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International trade fairs are an important marketing tool for the expanding organic market. Although much output is marketed locally and there is a growing demand for the so-called zero miles products, Italian organic raw materials and processed foods are largely exported. For the first time, a direct survey has been conducted on 100 Italian firms attending BioFach 2000, to analyze aspects like activities performed, goals and expectations, forms of private – public partnerships. By using seven relevant variables, the firms are grouped into five categories. The same firms have been contacted after one month, to assess results, level of satisfaction and willingness to participate in a next edition.

The nesting whitefly Paraleyrodes minei Iaccarino 1990 (Homoptera: Aleyrodidae) was found on leaves of sour orange tree Citrus aurantium L., 1758, in the Gussone Park of the Faculty of Agriculture, of the U niversity of N aples “Federico II”, at Portici, Italy.

O presente trabalho foi desenvolvido na Estação de Produção e Uso de Biogás, da Universidade Federal de Lavras. O objetivo foi desenvolver e avaliar um sistema de alimentação de um motor estacionário com biogás. Foi utilizado um motor de 4 tempos com 4077 W de potência nominal à rotação de 4200 rpm acoplado a um gerador de 2400 W com rotação operacional de 3600 rpm. O biogás utilizado foi produzido em um biodigestor modelo indiano a partir de esterco bovino, contendo, em média, 63 % de metano. Os experimentos foram realizados com o motor operando, inicialmente, a gasolina na sua versão original e, posteriormente, com o sistema de alimentação a biogás. Para os ensaios foram medidas as rotações e o consumo energético do motor, além da tensão e corrente gerada para alimentar três potências resistivas iguais a 1000, 1200 e 1500 W. Para o conjunto motogerador alimentado a gasolina na sua versão original, observou-se uma potência média gerada de 840,2 W com tensão de 109 volts e eficiência do conjunto de 12,14 %. Para o conjunto motogerador operando a biogás utilizando o sistema de alimentação desenvolvido, observou-se uma potência média de 934,59 W, tensão 100,90 volts e eficiência de 8,22 %. Foi possível operar estavelmente o conjunto motogerador com combustível alternativo, biogás, utilizando o sistema de alimentação desenvolvido, substituindo em 100% o uso de gasolina.The present work was conducted at the Unity of Production and Use of Biogas of Lavras Federal University. Its objective was to develop and to evaluate the biogas powering system in a stationary motor using biogas. A four-timing stationary motor with 4077 W nominal potency at a 4200 rpm rotation was used coupled to a generator of 2400 W with operational rotation of 3600 rpm. The biogas used in the experiment was produced in an Indian model biodigestor operated with bovine manure, containing an average of 63% methane. The experiments were accomplished with a gasoline operating motor on its original version adapted for biogas. For the experiment, the rotation and the energy consumption of the motor were measured, besides the tension and current generated to feed three resistance potencies of 1000, 1200, and, 1500 W. For the motogenerator group using the gasoline in its original version we observed an average potency of 840.2 W with an average tension of 109 volts and an average group efficiency of 12.14 %. For the motogerador group operating with biogas using the supply system developed here, we observed an average potency of 934.59 W, an average tension of 100.90 volts and an average efficiency of 8.22 %. It was possible to steadily operate the motogenerator with biogas as the alternative fuel, using the supply system developed substituting 100% of the gasoline used.

A delimitação adequada de bacias hidrográficas é fundamental por ser essa a unidade territorial para fins de planejamento e de gerenciamento dos recursos hídricos. Desse modo, este trabalho teve objetivo de analisar uma metodologia de delimitação automática de bacias hidrográficas. Foram utilizados, para tanto, dados do projeto Shuttle Radar Topographic Mission (SRTM) integrados e processados em Sistema de Informações Geográficas (SIG). Os resultados confirmam valores de área compatíveis aos obtidos com base em cartas topográficas. Assim, a automatização da delimitação de bacias hidrográficas, por meio de dados do SRTM e ambiente SIG, apresenta-se vantajosa em relação ao custo e benefício proporcionado, além de estabelecer a padronização do traçado e posterior minimização de conflitos quanto à fixação da unidade de gestão dos recursos hídricos.The adequate delimitation of watersheds is essential to be that the territorial unit for planning and management of water resources. Thus, this work aims to evaluate a methodology for automatic delimitation of drainage basins. The project Shuttle Radar Topographic Mission (SRTM) data were integrated and processed in the Geographic Information System (GIS). The results show values of area compatible to those obtained on the topographic data. So, the automation of delimitation of watersheds, through SRTM data and GIS environment, appears to be advantageous in relation to cost and benefits offered, in addition, permits the standardization of the sketch and minimization of conflicts during fixation of the elementary unit of water resources management.

