Effects of the herbicides acetochlor and metolachlor on antioxidant enzymes in soil bacteria

The aim of this study was to investigate the antioxidant responses of three bacteria (SD1. KD and K9) isolated from soil previously treated with the herbicides metolachlor and acetochlor. By 165 rRNA gene sequencing, we determined that SD1 is phylogenetically related to Enterobacter asburiae, while KD and K9 have divergent genomes that more closely resemble that of Enterobacter amnigenus. Decreased levels of lipid peroxidation were observed in SD1 and KD following treatment with 34 mM metolachlor or 62 mM acetochlor, respectively, indicating that both bacteria were able to adapt to an increase in ROS production. In the presence of 34 mM metolachlor or 62 mM acetochlor, all bacterial isolates exhibited increases in total catalase (CAT) activity (81% for SDI, 53% for KD and 59% for K9), whereas total SOD activity (assessed based on the profile and intensity of the bands) was slightly reduced when the bacteria were exposed to high concentrations of the herbicides (340 mM metolachlor or 620 mM acetochlor). This effect was due to a specific reduction in SOD IV (K9 and KD isolates) by 45% and 90%, respectively, and SOD V (SD1 isolate) isoenzymes by 60%. The most striking result was obtained in the SD1 isolate, where two novel isoenzymes of glutathione reductase (GR) that responded specifically to metolachlor were identified. In addition, acetochlor was shown to induce the expression of a new 57 kDa protein band in the K9 and KD isolates. The bacteria isolated from the herbicide-contaminated soil exhibited an efficient antioxidant system response at herbicide concentrations of up to 34 mM metolachlor or 62 mM acetochlor. These data suggest a mechanism for tolerance that may include the control of an imbalance in ROS production versus scavenging. The data suggest that specific isoenzymes of CAT and GR could be involved in this herbicide tolerance mechanism. (C) 2011 Elsevier Ltd. All rights reserved.

Antioxidant response of Nicotiana tabacum cv. Bright Yellow 2 cells to cadmium and nickel stress

Plant cell cultures are a suitable model system for investigation of the physiological mechanisms of tolerance to environmental stress. We have determined the effects of Cd (0.1 and 0.2 mM CdCl(2)) and Ni (0.075 and 0.75 mM NiCl(2)) on Nicotiana tabacum L. cv. Bright Yellow (TBY-2) cell suspension cultures over a 72-h period. Inhibition of growth, loss of cell viability and lipid peroxidation occurred, in general, only when the TBY-2 cells were grown at 0.2 mM CdCl(2) and at 0.75 mM NiCl(2). At 0.1 mM CdCl(2), a significant increase in growth was determined at the end of the experiment. Increases in the activities of all of the four enzymatic antioxidant defence systems tested, were induced by the two concentrations of Cd and Ni, but at different times during the period of metal exposure. Overall, the cellular antioxidant responses to Cd and Ni were similar and were apparently sufficient to avoid oxidative stress at the lower concentrations of Cd and Ni. The activities of glutathione reductase and glutathione S-transferase increased early but transiently, whereas the activities of catalase and guaiacol peroxidase increased in the latter half of the experimental period. Therefore it is likely that the metabolism of reduced glutathione was enhanced during the initial onset of the stress, while catalase and guaiacol-type peroxidase appeared to play a more important role in the antioxidant response once the stress became severe.

