Heterologous expression and biochemical characterization of an α1,2-mannosidase encoded by the Candida albicans MNS1 gene

Protein glycosylation pathways, commonly found in fungal pathogens, offer an attractive new area of study for the discovery of antifungal targets. In particular, these post-translational modifications are required for virulence and proper cell wall assembly in Candida albicans, an opportunistic human pathogen. The C. albicans MNS1 gene is predicted to encode a member of the glycosyl hydrolase family 47, with 1,2-mannosidase activity. In order to characterise its activity, we first cloned the C. albicans MNS1 gene into Escherichia coli, then expressed and purified the enzyme. The recombinant Mns1 was capable of converting a Man9GlcNAc2 N-glycan core into Man8GlcNAc2 isomer B, but failed to process a Man5GlcNAc2-Asn N-oligosaccharide. These properties are similar to those displayed by Mns1 purified from C. albicansmembranes and strongly suggest that the enzyme is an ±1,2-mannosidase that is localised to the endoplasmic reticulum and involved in the processing of N-linked mannans. Polyclonal antibodies specifically raised against recombinant Mns1 also immunoreacted with the soluble ±1,2-mannosidases E-I and E-II, indicating that Mns1 could share structural similarities with both soluble enzymes. Due to the high degree of similarity between the members of family 47, it is conceivable that these antibodies may recognise ±1,2-mannosidases in other biological systems as well.

Glyphosate behavior in a Rhodic Oxisol under no-till and conventional agricultural systems

The behavior of glyphosate in a Rhodic Oxisol, collected from fields under no-till and conventional management systems in Ponta Grossa, Parana state (Brazil) was investigated. Both agricultural systems had been in production for 23 years. Glyphosate mineralization, soil-bound forms, sorption and desorption kinetics, sorption/desorption batch experiments, and soil glyphosate phythoavailability (to Panicum maximum) were determined. The mineralization experiment was set up in a completely randomized design with a 2 x 2 factorial scheme (two management systems and two 14C radiolabelled positions in the glyphosate), with five replicates. 14CO2 evolution was measured in 7-day intervals during 63 days. The glyphosate sorption kinetics was investigated in a batch experiment, employing a glyphosate concentration of 0.84 mg L-1. The equilibration solution was 0.01 mol L-1 CaCl2 and the equilibration times were 0, 10, 30, 60, 120, 240, and 360 min. Sorption/desorption of glyphosate was also investigated using equilibrium batch experiments. Five different concentrations of the herbicide were used for sorption (0.42, 0.84, 1.68, 3.36, and 6.72 mg L-1) and one concentration for desorption. Glyphosate phytoavailability was analyzed in a 2 x 5 factorial scheme with two management systems and five glyphosate concentrations added to soil (0, 4.2, 8.4, 42.0, and 210.0 µg g-1) in a completely randomized design. Phytotoxicity symptoms in P. maximum were evaluated for different periods. The soil under both management systems showed high glyphosate sorption, which impeded its desorption and impaired the mineralization in the soil solution. Practically the total amount of the applied glyphosate was quickly sorbed (over 90 % sorbed within 10 min). Glyphosate bound to residues did not have adverse effects on P. maximum growth. The mineralization of glyphosate was faster under no-till and aminomethylphosphonic acid was the main glyphosate metabolite.

Influence of the temperature and type of salt on the phase equilibrium of peg 1500 + potassium phosphate and peg 1500 + sodium citrate aqueous two-phase systems

The present work analyzed the effect of the temperature and type of salt on the phase equilibrium of aqueous two-phase systems (ATPS) formed by poly (ethylene glycol) (PEG) 1500 + potassium phosphate, from (278.15 to 318.15) K, and PEG 1500 + sodium citrate, from (278.15 to 298.15) K. The rise of the temperature normally increased the slope of the tie line (STL). With respect to the influence of the type of salt, sodium citrate showed better capability to induce phase separation, when compared to potassium phosphate.

