Crystallization and preliminary X-ray diffraction analysis of recombinant chlorocatechol 1,2-dioxygenase from Pseudomonas putida

Chlorocatechol 1,2-dioxygenase from the Gram-negative bacterium Pseudomonas putida (Pp 1,2-CCD) is considered to be an important biotechnological tool owing to its ability to process a broad spectrum of organic pollutants. In the current work, the crystallization, crystallographic characterization and phasing of the recombinant Pp 1,2-CCD enzyme are described. Reddish-brown crystals were obtained in the presence of polyethylene glycol and magnesium acetate by utilizing the vapour-diffusion technique in sitting drops. Crystal dehydration was the key step in obtaining data sets, which were collected on the D03B-MX2 beamline at the CNPEM/MCT - LNLS using a MAR CCD detector. Pp 1,2-CCD crystals belonged to space group P6(1)22 and the crystallographic structure of Pp 1,2-CCD has been solved by the MR-SAD technique using Fe atoms as scattering centres and the coordinates of 3-chlorocatechol 1,2-dioxygenase from Rhodococcus opacus (PDB entry type=""url"" id=""http://scripts.iucr.org/cgi-bin/explore.cgi?pdbid=2boy"">2boy) as the search model. The initial model, which contains three molecules in the asymmetric unit, has been refined to 3.4 A resolution.

Schistosoma mansoni Tegument Protein Sm29 Is Able to Induce a Th1-Type of Immune Response and Protection against Parasite Infection

Background: Schistosomiasis continues to be a significant public health problem. This disease affects 200 million people worldwide and almost 800 million people are at risk of acquiring the infection. Although vaccine development against this disease has experienced more failures than successes, encouraging results have recently been obtained using membrane-spanning protein antigens from the tegument of Schistosoma mansoni. Our group recently identified Sm29, another antigen that is present at the adult worm tegument surface. In this study, we investigated murine cellular immune responses to recombinant (r) Sm29 and tested this protein as a vaccine candidate. Methods and Findings: We first show that Sm29 is located on the surface of adult worms and lung-stage schistosomula through confocal microscopy. Next, immunization of mice with rSm29 engendered 51%, 60% and 50% reduction in adult worm burdens, in intestinal eggs and in liver granuloma counts, respectively (p<0.05). Protective immunity in mice was associated with high titers of specific anti-Sm29 IgG1 and IgG2a and elevated production of IFN-gamma, TNF-alpha and IL-12, a typical Th1 response. Gene expression analysis of worms recovered from rSm29 vaccinated mice relative to worms from control mice revealed a significant (q<0.01) down-regulation of 495 genes and up-regulation of only 22 genes. Among down-regulated genes, many of them encode surface antigens and proteins associated with immune signals, suggesting that under immune attack schistosomes reduce the expression of critical surface proteins. Conclusion: This study demonstrates that Sm29 surface protein is a new vaccine candidate against schistosomiasis and suggests that Sm29 vaccination associated with other protective critical surface antigens is the next logical strategy for improving protection.

Systematic review and cladistic analysis of the genus Eusarcus Perty 1833 (Arachnida, Opiliones, Gonyleptidae)

