Differential gene expression between the biotrophic-like and saprotrophic mycelia of the witches` broom pathogen Moniliophthora perniciosa

Moniliophthora perniciosa is a hemibiotrophic fungus that causes witches` broom disease (WBD) in cacao. Marked dimorphism characterizes this fungus, showing a monokaryotic or biotrophic phase that causes disease symptoms and a later dikaryotic or saprotrophic phase. A combined strategy of DNA microarray, expressed sequence tag, and real-time reverse-transcriptase polymerase chain reaction analyses was employed to analyze differences between these two fungal stages in vitro. In all, 1,131 putative genes were hybridized with cDNA from different phases, resulting in 189 differentially expressed genes, and 4,595 reads were clusterized, producing 1,534 unigenes. The analysis of these genes, which represent approximately 21% of the total genes, indicates that the biotrophic-like phase undergoes carbon and nitrogen catabollite repression that correlates to the expression of phytopathogenicity genes. Moreover, downregulation of mitochondrial oxidative phosphorylation and the presence of a putative ngr1 of Saccharomyces cerevisiae could help explain its lower growth rate. In contrast, the saprotrophic mycelium expresses genes related to the metabolism of hexoses, ammonia, and oxidative phosphorylation, which could explain its faster growth. Antifungal toxins were upregulated and could prevent the colonization by competing fungi. This work significantly contributes to our understanding of the molecular mechanisms of WBD and, to our knowledge, is the first to analyze differential gene expression of the different phases of a hemibiotrophic fungus.

Modeling soil organic carbon dynamics in Oxisols of Ibiruba (Brazil) with the Century Model

introduction of conservation practices in degraded agricultural land will generally recuperate soil quality, especially by increasing soil organic matter. This aspect of soil organic C (SOC) dynamics under distinct cropping and management systems can be conveniently analyzed with ecosystem models such as the Century Model. In this study, Century was used to simulate SOC stocks in farm fields of the Ibiruba region of north central Rio Grande do Sul state in Southern Brazil. The region, where soils are predominantly Oxisols, was originally covered with subtropical woodlands and grasslands. SOC dynamics was simulated with a general scenario developed with historical data on soil management and cropping systems beginning with the onset of agriculture in 1900. From 1993 to 2050, two contrasting scenarios based on no-tillage soil management were established: the status quo scenario, with crops and agricultural inputs as currently practiced in the region and the high biomass scenario with increased frequency of corn in the cropping system, resulting in about 80% higher biomass addition to soils. Century simulations were in close agreement with SOC stocks measured in 2005 in the Oxisols with finer texture surface horizon originally under woodlands. However, simulations in the Oxisols with loamy surface horizon under woodlands and in the grassland soils were not as accurate. SOC stock decreased from 44% to 50% in fields originally under woodland and from 20% to 27% in fields under grasslands with the introduction of intensive annual grain crops with intensive tillage and harrowing operations. The adoption of conservation practices in the 1980s led to a stabilization of SOC stocks followed by a partial recovery of native stocks. Simulations to 2050 indicate that maintaining status quo would allow SOC stocks to recover from 81% to 86% of the native stocks under woodland and from 80% to 91 % of the native stocks under grasslands. Adoption of a high biomass scenario would result in stocks from 75% to 95% of the original stocks under woodlands and from 89% to 102% in the grasslands by 2050. These simulations outcomes underline the importance of cropping system yielding higher biomass to further increase SOC content in these Oxisols. This application of the Century Model could reproduce general trends of SOC loss and recovery in the Oxisols of the Ibiruba region. Additional calibration and validation should be conducted before extensive usage of Century as a support tool for soil carbon sequestration projects in this and other regions can be recommended. (C) 2009 Elsevier B.V. All rights reserved.