Biological and morphometric aspects and rearing of Syssphinx molina (Cramer) (Lepidoptera: Saturniidae), a defoliator of parica tree

The objective of this research was to describe the biological and morphometric aspects of the parica tree defoliator, Syssphinx molina (Cramer), and make recommendations about the insect rearing. The life cycle was 62.9 days with mean periods for the egg, larval, pre-pupal and pupal stages of 5.6, 31.1, 2.2 and 16.6 days respectively. The pupal viability was 60.5% for females and 48.6% for males. The sexual ratio was 0.5 with mean production of 182.3 ± 2.2 eggs per female and egg viability of 24.3%. The mean longevity was 7.9 ± 2 and 8.1 ± 3 days for females and males respectively. Other parameters were also observed and compared with description of other Saturniidae species.

Compared nymphal development of Tingis americana (Hemiptera, Tingidae) in two Handroanthus species (Bignoniaceae) and reproductive parameters in seedlings of Handroanthus heptaphyllus

Nymphs and adults of Tingis americana Drake, 1922 were found feeding on leaves of Handroanthus heptaphyllus (Vell.) Mattos and Handroanthus chrysotrichus (Mart. ex A. DC.) Mattos in the Botanic Garden, Porto Alegre, state of Rio Grande do Sul, Brazil. This is the first record of T. americana on these host plants and in the southern Brazil. We aimed to compare the nymphal development on both hosts and to analyze the reproductive parameters on H. heptaphyllus (25 ± 1ºC; 60 ± 10% RH; 16 h photophase). The mean nymphal period (days) was shorter in individuals reared on H. heptaphyllus (12.69 ± 0.076) than on H. chrysotrichus (19.11 ± 0.208) (P < 0.0001), however, nymph viability was similar. On H. heptaphyllus, the mean embryonic period lasted 12.32 ± 0.274 days and the egg viability was 92%. The mean total and daily fecundity were 310.0 ± 19.40 eggs/female and 7.46 ± 0.302 eggs/female/day, respectively. Paired males and females showed similar longevity (P = 0.0691), while unpaired females lived longer than unpaired males (P = 0.0460).

Coadaptation and malaria control

Malaria emerges from a disequilibrium of the system 'human-plasmodium-mosquito' (HPM). If the equilibrium is maintained, malaria does not ensue and the result is asymptomatic plasmodium infection. The relationships among the components of the system involve coadaptive linkages that lead to equilibrium. A vast body of evidence supports this assumption, including the strategies involved in the relationships between plasmodium and human and mosquito immune systems, and the emergence of resistance of plasmodia to antimalarial drugs and of mosquitoes to insecticides. Coadaptive strategies for malaria control are based on the following principles: (1) the system HPM is composed of three highly complex and dynamic components, whose interplay involves coadaptive linkages that tend to maintain the equilibrium of the system; (2) human and mosquito immune systems play a central role in the coadaptive interplay with plasmodium, and hence, in the mainten-ance of the system's equilibrium; the under- or overfunction of human immune system may result in malaria and influence its severity; (3) coadaptation depends on genetic and epigenetic phenomena occurring at the interfaces of the components of the system, and may involve exchange of infectrons (genes or gene fragments) between the partners; (4) plasmodia and mosquitoes have been submitted to selective pressures, leading to adaptation, for an extremely long while and are, therefore, endowed with the capacity to circumvent both natural (immunity) and artificial (drugs, insecticides, vaccines) measures aiming at destroying them; (5) since malaria represents disequilibrium of the system HPM, its control should aim at maintaining or restoring this equilibrium; (6) the disequilibrium of integrated systems involves the disequilibrium of their components, therefore the maintenance or restoration of the system's equilibrium depend on the adoption of integrated and coordinated measures acting on all components, that means, panadaptive strategies. Coadaptive strategies for malaria control should consider that: (1) host immune response has to be induced, since without it, no coadaptation is attained; (2) the immune response has to be sustained and efficient enough to avoid plasmodium overgrowth; (3) the immune response should not destroy all parasites; (4) the immune response has to be well controlled in order to not harm the host. These conditions are mostly influenced by antimalarial drugs, and should also be taken into account for the development of coadaptive malaria vaccines.

