Culture-Independent Assessment of Rhizobiales-Related Alphaproteobacteria and the Diversity of Methylobacterium in the Rhizosphere and Rhizoplane of Transgenic Eucalyptus

The rhizosphere is an ecosystem exploited by a variety of organisms involved in plant health and environmental sustainability. Abiotic factors influence microorganism-plant interactions, but the microbial community is also affected by expression of heterologous genes from host plants. In the present work, we assessed the community shifts of Alphaproteobacteria phylogenetically related to the Rhizobiales order (Rhizobiales-like community) in rhizoplane and rhizosphere soils of wild-type and transgenic eucalyptus. A greenhouse experiment was performed and the bacterial communities associated with two wild-type (WT17 and WT18) and four transgenic (TR-9, TR-15, TR-22, and TR-23) eucalyptus plant lines were evaluated. The culture-independent approach consisted of the quantification, by real-time polymerase chain reaction (PCR), of a targeted subset of Alphaproteobacteria and the assessment of its diversity using PCR-denaturing gradient gel electrophoresis (DGGE) and 16S rRNA gene clone libraries. Real-time quantification revealed a lesser density of the targeted community in TR-9 and TR-15 plants and diversity analysis by principal components analysis, based on PCR-DGGE, revealed differences between bacterial communities, not only between transgenic and nontransgenic plants, but also among wild-type plants. The comparison between clone libraries obtained from the transgenic plant TR-15 and wild-type WT17 revealed distinct bacterial communities associated with these plants. In addition, a culturable approach was used to quantify the Methylobacterium spp. in the samples where the identification of isolates, based on 16S rRNA gene sequences, showed similarities to the species Methylobacterium nodulans, Methylobacterium isbiliense, Methylobacterium variable, Methylobacterium fujisawaense, and Methylobacterium radiotolerans. Colonies classified into this genus were not isolated from the rhizosphere but brought in culture from rhizoplane samples, except for one line of the transgenic plants (TR-15). In general, the data suggested that, in most cases, shifts in bacterial communities due to cultivation of transgenic plants are similar to those observed when different wild-type cultivars are compared, although shifts directly correlated to transgenic plant cultivation may be found.

Characterization of nicosulfuron availability in aged soils

Sorption-desorption interactions of pesticides with soil determine their availability for transport, plant uptake, and microbial degradation. These interactions are affected by the physical-chemical properties of the pesticide and soil, and for some pesticides, their residence time in the soil. This research evaluated changes in sorption/availability of nicosulfuron (2-[[[[(4,6-dimethoxy-2-pyrimidinyl]aminolcarbonyl]amino]sulfonyl]-N,N-dimethyl-3-pyridinecarboxamide) herbicide with aging in different soils, using a radiolabeled (C-14) tracer. Aging significantly increased sorption. For instance, after the 41-day incubation, calculated K-d,K-app increased by a factor of 2 to 3 in Mollisols from the Midwestern United States and by a factor of 5 to 9 in Oxisols from Brazil and Hawaii, as compared to freshly treated soils. In view of this outcome, potential transport of nicosulfuron would be overpredicted if freshly treated soil Kd values were used to predict transport. The fact that the nicosulfuron solution concentration decreased faster than the soil concentration with time suggested that the increase in sorption was because the rate of degradation in solution and on labile sites was faster than the rate of desorption of the neutral species from the soil particles. It may have also been due to nicosulfuron anion diffusion to less accessible sites with time, leaving the more strongly bound neutral molecules for the sorption characterization. Regardless of the mechanism, these results are further evidence that increases in sorption during pesticide aging should be taken into account during the characterization of the sorption process for mathematical models of pesticide degradation and transport.

