Long-Chain Acyl-Homoserine Lactones from Methylobacterium mesophilicum: Synthesis and Absolute Configuration

The acyl-homoserine lactones (acyl-HSLs) produced by Methylobacterium mesophilicum isolated from orange trees infected with the citrus variegated chlorosis (CVC) disease have been studied, revealing the occurrence of six long-chain acyl-HSLs, i.e., the saturated homologues (S)-N-dodecanoyl (1) and (S)-N-tetradecanoyl-HSL (5), the uncommon odd-chain N-tridecanoyl-HSL (3), the new natural product (S)-N-(2E)-dodecenoyl-HSL (2), and the rare unsaturated homologues (S)-N-(7Z)-tetradecenoyl (4) and (S)-N-(2E,7Z)-tetradecadienyl-HSL (6). The absolute configurations of all HSLs were determined as 3S. Compounds 2 and 6 were synthesized for the first time. Antimicrobial assays with synthetic acyl-HSLs against Gram-positive bacterial endophytes co-isolated with M. mesophilicum from CVC-infected trees revealed low or no antibacterial activity.

Relationship between occurrence of Panama disease in banana trees of cv. Nanicao and nutrients in soil and leaves

Relationship between occurrence of Panama disease in banana trees of cv. Nanicao and nutrients in soil and leaves The objective of the present work was to verify if the incited symptoms in banana trees cv. Nanicao, belonging to the subgroup Cavendish, in Vale do Ribeira, are related to levels of nutrients in soil and leaves. Sixteen areas in Vale do Ribeira were selected, one half with symptomatic plants and the other with healthy plants. In those areas the third leaf of five plants and the soil near those plants were collected, at depths from 0 to 20 cm and from 20 to 40 cm. At both depths of the sampled soil, levels of Ca, Mg, PO(4)(-3), S and cationic exchange capacity (CEC) were significantly different among the areas, and the low values of these elements were present in the areas containing symptomatic plants. At both depths, Mg, Al and H in relation to CEC were significantly different among the areas, and the low values of Mg and high of Al and H were present in the areas with symptomatic plants. The N, K and S in the leaves were significantly different among the areas. These elements showed low values in the areas containing symptomatic plants. Despite the fact that some amounts of macronutrients of the soil and of the leaves are present only in the areas containing plants of Nanicao with symptoms similar to fusariosis, proof of a possible occurrence of race of the pathogen should be looked for in Vale do Ribeira.

Hereditary hemochromatosis: Mutations in genes involved in iron homeostasis in Brazilian patients

Background: p.C282Y mutation and rare variants in the HFE gene have been associated with hereditary hemochromatosis (HH). HH is also caused by mutations in other genes, such as the hemojuvelin (HJV), hepcidin (HAMP), transferrin receptor 2 (TFR2) and ferroportin (SLC40A1). The low rate homozygous p.C282Y mutation in Brazil is suggestive that mutations in non-HFE genes may be linked to HH phenotype. Aim: To screen exon-by-exon DNA sequences of HFE, HJV, HAMP, TFR2 and SLC40A1 genes to characterize the molecular basis of HH in a sample of the Brazilian population. Materials and methods: Fifty-one patients with primary iron overload (transferrin saturation >= 50% in females and >= 60% in males) were selected. Subsequent bidirectional DNA sequencing of HFE, HJV, HAMP, TFR2 and SLC40A1 exons was performed. Results: Thirty-seven (72.5%) out of the 51 patients presented at least one HFE mutation. The most frequent genotype associated with HH was the homozygous p.C282Y mutation (n = 11, 21.6%). In addition, heterozygous HFE p.S65C mutation was found in combination with p.H63D in two patients and homozygous HFE p.H63D was found in two patients as well. Sequencing in the HJV and HAMP genes revealed HJV p.E302K, HJV p.A310G, HJV p.G320V and HAMP p.R59G alterations. Molecular and clinical diagnosis of juvenile hemochromatosis (homozygous form for the HJV p.G320V) was described for the first time in Brazil. Three TFR2 polymorphisms (p.A75V, p.A617A and p.R752H) and six SLC40A1 polymorphisms (rs13008848, rs11568351, rs11568345, rs11568344, rs2304704, rs11568346) and the novel mutation SLC40A1 p.G204S were also found. Conclusions: The HE p.C282Y in homozygosity or in heterozygosity with p.H63D was the most frequent mutation associated with HH in this sample. The HJV p.E302K and HAMP p.R59G variants, and the novel SLC40A1 p.G2045 mutation may also be linked to primary iron overload but their role in the pathophysiology of HH remain to be elucidated. (C) 2011 Elsevier Inc. All rights reserved.

