Protection conferred by heterologous vaccination against tuberculosis is dependent on the ratio of CD4(+)/CD4(+) Foxp3(+) cells

CD4(+) Foxp3(+) regulatory T cells inhibit the production of interferon-?, which is the major mediator of protection against Mycobacterium tuberculosis infection. In this study, we evaluated whether the protection conferred by three different vaccines against tuberculosis was associated with the number of spleen and lung regulatory T cells. We observed that after homologous immunization with the 65 000 molecular weight heat-shock protein (hsp 65) DNA vaccine, there was a significantly higher number of spleen CD4(+) Foxp3(+) cells compared with non-immunized mice. Heterologous immunization using bacillus Calmette Guerin (BCG) to prime and DNA-hsp 65 to boost (BCG/DNA-hsp 65) or BCG to prime and culture filtrate proteins (CFP)-CpG to boost (BCG/CFP-CpG) induced a significantly higher ratio of spleen CD4(+)/CD4(+) Foxp3(+) cells compared with non-immunized mice. In addition, the protection conferred by either the BCG/DNA-hsp 65 or the BCG/CFP-CpG vaccines was significant compared with the DNA-hsp 65 vaccine. Despite the higher ratio of spleen CD4(+)/CD4(+) Foxp3(+) cells found in BCG/DNA-hsp 65-immunized or BCG/CFP-CpG-immunized mice, the lungs of both groups of mice were better preserved than those of DNA-hsp 65-immunized mice. These results confirm the protective efficacy of BCG/DNA-hsp 65 and BCG/CFP-CpG heterologous prime-boost vaccines and the DNA-hsp 65 homologous vaccine. Additionally, the prime-boost regimens assayed here represent a promising strategy for the development of new vaccines to protect against tuberculosis because they probably induce a proper ratio of CD4(+) and regulatory (CD4(+) Foxp3(+)) cells during the immunization regimen. In this study, this ratio was associated with a reduced number of regulatory cells and no injury to the lungs.

Taxonomy, Hemipenial Morphology, and Natural History of Two Poorly Known Species of Anadia (Gymnophthalmidae) from Northern South America

Anadia pariaensis Rivas, La Marca, and Oliveros, 1999, and Anadia steyeri Nieden, 1914, are two particularly rare and poorly known lizards described from single specimens. In the case of A. pariaensis, it remains known from the holotype, whereas A. steyeri is known from three additional specimens reported in the literature after the original description of the species. A single new specimen of A. pariaensis and five of A. steyeri, including the first adult males recorded for both species, make possible a more representative description of both species, including descriptions of the hemipenes. Despite both species presenting some similar morphological characteristics, the examination of the hemipenial morphology revealed very different organs. The hemipenis of A. steyeri presents some characteristics that resemble the organs of two species from the Santa Marta Mountain Range in the "bitaeniata-group" (Anadia pulchella and Anadia altaserrania). On the other hand, the hemipenes of A. pariaensis are unique morphologically and cannot be associated with the hemipenes known from other species in the genus. We describe variation within both species, and we comment on possible sexual dimorphism (number and arrangement of the femoral pores), natural history, and the known geographic distribution of the species. We also comment on Anadia bumanguesa Rueda-Almonacid and Caicedo 2004 based on a new specimen, the second known. This species may be a synonym of A. steyeri.

Joint NOD2/RIPK2 Signaling Regulates IL-17 Axis and Contributes to the Development of Experimental Arthritis