Plant availability of phosphorus in four superphosphate fertilizers varying in water-insoluble phosphate compounds

There is concern that the use of lower quality phosphate rock can result in elevated amounts of Fe-Al-P water-insoluble compounds in fertilizers and, consequently, low agronomic effectiveness. Therefore, studies were conducted to evaluate the effect of some of these compounds on plant growth. Four commercial superphosphates varying in chemical composition (two single and two triple superphosphates) were selected for the study. Fertilizer impurities were collected as water-insoluble residues by washing each P source with deionized water. A modal analysis, based primarily on elemental chemical analysis and x-ray diffractometry, was used to estimate the chemical composition of each P source. Water-soluble monocalcium phosphate (MCP) and the water-leached fertilizer residues were prepared to give a range of fertilizers in terms of water-soluble phosphorus (WSP) (0-100% of the available P as MCP). The water-leached fractions, MCP, and the mixtures of MCP with water-leached fractions were applied to supply 40 mg available P kg(1) to a thermic Rhodic Kanhapludult with pH values of 5.2 +/- 0.05 (unlimed) and 6.4 +/- 0.08 (limed). Wheat (Triticum aestivum L.) grown in a greenhouse for 101 d served as the test crop. The requirement for WSP was source and pH dependent. At a soil pH of 5.2, the fertilizers required 73 to 95% WSP to reach the maximum dry-matter yield, while they required 60 to 86% WSP at pH 6.4. To reach 90% of the maximum yield, all superphosphate fertilizers required <50% WSP. These results show that it is not always necessary to have high water solubility as required by legislation in many countries.

Foundation and Perspectives of the Use of Plant Extracts as Performance Enhancers in Broilers

Feed is responsible for about 70% of broilers production costs, leading to an increasing number of studies on alternative dietary products that benefit bird performance and lower production costs. Since the 1950s, antimicrobial additives are the most frequently used performance enhancers in animal production and their positive results are observed even in high-challenge conditions. Since the 1990s, due to the ban of the use of some antibiotics as growth promoters and the growing trend of the public to consume natural products, plant extracts have been researched as alternatives to antibiotic growth promoters. The first study that evaluated the antibacterial activities of plant extracts was carried out in 1881; however, they started to be used as flavor enhancers only during the next decades. With the emergence of antibiotics in the 1950s, the use of plant extracts as antimicrobial agents almost disappeared. There are several studies in literature assessing the use of plant extracts, individually or in combination, as antimicrobials, antioxidants, or digestibility enhancers in animal feeds. Research results on the factors affecting their action, such as plant variety, harvest time, processing, extraction, as well as the technology employed to manufacture the commercial product and dietary inclusion levels show controversial results, warranting the need of further research and standardization for the effective use of plant extracts as performance enhancers, when added to animal feeds. This article aims at presenting plant extracts as alternatives to antibiotics, explaining their main modes of action as performance enhancers in broiler production.

Digestibility of plant protein-based diets by largemouth bass Micropterus salmoides

Commercial farming of carnivorous fish demands the reduction of environmental impact of feeds; that requires minimal use of dietary animal protein. This study investigated the digestibility of diets formulated exclusively out of plant protein, added feed attractants, by the carnivore largemouth bass, Micropterus salmoides. Juvenile largemouth bass (14.0 +/- 1.0 cm) conditioned to accept artificial, dry feed were confined in polypropylene cages and fed ad libitum in three daily meals, seven experimental diets containing varying levels of vegetable and animal protein sources, added of different feed stimulants. After last daily meal, cages were transferred to cylindrical-conical-bottomed, 200-L aquaria, where faeces were collected by sedimentation into refrigerated containers, preserved and later analysed for chemical composition. Soybean meal can be used as partial substitute of animal protein in diets for largemouth bass; the poultry by-product meal shows as a good option as animal protein source in these rations. Control treatment - 50PP : 50AP - yielded best performances; the need for the use of fish meal in the formulation for carnivorous diets is, at least, questionable. Results of the digestibility trials demonstrated the importance of determining the diet digestibility, if precision in the formulation of least-cost feeds for carnivorous fish is the ultimate goal.