Nutrient supply by mass flow and diffusion to maize plants in response to soil aggregate size and water potential

Nutrients are basically transported to the roots by mass flow and diffusion. The aim of this study was to quantify the contribution of these two mechanisms to the acquisition of macronutrients (N, P, K, Ca, Mg, and S) and cationic micronutrients (Fe, Mn, Zn, and Cu) by maize plants as well as xylem exudate volume and composition in response to soil aggregate size and water availability. The experiment was conducted in a greenhouse with samples of an Oxisol, from under two management systems: a region of natural savanna-like vegetation (Cerradão, CER) and continuous maize under conventional management for over 30 years (CCM). The treatments were arranged in a factorial [2 x (1 + 2) x 2] design, with two management systems (CER and CCM), (1 + 2) soil sifted through a 4 mm sieve and two aggregate classes (< 0.5 mm and 0.5 - 4.0 mm) and two soil matric potentials (-40 and -10 kPa). These were evaluated in a randomized block design with four replications. The experiment was conducted for 70 days after sowing. The influence of soil aggregate size and water potential on the nutrient transport mechanisms was highest in soil samples with higher nutrient concentrations in solution, in the CER system; diffusion became more relevant when water availability was higher and in aggregates < 0.5 mm. The volume of xylem exudate collected from maize plants increased with the decrease in aggregate size and the increased availability of soil water in the CER system. The highest Ca and Mg concentrations in the xylem exudate of plants grown on samples from the CER system were related to the high concentrations of these nutrients in the soil solution of this management system.

ESTABLISHMENT TECHNIQUES FOR TROPICAL LEGUMES IN THE UNDERSTORY OF A EUCALYPTUS PLANTATION

This study evaluated establishment methods for a mixture of herbaceous forage legumes [Centrosema acutifolium, Clitoria ternatea, Pueraria phaseoloides, Stylosanthes Campo Grande (Stylosanthes capitata + S. macrocephala), Calopogonium mucunoides, Lablab purpureus, Arachis pintoi, and Aeschynomene villosa] under the shade of an Eucalyptus grandis plantation submitted to thinning (40%) 8 years after planting in Anhembi, São Paulo (22°40'S, 48°10'W, altitude of 455 m). The experiment started in December 2008 and consisted of the comparison of the following four types of seed incorporation by light disc harrowing: (1) broadcast sowing without seed incorporation; disc harrowing before (2) or after (3) planting, and (4) disc harrowing before and after planting. Ninety days after planting, the number of legume plants/m2 and the percentage of ground cover by the plants varied between the treatments tested; however, the treatments had no effect on the dry matter accumulation of forage legumes. Disc harrowing before planting yielded superior results compared to the treatments without disc harrowing and disc harrowing after planting. At the end of the experimental period, the plots contained Arachis, Centrosema, Stylosanthes, and Pueraria. The dry matter accumulated by Centrosema corresponded to 73% of total dry matter yield of the plots. The participation of Arachis, Centrosema and Stylosanthes in final dry matter composition of the plots varied according to establishment method. The advantages of the use of species mixtures rather than monocultures in the understory of forest plantations were discussed.

Evolution in the profile of thyroid cancer cases treated in an oncology reference service: what changed in the last 20 years

Objective: To evaluate the characteristics of thyroid carcinoma cases treated at a reference hospital for cancer between 2008 and 2010.Methods: we studied 807 cases and analyzed the following clinicopathologic variables: symptoms, risk factors, diagnostic tests, staging, histological type, treatment performed and complications.Results: Females were more affected, with 660 cases (82%). The average age at diagnosis was 44.5 years. Prior exposure to ionizing radiation was reported by 22 (3%) patients, a family history of thyroid cancer by 89 (11%), and 289 (36%) individuals reported other types of cancer in the family. The fine needle aspiration biopsy was the main parameter for surgical indication and was suggestive of carcinoma in 463 patients (57%). Papillary carcinoma was the most common histological type, with 780 cases (96.6%). There were 728 (90%) total thyroidectomies, 43 (5.3%) reoperations or partial thyroidectomies followed by totalization, 23 (2.8%) extended thyroidectomies and only 13 (1.6%) partial thyroidectomies (lobectomy with isthmectomy). Neck dissection associated with thyroidectomy was done in 158 patients (19.5%). We observed a predominance of tumors classified as T1 in 602 (74.6%) patients. Transient hypocalcemia was the most frequent complication.Conclusion: The results show that the worldwide increase in the incidence of thyroid cancer has changed the profile of patients seen at a referral service. In addition, there were changes in the type of surgical treatment used, with increased use of total thyroidectomy in relation to partial and subtotal ones, and decreased use of elective neck dissections.