In this paper an analysis of different indirect methods for the calculation of the boundary layer height (BLH) using sodar, ultrasonic anemometer and a prognostic model based on single point surface measurements is presented. In particular the automatic spectral routine developed for Remtech sodar is compared with the results obtained with the parameterization of the vertical velocity variance of a minisodar, with the calculation of a prognostic model, with a parameterization based on horizontal velocity spectra and with the BLH evaluated from the intensity of minisodar echoes in stable conditions. The data of a radiosonde system taken in a nearby site was also analysed to get an independent evaluation of BLH for comparison. There is a significant scatter in the data for both the evaluation through the variance of vertical wind speed and the spectral analysis of the horizontal wind velocity although created by different effects. In unstable conditions the different methods give a similar pattern even if the prognostic model in some days predicts a significantly higher BLH with respect to the other methods. In stable nocturnal conditions the performances of the Remtech routine are worse than those in unstable conditions with an evident overestimation of the BLH that it is likely related to the overestimation of vertical turbulence and to the use of multiple range gates in the algorithm. Taking as reference the evaluation of BLH of the sodar, the spectral method applied to ultrasonic anemometer data seems to be affected by the lowest biases and it is a possible candidate, for the development of automatic routines for operational evaluation of BLH possibly with a different parameterisation for stable and unstable cases.

We characterized sequences of a novel SSS139 RsaI satellite DNA family in Drosophila gouveai and Drosophila seriema, two members of the Drosophila buzzatii cluster (D. repleta group). The sequences were AT-rich (69%) with a monomer unit length of about 139 bp and contained two direct subrepeats of 14 bp and 16 bp, suggesting that it might have originated by the duplication of smaller sequences. Southern and dot-blot hybridization analyses also detected SSS139 in other Drosophila buzzatii cluster species (D. koepferae, D. antonietae, D. borborema and D. serido) but not in D. buzzatii. These results agree with the marginal phylogenetic position of D. buzzatii within the D. buzzatii cluster.

Antecedentes: aunque el ecocardiograma y el cateterismo cardiaco se utilizan como primera instancia en el diagnóstico de cardiopatías congénitas, la angiografía por tomografía computarizada es un examen mínimamente invasivo que, mediante imágenes en dos/tres dimensiones en tiempo real, favorece el adecuado abordaje diagnóstico de pacientes con este tipo de patologías, en quienes se requiere una evaluación rápida y precisa de su anatomía extracardiaca. Objetivo: describir la experiencia institucional (agosto 2005-2006) en el uso de la angiografía por tomografía computarizada como método diagnóstico complementario en la evaluación de los pacientes pediátricos con cardiopatías congénitas. Método: estudio descriptivo - serie de casos. Se evaluaron pacientes pediátricos con diagnóstico clínico y ecocardiográfico de cardiopatías congénitas que se estudiaron con el escanógrafo General Electric Multislice Lightspeed/16 cortes. Resultados: mediante angiografía por tomografía, se evaluaron 58 pacientes de 2,4 ± 4,03 años de edad, con historia de cardiopatías congénitas. El 33,8% tuvo diagnóstico de atresia pulmonar, el 6,7% de atresia tricúspide, el 13,5% de doble tracto de salida del ventrículo derecho, el 11, 8% de tetralogía de Fallot, el 11,8% de coartación aórtica, el 3,3% de arco aórtico interrumpido; el 10,3% de ductus arterioso persistente, el 6,7% de drenaje venoso anómalo y el 5,1% de transposición de grandes arterias. Se obtuvieron imágenes de alta calidad que permitieron evaluar de forma precisa la anatomía vascular. Conclusiones: en los pacientes descritos, la angiografía por tomografía resultó ser una herramienta útil en el abordaje diagnóstico ya que permitió reconstruir su anatomía de manera tridimensional. Se requieren nuevos estudios que permitan evaluar la sensibilidad, la especificidad y el nivel de concordancia de esta técnica frente a otros métodos diagnósticos invasivos disponibles para el diagnóstico de este tipo de enfermedades.Antecedents: although echocardiogram and cardiac catheterization are used as first option tools for congenital heart diseases diagnosis, computed tomography angiography is a minimally invasive exam that through two to three dimensional images in real time gives an adequate approach to patients having this type of pathologies that require a rapid and precise evaluation of its extracardiac anatomy. Objective: describe the institutional experience from August 2005 to August 2006 in the use of angiography by tomography as a complementary diagnostic method in the evaluation of pediatric patients with congenital heart diseases. Method: serial descriptive study. 58 pediatric patients with clinical and echocardiographic diagnosis of congenital heart diseases were evaluated through the General Electric Multislice Lightpeed/16 scanner. Results: 58 patients with history of congenital heart disease were evaluated through CT angiography. Mean age was 2.4 ± 4.03 years. Twenty (33.8%) had diagnosis of pulmonary atresia, four (6.7%) had tricuspid atresia, eight (13.5%) had double-outlet right ventricle, seven (11.8%) had tetralogy of Fallot, nine (15.2%) had alterations of the aortic arch, seven (11.8%) had coarctation of the aorta, two (3.3%) had interrupted aortic arch, six (10.3%) had persistent ductus arteriosus, four (6.7%) had anomalous venous drainage and three (5.1%) had transposition of the great arteries. High quality images that allowed assessing the precise vascular anatomy were obtained. Conclusions: computed tomography angiography turned out to be a useful tool in the diagnostic approach of congenital heart diseases, because it allowed a tridimensional anatomic reconstruction. New studies that may permit the assessment of sensitivity, specificity and concordance level of this technique with other invasive diagnostic methods available for the diagnosis of this type of diseases, are required.

