Perimicrovillar membrane assembly: the fate of phospholipids synthesised by the midgut of Rhodnius prolixus

In this study, we describe the fate of fatty acids that are incorporated from the lumen by the posterior midgut epithelium of Rhodnius prolixus and the biosynthesis of lipids. We also demonstrate that neutral lipids (NL) are transferred to the haemolymphatic lipophorin (Lp) and that phospholipids remain in the tissue in which they are organised into perimicrovillar membranes (PMMs). 3H-palmitic acid added at the luminal side of isolated midguts of R. prolixus females was readily absorbed and was used to synthesise phospholipids (80%) and NL (20%). The highest incorporation of 3H-palmitic acid was on the first day after a blood meal. The amounts of diacylglycerol (DG) and triacylglycerol synthesised by the tissue decreased in the presence of Lp in the incubation medium. The metabolic fates of 3H-lipids synthesised by the posterior midgut were followed and it was observed that DG was the major lipid released to Lp particles. However, the majority of phospholipids were not transferred to Lp, but remained in the tissue. The phospholipids that were synthesised and accumulated in the posterior midgut were found to be associated with Rhodnius luminal contents as structural components of PMMs.

Examining ERBB2 as a candidate gene for susceptibility to leprosy (Hansen’s disease) in Brazil

Leprosy remains prevalent in Brazil. ErbB2 is a receptor for leprosy bacilli entering Schwann cells, which mediates Mycobacterium leprae-induced demyelination and the ERBB2 gene lies within a leprosy susceptibility locus on chromosome 17q11-q21. To determine whether polymorphisms at the ERBB2 locus contribute to this linkage peak, three haplotype tagging single nucleotide polymorphisms (tag-SNPs) (rs2517956, rs2952156, rs1058808) were genotyped in 72 families (208 cases; 372 individuals) from the state of Pará (PA). All three tag-SNPs were associated with leprosy per se [best SNP rs2517959 odds ratio (OR) = 2.22; 95% confidence interval (CI) 1.37-3.59; p = 0.001]. Lepromatous (LL) (OR = 3.25; 95% CI 1.37-7.70; p = 0.007) and tuberculoid (TT) (OR = 1.79; 95% CI 1.04-3.05; p = 0.034) leprosy both contributed to the association, which is consistent with the previous linkage to chromosome 17q11-q21 in the population from PA and supports the functional role of ErbB2 in disease pathogenesis. To attempt to replicate these findings, six SNPs (rs2517955, rs2517956, rs1810132, rs2952156, rs1801200, rs1058808) were genotyped in a population-based sample of 570 leprosy cases and 370 controls from the state of Rio Grande do Norte (RN) and the results were analysed using logistic regression analysis. However, none of the associations were replicated in the RN sample, whether analysed for leprosy per se, LL leprosy, TT leprosy, erythema nodosum leprosum or reversal reaction conditions. The role of polymorphisms at ERBB2 in controlling susceptibility to leprosy in Brazil therefore remains unclear.

Construction of three new Gateway® expression plasmids for Trypanosoma cruzi

We present here three expression plasmids for Trypanosoma cruzi adapted to the Gateway® recombination cloning system. Two of these plasmids were designed to express trypanosomal proteins fused to a double tag for tandem affinity purification (TAPtag). The TAPtag and Gateway® cassette were introduced into an episomal (pTEX) and an integrative (pTREX) plasmid. Both plasmids were assayed by introducing green fluorescent protein (GFP) by recombination and the integrity of the double-tagged protein was determined by western blotting and immunofluorescence microscopy. The third Gateway adapted vector assayed was the inducible pTcINDEX. When tested with GFP, pTcINDEX-GW showed a good response to tetracycline, being less leaky than its precursor (pTcINDEX).

DNA barcodes for soil animal taxonomy

The biodiversity of soil communities remains very poorly known and understood. Soil biological sciences are strongly affected by the taxonomic crisis, and most groups of animals in that biota suffer from a strong taxonomic impediment. The objective of this work was to investigate how DNA barcoding - a novel method using a microgenomic tag for species identification and discrimination - permits better evaluation of the taxonomy of soil biota. A total of 1,152 barcode sequences were analyzed for two major groups of animals, collembolans and earthworms, which presented broad taxonomic and geographic sampling. Besides strongly reflecting the taxonomic impediment for both groups, with a large number of species-level divergent lineages remaining unnamed so far, the results also highlight a high level (15%) of cryptic diversity within known species of both earthworms and collembolans. These results are supportive of recent local studies using a similar approach. Within an impeded taxonomic system for soil animals, DNA-assisted identification tools can facilitate and improve biodiversity exploration and description. DNA-barcoding campaigns are rapidly developing in soil animals and the community of soil biologists is urged to embrace these methods.

Is tagging with visual implant elastomer a reliable technique for marking earthworms?