Intracellular pattern recognition receptors such as the nucleotide-binding oligomerization domain (NOD)-like receptors family members are key for innate immune recognition of microbial infection and may play important roles in the development of inflammatory diseases, including rheumatic diseases. In this study, we evaluated the role of NOD1 and NOD2 on development of experimental arthritis. Ag-induced arthritis was generated in wild-type, NOD1(-/-)!, NOD2(-/-), or receptor-interacting serine-threonine kinase 2(-/-) (RIPK2(-/-)) immunized mice challenged intra-articularly with methylated BSA. Nociception was determined by electronic Von Frey test. Neutrophil recruitment and histopathological analysis of proteoglycan lost was evaluated in inflamed joints. Joint levels of inflammatory cytokine/chemokine were measured by ELISA. Cytokine (IL-6 and IL-23) and NOD2 expressions were determined in mice synovial tissue by RT-PCR. The NOD2(-/-) and RIPK2(-/-), but not NOD1(-/-), mice are protected from Ag-induced arthritis, which was characterized by a reduction in neutrophil recruitment, nociception, and cartilage degradation. NOD2/RIPK2 signaling impairment was associated with a reduction in proinflammatory cytokines and chemokines (TNF, IL-1 beta, and CXCL1/KC). IL-17 and IL-17 triggering cytokines (IL-6 and IL-23) were also reduced in the joint, but there is no difference in the percentage of CD4(+) IL-17(+) cells in the lymph node between arthritic wild-type and NOD2(-/-) mice. Altogether, these findings point to a pivotal role of the NOD2/RIPK2 signaling in the onset of experimental arthritis by triggering an IL-17-dependent joint immune response. Therefore, we could propose that NOD2 signaling is a target for the development of new therapies for the control of rheumatoid arthritis. The Journal of Immunology, 2012, 188: 5116-5122.

Ectopic expression of a fruit phytoene synthase from Citrus paradisi Macf. promotes abiotic stress tolerance in transgenic tobacco

Abscisic acid (ABA) is an important regulator of plant responses to environmental stresses and an absolute requirement for stress tolerance. Recently, a third phytoene synthase (PSY3) gene paralog was identified in monocots and demonstrated to play a specialized role in stress-induced ABA formation, thus suggesting that the first committed step in carotenogenesis is a key limiting step in ABA biosynthesis. To examine whether the ectopic expression of PSY, other than PSY3, would similarly affect ABA level and stress tolerance, we have produced transgenic tobacco containing a fruit-specific PSY (CpPSY) of grapefruit (Citrus paradisi Macf.). The transgenic plants contained a single- or double-locus insertion and expressed CpPSY at varying transcript levels. In comparison with the wild-type plants, the CpPSY expressing transgenic plants showed a significant increase on root length and shoot biomass under PEG-, NaCl- and mannitol-induced osmotic stress. The enhanced stress tolerance of transgenic plants was correlated with the increased endogenous ABA level and expression of stress-responsive genes, which in turn was correlated with the CpPSY copy number and expression level in different transgenic lines. Collectively, these results provide further evidence that PSY is a key enzyme regulating ABA biosynthesis and that the altered expression of other PSYs in transgenic plants may provide a similar function to that of the monocot's PSY3 in ABA biosynthesis and stress tolerance. The results also pave the way for further use of CpPSY, as well as other PSYs, as potential candidate genes for engineering tolerance to drought and salt stress in crop plants.

Alkaline sulfite/anthraquinone pretreatment followed by disk refining of Pinus radiata and Pinus caribaea wood chips for biochemical ethanol production

BACKGROUND: Alkaline sulfite/anthraquinone (ASA) cooking of Pinus radiata and Pinus caribaea wood chips followed by disk refining was used as a pretreatment for the production of low lignified and high fibrillated pulps. The pulps produced with different delignification degrees and refined at different energy inputs (250, 750 and 1600 Wh) were saccharified with cellulases and fermented to ethanol with Saccharomyces cerevisiae using separated hydrolysis and fermentation (SHF) or semi-simultaneous saccharification and fermentation (SSSF) processes. RESULTS: Delignification of ASA pulps was between 25% and 50%, with low glucans losses. Pulp yield was from 70 to 78% for pulps of P. radiata and 60% for the pulp of P. caribaea. Pulps obtained after refining were evaluated in assays of enzymatic hydrolysis. Glucans-to-glucose conversion varied from 20 to 70%, depending on the degree of delignification and fibrillation of the pulps. The best ASA pulp of P. radiata was used in SHF and SSSF experiments of ethanol production. Such experiments produced maximum ethanol concentration of 20 g L-1, which represented roughly90% of glucose conversion and an estimated amount of 260 L ethanol ton(-1) wood. P. caribaea pulp also presented good performance in the enzymatic hydrolysis and fermentation but, due to the low amount of cellulose present, only 140 L ethanol would be obtained from each ton of wood. CONCLUSION: ASA cooking followed by disk refining was shown to be an efficient pretreatment process, which generated a low lignified and high-fibrillated substrate that allowed the production of ethanol from the softwoods with high conversion yields. (C) 2012 Society of Chemical Industry