The remains of Mesozoic carbonate platforms outcrop along the margins of the Adriatic Sea and the mountain chains surrounding them in Italy, Slovenia and Croatia. Despite the fact that the platforms were far from continents, abundant evidence of terrestrial vertebrates was recently found, mainly in the Norian, Lower Jurassic and Cretaceous, and is here reviewed. The presence of populations of large terrestrial vertebrates (mainly dinosaurs) on these carbonate platforms, and in particular the discovery of late Hauterivian-early Barremian sauropods and late Santonian hadrosaurids, has important paleoecological and paleogeographical implications. KEY WORDS. Peri-Adriatic carbonate platforms. Mesozoic Tethysian paleogeography. Mesozoic terrestrial vertebrates. Paleoichnology. Reptilia. Dinosauria. Theropoda. Sauropoda. Ornithopoda. Hadrosauridae.

Overexpression of efflux transporters, in human cells, is a mechanism of resistance to drug and also to chemotherapy. We found that multidrug resistance protein-4 (MRP4) overexpression has a role in reducing aspirin action in patients after bypass surgery and, very recently, we found that aspirin enhances platelet MRP4 levels through peroxisome proliferator activated receptor-α (PPARα). In the present paper, we verified whether exposure of human embryonic kidney-293 cells (Hek-293) to aspirin modifies MRP4 gene expression and its correlation with drug elimination and cell toxicity. We first investigated the effect of high-dose aspirin in Hek-293 and we showed that aspirin is able to increase cell toxicity dose-dependently. Furthermore, aspirin effects, induced at low dose, already enhance MRP4 gene expression. Based on these findings, we compared cell viability in Hek-293, after high-dose aspirin treatment, in MRP4 overexpressing cells, either after aspirin pretreatment or in MRP4 transfected cells; in both cases, a decrease of selective aspirin cell growth inhibition was observed, in comparison with the control cultures. Altogether, these data suggest that exposing cells to low nontoxic aspirin dosages can induce gene expression alterations that may lead to the efflux transporter protein overexpression, thus increasing cellular detoxification of aspirin.

Endo-β-1, 4-mannanase from Thermotoga petrophila (TpMan) is a modular hyperthermostable enzyme involved in the degradation of mannan-containing polysaccharides. The degradation of these polysaccharides represents a key step for several industrial applications. Here, as part of a continuing investigation of TpMan, the region corresponding to the GH5 domain (TpManGH5) was characterized as a function of pH and temperature. The results indicated that the enzymatic activity of the TpManGH5 is pH-dependent, with its optimum activity occurring at pH 6. At pH 8, the studies demonstrated that TpManGH5 is a molecule with a nearly spherical tightly packed core displaying negligible flexibility in solution, and with size and shape very similar to crystal structure. However, TpManGH5 experiences an increase in radius of gyration in acidic conditions suggesting expansion of the molecule. Furthermore, at acidic pH values, TpManGH5 showed a less globular shape, probably due to a loop region slightly more expanded and flexible in solution (residues Y88 to A105). In addition, molecular dynamics simulations indicated that conformational changes caused by pH variation did not change the core of the TpManGH5, which means that only the above mentioned loop region presents high degree of fluctuations. The results also suggested that conformational changes of the loop region may facilitate polysaccharide and enzyme interaction. Finally, at pH 6 the results indicated that TpManGH5 is slightly more flexible at 65°C when compared to the same enzyme at 20°C. The biophysical characterization presented here is well correlated with the enzymatic activity and provide new insight into the structural basis for the temperature and pH-dependent activity of the TpManGH5. Also, the data suggest a loop region that provides a starting point for a rational design of biotechnological desired features.

En estos momentos es posible hacer recomendaciones sobre las indicaciones de la endarterectomía carotídea, basadas en los ensayos clínicos publicados y en la revisión de grandes series quirúrgicas. Las indicaciones para la cirugía dependerán del grado de estenosis y de la morfología de la placa, así como del estado clínico del paciente y de la morbi-mortalidad del equipo quirúrgico. Este al igual que otros estudios con grandes series de casos, evidencia que la endarterectomía carotídea y/o el manejo endovascular, realizado por equipos expertos, es un procedimiento seguro en pacientes con indicación quirúrgica.It is now possible to recommend indications for aortic endarterectomy, based on published clinical essays and revision of large surgical series. Surgery indications will depend on the stenosis degree and the plaque morphology, as well as on the patient's clinical state and the morbid-mortality in this surgical team. As other studies with large case series, evidences that carotid endarterectomy and/or endovascular management, when realized by expert teams, is a safe procedure in patients with surgical indication.