HFE gene mutations in patients with primary iron overload: Is there a significant improvement in molecular diagnosis yield with HFE sequencing?

Rare HFE variants have been shown to be associated with hereditary hemochromatosis (HH), an iron overload disease. The low frequency of the HFE p.C282Y mutation in HH-affected Brazilian patients may suggest that other HFE-related mutations may also be implicated in the pathogenesis of HH in this population. The main aim was to screen for new HFE mutations in Brazilian individuals with primary iron overload and to investigate their relationship with HH. Fifty Brazilian patients with primary iron overload (transferrin saturation >50% in females and 60% in males) were selected. Subsequent bidirectional sequencing for each HFE exon was performed. The effect of HFE mutations on protein structure were analyzed by molecular dynamics simulation and free binding energy calculations. p.C282Y in homozygosis or in heterozygosis with p.H63D were the most frequent genotypic combinations associated with HH in our sample population (present in 17 individuals, 34%). Thirty-six (72.0%) out of the 50 individuals presented at least one HFE mutation. The most frequent genotype associated with HH was the homozygous p.C282Y mutation (n = 11, 22.0%). One novel mutation (p.V256I) was indentified in heterozygosis with the p.H63D mutation. In silico modeling analysis of protein behavior indicated that the p.V256I mutation does not reduce the binding affinity between HFE and beta 2-microglobulin ((beta 2M) in the same way the p.C282Y mutation does compared with the native HFE protein. In conclusion, screening of HFE through direct sequencing, as compared to p.C282Y/p.H63D genotyping, was not able to increase the molecular diagnosis yield of HH. The novel p.V256I mutation could not be implicated in the molecular basis of the HH phenotype, although its role cannot be completely excluded in HH-phenotype development. Our molecular modeling analysis can help in the analysis of novel, previously undescribed, HFE mutations. (C) 2010 Elsevier Inc. All rights reserved.

Dissemination of bla(IMP-1)-carrying integron In86 among Klebsiella pneumoniae isolates harboring a new trimethoprim resistance gene dfr23

The genetic context of the bla(IMP-1) gene was evaluated in 9 Klebsiella pneumoniae isolates recovered from 2 hospitals in Sao Paulo, Brazil. All isolates harbored a copy of In86 carrying bla(Imp-1), aac(6`)-31, and aadAl. Eight strains from the same hospital also carried another class I integron harboring a new trimethoprim resistance gene (dfr23) that was chromosomally embedded. In86 was likely to be in a 30-kb nontransferable plasmid and was flanked upstream by a sequence identical to one identified in an IMP-1-producing Pseudomonas putida isolate. The bla(IMP-1)-carrying integron In86 was recently reported from nonfermentative bacilli isolated in Sao Paulo. These isolates appear to be the Source of this integron now acquired by K. pneumoniae strains from different hospitals in the same city. Metallo-beta-lactamase production is still rare among Enterobacteriaceae isolates in Brazil, but the acquisition of genetic structures carrying these mobile resistance determinants is worrisome and could lead to an increase in the prevalence of these phenotypes of resistance. (C) 2008 Elsevier Inc. All rights reserved.