Test procedure for variable rate fertilizer on coffee

The objective was to develop and test a procedure for applying variable rates of fertilizers and evaluate yield response in coffee (Coffea arabica L.) with regard to the application of phosphorus and potassium. The work was conducted during the 2004 season in a 6.4 ha field located in central Sao Paulo state. Two treatments were applied with alternating strips of fixed and variable rates during the whole season: one following the fertilizing procedures recommended locally, and the other based on a grid soil sampling. A prototype pneumatic fertilizer applicator was used, carrying two conveyor belts, one for each row. Harvesting was done with a commercial harvester equipped with a customized volumetric yield monitor, separating the two treatments. Data were analyzed based on geostatistics, correlations and regressions. The procedure showed to be feasible and effective. The area that received fertilizer applications at a variable rate showed a 34% yield increase compared to the area that received a fixed rate. The variable rate fertilizer resulted in a savings of 23% in phosphate fertilizer and a 13% increase in potassium fertilizer, when compared to fixed rate fertilizer. Yield in 2005, the year after the variable rate treatments, still presented residual effect from treatments carried out during the previous cycle.

Large-scale ecological restoration of high-diversity tropical forests in SE Brazil

The complex interactions among endangered ecosystems, landowners` interests, and different models of land tenure and use, constitute an important series of challenges for those seeking to maintain and restore biodiversity and augment the flow of ecosystem services. Over the past 10 years, we have developed a data-based approach to address these challenges and to achieve medium and large-scale ecological restoration of riparian areas on private lands in the state of Sao Paulo, southeastern Brazil. Given varying motivations for ecological restoration, the location of riparian areas within landholdings, environmental zoning of different riparian areas, and best-practice restoration methods were developed for each situation. A total of 32 ongoing projects, covering 527,982 ha, were evaluated in large sugarcane farms and small mixed farms, and six different restoration techniques have been developed to help upscale the effort. Small mixed farms had higher portions of land requiring protection as riparian areas (13.3%), and lower forest cover of riparian areas (18.3%), than large sugarcane farms (10.0% and 36.9%, respectively for riparian areas and forest cover values). In both types of farms, forest fragments required some degree of restoration. Historical anthropogenic degradation has compromised forest ecosystem structure and functioning, despite their high-diversity of native tree and shrub species. Notably, land use patterns in riparian areas differed markedly. Large sugarcane farms had higher portions of riparian areas occupied by highly mechanized agriculture, abandoned fields, and anthropogenic wet fields created by siltation in water courses. In contrast, in small mixed crop farms, low or non-mechanized agriculture and pasturelands were predominant. Despite these differences, plantations of native tree species covering the entire area was by far the main restoration method needed both by large sugarcane farms (76.0%) and small mixed farms (92.4%), in view of the low resilience of target sites, reduced forest cover, and high fragmentation, all of which limit the potential for autogenic restoration. We propose that plantations should be carried out with a high-diversity of native species in order to create biologically viable restored forests, and to assist long-term biodiversity persistence at the landscape scale. Finally, we propose strategies to integrate the political, socio-economic and methodological aspects needed to upscale restoration efforts in tropical forest regions throughout Latin America and elsewhere. (C) 2010 Elsevier BA/. All rights reserved.

Annual growth rings in the mangrove Laguncularia racemosa (Combretaceae)

Stem discs from trees of known age were used to determine the periodic nature of the growth rings formed in Laguncularia racemosa and to describe the anatomical features of these rings. The growth rings were scarcely distinct on microscopic examination, but they were well distinguishable macroscopically, with alternating light brown and dark brown layers. Cross-dating analysis revealed the occurrence of annual growth rings in L. racemosa. The existence of annual growth rings in L. racemosa suggests that it may have great potential for dendrochronology and should encourage age-related studies on the dynamics of mangrove forests. These studies can be important for the evaluation of climate change impact on mangrove ecosystems, as well as for the analysis of effects related to climate variability on plant communities.