En comparación con el tratamiento convencional, la terapia endovascular en aneurisma de aorta torácica, presenta los mejores resultados, por lo que se convierte en el tratamiento de elección para la patología de aorta torácica descendente endovascular, por su baja morbimortalidad perioperatoria. El tratamiento quirúrgico por vía retroperitoneal y/o endovascular para aneurisma de aorta abdominal infrarrenal, resulta ser especialmente seguro en pacientes octogenarios o con alta morbilidad. Esta cohorte institucional presenta resultados perioperatorios y en el seguimiento, similares a los reportados en la literatura mundial.Compared with the conventional treatment, endovascular therapy in thoracic aortic aneurysm shows the best results, being the election treatment for the pathology of the descending thoracic aorta, due to its low peri-operative morbid-mortality. Surgical treatment by retro-peritoneal route and/or endovascular for infra-renal abdominal aortic aneurysm is especially safe in octogenarian patients or in those with a high mortality rate. This institutional cohort show peri-operative and follow-up results similar to those reported in the world literature.

The repleta group is one of the largest and most complex species groups in the genus Drosophila, representing an important biological model for the elaboration of evolutionary and biogeographical hypotheses on the American Continent. It is divided into six subgroups, of which the fasciola subgroup is the only one with most of its species found in the humid forests of Central and South America. With the aim of understanding the origin and evolution of the fasciola subgroup, and consequently adding information about the repleta group, we analyzed partial sequences of the mitochondrial cytochrome oxidase subunit I (COI) gene of nine Drosophila species (D. carolinae, D. coroica, D. ellisoni, D. fascioloides, D. ivai, D. moju, D. onca, D. rosinae and D. senei). The phylogenetic hypotheses obtained neither defined the relationships between the subgroups of the repleta group nor defined all the interspecific relationships in the fasciola subgroup. We found that the species D. carolinae, D. coroica, D. onca, D. rosinae and D. senei comprised a clade in which D. carolinae, D. onca and D. senei appeared together at a polytomy while D. fascioloides and D. ellisoni comprised another clade with a high bootstrap value.

Estudos foram desenvolvidos para dimensionar e adaptar o injetor do queimador principal de um aquecedor de água tipo acumulação de 75 L. O diâmetro do injetor foi redimensionado em função da pressão de serviço de 100 mm H2O e poder calorífico inferior do biogás de 21.600 kJ m-3 n, garantindo a manutenção da potência calorífica do equipamento de 20.900 kJ h-1. Os resultados demonstraram que o queimador adaptado operou com biogás adequadamente, com chama estável. A eficiência média do aquecedor foi de 68%, para ganho térmico de 36,7 ºC, correspondendo à temperatura final da água igual a 62,7 ºC, sendo consumido 0,796 m³n de biogás, aquecendo 75 L de água em 72 minutos.Studies had been developed to project and to adapt the injector of the main burner of water heater accumulated type of 75 L. The diameter of the injector was project in function of the pressure of service of 100 mm H2O and inferior calorific power of biogas of 21,600 kJ m-3 n, having guaranteed the maintenance of the calorific power of the equipment of 20,900 of kJ h-1 .The results had demonstrated that the adapted burner to operate with biogas operated adequately with a steady flame. The average efficiency of the heater was of 68%, for a thermal profit of 36.7 ºC, corresponding the final temperature of the water of 62.7 ºC being consumed 0.796 m³ n of biogas, heating 75 L of water in 72 minutes.

Experimental colitis induced by acetic acid has been used extensively as a model for intestinal inflammatory disease. Colonic tissue lesions of intestinal inflammatory disease patients seem to be related to the increased local production of pro-inflammatory cytokines (IL-1, IL-6, TNF-alpha, and IFN-gamma). PURPOSE: To assess the cytokine expression pattern identified through immunohistochemistry in colonic mucosa after experimental colitis induced by acetic acid and establish the relationship between this pattern and the presence of macroscopic lesions. MATERIALS AND METHODS: Adult male Wistar rats (n = 39) were divided at random into 4 groups: NC45 and NC24 (control without colitis; sacrificed at 45 minutes and 24 hours, respectively); and WC45 and WC24 (with experimental colitis induced by acetic acid; sacrificed at 45 minutes and 24 hours, respectively). Macroscopic and microscopic alterations in colonic tissue were evaluated, and cytokine expression was assessed through immunohistochemistry. RESULTS: After 24 hours, IL-1 expression was greater in the groups with colitis when compared to the groups without colitis. IL-4 expression was higher in the WC45 group. There was an increase in both INF-gamma and IL-6 related to the presence of necrosis of the colonic mucosa in the groups with colitis for both periods evaluated. CONCLUSION: The immunohistochemical technique was efficient for the analysis of various cytokine expressions in the colonic tissue. There was an increase in the IL-1 pro-inflammatory cytokines as well as in IL-6 and IFN-gamma associated with the presence of colonic necrosis. Experimental colitis induced by acetic acid is a useful model for the development of studies assessing the role of cytokines in the inflammation of mucosa as well as anti-cytokine therapies.O modelo de colite experimental induzida por ácido acético (CEAA) vem sendo extensamente utilizado em estudos sobre doenças inflamatórias intestinais (DII). Lesões no tecido colônico em portadores de DII parecem estar relacionados à produção local aumentada de citocinas pró-inflamatórias (IL-1, IL-6, TNF-alfa e IFN-gama). OBJETIVO: Avaliar o padrão de expressão de citocinas identificadas por imunohistoquímica em tecido colônico após CEAA e relacioná-lo à presença de lesões macroscópicas. MATERIAL E MÉTODOS: Ratos machos Wistar adultos (n=39) foram submetidos ou não à CEAA e sacrificados para retirada do tecido colônico em dois períodos distintos, perfazendo 4 grupos aleatórios: SC45 e SC24 (sem colite; sacrifício 45 minutos e 24 horas, respectivamente); CC45 e CC24 (com colite; sacrifício 45 minutos e 24 horas, respectivamente). Avaliaram-se alterações macro e microscópicas do cólon e sua expressão de citocinas foi avaliada por imunohistoquímica. RESULTADOS: Após 24 horas, a expressão de IL-1 foi maior no grupo com colite, em relação ao sem colite. IL-4 foi mais expressa no grupo CC45. Houve aumento de INF-gama e IL-6, relacionados à presença de necrose da mucosa colônica, nos grupos com colite, em ambos os períodos avaliados. CONCLUSÃO: A técnica de imunohistoquímica foi eficiente para a análise da expressão de citocinas na mucosa colônica. Houve aumento da expressão das citocinas pró-inflamatórias IL-1 e de IL-6 e IFN-gama associado à presença de necrose colônica. A CEAA é um bom modelo para o desenvolvimento de estudos destinados a avaliar o papel das citocinas na inflamação da mucosa e terapias anti-citocinas.