The distribution and abundance of small mammals in agroecosystems of southeastern Brazil

The conservation of biodiversity in agricultural landscapes depends on information about the ways in which species are affected by the conversion of native habitats into novel anthropogenic environments and the strategies that the species use to persist in these altered ecosystems. Here, we investigate how small mammals occupy the different agroecosystems of an agricultural landscape in the state of Sao Paulo, Brazil. From August 2003 through January 2005, we surveyed small mammals using Sherman traps at 16 sampling sites in each of the four predominant environments of the local agricultural landscape: remnant fragments of semideciduous forest, Eucalyptus plantations, sugarcane plantations, and pastures. With a total effort of 23,040 trap-nights and a capture success of 0.8%, we captured 177 rodents and marsupials belonging to eight species. The assemblage represented by these mammals is essentially composed of generalist species, which are common in degraded areas. Sugarcane plantations had the highest abundance, whereas pastures had the lowest species richness. Our results suggest that the loss of forest species can be related to the loss of native forest. The results also indicate that to improve the conservation value of agricultural landscapes, native forest fragments should be conserved, extensive monocultures should be avoided and agricultural impacts should be mitigated.

Toward Global Standards for Comparator Pharmaceutical Products: Case Studies of Amoxicillin, Metronidazole, and Zidovudine in the Americas

This study compared in vitro dissolution characteristics and other quality measures of different amoxicillin, metronidazole, and zidovudine products purchased in the Americas to a comparator pharmaceutical product (CPP). These three drugs are classified as Biopharmaceutics Classification System Class I drugs with the possibility that dissolution findings might be used to document bioequivalence. All investigated zidovudine products were found to be in vitro equivalent to the CPP. Only 3 of 12 tested amoxicillin products were found to be in vitro equivalent to the CPP. None of the tested metronidazole products were in vitro equivalent to the CPP. These findings suggest but do not confirm bioinequivalence where in vitro comparisons failed, given that an in vivo blood level study might have confirmed bioequivalence. At times, identifying a CPP in one of the selected markets proved difficult. The study demonstrates that products sold across national markets may not be bioequivalent. When coupled with the challenge of identifying a CPP in different countries, the results of this study suggest the value of an international CPP as well as increased use of BCS approaches as means of either documenting bioequivalence or signaling the need for further in vivo studies. Because of increased movement of medicines across national borders, practitioners and patients would benefit from these approaches.

The antioxidative effect of de novo generated vitamin B-6 in Plasmodium falciparum validated by protein interference

The malaria parasite Plasmodium falciparum is able to synthesize de novo PLP (pyridoxal 5'-phosphate), the active form of vitamin B-6. In the present study, we have shown that the de novo synthesized PLP is used by the parasite to detoxify O-1(2) (singlet molecular oxygen), a highly destructive reactive oxygen species arising from haemoglobin digestion. The formation of O-1(2) and the response of the parasite were monitored by live-cell fluorescence microscopy, by transcription analysis and by determination of PLP levels in the parasite. Pull-down experiments of transgenic parasites overexpressing the vitamin B-6-biosynthetic enzymes PfPdx1 and PfPdx2 clearly demonstrated an interaction of the two proteins in vivo which results in an elevated PLP level from 12.5 mu M in wild-type parasites to 36.6 mu M in the PfPdx1/PfPdx2-overexpressing cells and thus to a higher tolerance towards O-1(2). In contrast, by applying the dominant-negative effect on the cellular level using inactive mutants of PfPdx1 and PfPdx2, P. falciparum becomes susceptible to O-1(2). Our results demonstrate clearly the crucial role of vitamin B-6 biosynthesis in the detoxification of O-1(2) in P falciparum. Besides the known role of PLP as a cofactor of many essential enzymes, this second important task of the vitamin B-6 de novo synthesis as antioxidant emphasizes the high potential of this pathway as a target of new anti-malarial drugs.