Eusarcus Perty 1833 is one of the oldest described genera of Pachylinae, comprising 36 species distributed from northeastern to southern Brazil (including the central west region), northeastern Argentina, eastern Paraguay and Uruguay. The genus is reviewed and a new classification is proposed based on a cladistic analysis. A cladistic analysis was performed with the 34 valid species of Eusarcus and 11 species belonging to certain Gonyleptidae subfamilies. The data matrix has 67 characters: 14 from dorsal scutum and pedipalp, 38 from male legs and 15 from male genitalia. Two equally parsimonious trees were found (L=319; C. I.=0.26, R. I.=0.61). Pygophalangodus gemignanii uruguayensis Ringuelet 1955a and Pygophalangodus gemignanii gemignanii Mello-Leitao 1931b are here elevated to the category of species, and the following new combinations are proposed: E. catharinensis (Mello-Leitao 1927); E. berlae (Mello-Leitao 1932); E. gemignanii (Mello-Leitao 1931b); E. signatus(Roewer 1949); E. sooretamae (Soares & Soares 1946a); E. uruguayensis (Ringuelet 1955a). The following generic synonymies are proposed: Eusarcus Perty 1833 (type species E. armatus Perty 1833) = Metagraphinotus Mello-Leitao 1927 (type species M. catharinensis Mello-Leitao 1927), Pareusarcus Roewer 1929 (type species P. corniculatus Roewer 1929), Pygophalangodus Mello-Leitao 1931b (type species P. gemignanii-gemignanii Mello-Leitao 1931b) and Antetriceras Roewer 1949 (type species A. signatus Roewer 1949). The following specific synonymies are proposed: Eusarcus hastatus Sorensen 1884 = Pucrolioides argentina Roewer 1913, E. guimaraensi H. Soares 1945, Jacarepaguana pectinifemur Piza 1943, Canestrinia canalsi Mello-Leitao 1931a, and E. maquinensis H. Soares 1966b; E. armatus Perty 1833 = E. curvispinosus Mello-Leitao 1923b, and Enantiocentron montis Mello-Leitao 1936; Eusarcus catharinensis (Mello-Leitao 1927) = E. antoninae Mello-Leitao 1936, E. perpusillus Mello-Leitao 1945, E. tripos Mello-Leitao 1940, and Metagraphinotus trochanterspinosus Soares & Soares 1947b; E. nigrimaculatus Mello-Leitao 1924 = Pareusarcus centromelos Mello-Leitao 1935a, E. furcatus Roewer 1929, Orguesia armata Roewer 1913, and Pareusarcus corniculatus Roewer 1929; E. oxyacanthus Kollar in Koch 1839a = Enantiocentron doriphorus Mello-Leitao 1932, and E. spinimanu Mello-Leitao 1932; E. pusillus Sorensen 1884 = E. vervloeti B. Soares 1944c; E. berlae Mello-Leitao 1932 = Metagraphinotus arlei Mello-Leitao 1935a. Metapucrolia armata (Sorensen 1895) is revalidated, transferred to Eusarcus and considered as a species inquirenda. A new name, Eusarcus metapucrolia is proposed for this species to avoid homonymy with the type species of Eusarcus, E. armatus Perty 1833. Eusarcus aberrans Mello-Leitao 1939a is considered as a species inquirenda. The male of E. teresincola Soares & Soares 1946a is described. Female of the following species are described: E. bifidus Roewer 1929; E. dubius B. Soares 1943b; E. insperatus B. Soares 1944a; E. schubarti Soares & Soares 1946a; E. sooretamae (Soares & Soares 1946a). The following new species are described from Brazil: E. acrophthalmus (type locality: Bahia, Ilheus, Parataquice); E. alpinus (Rio de Janeiro, Santa Maria Madalena, Parque Estadual do Desengano); E. caparaoensis (Minas Gerais, Alto Caparao, Parque Nacional do Caparao); E. cavernicola (Goias, Sao Domingos, Parque Estadual de Terra Ronca, Lapa da Angelica); E. didactylus (Rio de Janeiro, Teresopolis, Parque Nacional Serra dos Orgaos); E. garibaldiae (Santa Catarina, Itajai); E. geometricus (Rio de Janeiro, Teresopolis, Parque Nacional Serra dos Orgaos); E. manero (Rio de Janeiro, Marica, Itaipuacu); E. matogrossensis (Mato Grosso, Chapada dos Guimaraes); E. mirabilis (Minas Gerais, Marlieria, Parque Estadual Rio Doce); E. sergipanus (Sergipe, Itabaiana, Parque Nacional de Itabaiana) and E. tripectinatus (Minas Gerais, Rio Preto). The holotype of E. curvispinosus is proposed as the neotype of E. armatus Perty 1833, the type material of which has been lost. Lectotypes for the following species were designated: E. aduncus; E. hastatus; E. oxyacanthus.

New familial assignments for three species of Neotropical harvestmen based on cladistic analysis (Arachnida: Opiliones: Laniatores)

We herein propose transfer of the two monotypic genera Globibunus (type-species G. rubrofemoratus Roewer, 1912) and Rivetinus (type-species R. minutus Roewer, 1919) to the subfamily Zamorinae (Agoristenidae) and Ramonus (type-species R. conifrons Roewer, 1956), previous placed in Agoristenidae, to the Prostygninae (Cranaidae) based on several characteristics of male genitalia. These transfers are corroborated by a cladistic analysis, which also recovered the three currently recognized agoristenid subfamilies.