Horticultural performance of `Folha Murcha` sweet orange onto twelve rootstocks

Despite its outstanding position, the Brazilian citriculture is established on a very limited pool of varieties that limits its expansion and restricts the fruit availability throughout the year. This situation determines the urgent necessity of developing alternative scion and rootstock cultivars, with good performance under local conditions. `Folha Murcha` sweet orange (Citrus sinensis (L.) Osbeck) is a late-harvest cultivar, suitable both for the juice processing industry and the fresh fruit market, being described as tolerant to citrus canker (Xanthomonas citri subsp. citri Schaad et al.), and less affected by citrus variegated chlorosis (Xylella fastidiosa Wells et al.). A study was conducted in Bebedouro, Sao Paulo State, Brazil, to evaluate the horticultural performance of `Folha Murcha` sweet orange budded onto 12 rootstocks: the citrandarin `Changsha` mandarin (Citrus reticulata Blanco) x Poncirus trifoliata `English Small`: the hybrid `Rangpur` lime (Citrus limonia Osbeck) x `Swingle` citrumelo (P. trifoliata (L.) Raf x Citrus paradisi Macfad.); the trifoliates (P. trifoliata (L.) Raf.)`Rubidoux`, `FCAV`, and `Flying Dragon` (P. trifoliata var. monstrosa); the `Sun Chu Sha Kat` mandarin (C. reticulata Blanco); the `Sunki` mandarin (Citrus sunki (Hayata) Hart. ex. Tanaka); the `Rangpur` limes (C. limonia Osbeck) `Cravo Limeira` and `Cravo FCAV`; `Carrizo` citrange (C. sinensis x P. trifoliata), `Swingle` citrumelo (P. trifoliata x C. paradisi), and `Orlando` tangelo (C. paradisi x Citrus tangerina cv. `Dancy`). The experimental grove was planted in 2001, using a 7 m x 4 m spacing, in a randomized block design, with five replications and two plants per plot. No supplementary irrigation was applied. Fruit yield, canopy volume, tree tolerance to drought and to citrus variegated chlorosis, and fruit quality were assessed for each rootstock. Trees grafted onto the `Flying Dragon` trifoliate were smaller in size, but had largest yield efficiency when compared to those grafted onto other rootstocks. Lower alternate bearing index was observed on trees budded onto `Cravo FCAV` `Rangpur` lime. Both `Rangpur` lime rootstocks and the `Sunki` mandarin induced higher tree tolerance to drought. The `Flying Dragon` trifoliate induced better fruit quality and higher tolerance to citrus variegated chlorosis (CVC) to `Folha Murcha` trees. A cluster multivariate analysis identified three groups of rootstocks with similar effects on `Folha Murcha` tree performance. Among the 12 evaluated rootstocks, the `Flying Dragon` trifoliate has a unique effect on plant growth, tolerance to drought and CVC, fruit yield and fruit quality of `Folha Murcha` trees, and may be better suited for high-density plantings. (C) 2011 Elsevier B.V. All rights reserved.

Detection of Sourgrass (Digitaria insularis) Biotypes Resistant to Glyphosate in Brazil

Sourgrass is a perennial weed infesting annual and perennial crops in Brazil. Three biotypes (R1, R2, and R3) of sourgrass suspected to be glyphosate-resistant (R) and another one (S) from a natural area without glyphosate application, in Brazil, were tested for resistance to glyphosate based on screening, dose-response, and shikimic acid assays. Both screening and dose-response assays confirmed glyphosate resistance in the three sourgrass biotypes. Dose-response assay indicated a resistance factor of 2.3 for biotype RI and 3.9 for biotypes R2 and R3. The hypothesis of a glyphosate resistance was corroborated on the basis of shikimic acid accumulation, where the S biotype accumulated 3.3, 5.0, and 5.7 times more shikimic acid than biotypes R1, R2, and R3, respectively, 168 h after treatment with 157.50 g ae ha(-1) of glyphosate. There were no differences in contact angle of spray droplets on leaves and spray retention, indicating that differential capture of herbicide by leaves was not responsible for resistance in these biotypes. The results confirmed resistance of sourgrass to glyphosate in Brazil.

Can native vegetation recover after slash pine cultivation in the Brazilian Savanna?