Magnetic and optical investigation of 40SiO(2)center dot 30Na(2)O center dot 1Al(2)O(3)center dot(29-x) B2O3 center dot xFe(2)O(3) glass matrix

Samples of 40SiO(2)center dot 30Na(2)O center dot 1Al(2)O(3)center dot(29 - x)B2O3 center dot xFe(2)O(3) (mol%), with 0.0 <= x <= 17.5, were prepared by the fusion method and investigated by electron paramagnetic resonance (EPR), optical absorption (OA) and Mossbauer spectroscopy (MS). The EPR spectra of the as-synthesized samples exhibit two well-defined EPR signals around g = 4.27 and g = 2.01 and a visible EPR shoulder around g = 6.4, assigned to isolated Fe3+ ion complexes (g = 4.27 and g = 6.4) and Fe3+-based clusters (g = 2.01). Analyses of both EPR line intensity and line width support the model picture of Fe3+-based clusters built in from two sources of isolated ions, namely Fe2+ and Fe3+; the ferrous ion being used to build in iron-based clusters at lower x-content (below about x = 2.5%) whereas the ferric ion is used to build in iron-based clusters at higher x-content (above about x = 2.5%). The presence of Fe2+ ions incorporated within the glass template is supported by OA data with a strong band around 1100 nm due to the spin-allowed E-5(g)-T-5(2g) transition in an octahedral coordination with oxygen. Additionally, Mossbauer data (isomer shift and quadrupole splitting) confirm incorporation of both Fe2+ and Fe3+ ions within the template, more likely in tetrahedral-like environments. We hypothesize that ferrous ions are incorporated within the glass template as FeO4 complex resulting from replacing silicon in non-bridging oxygen (SiO3O-) sites whereas ferric ions are incorporated as FeO4 complex resulting from replacing silicon in bridging-like oxygen silicate groups (SiO4). (C) 2012 Elsevier Masson SAS. All rights reserved.

Staining of the Internal Limiting Membrane with the Use of Heavy Brilliant Blue G

Background: Brilliant blue G (BBG) is frequently used in chromovitrectomy to facilitate internal limiting membrane (ILM) peeling. A study was initiated to evaluate if heavy BBG is safe and effective in staining the ILM. Methods: We studied 30 eyes, 23 with idiopathic macular holes and 7 of patients with diabetic macular edema. Removal of the ILMs was assisted by heavy BBG staining. In cases with histopathological correlation the ILMs were evaluated with hematoxylin and eosin, Masson's trichrome, periodic acid-Schiff and glial fibrillary acidic protein staining. In addition, immunohistochemistry was also performed using specific antibodies for vimentin, neuron-specific enolase, factor VIII and CD68. Using the Image-Pro Plus software of Media Cybernetics Co. we found an average thickness in ILMs. Results: Of the ILM specimens sent, 19/30(63.33%) could not be processed properly because of the limited sample material, recognizing only fragments of dispersed fibrillar material. In macular hole ILMs we found an average thickness of 1.3 +/- 0.65 mu m, and in diabetic macular edema ILMs an average thickness of 6.2 +/- 1.4 mu m. Conclusions: In heavy BBG-assisted ILM peeling we observed no intraoperative or postoperative complications after a mean follow-up of 12 months. Heavy BBG could be an effective and safe vehicle for staining the ILM. Copyright (C) 2012 S. Karger AG, Basel

