Monitoring and molecular characterization of group Drotavirus in Brazilian poultry farms

Abstract: Rotaviruses are etiological agents of diarrhea both in humans and in several animal species. Data on avian Group D rotaviruses (RVD) are scarce, especially in Brazil. We detected RVD in 4 pools of intestinal contents of broilers, layer and broiler breeders out of a total of 111 pools from 8 Brazilian states, representing an occurrence of 3.6%, by a specific RVD RT-PCR targeting the VP6 gene. Phylogenetic tree confirmed that the Brazilian strains belong to group D and 3 of the sequences were identical in terms of amino acid whereas one showed 99.5% identity with the others. The sequences described in this study are similar to other sequences previously detected in Brazil, confirming the conserved nature of the VP6 protein.

Germination and morphological characterization of the fruits, seeds, and seedlings of Pilosocereus gounellei

The present work examined the germination and morphology of the fruits and seeds of the cactus Pilosocereus gounellei (xique-xique) and describeb the different stages of seedling growth. Germination tests examined the effects of combinations of two illumination regimes (a 12 hour photoperiod and total darkness) and three temperature regimes (25, 30, and 20-30 ºC) on the seeds of this species. Germination counts were made daily until the 16th day after sowing and the germination percentage and velocity index, and average germination time were evaluated. Descriptions of seed morphology considered both their external and internal aspects. The processes of germination and seedling growth were evaluated during 150 days and eight developmental stages were identified based on morphological changes. The fruits of the xique-xique cactus are 40.67 ± 4.40 mm long and 48.09 ± 3.23 mm in diameter, weigh 53.85 ± 10.03 g, and contain 3713 ± 689.50 seeds. Alternating temperatures of 20-30 ºC together with a photoperiod of 12 hours of light resulted in the highest seed germination rate. The seeds are positively photoblastic and germination is epigeal and fanerocotyledonary. Seedling growth is slow, but at 150 days after sowing the plants had developed epicotyls, large numbers of thorns, and the whole plant averaged 59.56 mm in height.

Proteolytic release and partial characterization of human sperm-surface glycopeptides

Sperm-surface glycopeptides were obtained from intact sperm membranes after proteolytic release by different enzymatic treatments such as autoproteolysis, trypsin, papain and pronase. Glycopeptides were isolated, their properties and composition were examined, and their monosaccharide and amino acid constituents were characterized. The monosaccharides identified were fucose, mannose, galactose, N-acetylglucosamine, and N-acetylgalactosamine, which form part of more than one type of oligosaccharide units. Autoproteolytic treatment mainly provided O-glycosidic type oligosaccharides, while a mixture of O- and N-glycosidic oligosaccharides was obtained in variable proportions when treated with trypsin, papain or pronase. The highest degree of peptide cleavage was obtained with pronase. Despite the higher yields reached with trypsin, these glycopeptides contain the lowest percentage of oligosaccharide chains. Proteolytic treatment provides a simple, rapid procedure for the isolation of glycopeptides from the sperm surface

Anticonvulsant and proconvulsant roles of nitric oxide in experimental epilepsy models

The effect of acute (120 mg/kg) and chronic (25 mg/kg, twice a day, for 4 days) intraperitonial injection of the nitric oxide (NO) synthase (NOS) inhibitor NG-nitro-L-arginine (L-NOARG) was evaluated on seizure induction by drugs such as pilocarpine and pentylenetetrazole (PTZ) and by sound stimulation of audiogenic seizure-resistant (R) and audiogenic seizure-susceptible (S) rats. Seizures were elicited by a subconvulsant dose of pilocarpine (100 mg/kg) only after NOS inhibition. NOS inhibition also simultaneously potentiated the severity of PTZ-induced limbic seizures (60 mg/kg) and protected against PTZ-induced tonic seizures (80 mg/kg). The audiogenic seizure susceptibility of S or R rats did not change after similar treatments. In conclusion, proconvulsant effects of NOS inhibition are suggested to occur in the pilocarpine model and in the limbic components of PTZ-induced seizures, while an anticonvulsant role is suggested for the tonic seizures induced by higher doses of PTZ, revealing inhibitor-specific interactions with convulsant dose and also confirming the hypothesis that the effects of NOS inhibitors vary with the model of seizure

Clinical and molecuar characterization of Brazilian patients with growth hormone gene deletions