Hydrological processes obtained on the plot scale under four simulated rainfall tests during the cycle of different crop systems

The cropping system influences the interception of water by plants, water storage in depressions on the soil surface, water infiltration into the soil and runoff. The aim of this study was to quantify some hydrological processes under no tillage cropping systems at the edge of a slope, in 2009 and 2010, in a Humic Dystrudept soil, with the following treatments: corn, soybeans, and common beans alone; and intercropped corn and common bean. Treatments consisted of four simulated rainfall tests at different times, with a planned intensity of 64 mm h-1 and 90 min duration. The first test was applied 18 days after sowing, and the others at 39, 75 and 120 days after the first test. Different times of the simulated rainfall and stages of the crop cycle affected soil water content prior to the rain, and the time runoff began and its peak flow and, thus, the surface hydrological processes. The depth of the runoff and the depth of the water intercepted by the crop + soil infiltration + soil surface storage were affected by the crop systems and the rainfall applied at different times. The corn crop was the most effective treatment for controlling runoff, with a water loss ratio of 0.38, equivalent to 75 % of the water loss ratio exhibited by common bean (0.51), the least effective treatment in relation to the others. Total water loss by runoff decreased linearly with an increase in the time that runoff began, regardless of the treatment; however, soil water content on the gravimetric basis increased linearly from the beginning to the end of the rainfall.

Extended-spectrum β-lactamase-producing Salmonella enterica serovar Oranienburg (CTX-M-2 group) in a pediatric hospital in Tucumán, Argentina

INTRODUCTION: Salmonella sp infections have been reported over recent years in hospitals in Argentina and other countries due to multiresistant strains. The aim of this study was to characterize the extended-spectrum β-lactamases in third-generation cephalosporin-resistant strains of Salmonella enterica serovar Oranienburg. METHODS: We studied 60 strains isolated from children with gastroenteritis and/or extraintestinal complications. The antibiotic susceptibility patterns of the isolates were analyzed and the β-lactamases were characterized using phenotyping and genotyping methods. RESULTS: All the strains were resistant to ampicillin, cefotaxime, cefepime and aztreonam and partially susceptible to ceftazidime, thus corresponding well with the resistance phenotype conferred by CTX-M-type β-lactamases. An isoelectric point enzyme (pI = 7.9) was detected in all of the strains, and this was confirmed by PCR as a member of the CTX-M-2 group. CONCLUSIONS: This is the first report of Salmonella enterica serovar Oranienburg producing β-lactamases of the CTX-M-2 group in a pediatric hospital in Tucumán, Argentina.

blaCTX-M-2 and blaCTX-M-28 extended-spectrum β-lactamase genes and class 1 integrons in clinical isolates of Klebsiella pneumoniae from Brazil

Twenty-eight Klebsiella pneumoniae clinical isolates that exhibited an extended-spectrum cephalosporin-resistance profile from a city in the Northeast of Brazil were analysed by PCR and DNA sequencing in order to determine the occurrence of blaCTX-M genes and class 1 integrons. We determined the occurrence of the blaCTX-M-2 gene in six K. pneumoniae isolates and describe the first detection of the blaCTX-M-28 gene in South America. Seven isolates carried class 1 integrons. Partial sequencing analysis of the 5'-3'CS variable region in the class 1 integrons of three isolates revealed the presence of aadA1, blaOXA-2 and dfr22 gene cassettes.