There is a widespread view that forest plantations with exotic species are green deserts, unable to sustain biodiversity. Few studies have demonstrated, however, that planted stands of exotic trees have a greater negative effect on the plant diversity of savanna vegetation. We compared the native woody flora under four stands of slash pine of about 45 years old with four stands where the previously existing native Cerrado vegetation was preserved and protected from disturbances for the same period, has changed into dense vegetation - the ""cerradao"", at Assis municipality, Sao Paulo State, Brazil. Aiming at understanding the potential ecological filters driving these communities, we assessed air and soil humidity, light availability and classified the native species on the basis of shade tolerance, dispersal syndrome and biomes in which they occur (Atlantic Forest or Cerrado). We recorded an average of 70 (+/- 13) species under pine stands and 54 (+/- 16) species in cerradao. Of the total of 136 species recorded, 78 occurred in both habitats, eight were exclusive to the ""cerradao"" (shade tolerant and also occurring in forest ecosystems) and 18 were recorded only under pine stands (82% heliophytic, exclusive to the Cerrado biome). Among the functional attributes and abiotic variables analyzed, only light availability explained the floristic differences found. Since richness was higher under pine, we refuted the hypothesis that exotic species constrain the establishment of the native species richness in the understory. On the other hand, the dark environment under the closed-canopy of the ""cerradao"" acts as a filter inhibiting the establishment of typical Cerrado species. Since pine stands, if managed in long cycle, maintain a reasonable pool of Cerrado endemic species in the understory pine plantations may be a good starting point for savanna restoration. (C) 2011 Elsevier B.V. All rights reserved.

Influence of Urea and Ammonium Sulfate on Soil Acidity Indices in Lowland Rice Production

Urea and ammonium sulfate are principal nitrogen (N) sources for crop production. Two field experiments were conducted during three consecutive years to evaluate influence of urea and ammonium sulfate application on grain yield, soil pH, calcium (Ca) saturation, magnesium (Mg) saturation, base saturation, aluminum (Al) saturation, and acidity (H + Al) saturation in lowland rice production. Grain yield was significantly influenced by urea as well as ammonium sulfate fertilization. Soil pH linearly decreased with the application of N by ammonium sulfate and urea fertilizers. However, the magnitude of the pH decrease was greater by ammonium sulfate than by urea. The Ca and Mg saturations were decreased at the greater N rates compared to low rates of N by both the fertilizer sources. The Al and acidity saturation increased with increasing N rates by both the fertilizer sources. However, these acidity indices were increased more with the application of ammonium sulfate compared with urea. Rice grain yield had negative associations with pH, Ca saturation, Mg saturation, and base saturation and positive associations with Al and acidity saturation. This indicates that rice plant is tolerant to soil acidity.

Overwintering of Sclerotium rolfsii and S-rolfsii var. delphinii in different latitudes of the United States

Previously known only from the southern United States, hosta petiole rot recently appeared in the northern United States. Sclerotium rolfsii var. delphinii is believed to be the predominant petiole rot pathogen in the northern United States, whereas S. rolfsii is most prevalent in the southern United States. In order to test the hypothesis that different tolerance to climate extremes affects the geographic distribution of these fungi, the survival of S. rolfsii and S. rolfsii var. delphinii in the northern and southeastern United States was investigated. At each of four locations, nylon screen bags containing sclerotia were placed on the surface of bare soil and at 20-cm depth. Sclerotia were recovered six times from November 2005 to July 2006 in North Dakota and Iowa, and from December 2005 to August 2006 in North Carolina and Georgia. Survival was estimated by quantifying percentage of sclerotium survival on carrot agar. Sclerotia of S. rolfsii var. delphinii survived until at least late July in all four states. In contrast, no S. rolfsii sclerotia survived until June in North Dakota or Iowa, whereas 18.5% survived until August in North Carolina and 10.3% survived in Georgia. The results suggest that inability to tolerate low temperature extremes limits the northern range of S. rolfsii.