Hypoxia Inducible Factor-Dependent Regulation of Angiogenesis by Nitro-Fatty Acids

Objective-Nitro-fatty acids (NO(2)-FAs) are emerging as a new class of cell signaling mediators. Because NO(2)-FAs are found in the vascular compartment and their impact on vascularization remains unknown, we aimed to investigate the role of NO(2)-FAs in angiogenesis. Methods and Results-The effects of nitrolinoleic acid and nitrooleic acid were evaluated on migration of endothelial cell (EC) in vitro, EC sprouting ex vivo, and angiogenesis in the chorioallantoic membrane assay in vivo. At 10 mu mol/L, both NO(2)-FAs induced EC migration and the formation of sprouts and promoted angiogenesis in vivo in an NO-dependent manner. In addition, NO(2)-FAs increased intracellular NO concentration, upregulated protein expression of the hypoxia inducible factor-1 alpha (HIF-1 alpha) transcription factor by an NO-mediated mechanism, and induced expression of HIF-1 alpha target genes, such as vascular endothelial growth factor, glucose transporter-1, and adrenomedullin. Compared with typical NO donors such as spermine-NONOate and deta-NONOate, NO(2)-FAs were slightly less potent inducers of EC migration and HIF-1 alpha expression. Short hairpin RNA-mediated knockdown of HIF-1 alpha attenuated the induction of vascular endothelial growth factor mRNA expression and EC migration stimulated by NO(2)-FAs. Conclusion-Our data disclose a novel physiological role for NO(2)-FAs, indicating that these compounds induce angiogenesis in an NO-dependent mechanism via activation of HIF-1 alpha. (Arterioscler Thromb Vasc Biol. 2011;31:1360-1367.)

Molecular Modeling Suggests Cruzain Specificity for Peptide Primaquine Prodrugs

Chagas` disease, infection caused by the protozoan Trypanosoma cruzi, is an important, social and medical ailment in the Latin America. This disease is endemic in 21 countries, mostly Latin America countries, with more than 300,000 new cases every year and about 16-18 million infected people. Current therapy is not effective in the chronic phase of the disease. Thus, new and better drugs are urgently needed. In this sense, the in vitro activity of primaquine (PQ) was reported. Based on this, peptide prodrugs of primaquine containing dipeptides - lysine-arginine (LysArg), phenylalanine-alanine (PheAla) and phenylalanine-arginine (PheArg) -- as carriers, were designed to be selectively cleaved by cruzain, a specific cysteine protease of T. cruzi. The prodrugs have shown to be active against tripomastigote forms according to this order: LysArg-PQ> PheAla-PQ> PheArg-PQ. The molecular mechanism of action considered a probable nucleophilic attack of the catalytic residue of cruzain (Cys25) on the respective prodrug amide carbonyl carbon, releasing PQ. In order to test this hypothesis, molecular modeling studies were performed, physicochemical parameters and stereoelectronic features calculated by using the AM1 semi-empirical method suggest that the amide carbonyl carbon is favorable for cleavage, where the LysArg showed the most electronic reactive and sterically disposable, leading to the prodrug release and action. In addition, the docking study indicates the occurrence of specific interactions between prodrugs and the pockets S1 and S2 of cruzain through the dipeptides carriers, being the distance between cruzain Cys25 and the amide carbonyl group related to the biological activity of the prodrugs.

Enantioselective Arylations Catalyzed by Carbohydrate-Based Chiral Amino Alcohols

The application of carbohydrate-derived amino alcohols in the asymmetric arylation of aldehydes by using arylboronic acids as the source of transferable aryl groups is described. The best ligand is derived from the readily available sugar D-xylose and it mediates the addition of a range of arylboronic acids to various aromatic aldehydes in excellent yields and high enantiomeric excesses.

Cruzain inhibition by hydroxymethylnitrofurazone and nitrofurazone: investigation of a new target in Trypanosoma cruzi

Nitrofurazone (NF) and its derivative, hydroxymethylnitrofurazone (NFOH), have presented antichagasic activity. NFOH has higher activity and lower mutagenicity. The aim of this work was to assess whether NF and its derivative NFOH would also be inhibitors of cruzain, besides their trypanothione reductase inhibitory activity. In vitro cruzain inhibition tests were performed for both compounds, and the 50% inhibitory concentration (IC(50)) for NF and NFOH presented values of 22.83 +/- 1.2 mu M and 10.55 +/- 0.81 mu M, respectively. AM1 semi-empirical molecular modeling studies were performed to understand the activity of the compounds, corroborating the observed cruzain inhibitory activity.