Nutritional status of selenium in Alzheimer`s disease patients

Studies have shown that various antioxidants are decreased in different age-related degenerative diseases and thus, oxidative stress would have a central role in the pathogenesis of many disorders that involve neuronal degeneration, including Alzheimer`s disease (AD). The present study aimed to assess the nutritional status of Se in AD patients and to compare with control subjects with normal cognitive function. The case control study was carried out on a group of elderly with AD (n 28) and compared with a control group (n 29), both aged between 60 and 89 years. Se intake was evaluated by using a 3-d dietary food record. Se was evaluated in plasma, erythrocytes and nails by using the method of hydride generation atomic absorption spectroscopy. Deficient Se intake was largely observed in the AD group. AD patients showed significantly lower Se levels in plasma, erythrocytes and nails (32.59 mu g/l, 43.74 mu g/l and 0.302 mu g/g) when compared with the control group (50.99 mu g/l, 79.16 mu g/l and 0.400 mu g/g). The results allowed us to suggest that AD has an important relation with Se deficiency.

Hand position on the bunch and source-sink ratio influence the banana fruit susceptibility to crown rot disease

The postharvest development of crown rot of bananas depends notably on the fruit susceptibility to this disease at harvest. It has been shown that fruit susceptibility to crown rot is variable and it was suggested that this depends on environmental preharvest factors. However, little is known about the preharvest factors influencing this susceptibility. The aim of this work was to evaluate the extent to which fruit filling characteristics during growth and the fruit development stage influence the banana susceptibility to crown rot. This involved evaluating the influence of (a) the fruit position at different levels of the banana bunch (hands) and (b) changing the source-sink ratio (So-Si ratio), on the fruit susceptibility to crown rot. The fruit susceptibility was determined by measuring the internal necrotic surface (INS) after artificial inoculation of Colletotrichum musae. A linear correlation (r = -0.95) was found between the hand position on the bunch and the INS. The So-Si ratio was found to influence the pomological characteristics of the fruits and their susceptibility to crown rot. Fruits of bunches from which six hands were removed (two hands remaining on the bunch) proved to be significantly less susceptible to crown rot (INS = 138.3 mm 2) than those from bunches with eight hands (INS = 237.9 mm 2). The banana susceptibility to crown rot is thus likely to be influenced by the fruit development stage and filling characteristics. The present results highlight the importance of standardising hand sampling on a bunch when testing fruit susceptibility to crown rot. They also show that hand removal in the field has advantages in the context of integrated pest management, making it possible to reduce fruit susceptibility to crown rot while increasing fruit size.

In Vitro Simultaneous Transfer of Lipids to HDL in Coronary Artery Disease and in Statin Treatment

The exchange of lipids with cells and other lipoproteins is a crucial process in HDL metabolism and for HDL antiatherogenic function. Here, we tested a practical method to quantify the simultaneous transfer to HDL of phospholipids, free-cholesterol, esterified cholesterol and triacylglycerols and to verify the lipid transfer in patients with coronary artery disease (CAD) or undergoing statin treatment. Twenty-eight control subjects without CAD, 27 with CAD and 25 CAD patients under simvastatin treatment were studied. Plasma samples were incubated with a donor nanoemulsion prepared by ultrasonication of the constituent lipids and labeled with radioactive lipids; % lipids transferred to HDL were quantified in the HDL-containing supernatant after chemical precipitation of non-HDL fractions and the nanoemulsion. The assay was precise and reproducible. Increase of temperature (4-37 A degrees C), of incubation period (5 min to 2 h), of HDL-cholesterol concentration (33-244 mg/dL) and of mass of nanoemulsion lipids (0.075-0.3 mg/mu L) resulted in increased lipid transfer from the nanoemulsion to HDL. In contrast, increasing pH (6.5-8.5) and albumin concentration (3.5-7.0 g/dL) did not affect lipid transfer. There was no difference between CAD and control non-CAD with regard to the lipid transfer, but statin treatment reduced the transfer to HDL of all four lipids. The test herein described is a valid and practical tool for exploring an important aspect of HDL metabolism.