EVALUATION OF RED CURRANTS (RIBES RUBRUM L.), BLACK CURRANTS (RIBES NIGRUM L.), RED AND GREEN GOOSEBERRIES (RIBES UVA-CRISPA) FOR POTENTIAL MANAGEMENT OF TYPE 2 DIABETES AND HYPERTENSION USING IN VITRO MODELS

Red currants (Ribes rubrum L.), black currants (Ribes nigrum L.), red and green gooseberries (Ribes uva-crispa) were evaluated for the total phenolics, antioxidant capacity based on 2, 2-diphenyl-1-picrylhydrazyl radical scavenging assay and functionality such as in vitro inhibition of alpha-amylase, alpha-glucosidase and angiotensin I-converting enzyme (ACE) relevant for potential management of hyperglycemia and hypertension. The total phenolics content ranged from 3.2 (green gooseberries) to 13.5 (black currants) mg/g fruit fresh weight. No correlation was found between total phenolics and antioxidant activity. The major phenolic compounds were quercetin derivatives (black currants and green gooseberries) and chlorogenic acid (red currants and red gooseberries). Red currants had the highest alpha-glucosidase, alpha-amylase and ACE inhibitory activities. Therefore red currants could be good dietary sources with potential antidiabetes and antihypertension functionality to compliment overall dietary management of early stages of type 2 diabetes.

Evaluation of the role of environmental factors in the human gastrointestinal tract on the behaviour of probiotic cultures of Lactobacillus casei Shirota and Lactobacillus casei LC01 by the use of a semi-dynamic in vitro model

This study evaluated the influence of gastrointestinal environmental factors (pH, digestive enzymes, food components, medicaments) on the survival of Lactobacillus casei Shirota and Lactobacillus casei LC01, using a semi-dynamic in vitro model that simulates the transit of microorganisms through the human GIT. The strains were first exposed to different simulated gastric juices for different periods of time (0, 30, 60 and 120 min), and then to simulated intestinal fluids for zero, 120, 180 and 240 min, in a step-wise format. The number of viable cells was determined after each step. The influence of food residues (skim milk) in the fluids and resistance to medicaments commonly used for varied therapeutic purposes (analgesics, antiarrhythmics, antibiotics, antihistaminics, proton pump inhibitors, etc.) were also evaluated. Results indicated that survival of both cultures was pH and time dependent, and digestive enzymes had little influence. Milk components presented a protective effect, and medicaments, especially anti-inflammatory drugs, influenced markedly the viability of the probiotic cultures, indicating that the beneficial effects of the two probiotic cultures to health are dependent of environmental factors encountered in the human gastrointestinal tract.

Effects of glutamine on the nuclear factor-kappaB signaling pathway of murine peritoneal macrophages

The aim of this study was to evaluate the effect of glutamine on the expression of proteins involved in the nuclear factor-kappaB (NF-kappa B) signaling pathway of murine peritoneal macrophages. Since glutamine is essential for the normal functioning of macrophages, it was hypothesized that in vitro glutamine supplementation would increase NF-kappa B activation. Peritoneal macrophages were pretreated with glutamine (0, 0.6, 2 and 10 mM) before incubation with lipopolysaccharide (LPS), and the effects of glutamine on the production of tumor necrosis factor-alpha and on the expression and activity of proteins involved in the NF-kappa B signaling pathway were studied by an enzyme linked immuno-sorbent assay, Western blotting, and an electrophoretic mobility shift assay. Glutamine treatment (2 and 10 mM) increased the activation of NF-kappa B in LPS-stimulated peritoneal macrophages (P < 0.05). In non-stimulated cells, glutamine treatment (2 and 10 mM) significantly reduced I kappa B-alpha protein expression (P < 0.05). Glutamine modulates NF-kappa B signaling pathway by reducing the level of I kappa B-alpha, leading to an increase in NF-kappa B within the nucleus in peritoneal macrophages.