Nitrate and ammonium in soil solution in tobacco management systems

Tobacco farmers of southern Brazil use high levels of fertilizers, without considering soil and environmental attributes, posing great risk to water resources degradation. The objective of this study was to monitor nitrate and ammonium concentrations in the soil solution of an Entisol in and below the root zone of tobacco under conventional tillage (CT), minimum tillage (MT) and no-tillage (NT). The study was conducted in the small-watershed Arroio Lino, in Agudo, State of Rio Grande do Sul, Brazil. A base fertilization of 850 kg ha-1 of 10-18-24 and topdressing of 400 kg ha-1 of 14-0-14 NPK fertilizer were applied. The soil solution was sampled during the crop cycle with a tension lysimeter equipped with a porous ceramic cup. Ammonium and nitrate concentrations were analyzed by the distillation and titration method. Nitrate concentrations, ranging from 8 to 226 mg L-1, were highest after initial fertilization and decreased during the crop cycle. The average nitrate (N-NO3-) concentration in the root zone was 75 in NT, 95 in MT, and 49 mg L-1 in CT. Below the root zone, the average nitrate concentration was 58 under NT, 108 under MT and 36 mg L-1 under CT. The nitrate and ammonium concentrations did not differ significantly in the management systems. However, the nitrate concentrations measured represent a contamination risk to groundwater of the watershed. The ammonium concentration (N-NH4+) decreased over time in all management systems, possibly as a result of the nitrification process and root uptake of part of the ammonium by the growing plants.

Nitrogen symbiotically fixed by cowpea and gliricidia in traditional and agroforestry systems under semiarid conditions

The objective of this work was to estimate the amounts of N fixed by cowpea in a traditional system and by cowpea and gliricidia in an agroforestry system in the Brazilian Northeast semiarid. The experiment was carried out in a randomized complete block design, in a split-plot arrangement, with four replicates, in the semiarid region of the state of Paraíba, Brazil. Plots consisted of agroforestry and traditional systems (no trees), and split-plots of the three crops planted between the tree rows in the agroforestry system. To estimate N fixation, plant samples were collected in the fourth growth cycle of the perennial species and in the fourth planting cycle of the annual species. In the agroforestry system with buffel grass and prickly-pear cactus, gliricidia plants symbiotically fix high proportions of N (>50%) and contribute with higher N amounts (40 kg ha-1 in leaves) than in the traditional system (11 kg ha-1 in grain and 18 kg ha-1 in straw). In the agroforestry system with maize and cowpea, gliricidia plants do not fix nitrogen, and N input is limited to the fixation by cowpea (2.7 kg ha-1), which is lower than in the traditional system due to its lower biomass production.

Effect of medroxy-progesterone acetate on follicular growth and endometrial cycloxygenase-2 (COX-2) expression during the bovine estrous cycle

The objective of this study was to evaluate the effect of medroxy-progesterone acetate (MAP) with or without estradiol benzoate (EB) on follicular growth during the estrous cycle in cattle. In the first experiment, Hereford cows were synchronized with a synthetic analogue of PGF2 alpha and were treated with two different doses of MAP (250 or 500 mg) with or without EB for 7 days starting on day 8 of the estrous cycle. Follicular growth was inhibited (P<0.05) in all cows except controls and those receiving 250mg MAP without EB. Seventy-five percent of the animals (15/20) showed estrus on days 21 and 22 of the cycle rather than at MAP withdrawal, demonstrating that these treatments did not induce estrus. To determine whether the EB treatment altered endometrial sensitivity to oxytocin and thus the luteolytic cascade, multiparous pre-synchronized cows received 5 mg of EB followed 6 hours later with 50 IU of oxytocin (OT; n=9). Eight hours after EB injection, endometrial fragments were collected from the cows on days 4, 13 and 17 of the estrous cycle and COX-2 gene expression was measured by PCR. EB increased COX-2 mRNA levels only on day 17 of the estrous cycle (P<0.05). In conclusion, MAP alone or associated with EB is able to suppress bovine follicular growth. However, EB in the presence of MAP is not efficient to induce luteolysis in cows when injected on day 8 of the estrous cycle.