A new set of microsatellite markers for the genetic characterization of Ceratocystis fimbriata, an economically important plant pathogen

Ceratocystis fimbriata is a fungal pathogen which attacks several economically important plants, but occurs in host-associated, morphologically indistinguishable forms. In Brazil, this fungus seriously attacks mango trees (Mangifera indica), causing severe loss of yield. This work aimed to develop and characterize a novel set of microsatellite markers for this important pathogen, providing researchers with new molecular tools for the characterization of isolates. Twenty polymorphic primer pairs were designed from a microsatellite-enriched library. We tested the usefulness of these markers through genotyping thirteen isolates of the fungus. On average, 6.65 alleles per locus were detected, revealing the ability of this set of markers to characterize C. fimbriata isolates associated to mango and to other plant species.

Yield, Potassium Uptake, and Use Efficiency in Upland Rice Genotypes

Potassium (K) is an essential nutrient for higher plants. Information on K uptake and use efficiency of upland rice under Brazilian conditions is limited. A greenhouse experiment was conducted with the objective to evaluate influence of K on yield, K uptake, and use efficiency of six upland rice genotypes grown on Brazilian Oxisol. The K rate used was zero (natural soil level) and 200 mg K kg-1 of soil. Shoot dry weight and grain yield were significantly influenced by K level and genotype treatments. However, K x genotype interactions were not significant, indicating similar responses of genotypes at two K levels for shoot dry weight and grain yield. Genotypes produced grain yield in the order of BRS Primavera BRA 01596 BRSMG Curinga BRS 032033 BRS Bonanca BRA 02582. Potassium concentration in shoot was about sixfold greater compared to grain, across two K levels and six genotypes. However, K utilization efficiency ratio (KUER) (mg shoot or grain yield / mg K uptake in shoot or root) was about 6.5 times greater in grain compared to shoot, across two K level and six genotypes. Potassium uptake in shoot and grain and KUER were significantly and positively associated with grain yield. Soil calcium (Ca), K, base saturation, acidity saturation, Ca saturation, K saturation, Ca/K ratio, and magnesium (Mg)/K ratio were significantly influenced by K application rate.

Oviposition by a moth suppresses constitutive and herbivore-induced plant volatiles in maize

Plant volatiles function as important signals for herbivores, parasitoids, predators, and neighboring plants. Herbivore attack can dramatically increase plant volatile emissions in many species. However, plants do not only react to herbivore-inflicted damage, but also already start adjusting their metabolism upon egg deposition by insects. Several studies have found evidence that egg deposition itself can induce the release of volatiles, but little is known about the effects of oviposition on the volatiles released in response to subsequent herbivory. To study this we measured the effect of oviposition by Spodoptera frugiperda (J.E. Smith) (Lepidoptera: Noctuidae) moths on constitutive and herbivore-induced volatiles in maize (Zea mays L.). Results demonstrate that egg deposition reduces the constitutive emission of volatiles and suppresses the typical burst of inducible volatiles following mechanical damage and application of caterpillar regurgitant, a treatment that mimics herbivory. We discuss the possible mechanisms responsible for reducing the plant`s signaling capacity triggered by S. frugiperda oviposition and how suppression of volatile organic compounds can influence the interaction between the plant, the herbivore, and other organisms in its environment. Future studies should consider oviposition as a potential modulator of plant responses to insect herbivores.

Isolation of micropropagated strawberry endophytic bacteria and assessment of their potential for plant growth promotion

Twenty endophytic bacteria were isolated from the meristematic tissues of three varieties of strawberry cultivated in vitro, and further identified, by FAME profile, into the genera Bacillus and Sphingopyxis. The strains were also characterized according to indole acetic acid production, phosphate solubilization and potential for plant growth promotion. Results showed that 15 strains produced high levels of IAA and all 20 showed potential for solubilizing inorganic phosphate. Plant growth promotion evaluated under greenhouse conditions revealed the ability of the strains to enhance the root number, length and dry weight and also the leaf number, petiole length and dry weight of the aerial portion. Seven Bacillus spp. strains promoted root development and one strain of Sphingopyxis sp. promoted the development of plant shoots. The plant growth promotion showed to be correlated to IAA production and phosphate solubilization. The data also suggested that bacterial effects could potentially be harnessed to promote plant growth during seedling acclimatization in strawberry.