Visual implant elastomer (VIE) has recently been employed to investigate different aspects of earthworm ecology. However, a number of fundamental questions relating to the detection and positioning of the tag, its persistence and potential effects on earthworms remain unknown. Seven earthworm species belonging to three ecological groupings, with different pigmentation and burrowing behaviour, were tagged using different coloured VIE. External inspection after two days, one week and 1, 10 and 27 months were followed by preservation, dissection and internal inspection. Tags could be seen in living specimens to 27 months, and dissection revealed that in most cases they were lodged in the coelomic cavity, held in place by septa. However, over longer time periods (more than two years), the chlorogogenous tissue tended to bind to the tags and made external observation increasingly difficult. Migration of the VIE material towards the posterior of the earthworm and potential loss of the tag were only observed on rare occasions, and a recovery rate in excess of 98% was recorded. By introducing a reasonable amount of VIE into segments, just after the clitellum, this technique can become a valuable tool in earthworm ecology and life history studies, particularly in short-medium term laboratory and field experiments.

Dual-purpose wheat grain and animal production under different grazing periods

The objective of this work was to evaluate the influence of different grazing periods on beef animal production and on wheat forage and grain yield. The experiment was carried out in Pato Branco, PR, Brazil. Six grazing periods were evaluated (0, 21, 42, 63, 84, and 105 days) on dual-purpose wheat cultivar BRS Tarumã. Purunã steers, with average live weight of 162 kg and ten months of age, were kept under continuous grazing using a variable stocking rate, in order to maintain the established sward height of 25 cm. Greater increases in total animal gain (TAG) occurred with longer grazing periods. However, there was little increase after 63 days (490 kg ha-1), and TAG decreased from 552 to 448 kg ha-1 between 84 and 105 days. Grain yield decreased from 2,830 to 610 kg ha-1 when the grazing period increased from 0 to 105 days, but there was little change after 63 days (750 kg ha-1). Cultivar BRS Tarumã shows excellent animal production potential, and the decision on how long wheat pastures should be grazed must be based on relative prices of grain and livestock.

Microsatellite markers in analysis of resistance to coffee leaf miner in Arabica coffee

The objective of this work was to analyze coffee (Coffea arabica) genotypes resistant to the coffee leaf miner (Leucoptera coffeella) using microsatellite markers. Sixty-six loci were evaluated, of which 63 were obtained from the Brazilian Coffee Expressed Sequence Tag (EST) database. These loci were amplified in bulks of individuals from F5 progenies of 'Siriema' (C. arabica x C. racemosa) resistant and susceptible to the insect, in eight samples of C. racemosa, and in a F6 population of 'Siriema' with 91 individuals segregating for resistance to the leaf miner. Polymorphisms were verified for two simple sequence repeat (SSR) loci in bulks of the susceptible progenies. The two polymorphic alleles were present in around 70% of the susceptible genotypes in F5 and in approximately 90% of the susceptible individuals in F6. However, the polymorphic EST-SSR markers among populations contrasting for resistance to leaf miner were not correlated to the evaluated characteristics. SSR markers show inter- and intraspecific polymorphism in C. arabica and C. racemosa.

Lipídios estruturados obtidos a partir da mistura de gordura de frango, sua estearina e triacilgliceróis de cadeia média: I- Composição em ácidos graxos e em triacilgliceróis

The purpose of this study is to analyze the interactions that occur in binary and ternary fat blends between medium and long chain triacylglycerols and their structured lipids obtained by chemical interesterification through the analysis of their physico-chemical properties. The synthesized structured triacylglycerols presented from 14.8 to 58.4% medium chain fatty acids, from 15.7 to 37.2% saturated fatty acids, from 19.2 to 47.5% monounsaturated fatty acids, and from 6.7 to 15.2% essential fatty acids. Chemical interesterification modified the behavior of binary and ternary mixtures and new types of triacylglycerol groups were formed.

Interesterificação química de óleo de soja e óleo de soja totalmente hidrogenado: influência do tempo de reação

Chemical interesterification is an important alternative to produce zero trans fats. In practice, however, excessive reaction times are used to ensure complete randomization. This work evaluated the influence of the reaction time on the interesterification of soybean oil/fully hydrogenated soybean oil blend, carried out in the following conditions: 100 ºC, 500 rpm stirring speed, 0.4% (w/w) sodium methoxide catalyst. The triacylglycerol composition, solid fat content and melting point analysis showed that the reaction was very fast, reaching the equilibrium within 5 min. This result suggests the interesterification can be performed in substantially lower times, with reduction in process costs.

Produção de monoacilgliceróis e diacilgliceróis via glicerólise enzimática e destilação molecular

Monoacilglycerides and diacilglycerides are produced through lipase-catalyzed glycerolysis of soybean oil using Candida antarctica B in a solvent-free system. The reaction was carried out at a glycerol to triacylglycerol molar ratio of 8:1 with 2% of lipase. Acylglycerides, free fatty acids (FFA) and glycerol produced were separated employing the molecular distillation process. Starting from a product of enzymatic reaction 25.06% of triacylglycerols, 46.63% of diacylglycerides, 21.72% of monoacylglycerides, 5.38% of FFA and 1.21% of glycerol and after consecutively distillations, monoacylglycerides with 80% of purity was obtained and also oil with 54% of diacylglycerides to be used in human dietary.