Analysis of pelvic floor musculature function during pregnancy and postpartum: a cohort study

Aim. To compare the measurements of women`s pelvic floor musculature strength (PFMS) during pregnancy and postpartum period. Background. Pregnancy and childbirth can have an influence on the muscles and pelvic floor and can cause morbidities of women`s genito-urinary tract. Design. A prospective cohort study. Methods. There were included 226 primigravidae women, attended by community health services in the city of Itapecerica da Serra, Sao Paulo, Brazil. The participants were followed in four stages: (1) within 12 weeks of pregnancy; (2) between 36-40 weeks of pregnancy; (3) within 48 hours after childbirth; (4) 42-60 days after childbirth. Data were collected from February 2007-August 2008. The pelvic floor musculature strength was evaluated by perineometry and digital vaginal palpation in stages 1, 2 and 4. The final sample included 110 women who completed all four stages of the study. Results. The pelvic floor musculature strength of the women did not change significantly during pregnancy or after delivery (anova: p = 0 center dot 78). In all three examined stages, a low-intensity pelvic floor musculature strength was prevalent (in mmHg: stage 1 = 15 center dot 9; stage 2 = 15 center dot 2, stage 4 = 14 center dot 7), with scores from 0-3 on the Oxford scale. The pelvic floor musculature strength did not differ in relation to maternal age, skin colour, conjugal status, dyspareunia, stool characteristics, type of delivery, or conditions of the perineum. An interaction between maternal nutritional state and newborn`s weight may affect the pelvic floor musculature strength (manova: p = 0 center dot 04). Conclusion. Pregnancy and childbirth did not reduce significantly pelvic floor musculature strength. The perineometry and digital vaginal palpation used to assess the pelvic floor musculature strength were well accepted by the women. Relevance to clinical practice. In clinical practice, digital vaginal palpation is effective for supporting the diagnosis of urinary, intestinal and sexual dysfunctions. Perineometry use is particularly important together with the performance of perineal exercises with biofeedback in the treatment these disorders.

Involvement of L-type calcium channel and SERCA2a in myocardial dysfunction induced by obesity

Obesity has been shown to impair myocardial performance. Nevertheless, the mechanisms underlying the participation of calcium (Ca(2+)) handling on cardiac dysfunction in obesity models remain unknown. L-type Ca(2+) channels and sarcoplasmic reticulum (SR) Ca(2+)-ATPase (SERCA2a), may contribute to the cardiac dysfunction induced by obesity. The purpose of this study was to investigate whether myocardial dysfunction in obese rats is related to decreased activity and/or expression of L-type Ca(2+) channels and SERCA2a. Male 30-day-old Wistar rats were fed standard (C) and alternately four palatable high-fat diets (Ob) for 15 weeks. Obesity was determined by adiposity index and comorbidities were evaluated. Myocardial function was evaluated in isolated left ventricle papillary muscles under basal conditions and after inotropic and lusitropic maneuvers. L-type Ca(2+) channels and SERCA2a activity were determined using specific blockers, while changes in the amount of channels were evaluated by Western blot analysis. Phospholamban (PLB) protein expression and the SERCA2a/PLB ratio were also determined. Compared with C rats, the Ob rats had increased body fat, adiposity index and several comorbidities. The Ob muscles developed similar baseline data, but myocardial responsiveness to post-rest contraction stimulus and increased extracellular Ca(2+) was compromised. The diltiazem promoted higher inhibition on developed tension in obese rats. In addition, there were no changes in the L-type Ca(2+) channel protein content and SERCA2a behavior (activity and expression). In conclusion, the myocardial dysfunction caused by obesity is related to L-type Ca(2+) channel activity impairment without significant changes in SERCA2a expression and function as well as L-type Ca(2+) protein levels. J. Cell. Physiol. 226: 2934-2942, 2011. (C) 2011 Wiley-Liss, Inc.

A review of time-motion analysis and combat development in mixed martial arts matches at regional level tournaments

Mixed martial arts (MMA) have become a fast-growing worldwide expansion of martial arts competition, requiring high level of skill, physical conditioning, and strategy, and involving a synthesis of combat while standing or on the ground. This study quantified the effort-pause ratio (EP), and classified effort segments of stand-up or groundwork development to identify the number of actions performed per round in MMA matches. 52 MMA athletes participated in the study (M age = 24 yr., SD = 5; average experience in MMA = 5 yr., SD = 3). A one-way analysis of variance with repeated measurements was conducted to compare the type of action across the rounds. A chi-squared test was applied across the percentages to compare proportions of different events. Only one significant difference (p < .05) was observed among rounds: time in groundwork of low intensity was longer in the second compared to the third round. When the interval between rounds was not considered, the EP ratio (between high-intensity effort to low-intensity effort plus pauses) WE S 1:2 to 1:4. This ratio is between ratios typical for judo, wrestling, karate, and taekwondo and reflects the combination of ground and standup techniques. Most of the matches ended in the third round, involving high-intensity actions, predominantly executed during groundwork combat.