Inhibitory effects of Solidago chilensis Meyen hydroalcoholic extract on acute inflammation

Aim of the study: Alcoholic or hydroalcoholic preparations of the plant Solidago chilensis Meyen (Asteraceae) are employed in popular medicines to treat inflammation. The anti-inflammatory effects of the hydroalcoholic extract of aerial parts of the plant (93% ethanol) were investigated and the main components of the extract were identified. Materials and methods: Ear oedema was induced in male Wistar rats by topical application of the chloroform fraction of latex-extract from Euphorbia milii. Leukocyte mobilisation was quantified after air-pouch inflammation evoked by oyster glycogen. Leukocyte-endothelial interactions and mast cell degranulation were quantified by intravital microscopy. The extract itself was characterised via HPLC-DAD-MS and HPLC-MS/MS. Results: Topical (12.5-50 mg/kg) or intraperitoneal (25 or 50 mg/kg) administrations of the extract reduced ear oedema formation (>25% reduction). Intraperitoneal applications of 25 mg/kg of extract inhibited the migration of polymorphonuclear cells into the inflamed cavity (about 50%). In addition, the rolling behaviour and adherence of circulating leukocytes to postcapillary venules of the mesentery network was diminished (50%), but the mast cell degranulation in the perivascular area was not affected. The major components of the extract were identified as caffeoylquinic acid derivatives and the flavonoid rutin. Conclusions: The data presented herein show local and systemic anti-inflammatory effects of the hydroalcoholic extract of aerial parts of Solidago chilensis, and implicate the inhibition of leukocyte-endothelial interactions as an important mechanism of the extract`s action. (C) 2009 Elsevier Ireland Ltd. All rights reserved.

Gas Chromatographic Analysis of Dimethyltryptamine and beta-Carboline Alkaloids in Ayahuasca, an Amazonian Psychoactive Plant Beverage

Introduction - Ayahuasca is obtained by infusing the pounded stems of Banisteriopsis caapi in combination with the leaves of Psychotria viridis. P. viridis is rich in the psychedelic indole N,N-dimethyltryptamine, whereas B. caapi contains substantial amounts of beta-carboline alkaloids, mainly harmine, harmaline and tetrahydroharmine, which are monoamine-oxidase inhibitors. Because of differences in composition in ayahuasca preparations, a method to measure their main active constituents is needed. Objective - To develop a gas chromatographic method for the simultaneous determination of dimethyltryptamine and the main beta-carbolines found in ayahuasca preparations. Methodology - The alkaloids were extracted by means of solid phase extraction (C(18)) and detected by gas chromatography with nitrogen/phosphorous detector. Results - The lower limit of quantification (LLOQ) was 0.02 mg/mL for all analytes. The calibration curves were linear over a concentration range of 0.02-4.0 mg/mL (r(2) > 0.99). The method was also precise (RSD < 10%). Conclusion - A simple gas chromatographic method to determine the main alkaloids found in ayahuasca was developed and validated. The method can be useful to estimate administered doses in animals and humans for further pharmacological and toxicological investigations of ayahuasca. Copyright (C) 2009 John Wiley & Sons, Ltd.

Optimization of Pothomorphe umbellata (L.) Miquel topical formulations using experimental design

Pothomorphe umbellata is a native plant widely employed in the Brazilian popular medicine. This plant has been shown to exert a potent antioxidant activity on the skin and to delay the onset and reduce the incidence of UVB-induced skin damage and photoaging. The aim of this work was to optimize the appearance, the centrifuge stability and the permeation of emulsions containing R umbellata (0. 1% 4-nerolidylchatecol). Experimental design was used to study ternary mixtures models with constraints and graphical representation by phase diagrams. The constraints reduce the possible experimental domain, and for this reason, this methodology offers the maximum information while requiring the minimum investment. The results showed that the appearance follows a linear model, and that the aqueous phase was the principal factor affecting the appearance; the centrifuge stability parameter followed a mathernatic quadratic model and the interactions between factors produced the most stable emulsions; skin permeation was improved by the oil phase, following a linear model generated by data analysis. We propose as optimized P. umbellata formulation: 68.4% aqueous phase, 26.6% oil phase and 5.0% of self-emulsifying phase. This formulation displayed an acceptable compromise between factors and responses investigated. (c) 2007 Elsevier B.V. All rights reserved.