ImportanceGlioblastoma is the most lethal primary brain cancer. Clinical outcomes for glioblastoma remain poor, and new treatments are needed.ObjectiveTo investigate whether adding autologous tumor lysate-loaded dendritic cell vaccine (DCVax-L) to standard of care (SOC) extends survival among patients with glioblastoma.Design, setting, and participantsThis phase 3, prospective, externally controlled nonrandomized trial compared overall survival (OS) in patients with newly diagnosed glioblastoma (nGBM) and recurrent glioblastoma (rGBM) treated with DCVax-L plus SOC vs contemporaneous matched external control patients treated with SOC. This international, multicenter trial was conducted at 94 sites in 4 countries from August 2007 to November 2015. Data analysis was conducted from October 2020 to September 2021.InterventionsThe active treatment was DCVax-L plus SOC temozolomide. The nGBM external control patients received SOC temozolomide and placebo; the rGBM external controls received approved rGBM therapies.Main outcomes and measuresThe primary and secondary end points compared overall survival (OS) in nGBM and rGBM, respectively, with contemporaneous matched external control populations from the control groups of other formal randomized clinical trials.ResultsA total of 331 patients were enrolled in the trial, with 232 randomized to the DCVax-L group and 99 to the placebo group. Median OS (mOS) for the 232 patients with nGBM receiving DCVax-L was 19.3 (95% CI, 17.5-21.3) months from randomization (22.4 months from surgery) vs 16.5 (95% CI, 16.0-17.5) months from randomization in control patients (HR = 0.80; 98% CI, 0.00-0.94; P = .002). Survival at 48 months from randomization was 15.7% vs 9.9%, and at 60 months, it was 13.0% vs 5.7%. For 64 patients with rGBM receiving DCVax-L, mOS was 13.2 (95% CI, 9.7-16.8) months from relapse vs 7.8 (95% CI, 7.2-8.2) months among control patients (HR, 0.58; 98% CI, 0.00-0.76; P

The Argentinean Chaco region has been considered the center of origin of Drosophila buzzatii in South America because it contains most of the chromosomal polymorphism detected in natural populations. Two hypotheses have been put forward to explain the distribution of D. buzzatii in Brazil, one proposing that it has only recently passively colonized Brazil via human dispersal and the other suggesting that D. buzzatii has actively migrated to Brazil some time ago. Data from chromosomal inversions support recent colonization, whereas data from allozymes and mtDNA variation indicate that D. buzzatii has been in Brazil longer, favoring an active dispersal hypothesis. In our present work we analyzed data on 56 South American flies, mostly from Brazil, sequenced for the 5’ end of the mtDNA COI gene. The combined use of many neutrality tests and phylogeographic methods (e.g. nested clade analysis) indicated high gene flow throughout most of the range of D. buzzatii, although significant population structure was still detected. The high nucleotide diversity in the Northeast region of Brazil and the results from the nested clade analysis suggest that D. buzzatii has been in Brazil longer than proposed by the passive dispersal hypothesis. Our data indicate that D. buzzatii has been distributed throughout Brazil and Argentina since the Quaternary, though more data from different localities and markers need to be gathered to determine how the occupation of South America by D. buzzatii has occurred.