Analysis of Bacteria, Parasites, and Heavy Metals in Lettuce (Lactuca sativa) and Rocket Salad (Eruca sativa L.) Irrigated with Treated Effluent from a Biological Wastewater Treatment Plant

This study aimed to evaluate the viability of using treated residuary water from the Biological Wastewater Treatment Plant of Ribeiro Preto to grow vegetables, through the characterization and quantification of parasites, coliforms, and heavy metals. Three equal cultivation areas were prepared. The first was irrigated with treated/chlorinated (0.2 mg L(-1)) wastewater, the second one with treated wastewater without chlorination, and the third site with potable water, which was the control group. The presence of Hymenolepis nana, Enterobius vermicularis, nematode larvae, and Entamoeba coli was verified in lettuce (Lactuca sativa) samples. Although nematode larvae were observed in rocket salad (Eruca sativa L.), no significant differences were found between the number of parasites and type of irrigation water used. No significant differences were found between the number of fecal coliforms in vegetables and the different types of irrigation. However, the vegetables irrigated with treated effluent without chlorination showed higher levels of fecal coliforms. The risk of pathogens is reduced with bleach addition to the treated effluent at 0.2 mg/L. Concentration of heavy metals in vegetables does not mean significant risks to human health, according with the parameters recommended by the World Health Organization.

Non-linear boundary element formulation applied to contact analysis using tangent operator

This work presents a non-linear boundary element formulation applied to analysis of contact problems. The boundary element method (BEM) is known as a robust and accurate numerical technique to handle this type of problem, because the contact among the solids occurs along their boundaries. The proposed non-linear formulation is based on the use of singular or hyper-singular integral equations by BEM, for multi-region contact. When the contact occurs between crack surfaces, the formulation adopted is the dual version of BEM, in which singular and hyper-singular integral equations are defined along the opposite sides of the contact boundaries. The structural non-linear behaviour on the contact is considered using Coulomb`s friction law. The non-linear formulation is based on the tangent operator in which one uses the derivate of the set of algebraic equations to construct the corrections for the non-linear process. This implicit formulation has shown accurate as the classical approach, however, it is faster to compute the solution. Examples of simple and multi-region contact problems are shown to illustrate the applicability of the proposed scheme. (C) 2011 Elsevier Ltd. All rights reserved.

Dual boundary element formulation applied to analysis of multi-fractured domains

This paper deals with analysis of multiple random crack propagation in two-dimensional domains using the boundary element method (BEM). BEM is known to be a robust and accurate numerical technique for analysing this type of problem. The formulation adopted in this work is based on the dual BEM, for which singular and hyper-singular integral equations are used. We propose an iterative scheme to predict the crack growth path and the crack length increment at each time step. The proposed scheme able us to simulate localisation and coalescence phenomena, which is the main contribution of this paper. Considering the fracture mechanics analysis, the displacement correlation technique is applied to evaluate the stress intensity factors. The propagation angle and the equivalent stress intensity factor are calculated using the theory of maximum circumferential stress. Examples of simple and multi-fractured domains, loaded up to the rupture, are considered to illustrate the applicability of the proposed scheme. (C) 2010 Elsevier Ltd. All rights reserved.

Analysis of a semi-rigid connection for precast concrete

This paper presents a study of a specific type of beam-to-column connection for precast concrete structures. Furthermore, an analytical model to determine the strength and the stiffness of the connection, based on test results of two prototypes, is proposed. To evaluate the influence of the strength and stiffness of the connection on the behaviour of the structure, the results of numerical simulations of a typical multi-storey building with semi-rigid connections are also presented and compared with the results using pinned and rigid connections. The main conclusions are: (a) the proposed design model can reasonably evaluate the studied connection strength; (b) the evaluation of strength is more accurate than that of stiffness; (c) for a typical structure, it is possible to increase the number of storeys of the structure from two to four with lower horizontal displacement at the top, and only a small increase of the column base bending moment by replacing the pinned connections with semi-rigid ones; and (d) although there is significant uncertainty in the connection stiffness, the results show that the displacements at the top of the structure, and the column base moments present low susceptibility deviations to this parameter.

Analysis of the behavior of transverse walls of socket base connections

This paper presents a theoretical and experimental analysis of socket base connections of precast concrete structures with regard to the behavior of transverse walls. The experimental program included seven specimens, for which the type of interface in contact with cast-in-place concrete, the load eccentricities and the embedded lengths were all varied, A design model was proposed to calculate the reinforcements of the transverse walls. Based on the obtained results, some conclusions can be drawn: (a) The top of the transverse wall on the compression side of the smooth connections and the top of the two transverse walls of the rough connections are submitted to a bending-tension and this tension prevails on the bending; (b) The design model proposed for the calculation of the reinforcement of the transverse wall on the compression side provides the best prediction of the experimental results for all specimens when compared to the current design models; (c) For rough interfaces, the top of the transverse wall on the tension side is more requested than the top of transverse wall on the compression side; (d) The results of the proposed design model for the reinforcement of the transverse wall on the tension side of rough connections were in close agreement with the experimental results. (C) 2008 Elsevier Ltd. All rights reserved.