WHO comparative evaluation of serologic assays for Chagas disease

Evaluation of commercially available test kits for Chagas disease for use in blood bank screening is difficult due to a lack of large and well-characterized specimen panels. This study presents a collaborative effort of Latin American blood centers and the World Health Organization (WHO) to establish such a panel. A total of 437 specimens, from 10 countries were collected and sent to the WHO Collaborating Center in Sao Paulo and used to evaluate 19 screening assays during 2001 through 2005. Specimens were assigned a positive or negative status based on concordant results in at least three of the four confirmatory assays (indirect immunofluorescence, Western blot, radioimmunoprecipitation assay, and recombinant immunoblot). Of the 437 specimens, 168 (39%) were characterized as positive, 262 (61%) were characterized as negative, and 7 (2%) were judged inconclusive and excluded from the analysis. Sensitivity and specificity varied considerably: 88 to 100 and 60 to 100 percent, respectively. Overall, enzyme immunoassays (EIAs) performed better than the other screening assays. Four EIAs had both parameters higher than 99 percent. Of the four confirmatory assays, only the RIPA gave a 100 percent agreement with the final serologic status of the specimens. The sensitivities and specificities of at least four of the commercially available EIAs for Chagas disease are probably high enough to justify their use for single-assay screening of blood donations. Our data suggest that the majority of commercially available indirect hemagglutination assays should not be used for blood donor screening and that the RIPA could be considered a gold standard for evaluating the performance of other assays.

Sigatoka disease reduces the greenlife of bananas

Sigatoka disease (SD) of bananas is caused by the pathogenic fungus Mycosphaerella musicola Leach. This disease provokes necrotic lesions on leaves and serious infestations can lead to a substantial reduction in the leaf area of infected plants and thus to yield losses. In addition to these effects on yield, SO was found to have an impact on fruit quality, especially because exported bananas ripen prematurely. In the present work, a plantation survey and experiments have been conducted in Guadeloupe (FWI) to assess the effect of this disease on the greenlife of bananas harvested at a constant physiological age, as measured in degree-days (dd). Our results revealed that bananas harvested at 900 dd from plants with high Sigatoka disease severity had normal diameter growth, but a shorter greenlife (GL) than bananas harvested from uninfected plants. These results indicate that SD is directly responsible for the reduction of banana greenlife since the reduction of GL could not be attributed to the harvest of fruits at a more advanced physiological age (dd). Furthermore, a correlation was noted between SO severity and GL The potential physiological mechanisms involved are also discussed. (C) 2008 Elsevier Ltd. All rights reserved.

Nitric oxide donor trans-[RuCl([15]aneN(4))NO]2+as a possible therapeutic approach for Chagas` disease

Background and purpose: Benznidazole (Bz) is the therapy currently available for clinical treatment of Chagas` disease. However, many strains of Trypanosoma cruzi parasites are naturally resistant. Nitric oxide (NO) produced by activated macrophages is crucial to the intracellular killing of parasites. Here, we investigate the in vitro and in vivo activities against T. cruzi, of the NO donor, trans-[RuCl([15]aneN(4))NO]2+. Experimental approach: Trans-[RuCl([15]aneN(4))NO]2+ was incubated with a partially drug-resistant T. cruzi Y strain and the anti-proliferative (epimastigote form) and trypanocidal activities (trypomastigote and amastigote) evaluated. Mice were treated during the acute phase of Chagas` disease. The anti-T. cruzi activity was evaluated by parasitaemia, survival rate, cardiac parasitism, myocarditis and the curative rate. Key results: Trans-[RuCl([15]aneN(4))NO]2+ was 10- and 100-fold more active than Bz against amastigotes and trypomastigotes respectively. Further, trans-[RuCl([15]aneN(4))NO]2+ (0.1 mM) induced 100% of trypanocidal activity (trypomastigotes forms) in vitro. Trans-[RuCl([15]aneN(4))NO]2+ induced permanent suppression of parasitaemia and 100% survival in a murine model of acute Chagas` disease. When the drugs were given alone, parasitological cures were confirmed in only 30 and 40% of the animals treated with the NO donor (3.33 mu mol center dot kg-1 center dot day-1) and Bz (385 mu mol center dot kg-1 center dot day-1), respectively, but when given together, 80% of the animals were parasitologically cured. The cured animals showed an absence of myocarditis and a normalisation of cytokine production in the sera. In addition, no in vitro toxicity was observed at the tested doses. Conclusions and implications: These findings indicate that trans-[RuCl([15]aneN(4))NO]2+ is a promising lead compound for the treatment of human Chagas` disease. This article is commented on by Machado et al., pp. 258-259 of this issue. To view this commentary visit http://dx.doi.org/10.1111/j.1476-5381.2010.00662.x and to view a related paper in this issue by Silva et al. visit http://dx.doi.org/10.1111/j.1476-5381.2010.00524.x.