Endo-β-1,4-mannanase from Thermotoga petrophila (TpMan) is a hyperthermostable enzyme that catalyzes the hydrolysis of β-1,4-mannoside linkages in various mannan-containing polysaccharides. A recent study reported that TpMan is composed of a GH5 catalytic domain joined by a linker to a carbohydrate-binding domain. However, at this moment, there is no three-dimensional structure determined for TpMan. Little is known about the conformation of the TpMan as well as the role of the length and flexibility of the linker on the spatial arrangement of the constitutive domains. In this study, we report the first structural characterization of the entire TpMan by small-angle X-ray scattering combined with the three-dimensional structures of the individual domains in order to shed light on the low-resolution model, overall dimensions, and flexibility of this modular enzyme at different temperatures. The results are consistent with a linker with a compact structure and that occupies a small volume with respect to its large number of amino acids. Furthermore, at 20°C the results are consistent with a model where TpMan is a molecule composed of three distinct domains and that presents some level of molecular flexibility in solution. Even though the full enzyme has some degree of molecular flexibility, there might be a preferable conformation, which could be described by the rigid-body modeling procedure. Finally, the results indicate that TpMan undergoes a temperature-driven transition between conformational states without a significant disruption of its secondary structure. Our results suggest that the linker can optimize the geometry between the other two domains with respect to the substrate at high temperatures. These studies should provide a useful basis for future biophysical studies of entire TpMan.

We provide a thorough re-evaluation of the taxonomic diversity, phylogenetic relationships, and historical biogeography of the lambeosaurine hadrosaurids from the European Archipelago. Previously published occurrences of European Lambeosaurinae are reviewed and new specimens collected from upper Maastrichtian strata of the south-central Pyrenees are described. No support is found for the recognition of European saurolophines in the available hadrosaurid materials recovered so far from this area. A new genus and species of basal lambeosaurine, Canardia garonnensis, is described on the basis of cranial and appendicular elements collected from upper Maastrichtian strata of southern France. C. garonnensis differs from all other hadrosaurids, except Aralosaurus tuberiferus, in having maxilla with prominent subrectangular rostrodorsal flange; it differs from A. tuberiferus in a few maxillary and prefrontal characters. Together with A. tuberiferus, C. garonnensis integrates the newly recognized tribe Aralosaurini. Inference of lambeosaurine interrelationships via maximum parsimony analysis indicates that the other three known European lambeosaurines are representatives of two additional subclades (tribes) of these hadrosaurids: Tsintaosaurini (Pararhabdodon isonensis) and Lambeosaurini (the Arenysaurus ardevoli-Blasisaurus canudoi clade). The tribes Aralosaurini, Tsintaosaurini, Lambeosaurini, and Parasaurolophini are formally defined and diagnosed for the first time. Three event-based quantitative methods of ancestral range reconstruction were implemented to infer the historical biogeography of European lambeosaurines: Dispersal-Vicariance Analysis, Bayesian Binary MCMC, and Dispersal-Extinction-Cladogenesis. The results of these analyses, coupled with the absence of pre-Maastrichtian lambeosaurines in the Mesozoic vertebrate fossil record of Europe, favor the hypothesis that aralosaurins and tsintaosaurins were Asian immigrants that reached the Ibero-Armorican island via dispersal events sometime during the Maastrichtian. Less conclusive is the biogeographical history of European lambeosaurins; several scenarios, occurring sometime during the Maastrichtian, are possible, from vicariance leading to the splitting of Asian or North American from European ranges to a dispersal event from North America to the European Archipelago.

Cellulases play a key role in enzymatic routes for degradation of plant cell-wall polysaccharides into simple and economically-relevant sugars. However, their low performance on complex substrates and reduced stability under industrial conditions remain the main obstacle for the large-scale production of cellulose-derived products and biofuels. Thus, in this study a novel cellulase with unusual catalytic properties from sugarcane soil metagenome (CelE1) was isolated and characterized. The polypeptide deduced from the celE1 gene encodes a unique glycoside hydrolase domain belonging to GH5 family. The recombinant enzyme was active on both carboxymethyl cellulose and β-glucan with an endo-acting mode according to capillary electrophoretic analysis of cleavage products. CelE1 showed optimum hydrolytic activity at pH 7.0 and 50 °C with remarkable activity at alkaline conditions that is attractive for industrial applications in which conventional acidic cellulases are not suitable. Moreover, its three-dimensional structure was determined at 1.8 Å resolution that allowed the identification of an insertion of eight residues in the β8-α8 loop of the catalytic domain of CelE1, which is not conserved in its psychrophilic orthologs. This 8-residue-long segment is a prominent and distinguishing feature of thermotolerant cellulases 5 suggesting that it might be involved with thermal stability. Based on its unconventional characteristics, CelE1 could be potentially employed in biotechnological processes that require thermotolerant and alkaline cellulases.