Genomic DNA from 23 patients with isolated growth hormone (GH) deficiency (12 males and 11 females: heights -4.9 ± 1.4 SDS) was screened for GH gene deletions by restriction endonuclease analysis of polymerase chain reaction amplification products. Three unrelated patients had typical features of severe GH deficiency and deletions (6.7 kb in two and 7.6 kb in one) of the GH gene. The two patients with 6.7-kb deletions developed growth-attenuating anti-GH antibodies whereas the patient with the 7.6-kb deletion continued to grow with GH replacement therapy. Our finding that 3/23 (~13%) Brazilian subjects had GH gene deletions agrees with previous studies of severe isolated GH deficiency subjects in other populations. Two of three subjects (67%) with deletions developed blocking antibodies despite administration of exogenous GH at low doses. Interestingly, only 1/10 of cases with affected relatives or parental consanguinity had GH-1 gene deletions

Purification and partial characterization of Phaseolus vulgaris seed aminopeptidase

The aminopeptidase activity of Phaseolus vulgaris seeds was measured using L-Leu-p-nitroanilide and the L-aminoacyl-ß-naphthylamides of Leu, Ala, Arg and Met. A single peak of aminopeptidase activity on Leu-ß-naphthylamide was eluted at 750 µS after gradient elution chromatography on DEAE-cellulose of the supernatant of a crude seed extract. The effluent containing enzyme activity was applied to a Superdex 200 column and only one peak of aminopeptidase activity was obtained. SDS-polyacrylamide gel electrophoresis (10%) presented only one protein band with molecular mass of 31 kDa under reducing and nonreducing conditions. The aminopeptidase has an optimum pH of 7.0 for activity on all substrates tested and the highest Vmax/KM ratio for L-Leu-ß-naphthylamide. The enzyme activity was increased 40% by 0.15 M NaCl, inhibited 94% by 2.0 mM Zn2+, inhibited 91% by sodium p-hydroxymercuribenzoate and inhibited 45% by 0.7 mM o-phenanthroline and 30 µM EDTA. Mercaptoethanol (3.3 mM), dithioerythritol (1.7 mM), Ala, Arg, Leu and Met (70 µM), p-nitroaniline (0.25 mM) and ß-naphthylamine (0.53 mM) had no effect on enzyme activity when assayed with 0.56 mM of substrate. Bestatin (20 µM) inhibited 18% the enzyme activity. The aminopeptidase activity in the seeds decayed 50% after two months when stored at 4oC and room temperature. The enzyme is leucyl aminopeptidase metal- and thiol group-dependent.

Cloning, purification and comparative characterization of two digestive lysozymes from Musca domestica larvae

cDNA coding for two digestive lysozymes (MdL1 and MdL2) of the Musca domestica housefly was cloned and sequenced. MdL2 is a novel minor lysozyme, whereas MdL1 is the major lysozyme thus far purified from M. domestica midgut. MdL1 and MdL2 were expressed as recombinant proteins in Pichia pastoris, purified and characterized. The lytic activities of MdL1 and MdL2 upon Micrococcus lysodeikticus have an acidic pH optimum (4.8) at low ionic strength (μ = 0.02), which shifts towards an even more acidic value, pH 3.8, at a high ionic strength (μ = 0.2). However, the pH optimum of their activities upon 4-methylumbelliferyl N-acetylchitotrioside (4.9) is not affected by ionic strength. These results suggest that the acidic pH optimum is an intrinsic property of MdL1 and MdL2, whereas pH optimum shifts are an effect of the ionic strength on the negatively charged bacterial wall. MdL2 affinity for bacterial cell wall is lower than that of MdL1. Differences in isoelectric point (pI) indicate that MdL2 (pI = 6.7) is less positively charged than MdL1 (pI = 7.7) at their pH optima, which suggests that electrostatic interactions might be involved in substrate binding. In agreement with that finding, MdL1 and MdL2 affinities for bacterial cell wall decrease as ionic strength increases.