Precursor system of liquid crystalline phase containing propolis microparticles for the treatment of periodontal disease: Development and characterization

Precursor systems of liquid crystalline phase were prepared using the surfactant PPG-5-Ceteth-20, isopropyl myristate, and water; gelatin microparticles containing propolis were then added into these systems. Homogeneity of dispersion, the in-system microparticle morphology, and sedimentation behavior of each formulation were evaluated. The rheological and mechanical properties (hardness, compressibility, and adhesiveness), the work of syringing, and the propolis release profile were also evaluated. All the formulations exhibited pseudoplastic flow and thixotropy, and they displayed storage modulus, loss modulus, dynamic viscosity, and loss tangent that depended on temperature, frequency, and composition. Mechanical properties varied significantly among the formulations being affected by changes in the composition and temperature. Raising the concentration of surfactant and adding propolis microparticles significantly decreased the work of syringing. The drug release was non-Fickian (anomalous) and there was no significant difference between the tested systems in the times required for 10%, 30%, and 50% release of the initial drug loading.

(-)-Hinokinin-loaded poly(d,l-lactide-co-glycolide) microparticles for Chagas disease

The (-)-hinokinin display high activity against Trypanosoma cruzi in vitro and in vivo. (-)-Hinokinin-loaded poly(d,l-lactide-co-glycolide) microparticles were prepared and characterized in order to protect (-)-hinokinin of biological interactions and promote its sustained release for treatment of Chagas disease. The microparticles contain (-)-hinokinin were prepared by the classical method of the emulsion/solvent evaporation. The scanning electron microscopy, light-scattering analyzer were used to study the morphology and particle size, respectively. The encapsulation efficiency was determined, drug release studies were kinetically evaluated, and the trypanocidal effect was evaluated in vivo. (-)-Hinokinin-loaded microparticles obtained showed a mean diameter of 0.862 A mu m with smooth surface and spherical shape. The encapsulation efficiency was 72.46 A +/- 2.92% and developed system maintained drug release with Higuchi kinetics. The preparation method showed to be suitable, since the morphological characteristics, encapsulation efficiency, and in vitro release profile were satisfactory. In vivo assays showed significant reduction of mice parasitaemia after administration of (-)-hinokinin-loaded microparticles. Thus, the developed microparticles seem to be a promising system for sustained release of (-)-hinokinin for treatment of Chagas disease.

Trypanosoma cruzi: The effects of zinc supplementation in the immune response during the course of experimental disease

Zinc is an essential nutritional component required for normal development and maintenance of immune functions. The possible effects of zinc in upregulating the host immune response during the acute and chronic phases of experimental Chagas` disease were evaluated. In young, infected and Zn-supplemented animals, higher concentrations of IFN-gamma and NO were observed. During the chronic phase, decreased concentrations of NO and IFN-gamma were found for older infected animals that received Zn supplementation. For young animals, hearts from Zn-supplemented groups displayed reduced inflammatory infiltrate, heart weight and number of amastigote burdens. For older, infected and Zn-supplemented animals amastigote nests were absent with reduced inflammatory cell infiltrate. This study identifies a potentially novel therapeutic approach that could control the parasite load during acute phase of disease, consequently preventing the deleterious, parasite-elicited responses observed during chronic phase. (C) 2008 Elsevier Inc. All rights reserved.