Metagenomics has been widely employed for discovery of new enzymes and pathways to conversion of lignocellulosic biomass to fuels and chemicals. In this context, the present study reports the isolation, recombinant expression, biochemical and structural characterization of a novel endoxylanase family GH10 (SCXyl) identified from sugarcane soil metagenome. The recombinant SCXyl was highly active against xylan from beechwood and showed optimal enzyme activity at pH 6,0 and 45°C. The crystal structure was solved at 2.75 Å resolution, revealing the classical (β/α)8-barrel fold with a conserved active-site pocket and an inherent flexibility of the Trp281-Arg291 loop that can adopt distinct conformational states depending on substrate binding. The capillary electrophoresis analysis of degradation products evidenced that the enzyme displays unusual capacity to degrade small xylooligosaccharides, such as xylotriose, which is consistent to the hydrophobic contacts at the +1 subsite and low-binding energies of subsites that are distant from the site of hydrolysis. The main reaction products from xylan polymers and phosphoric acid-pretreated sugarcane bagasse (PASB) were xylooligosaccharides, but, after a longer incubation time, xylobiose and xylose were also formed. Moreover, the use of SCXyl as pre-treatment step of PASB, prior to the addition of commercial cellulolytic cocktail, significantly enhanced the saccharification process. All these characteristics demonstrate the advantageous application of this enzyme in several biotechnological processes in food and feed industry and also in the enzymatic pretreatment of biomass for feedstock and ethanol production.

Age is a powerful predictor of survival in glioblastoma multiforme (GBM) yet the biological basis for the difference in clinical outcome is mostly unknown. Discovering genes and pathways that would explain age-specific survival difference could generate opportunities for novel therapeutics for GBM. Here we have integrated gene expression, exon expression, microRNA expression, copy number alteration, SNP, whole exome sequence, and DNA methylation data sets of a cohort of GBM patients in The Cancer Genome Atlas (TCGA) project to discover age-specific signatures at the transcriptional, genetic, and epigenetic levels and validated our findings on the REMBRANDT data set. We found major age-specific signatures at all levels including age-specific hypermethylation in polycomb group protein target genes and the upregulation of angiogenesis-related genes in older GBMs. These age-specific differences in GBM, which are independent of molecular subtypes, may in part explain the preferential effects of anti-angiogenic agents in older GBM and pave the way to a better understanding of the unique biology and clinical behavior of older versus younger GBMs.

Due to the request of numerous patients to improve the aspect of the perioral area in combination with other types of cosmetic and reconstructive surgery, we started to use autologous fillers. In fact, there are numerous potential fillers that can be utilized during various operations executed in many bodily areas, such as the breast, abdomen, and face. The muscular fascia as well as the dense connective tissue which the surgeon encounters in various bodily areas during some stages of the operation, in fact, can be removed and replaced both by themselves or superimposed in order to increase their thickness. The insertion of the grafts is carried out by using a needle, but other methods can also be used with the same success. The consistency of the area treated, after a few days of edema, is very similar to the host area, and the volume obtained remains uniform in time (our followup is after 24 months). The time utilized for the removal and the insertion in the chosen area was only a few minutes. The result was extremely satisfactory in all the 30 patients treated, and there was no complication or side effects.

Plant feedstocks are at the leading front of the biofuel industry based on the potential to promote economical, social and environmental development worldwide through sustainable scenarios related to energy production. Penicillium echinulatum is a promising strain for the bioethanol industry based on its capacity to produce large amounts of cellulases at low cost. The secretome profile of P. echinulatum after grown on integral sugarcane bagasse, microcrystalline cellulose and three types of pretreated sugarcane bagasse was evaluated using shotgun proteomics. The comprehensive chemical characterization of the biomass used as the source of fungal nutrition, as well as biochemical activity assays using a collection of natural polysaccharides, were also performed. Our study revealed that the enzymatic repertoire of P. echinulatum is geared mainly toward producing enzymes from the cellulose complex (endogluganases, cellobiohydrolases and β-glucosidases). Glycoside hydrolase (GH) family members, important to biomass-to-biofuels conversion strategies, were identified, including endoglucanases GH5, 7, 6, 12, 17 and 61, β-glycosidase GH3, xylanases GH10 and GH11, as well as debranching hemicellulases from GH43, GH62 and CE2 and pectinanes from GH28. Collectively, the approach conducted in this study gave new insights on the better comprehension of the composition and degradation capability of an industrial cellulolytic strain, from which a number of applied technologies, such as biofuel production, can be generated.

A serrated tooth from the Coniacian-Santonian (Upper Cretaceous) Polazzo fossil site (Karst, NE Italy) is the first record of a notosuchian crocodyliform from Italy. Although it shares synapomorphies with teeth referred to the European genus Doratodon and with the Gondwanan genus Araripesuchus, it is distinct in the unusual combination of features, suggesting the presence of a yet unreported notosuchian taxon in the Adriatic-Dinaric Carbonate Platform located in the Tethys between the Afroarabian continent and the North European landmass during Late Cretaceous times. Notosuchians were typically terrestrial crocodyliforms, supporting the presence of emergent areas on the carbonate platform.