Effect of impeller type and stirring frequency on the behavior of an AnSBBR in the treatment of low-strength wastewater

The influence of impeller type and stirring frequency on the performance of a mechanically stirred anaerobic sequencing batch reactor containing immobilized biomass on an inert support (AnSBBR - Anaerobic Sequencing Batch Biofilm Reactor) was evaluated. The biomass was immobilized on polyurethane foam cubes placed in a stainless-steel basket inside a glass cylinder. Each 8-h batch run consisted of three stages: feed (10 min), reaction (460 min) and discharge (10 min) at 30 degrees C. Experiments were performed with four impeller types, i.e., helical, flat-blade, inclined-blade and curved-blade turbines, at stirring frequencies ranging from 100 to 1100 rpm. Synthetic wastewater was used in all experiments with an organic-matter concentration of 530 +/- 37 mg/L measured as chemical oxygen demand (COD). The reactor achieved an organic-matter removal efficiency of around 87% under all investigated conditions. Analysis of the four impeller types and the investigated stirring frequencies showed that mass transfer in the liquid phase was affected not only by the applied stirring frequency but also by the agitation mode imposed by each impeller type. The best reactor performance at all stirring frequencies was obtained when agitation was provided by the flat-blade turbine impeller. (C) 2010 Elsevier Ltd. All rights reserved.

Effect of impeller type and mechanical agitation on the mass transfer and power consumption aspects of ASBR operation treating synthetic wastewater

The effect of flow type and rotor speed was investigated in a round-bottom reactor with 5 L useful volume containing 2.0 L of granular biomass. The reactor treated 2.0 L of synthetic wastewater with a concentration of 800 mgCOD/L in 8-h cycles at 30 degrees C. Five impellers, commonly used in biological processes, have been employed to this end, namely: a turbine and a paddle impeller with six-vertical-flat-blades, a turbine and a paddle impeller with six-45 degrees-inclined-flat-blades and a three-blade-helix impeller. Results showed that altering impeller type and rotor speed did not significantly affect system stability and performance. Average organic matter removal efficiency was about 84% for filtered samples, total volatile acids concentration was below 20 mgHAc/L and bicarbonate alkalinity a little less than 400 mgCaCO(3)/L for most of the investigated conditions. However, analysis of the first-order kinetic model constants showed that alteration in rotor speed resulted in an increase in the values of the kinetic constants (for instance, from 0.57 h(-1) at 50 rpm to 0.84 h(-1) at 75 rpm when the paddle impeller with six-45 degrees-inclined-flat-blades was used) and that axial flow in mechanically stirred reactors is preferable over radial-flow when the vertical-flat-blade impeller is compared to the inclined-flat-blade impeller (for instance at 75 rpm, from 0.52 h(-1) with the six-flat-blade-paddle impeller to 0.84 h(-1) with the six-45 degrees-inclined-flat-blade-paddle impeller), demonstrating that there is a rotor speed and an impeller type that maximize solid-liquid mass transfer in the reaction medium. Furthermore, power consumption studies in this reduced reactor volume showed that no high power transfer is required to improve mass transfer (less than 0.6 kW/10(3) m(3)). (C) 2008 Elsevier Ltd. All rights reserved.

Expert system for the assessment of power transformer insulation condition based on type-2 fuzzy logic systems

An efficient expert system for the power transformer condition assessment is presented in this paper. Through the application of Duval`s triangle and the method of the gas ratios a first assessment of the transformer condition is obtained in the form of a dissolved gas analysis (DGA) diagnosis according IEC 60599. As a second step, a knowledge mining procedure is performed, by conducting surveys whose results are fed into a first Type-2 Fuzzy Logic System (T2-FLS), in order to initially evaluate the condition of the equipment taking only the results of dissolved gas analysis into account. The output of this first T2-FLS is used as the input of a second T2-FLS, which additionally weighs up the condition of the paper-oil system. The output of this last T2-FLS is given in terms of words easily understandable by the maintenance personnel. The proposed assessing methodology has been validated for several cases of transformers in service. (C) 2010 Elsevier Ltd. All rights reserved.