Effects of Protein-Energy Malnutrition on NF-KappaB Signalling in Murine Peritoneal Macrophages

Protein-energy malnutrition (PEM) is an important public health problem affecting millions of people worldwide. PEM decreases resistance to infection, impairing a number of physiological processes. In unstimulated cells, NF-kappa B is kept from binding to its consensus sequence by the inhibitor I kappa B alpha, which retains NF-kappa B in the cytoplasm. Upon various signals, such as lipopolysaccharide (LPS), I kappa B alpha is rapidly degraded and NF-kappa B is induced to translocate into the nucleus, where it activates expression of various genes that participate in the inflammatory response, including those involved in the synthesis of TNF-alpha. TRAF-6 is a cytoplasmic adapter protein that links the stimulatory signal from Toll like receptor-4 to NF-kappa B. The aim of this study was to evaluate the effect of malnutrition on induction of TNF-a by LPS in murine peritoneal macrophages. We evaluated peritoneal cellularity, the expression of MyD88, TRAF-6, IKK, I kappa B alpha and NF-kappa B, NF-kappa B activation and TNF-alpha mRNA and protein synthesis inmacrophages. Two-month-old male BALB/Cmice were submitted to PEM with a low-protein diet that contained 2% protein, compared to 12% protein in the control diet. When the experimental group had lost about 20% of the original body weight, it was used in the subsequent experiments. Malnourished animals presented anemia, leucopenia and severe reduction in peritoneal cavity cellularity. TNF-a mRNA and protein levels of macrophages stimulated with LPS were significantly lower in malnourished animals. PEM also decreased TRAF-6 expression and NF-kappa B activation after LPS stimulation. These results led us to conclude that PEM changes NF-kappa B signalling pathway in macrophages to LPS stimulus.

Two physically, functionally, and developmentally distinct peritoneal macrophage subsets

The peritoneal cavity (PerC) is a unique compartment within which a variety of immune cells reside, and from which macrophages (Mempty set) are commonly drawn for functional studies. Here we define two Mempty set subsets that coexist in PerC in adult mice. One, provisionally called the large peritoneal Mempty set (LPM), contains approximately 90% of the PerC Mempty set in unstimulated animals but disappears rapidly from PerC following lipopolysaccharide (LPS) or thioglycolate stimulation. These cells express high levels of the canonical Mempty set surface markers, CD11b and F4/80. The second subset, referred to as small peritoneal Mempty set (SPM), expresses substantially lower levels of CD11b and F4/80 but expresses high levels of MHC-II, which is not expressed on LPM. SPM, which predominates in PerC after LPS or thioglycolate stimulation, does not derive from LPM. Instead, it derives from blood monocytes that rapidly enter the PerC after stimulation and differentiate to mature SPM within 2 to 4 d. Both subsets show clear phagocytic activity and both produce nitric oxide (NO) in response to LPS stimulation in vivo. However, their responses to LPS show key differences: in vitro, LPS stimulates LPM, but not SPM, to produce NO; in vivo, LPS stimulates both subsets to produce NO, albeit with different response patterns. These findings extend current models of Mempty set heterogeneity and shed new light on PerC Mempty set diversity, development, and function. Thus, they introduce a new context for interpreting (and reinterpreting) data from ex vivo studies with PerC Mempty set.

Immunogenicity of a Whole-Cell Pertussis Vaccine with Low Lipopolysaccharide Content in Infants

