSI)

Assisted Reproductive Technologies (ART) offer a wide range of techniques that have the potential to augment efforts to conserve and manage endangered amphibians and improve wild and captive population numbers. Gametes and tissues of species nearing endangered or extinct status can be cryopreserved and stored in gene banks, to provide material that can be utilised in the future as ART methods are refined. The Spotted Grass Frog, Limnodynastes tasmaniensis, is an abundant amphibian species in South-Eastern Australia of the family Myobatrachidae, that is suitable for the development of ART systems that can be applied to the threatened and endangered myobatrachid and other amphibian species native to Australia. The aim of this study was to advance the understanding of ovulation, fertilisation and embryo nic development of Lim. tasmaniensis and in vitro manipulations of reproduction and development for use in the development of advanced ART procedures such as intracytoplasmic spermatozoon injection (ICSI), androgenesis and nuclear transfer. Ovulation in amphibians can be induced by protocols utilising natural or synthetic hormones. All protocols tested on Lim. tasmaniensis in this study required two injections and the most effective protocols continued to require a first injection of pituitary extracts to induce ovulation. The second injection was, however, successfully replaced by synthetic chorionic gonadotrophin at a threshold dosage of 100 iu and halved the number of cane toads required to source the pituitaries. A combination of LHRH and Pimozide offered a less effective protocol, that did not require the use of pituitary extracts, and avoided the risk of pathogen transfer associated with unsterilised pituitary extracts. Unfertilised eggs of Lim. tasmaniensis were exposed to media of various osmolalities to determine media effects on eggs and their surrounding jelly layers that might impact on egg viability and fertilisability. Osmolality had no effect upon the egg diameter, however, rapid swelling of the jelly layers occurred within 15 minutes of exposure to various media treatments and plateaued from 30-90 minutes without further expansion. Swelling of the jelly layers was increased in hypotonic media (2.5% SAR, H2O) and minimised in the isotonic media (100% SAR). The optimal conditions for the culture of Lim. tasmaniensis eggs were identified as a holding media of 100% SAR, followed by a medium change to 2.5% SAR at insemination. This sequence of media minimised the rate of swelling of the jelly layers prior to contact with the spermatozoa, and maximised the activation of spermatozoa and eggs throughout fertilisation and embryonic development. Embryos of Lim. tasmaniensis were cultured at four temperatures (13 C, 17 C, 23 C and 29 C), to determine the effect of temperature on cleavage and embryonic development rates. Embryonic development progressed through a sequence of stages that were not altered by changes in temperature. However cleavage rates were affected by changes in temperature as compared with normal embryonic growth at 23 C. Embryonic development was suspended at the lowest temperature (13 C) while embryonic viability was maintained. A moderate decrease in temperature (17 C) slowed cleavage, while the highest temperature (29 C) increased the cleavage rate, but decreased the embryo survival. Rates of embryonic development can be manipulated by changes in temperature and this method can be used to source blastomeres of a specific size/stage at a predetermined age or halt cleavage at specific stages for embryos or embryo derived cells to be included in ART procedures. This study produced the first report of the application of Intracytoplasmic Spermatozoon Injection (ICSI) in an Australian amphibian. Eggs that were activated by microinjection with a single spermatozoon (n=50) formed more deep, but abnormal, cleavage furrows post-injection (18/50, 36%), than surface changes (12/50, 24%). This result is in contrast to eggs injected without a spermatozoon (n=42), where the majority of eggs displayed limited surface changes (36/42, 86%), and few deep, abnormal furrows (3/42, 7%). Three advanced embryos (3/50, 6%) were produced by ICSI that developed to various stages within the culture system. Technical difficulties were encountered that prevented the generation of any metamorphs from ICSI tadpoles. Nevertheless, when these blocks to ICSI are overcome, the ICSI procedure will be both directly useful as an ART procedure in its own right, and the associated refinement of micromanipulation procedures will assist in the development of other ART procedures in Lim. tasmaniensis. A greater understanding of basic reproductive and developmental biology in Lim. tasmaniensis would greatly facilitate refinement of fertilisation by ICSI. Assisted Reproductive Technologies, in conjunction with gene banks may in the future regenerate extinct amphibian species, and assist in the recovery of declining amphibian populations nationally and worldwide.

Recovering Functional Integrity in Pastures Dominated by an Invasive Grass Species

Exotic grasses have been introduced in countries worldwide for pasture improvement, soil stabilisation and ornamental purposes. Some of these introductions have proven successful, but many have not (Cook & Dias 2006). In Australia, the Commonwealth Plant Introduction Scheme was initiated in 1929, and over-time introduced more than 5000 species of grasses, legumes and other forage and browse plants (Cook & Dias 2006). Lonsdale (1994) suggested that, in tropical Australia, 13% of introductions have become a problem, with only 5% being considered useful for agriculture. Low (1997) suggested that 5 out of 18 of Australia's worst tropical environmental weeds were intentionally introduced as pasture grasses. The spread and dominance of invasive grass species that degrade the quality of pastures for production can impact significantly on the livelihoods of small proprietors. Although Livestock grazing contributes only a small percentage to the world's GDP (1.5%), maintaining the long-term stability of this industry is crucial because of the high social and environmental consequence of a collapse. One billion of the world's poor are dependent on livestock grazing for food and income with this industry occupying more than 25% of the world's land base (Steinfeld et al. 2006). The ling-term sustainability of livestock grazing is also crucial for the environment. A recent FAO report attributed livestock production as a major cause of five of the most serious environmental problems: global warming, land degredation, air and water pollution, and the loss of biodiversity (Steinfeld et al. 2006). For these reasons, finding more effective approaches that guide the sustainable management of pastures is urgently needed. In Australia more than 55% of land use is for livestock grazing by sheelp and/or cattle. This land use dominate in the semi-arid and arid regions where rainfall and soil conditions are marginal for production (Commonwealth of Australia 2004). Although the level of agriculture production by conglomerates is increasing, the majority of livestock grazing within Australia remains family owned and operated (Commonwealth of Australia 2004). The sustainability of production from a grazed pasture is dependent on its botanical composition (Kemp & Dowling 1991, Kemp et al. 1996). In a grazed pasture, the dominance of an invasive grass species can impact on the functional integrity of the ecosystem, including production and nutrient cycling; wwhich will in turn, affect the income of proprietors and the ability of the system to recover from disturbance and environmental change. In Australia, $0.3 billion is spent on weed control in livestock production, but despite this substantial investment $1.9 billion is still lost in yield as a result of weeds (Sinden et al. 2004). In this paper, we adaprt a framework proposed for the restoration of degraded rainforest communities (Lamb & Gilmour 2003, Lamb et al. 2005) to compare and contrast options for recovering function integrity (i.e. a diverse set of desirable plant species that maintain key ecological processes necessary for sustainable production and nutrient cycling) within pasture communities dominated by an invasive grass species. To do this, we uase a case-study of the invasion of Eragrostis curvula (Africal lovegrss; hereafter, Lovegrass), a serious concern in Australian agricultural communities (Parsons and Cuthbertson 1992). The spread and dominance of Lovegrass is a problem because its low palatability, low nutritional content and competitiveness affect the livelihood of graziers by reducing the diversity of other plant species. We conclude by suggesting modifications to this framework for pasture ecosystems to help increase the effiency of strategies to protect functional integrity and balance social/economic and biodiversity values.