Effect of inulin as prebiotic and synbiotic interactions between probiotics to improve fermented milk firmness

Inulin behaved as a prebiotic to improve firmness of skim milk fermented by (a) pure cultures of Lactobacillus acidophilus (La), Lactobacillus rhamnosus (Lr), Lactobacillus bulgaricus (Lb) and Bifidobacterium lactis (Bl), (b) binary co-cultures of them with Streptococcus thermophilus (St), or (c) a cocktail containing all them. Inulin addition to co-cultures and cocktail enhanced products firmness, either after 1 day (D1) or 7 days (D7) of cold storage, likely due to the increase in microbial growth induced by metabolic interactions among lactic acid bacteria and partial inulin metabolization. Co-culture firmness did in fact range from 0.33 N without inulin (St-Lb) after D1 and 0.55 N with inulin (St-Lr) after D7. Also cocktail cultures exhibited high values of firmness, ranging, as an average, from 0.43 N (D1) to 0.46 N (D7), which suggests that they could have been potentiated by the reciprocal synergistic effects of microorganisms in complex mixture. (C) 2011 Elsevier Ltd. All rights reserved.

qNMR: An applicable method for the determination of dimethyltryptamine in ayahuasca, a psychoactive plant preparation

Ayahuasca is an Amazonian plant beverage obtained by infusing the pounded stems of Banisteriopsis caapi in combination with the leaves of Psychotria viridis. P. viridis contains the psychedelic indole N,N-dimethyltryptamine (DMT). This association has a wide range of use in religious rituals around the world. In the present work, an easy, fast and non-destructive method by Nuclear Magnetic Resonance of proton ((1)H NMR) for quantification of DMT in ayahuasca samples was developed and validated. 2,5-Dimethoxybenzaldehyde (DMBO) was used as internal standard (IS). For this purpose, the area ratios produced by protons of DMT (N(CH(3))(2)) at 2.70 ppm, singlet, (6H) and for DMBO (Ar(OCH(3))(2)) at 3.80 and 3.89 ppm, doublet, (6H) were used for quantification. The lower limit of quantification (LLOQ) was 12.5 mu g/mL and a good intra-assay precision was also obtained (relative standard deviation < 5.1%). The present (1)H NMR method is not time consuming and can be readily applied to monitor this tryptamine in plant preparations. We believe that qNMR can be used for identification and quantification of many plant-based products and metabolites with important advantages, while comparing with other analytical techniques. (C) 2010 Phytochemical Society of Europe. Published by Elsevier B. V. All rights reserved.

A survey on the sanitary condition of commercial foods of plant origin sold in Brazil

The objective of the study was to evaluate the hygienic conditions and practices of commercial foods of plant origin in establishments and street marketed by street vendors in cities in the State of Rio de Janeiro, Brazil. Forty different sales points were evaluated (establishments that prepared and sold fruit juices and street vendors that commercialized fresh coconut water, sugarcane juice and orange juice) using a questionnaire with 12 items, divided into three blocks (salesmen/handlers, operations, installations). The results indicated that the activities related to the commerce of fruit beverages in the cities of Sao Goncalo and Rio de Janeiro required the elaboration of a set of actions by the Sanitary Vigilance Agency in order to improve the hygienic and sanitary level and minimize health risk to consumers. Important requirements in the legislation relevant to this type of food are still not followed; adequate packaging and storage of the raw material, obtaining the raw material from registered suppliers, hygiene of the handlers and adequate management of wastes produced during the activities in question are amongst the main items deserving attention. (C) 2009 Elsevier Ltd. All rights reserved.