Rubus rosaefolius Extract as a Natural Preservative Candidate in Topical Formulations

Even though the synthetic preservatives may offer a high antimicrobial efficacy, they are commonly related to adverse reactions and regarded as having potentially harmful effects caused by chronic consumption. The development of natural preservatives provides a way of reducing the amount of synthetic preservatives normally used in pharmaceutical and cosmetic preparations. In addition, these agents have less toxic effects and represent a possible natural and safer alternative of the preservatives. The purpose of this research was to evaluate the Rubus rosaefolius Smith extract efficiency as a natural preservative in base formulations. Of the extract, 0.2% (w/w) was assayed for its effectiveness of antimicrobial protection in two different base formulations (emulsion and gel). The microbial challenge test was performed following the standard procedures proposed by The United States Pharmacopoeia 33nd, European Pharmacopoeia 6th, Japanese Pharmacopoeia 15th, and the Cosmetics, Toiletries, and Fragrance Association using standardized microorganisms. The results demonstrated that R. rosaefolius extract at the studied concentration reduced the bacterial inocula, satisfying the criterion in all formulations, even though it was not able to present an effective preservative behavior against fungi. Thus, the investigation of new natural substances with preservative properties that could be applied in pharmaceutical and cosmetic products is relevant due to the possibility of substituting or decreasing the concentration of synthetic preservatives, providing a way for the development of safer formulas for the use of consumers.

Prospects in Lyophilization of Bovine Pericardium

Almost 30 years after the introduction of heart valve prostheses patients worldwide are benefiting from the implant of these devices. Among the various types of heart valves, the ones made of treated bovine pericardium have become a frequently used replacement of the heart`s native valve. Lyophilization, also known as freeze-drying, is an extremely useful technique for tissue storage for surgical applications. This article gives a brief overview on the current bovine pericardium lyophilization development, including the chemical modification to improve physical-chemical characteristics and the advanced technologies used to guarantee a high-quality product. It was shown that lyophilization process can be successfully applied as a method of bovine pericardium preservation and also as a technological tool to prepare new materials obtained by chemical modification of native tissues.

Contribution of peripheral vanilloid receptor to the nociception induced by injection of spermine in mice

Polyamines (putrescine, spermidine and spermine) are important endogenous regulators of ion channels, such as vanilloid (TRPV1), glutamatergic (NMDA or AMPA/kainate) and acid-sensitive (ASIC) receptors. In the present study, we have investigated the possible nociceptive effect induced by polyamines and the mechanisms involved in this nociception in vivo. The subcutaneous (s.c.) injection of capsaicin (as positive control), spermine, spermidine or putrescine produced nociception with ED(50) of 0.16 (0.07-0.39) nmol/paw, 0.4 (0.2-0.7) mu mol/paw, 0.3 (0.1-0.9) mu mol/paw and 3.2 (0.9-11.5) mu mol/paw, respectively. The antagonists of NMDA (MK801, 1 nmol/paw), AMPA/kainate (DNQX, 1 nmol/paw) or ASIC receptors (amiloride, 100 nmol/paw) failed to reduce the spermine-trigged nociception. However, the TRPV1 antagonists capsazepine or SB366791 (1 nmol/paw) reduced spermine-induced nociception, with inhibition of 81 +/- 10 and 68 +/- 9%, respectively. The previous desensitization with resiniferatoxin (RTX) largely reduced the spermine-induced nociception and TRPV1 expression in the sciatic nerve, with reductions of 82 +/- 9% and 67 +/- 11%, respectively. Furthermore, the combination of spermine (100 nmol/paw) and RTX (0.005 fmol/paw), in doses which alone were not capable of inducing nociception, produced nociceptive behaviors. Moreover, different concentrations of spermine (3-300 mu M) enhanced the specific binding of [(3)H](center dot)-RTX to TRPV1 receptor. Altogether, polyamines produce spontaneous nociceptive effect through the stimulation of TRPV1, but not of ionotropic glutamate or ASIC receptors. (C) 2011 Elsevier Inc. All rights reserved.