Production and partial characterization of lipase from Penicillium verrucosum obtained by submerged fermentation of conventional and industrial media

The growing interest in lipase production is related to the potential biotechnological applications that these enzymes present. Current studies on lipase production by submerged fermentation involve the use of agro-industrial residues aiming at increasing economic attractiveness. Based on these aspects, the objective of this work was to investigate lipase production by Penicillium verrucosum in submerged fermentation using a conventional medium based on peptone, yeast extract, NaCl and olive oil, and an industrial medium based on corn steep liquor, Prodex Lac (yeast hydrolysate), NaCl and olive oil, as well as to characterize the crude enzymatic extracts obtained. Kinetics of lipase production was evaluated and the highest enzymatic activities, of 3.15 and 2.22 U.mL-1, were observed when conventional and industrial media were used, respectively. The enzymatic extract showed optimal activity in the range from 30 to 40 °C and at pH 7.0. Although the industrial medium presents economical advantages over the conventional medium, the presence of agro-industrial residues rich in nitrogen and other important nutrients seemed to contribute to a reduction in lipase activity.

Chemical and biochemical characterization of soybean produced under drought stress

Brazil is the second soybean (Glycine max L. Merrill) producer and exporter in the world. In 2005, soybean cultivated in the southeastern region of the country suffered drought stress imposed by adverse high temperatures and low humidity during its reproductive stage. Little information is available regarding the effect of drought stress on the quality of grains. In this study chemical and biochemical characteristics of five soybean samples belonging to three different cultivars grown under drought stress were evaluated. The samples did not meet standards for marketing and contained high amounts of green seeds. Grains were analyzed for appearance, 100 seed weight, humidity, water activity, proteins, lipids, lipoxygenase 1 activity, peroxides, and pigment contents after harvest and after 20 months of storage at room temperature. Acidity was measured also after 30 months of storage. The values of water activity and humidity were 0.6-0.7 and 8.7-11.9%, respectively, and they did not change during storage time, but there was an increase in acidity, which alludes to lipase activity. The activity of lipoxygenase 1 was greatly affected. Immediately after harvest, the green pigments were represented mainly by pheophytin a, followed by pheophytin b, small quantities of chlorophyll b and chlorophyll a, and traces of other chlorophyll derivatives. After 20 months of storage almost all green pigments had disappeared. Drought stress probably enhanced membrane permeability, which led to a lower pH and promoted transformation of chlorophylls to pheophytins.

Development and chemical characterization of flour obtained from the external mesocarp of "pequizeiro" fruit

The objective of this work was to develop a recommendation for the chemical peeling of pequi fruit and characterize the flour obtained from the external mesocarp of "Pequizeiro", pequi tree (Caryocar brasiliense Camb.). The technology applied to obtain the external mesocarp pequi flour included the epicarp removal with NaOH solution. The Response Surface Method was used to optimize the chemical peeling process by applying the Central Composite Rotatable Design, with eleven trials including three replicates at the central point, varying the NaOH aqueous solution concentration and fruit immersion time. The mass loss was evaluated through the analysis of variance and using bi and three dimensional graphs. The chemical characteristics of the external mesocarp pequi flour evaluated were: moisture content, ashes, proteins, lipids, total carbohydrates, dietary fiber, and some minerals. The best combination for an efficient removal of the fruit peel with the lowest mass loss was reached with 7.05 minutes of immersion in a 5.08 g.L-1 NaOH aqueous solution. This study indicated that the external mesocarp pequi flour is a food source rich in dietary fiber, carbohydrates, ashes, magnesium, calcium, manganese, and copper, but it is poor in lipids, zinc, and iron.

Macrocospic and physiochemical characterization of a sugarless and gluten-free cake enriched with fibers made from pumpkin seed (Cucurbita maxima, L.) flour and cornstarch

The Consumers' interest for products with caloric reduction has increased, and their development is a technological challenge. The consumption of cakes has grown in importance and the demand for dietary products has stimulated the use of sweeteners and the optimization of bakery products. The consumption of fibers is related to chronic diseases prevention. Pumpkin seeds (maximum Cucurbita, L.), rich in fibers, can be used as a source of fiber in food products. A gluten-free diet is not easy to follow since gluten free products are not always available. The objective of this work was to perform a physicochemical characterization of cakes prepared with flours blends (FB) based on Pumpkin Seed Flour (PSF). The cakes were elaborated with FB in the ratios of 30:70 (C30) and 40:60 (C40) of PSF and cornstarch (CS), respectively. The results showed gluten absence and near-neutral pH. The chemical analysis of C30 and B40 showed increase of ashes, lipids, proteins, and insoluble dietary fiber and a decrease in the content of carbohydrates and calories. The chemical composition of C40 presented the greatest content of lipids, proteins, and dietary fibers, the lowest content of calories, and the best physical parameters. Therefore, both products proved suitable for human consumption.