The lack of a clear correlation between the levels of antibody to pertussis antigens and protection against disease lends credence to the possibility that cell-mediated immunity provides primary protection against disease. This phase I comparative trial had the aim of comparing the in vitro cellular immune response and anti-pertussis toxin (anti-PT) immunoglobulin G (IgG) titers induced by a cellular pertussis vaccine with low lipopolysaccharide (LPS) content (wP(low) vaccine) with those induced by the conventional whole-cell pertussis (wP) vaccine. A total of 234 infants were vaccinated at 2, 4, and 6 months with the conventional wP vaccine or the wP(low) vaccine. Proliferation of CD3(+) T cells was evaluated by flow cytometry after 6 days of peripheral blood mononuclear cell culture with stimulation with heat-killed Bordetella pertussis or phytohemagglutinin (PHA). CD3(+), CD4(+), CD8(+), and T-cell receptor gamma delta-positive (gamma delta(+)) cells were identified in the gate of blast lymphocytes. Gamma interferon, tumor necrosis factor alpha, interleukin-4 (IL-4), and IL-10 levels in super-natants and serum anti-PT IgG levels were determined using enzyme-linked immunosorbent assay (ELISA). The net percentage of CD3(+) blasts in cultures with B. pertussis in the group vaccinated with wP was higher than that in the group vaccinated with the wP(low) vaccine (medians of 6.2% for the wP vaccine and 3.9% for the wP(low) vaccine; P = 0.029). The frequencies of proliferating CD4(+), CD8(+), and gamma delta(+) cells, cytokine concentrations in supernatants, and the geometric mean titers of anti-PT IgG were similar for the two vaccination groups. There was a significant difference between the T-cell subpopulations for B. pertussis and PHA cultures, with a higher percentage of gamma delta(+) cells in the B. pertussis cultures (P < 0.001). The overall data did suggest that wP vaccination resulted in modestly better specific CD3(+) cell proliferation, and gamma delta(+) cell expansions were similar with the two vaccines.

Effects of chlorogenic acid on neutrophil locomotion functions in response to inflammatory stimulus

Aim of the study: Species of Lychnophora are used in Brazilian folk medicine as analgesic and anti-inflammatory agents. Chlorogenic acid (CGA) and their analogues are important components of polar extracts of these species, as well in several European and Asian medicinal plants. Some of these phenolic compounds display anti-inflammatory effects. In this paper we report the isolation of CGA from Lychnophora salicifolia and its effects on functions involved in neutrophils locomotion. Materials and methods: LC-MS(n) data confirmed the presence of CGA in the plant. Actions of CGA were investigated on neutrophils obtained from peritoneal cavity of Wistar rats (4h after 1% oyster glycogen solution injection; 10 ml), and incubated with vehicle or with 50, 100 or 1000 mu M CGA in presence of lipopolysaccharide from Escherichia coil (LPS, 5 mu g/ml). Nitric oxide (NO; Griess reaction); prostaglandin E(2) (PGE(2)), interleukin-1 beta (IL-1 beta) and tumor necrosis factor-alpha [TNF-alpha; enzyme-linked immunosorbent assay (EIA)]; protein (flow cytometry) and gene (RT-PCR) expression of L-selectin, beta(2)integrin and platelet-endothelial cell adhesion molecule-1 (PECAM-1) were quantified. In vitro neutrophil adhesion to primary culture of microvascular endothelial cell (PMEC) and neutrophil migration in response to formyl-methionil-leucil-phenilalanine (fMLP, 10(-8)M, Boyden chamber) was determined. Results: CGA treatment did not modify the secretion of inflammatory mediators, but inhibited L-selectin cleavage and reduced beta(2) integrin, independently from its mRNA synthesis, and reduced membrane PECAM-1 expression: inhibited neutrophil adhesion and neutrophil migration induced by fMLP. Conclusions: Based on these findings, we highlight the direct inhibitory actions of CGA on adhesive and locomotion properties of neutrophils, which may contribute to its anti-inflammatory effects and help to explain the use of Lychnophora salicifolia as an anti-inflammatory agent. (C) 2011 Elsevier Ireland Ltd. All rights reserved.

Direct Zincation of Functionalized Aromatics and Heterocycles by Using a Magnesium Base in the Presence of ZnCl(2)

A wide range of polyfunctional aryl and heteroaryl zinc reagents were efficiently prepared in THF by using (TMP)(2)Mg center dot 2LiCl (TMP = 2,2,6,6-tetramethylpiperamidyl) in the presence of ZnCl(2). The possible pathways of this metalation procedure as well as possible reactive intermediates are discussed. This experimental protocol expands the tolerance of functional groups and allows an efficient zincation of sensitive heterocycles such as quinoxaline or pyrazine. The zincated arenes and heteroarenes react with various electrophiles providing the expected products in 60-95 % yield.