Effect of freeze-drying on the mechanical, physical and morphological properties of glutaraldehyde-treated bovine pericardium: evaluation of freeze-dried treated bovine pericardium properties

Purpose: Biomaterials have been widely used in the field of regenerative medicine. Bovine pericardium tissue has been successfully used as a bioprosthetic material in manufacturing heart valves, but studies concerning the tissue are ongoing in order to improve its storage, preservation and transportation. This article provides an overview of the characteristics of bovine pericardium tissue chemically treated after the freeze-drying process. These characteristics are essential to evaluate the changes or damage to the tissue during the process. Methods: The mechanical properties of the tissue were analyzed by three different methods due to its anisotropic characteristics. The physical properties were analyzed by a colorimetric method, while the morphological properties were evaluated by scanning electron microscopy (SEM). Results: The freeze-dried bovine pericardium showed no significant change in its mechanical properties. There was no significant change in the elasticity of the tissue (p > 0.05) and no color change. In addition, SEM analysis showed that the freeze-dried samples did not suffer structural collapse. Conclusions: It was concluded that glutaraldehyde-treated bovine pericardium tissue showed no significant change in its properties after the freeze-drying process.

Host-guest complexation of a nitroheterocyclic compound with cyclodextrins: a spectrofluorimetric and molecular modeling study

Nitroheterocyclic compounds (NC) were candidate drugs proposed for Chagas disease chemotherapy. In this study, we investigated the complexation of hydroxymethylnitrofurazone (NFOH), a potential antichagasic compound, with alpha-cyclodextrin (alpha-CD), beta-cyclodextrin (beta-CD), Hydroxypropyl-beta-cyclodextrin (HP-beta-CD), Dimethyl-beta-cyclodextrin (DM-beta-CD) and gamma-cyclodextrin (gamma-CD) by fluorescence spectroscopy and molecular modeling studies. Hildebrand-Benesi equation was used to calculate the formation constants of NFOH with cyclodextrins based on the fluorescence differences in the CDs solution. The complexing capacity of NFOH with different CDs was compared through the results of association constant according to the following order: DM-beta-CD > beta-CD > alpha-CD > HP-beta-CD > gamma-CD. Molecular modeling studies give support for the experimental assignments, in favor of the formation of an inclusion complex between cyclodextrins with NFOH. This is an important study to investigate the effects of different kinds of cyclodextrins on the inclusion complex formation with NFOH and to better characterize a potential formulations to be used as therapeutic options for the oral treatment of Chagas disease.

Mitochondrial function in the yeast form of the pathogenic fungus Paracoccidioides brasiliensis

Differences between the respiratory chain of the fungus Paracoccidioides brasiliensis and its mammalian host are reported. Respiration, membrane potential, and oxidative phosphorylation in mitochondria from P. brasiliensis spheroplasts were evaluated in situ, and the presence of a complete (Complex I-V) functional respiratory chain was demonstrated. In succinate-energized mitochondria, ADP induced a transition from resting to phosphorylating respiration. The presence of an alternative NADH-ubiquinone oxidoreductase was indicated by: (i) the ability to oxidize exogenous NADH and (ii) the lack of sensitivity to rotenone and presence of sensitivity to flavone. Malate/NAD(+)-supported respiration suggested the presence of either a mitochondrial pyridine transporter or a glyoxylate pathway contributing to NADH and/or succinate production. Partial sensitivity of NADH/succinate-supported respiration to antimycin A and cyanide, as well as sensitivity to benzohydroxamic acids, suggested the presence of an alternative oxidase in the yeast form of the fungus. An increase in activity and gene expression of the alternative NADH dehydrogenase throughout the yeast`s exponential growth phase was observed. This increase was coupled with a decrease in Complex I activity and gene expression of its subunit 6. These results support the existence of alternative respiratory chain pathways in addition to Complex I, as well as the utilization of NADH-linked substrates by P. brasiliensis. These specific components of the respiratory chain could be useful for further research and development of pharmacological agents against the fungus.