BackgroundThe study of ancient skeletal pathologies can be adopted as a key tool in assessing and tracing several diseases from past to present times. Skeletal fluorosis, a chronic metabolic bone and joint disease causing excessive ossification and joint ankylosis, has been only rarely considered in differential diagnoses of palaeopathological lesions. Even today its early stages are misdiagnosed in endemic areas.Methodology/principal findingsEndemic fluorosis induced by high concentrations of fluoride in water and soils is a major health problem in several countries, particularly in volcanic areas. Here we describe for the first time the features of endemic fluorosis in the Herculaneum victims of the 79 AD eruption, resulting from long-term exposure to high levels of environmental fluoride which still occur today.Conclusions/significanceOur observations on morphological, radiological, histological and chemical skeletal and dental features of this ancient population now suggest that in this area fluorosis was already endemic in Roman times. This evidence merged with currently available epidemiologic data reveal for the Vesuvius area population a permanent fluoride health hazard, whose public health and socio-economic impact is currently underestimated. The present guidelines for fluoridated tap water might be reconsidered accordingly, particularly around Mt Vesuvius and in other fluoride hazard areas with high natural fluoride levels.

Induced cell fusion has enabled several important discoveries, including the phenomenon of nuclear reprogramming and may yet be applied as a novel therapy for degenerative diseases. However, existing fusogens lack the efficiency required to enable investigation of the epigenetic modifications underlying nuclear reprogramming and the specificity required for clinical application. Here we present a chimeric measles hemagglutinin, Hα7, which specifically and efficiently mediates the fusion of diverse cell types with skeletal muscle both in vitro and in vivo. When compared directly to polyethylene glycol, Hα7 consistently generated a substantial increase in heterokaryon yield and exhibited insignificant levels of toxicity. Moreover, this increased fusion efficiency enabled detection of chromatin modifications associated with nuclear reprogramming following Hα7-mediated fusion of human fibroblasts and mouse myotubes. Finally, Hα7 was also capable of increasing the contribution of transplanted fibroblasts to skeletal muscle repair in vivo, suggesting that this strategy could be used for therapeutic gene delivery.

BACKGROUND: Trypanosoma cruzi has a single flagellum attached to the cell body by a network of specialized cytoskeletal and membranous connections called the flagellum attachment zone. Previously, we isolated a DNA fragment (clone H49) which encodes tandemly arranged repeats of 68 amino acids associated with a high molecular weight cytoskeletal protein. In the current study, the genomic complexity of H49 and its relationships to the T. cruzi calpain-like cysteine peptidase family, comprising active calpains and calpain-like proteins, is addressed. Immunofluorescence analysis and biochemical fractionation were used to demonstrate the cellular location of H49 proteins. METHODS AND FINDINGS: All of H49 repeats are associated with calpain-like sequences. Sequence analysis demonstrated that this protein, now termed H49/calpain, consists of an amino-terminal catalytic cysteine protease domain II, followed by a large region of 68-amino acid repeats tandemly arranged and a carboxy-terminal segment carrying the protease domains II and III. The H49/calpains can be classified as calpain-like proteins as the cysteine protease catalytic triad has been partially conserved in these proteins. The H49/calpains repeats share less than 60% identity with other calpain-like proteins in Leishmania and T. brucei, and there is no immunological cross reaction among them. It is suggested that the expansion of H49/calpain repeats only occurred in T. cruzi after separation of a T. cruzi ancestor from other trypanosomatid lineages. Immunofluorescence and immunoblotting experiments demonstrated that H49/calpain is located along the flagellum attachment zone adjacent to the cell body. CONCLUSIONS: H49/calpain contains large central region composed of 68-amino acid repeats tandemly arranged. They can be classified as calpain-like proteins as the cysteine protease catalytic triad is partially conserved in these proteins. H49/calpains could have a structural role, namely that of ensuring that the cell body remains attached to the flagellum by connecting the subpellicular microtubule array to it.

We studied growth and longevity of Lacerta agilis from a sample (34 adults and 2 small-sized juveniles) of a population living at high altitude in north-western Italy using skeletochronological method. Snout vent length (SVL) mean of males did not significantly differ from that of females although the latter were in average bigger (SVL ± SD, males: 69.3 ± 7.1 mm, n = 11; females: 73.9 ± 9.7 mm, n = 22; Mann-Whitney U-test, U = 1.76, P = 0.077). Age ranged from 2 to 4 years (mean age ± SD = 2.3 ± 0.2) in males and from 2 to 3 years in females (mean age ± SD = 2.59 ± 0.5 years). Age mean did not significantly differ between the sexes (Mann-Whitney U-test, U = 1.35, P = 0.174). The two juveniles were 30 and 32 mm in SVL and both were 1-2 months old. In both sexes, a significant positive correlation between SVL and age was recorded although weakly significant for males (Spearman’s correlation coefficient, males: rs = 0.70, P = 0.05; females: rs = 0.75, P < 0.001). Von Bertalanffy growth curves well fitted to the relationships between age and SVL and showed a different profile between males (asymptotics size, SVLmax = 81.9 mm; growth coefficient, k = 0.63) and females (SVLmax = 100 mm; k = 0.40). Results indicate that individuals of L. agilis studied by us are short-living when compared with other populations of the same species.