Production and partial characterization of alkaline polygalacturonase secreted by thermophilic Bacillus sp. SMIA-2 under submerged culture using pectin and corn steep liquor

Polygalacturonase production by the thermophilic Bacillus sp. SMIA-2 cultivated in liquid cultures containing 0.5% (w/v) apple pectin and supplemented with 0.3% (w/v) corn steep liquor, reached its maximum after 36 hours with levels of 39 U.mL-1. The increase in apple pectin and corn steep liquor concentrations in the medium from 0.5 and 0.3%, respectively, to 0.65%, markedly affected the production of polygalacturonase, whose activity increased four times, reaching a maximum of 150.3 U.mL-1. Studies on polygalacturonase characterization revealed that the optimum temperature of this enzyme was between 60-70 °C. Thermostability profile indicated that the enzyme retained about 82 and 63% of its activity at 60 and 70 °C, respectively, after 2 hours of incubation. The optimum pH of the enzyme was found to be 10.0. After incubation of crude enzyme solution at room temperature for 2 hours at pH 8.0, a decrease of about 29% on its original activity was observed. At pH 10.0, the decrease was 25%.

Physical-chemical, caloric and sensory characterization of light jambolan (Syzygium cumini Lamarck) jelly

In Brazil, several little economically explored fruits have good potential as raw material for the agro-industry. This study aimed to produce and determine the physical-chemical and sensory characteristics of light jambolan jelly. This fruit has intense purple color, which gave the jellies - both standard and light - a quite attractive visual aspect. The light jellies exhibited similar physical-chemical characteristics to the ones developed through the conventional method and; with the proportion of sweeteners used, the caloric values of the formulations were reduced to the range of 41 to 53%, attending the requirements of the Brazilian legislation for this type of product. The sensory profile obtained for the 4 light formulations developed, clearly showed the tasters' preference for the jelly elaborated with the association of cyclamate and saccharin. Thus, the results revealed good perspectives for the application of this fruit in the food industry.

Partial chemical and functional characterization of milk whey products obtained by different processes

Whey protein samples (S-1 to S-5) were tested in vivo and in vitro for nutritional properties and selected bioactivities. Weanling male Wistar rats fed modified AIN-93G (12 g protein.100 g-1) diets for 21 days were used the in vivo studies. The nutritional parameters did not differ among the protein diets tested. Erythrocyte glutathione content was considered high and was higher for S-3, but liver glutathione was the same for all dietary groups. For S-3, cytokine secretion (IL-10 and TNF-α) by human peripheral blood mononuclear cells (in RPMI-1640 medium) was higher in the absence of antigen than in the presence of BCG antigen. Interleukin-4 secretion was repressed in all treatments. The IC50, whey protein concentration required to inhibit 50% of the melanoma cell proliferation, was 2.68 mg.mL-1 of culture medium for the S-3 sample and 3.66 mg.mL-1 for the S-2 sample. Based on these results, it was concluded that S-3 (whey protein concentrate enriched with TGF-β and lactoferrin) produced better nutritional and immunological responses than the other products tested.

Physical and chemical characterization of the pulp of different varieties of avocado targeting oil extraction potential

The aim of this study was to evaluate the physicochemical properties of avocado pulp of four different varieties (Avocado, Guatemala, Dickinson, and Butter pear) and to identify which has the greatest potential for oil extraction. Fresh avocado pulp was characterized by moisture, protein, fat, ash, carbohydrates and energy contents were determined. The carotenoids and chlorophyll contents were determined by the organic solvent extraction method. The results showed significant differences in the composition of the fruit when varieties are compared. However, the striking feature in all varieties is high lipid content; Avocado and Dickinson are the most suitable varieties for oil extraction, taking into account moisture content and the levels of lipids in the pulp. Moreover, it could be said that the variety Dickinson is the most affected by the parameters evaluated in terms of overall quality. Chlorophyll and carotenoids, fat-soluble pigments, showed a negative correlation with respect to lipids since it could be related to its function in the fruit. The varieties Avocado and Dickinson are an alternative to oil extraction having great commercial potential to be exploited thus avoiding waste and increasing farmers’ income.