A intrigante bioquímica da niacina: uma revisão crítica

Niacin (nicotinamide, nicotinic acid) interferes on homeostasis, DNA regulation, signaling and longevity. Nicotinic acid reduces synthesis of lipoproteins-apo-B and increases HDL. Its antilipemic action in liver produces: 1) inhibition of DGAT2, with decreased triacylglycerol synthesis, 2) downregulation of the b-chain of adenosine triphosphate synthase, leading to reduced HDL-apo-A-I catabolism. Nicotinic acid could increase redox potential in vascular endothelium. HM74A receptor activation in macrophages would be responsible for the release of prostaglandins, causing flushing in epidermis. HM74A agonists could assist in identifying antilipemic agents. Extended release niacin in combination with statin appears to protect cardiovascular system of patients with low HDL.

Caracterização composicional e transesterificação de óleo de microalga: uma abordagem computacional

A procedure for compositional characterization of a microalgae oil is presented and applied to investigate a microalgae based biodiesel production process through process simulation. The methodology consists of: proposing a set of triacylglycerides (TAG) present in the oil; assuming an initial TAG composition and simulating the transesterification reaction (UNISIM Design, Honeywell) to obtain FAME characterization values (methyl ester composition); evaluating deviations of experimental from calculated values; minimizing the sum of squared deviations by a non-linear optimization algorithm, with TAG molar fractions as decision variables. Biodiesel from the characterized oil is compared to a rapeseed based biodiesel.

Expression of Grapevine leafroll-associated virus 3 coat protein gene in Escherichia coli and production of polyclonal antibodies

Grapevine leafroll-associated virus 3 (GLRaV-3), the main viral species of the grapevine leafroll complex, causes yield and quality reduction in grapes (Vitis spp.). The coat protein gene was RT-PCR-amplified from total RNA extracted from infected grapevine leaves and the amplified fragment was cloned and completely sequenced. The fragment was subsequently subcloned into the pRSET-C expression vector. The recombinant plasmid was used to transform Escherichia coli BL21:DE3 and express the capsid protein. The coat protein, fused to a 6 His-tag, was purified by affinity chromatography using an Ni-NTA resin. The identity of the purified protein was confirmed by SDS-PAGE and Western blot. The in vitro-expressed protein was quantified and used for rabbit immunizations. The antiserum was shown to be sensitive and specific for the detection of GLRaV-3 in grapevine extracts in Western blot and DAS-ELISA assays, with no unspecific or heterologous reactions against other non-serologically related viruses being observed.

Impact of Roux-en-Y gastric bypass on lipid and inflammatory profiles

Objective: To evaluate the behavior of acute phase proteins and lipid profile in patients undergoing Roux-en-Y gastric bypass. Methods : We conducted a prospective study, consisting of three moments: M1 - preoperative (24 hours before surgery); M2 - 30 days after surgery; and M3 - 180 days after surgery. We carried measured height and BMI, as well as determined the concentrations of acute phase proteins (C-reactive protein (CRP), albumin and Alpha-1-acid glycoprotein) and total cholesterol, LDL-c, HDL-c and triacylglycerol. Results : participants comprised 25 individuals, with a mean age of 39.28 ± 8.07, 72% female. At all times of the study there was statistically significant difference as for weight loss and BMI. We found a significant decrease in CRP concentrations between the moments M1 and M3 (p = 0.041) and between M2 and M3 (p = 0.018). There was decrease in Alpha-1-GA concentrations between M1 and M2 (p = 0.023) and between M1 and M3 (p = 0.028). The albumin values increased, but did not differ between times. Total cholesterol and triacylglycerol decreased significantly ay all times. LDL-c concentrations decreased and differed between M1 and M2 (p = 0.001) and between M1 and M3 (p = 0.001). HDL-c values increased, however only differing between M1 and M2 (p = 0.050). Conclusion : Roux-en-Y gastric bypass promoted a decrease in plasma concentrations of CRP and Alpha-1-acid glycoprotein, improving lipid and inflammatory profiles.

Prokaryotic expression of a truncated form of bovine herpesvirus 1 glycoprotein E (gE) and its use in an ELISA for gE antibodies

This article describes the expression of a truncated form of bovine herpesvirus 1 (BoHV-1) glycoprotein E (gE) for use as immunodiagnostic reagent. A 651 nucleotide fragment corresponding to the amino-terminal third (217 amino acids) of BoHV-1 gE - that shares a high identity with the homologous BoHV-5 counterpart - was cloned as a 6×His-tag fusion protein in an Escherichia coli expression vector. A soluble protein of approximately 25 kDa purified from lysates of transformed E. coli was recognized in Western blot (WB) by anti-6xHis-tag and anti-BoHV-1 gE monoclonal antibodies. In addition, the recombinant protein was specifically recognized in WB by antibodies present in the sera of cattle seropositive to BoHV-1 and BoHV-5. An indirect ELISA using the expressed protein as coating antigen performed comparably to a commercial anti-gE ELISA and was able to differentiate serologically calves vaccinated with a gE-deleted BoHV-5 strain from calves infected with BoHV-1. Thus, the truncated gE may be useful for serological tests designed to differentiate BoHV-1/BoHV-5 infected animals from those vaccinated with gE-negative marker vaccines.