Description of the female and new records of Triatoma baratai Carcavallo & Jurberg, 2000 (Hemiptera: Heteroptera: Reduviidae: Triatominae) from Mato Grosso do Sul, Brazil, with a key to the species of the Triatoma matogrossensis subcomplex

Triatoma baratai Carcavallo & Jurberg, 2000, a species similar to Triatoma williami Galvao, Souza & Lima, 1967 and belonging to the T. matogrossensis subcomplex, was described based on a male specimen collected in a sylvatic environment, near a cave, in Bonito county, Bodoquena mountain range, state of Mato Grosso do Sul, Brazil. In the present work we describe the female of T. baratai, captured in a chicken house, in Nioaque county, state of Mato Grosso do Sul, Brazil. Furthermore, we recorded the occurrence of T. baratai in domiciles and peridomestic environment in another four municipalities (Bodoquena, Bela Vista, Corumba, and Miranda), extending its geographical distribution. Finally, we present a key to the species of the Triatoma matogrossensis subcomplex.

Measurement of prompt J/psi and beauty hadron production cross sections at mid-rapidity in pp collisions at root s=7 TeV

The ALICE experiment at the LHC has studied J/psi production at mid-rapidity in pp collisions at root s = 7 TeV through its electron pair decay on a data sample corresponding to an integrated luminosity L-int = 5.6 nb(-1). The fraction of J/psi from the decay of long-lived beauty hadrons was determined for J/psi candidates with transverse momentum p(t) > 1,3 GeV/c and rapidity vertical bar y vertical bar < 0.9. The cross section for prompt J/psi mesons, i.e. directly produced J/psi and prompt decays of heavier charmonium states such as the psi(2S) and chi(c) resonances, is sigma(prompt J/psi) (p(t) > 1.3 GeV/c, vertical bar y vertical bar < 0.9) = 8.3 +/- 0.8(stat.) +/- 1.1 (syst.)(-1.4)(+1.5) (syst. pol.) mu b. The cross section for the production of b-hadrons decaying to J/psi with p(t) > 1.3 GeV/c and vertical bar y vertical bar < 0.9 is a sigma(J/psi <- hB) (p(t) > 1.3 GeV/c, vertical bar y vertical bar < 0.9) = 1.46 +/- 0.38 (stat.)(-0.32)(+0.26) (syst.) mu b. The results are compared to QCD model predictions. The shape of the p(t) and y distributions of b-quarks predicted by perturbative QCD model calculations are used to extrapolate the measured cross section to derive the b (b) over bar pair total cross section and d sigma/dy at mid-rapidity.

Decisions on Temporal Sampling Protocol Influence the Detection of Ecological Patterns

Within-site variability in species detectability is a problem common to many biodiversity assessments and can strongly bias the results. Such variability can be caused by many factors, including simple counting inaccuracies, which can be solved by increasing sample size, or by temporal changes in species behavior, meaning that the way the temporal sampling protocol is designed is also very important. Here we use the example of mist-netted tropical birds to determine how design decisions in the temporal sampling protocol can alter the data collected and how these changes might affect the detection of ecological patterns, such as the species-area relationship (SAR). Using data from almost 3400 birds captured from 21,000 net-hours at 31 sites in the Brazilian Atlantic Forest, we found that the magnitude of ecological trends remained fairly stable, but the probability of detecting statistically significant ecological patterns varied depending on sampling effort, time of day and season in which sampling was conducted. For example, more species were detected in the wet season, but the SAR was strongest in the dry season. We found that the temporal distribution of sampling effort was more important than its total amount, discovering that similar ecological results could have been obtained with one-third of the total effort, as long as each site had been equally sampled over 2 yr. We discuss that projects with the same sampling effort and spatial design, but with different temporal sampling protocol are likely to report different ecological patterns, which may ultimately lead to inappropriate conservation strategies.

The importance of landscape structure for seed dispersal in rain forest fragments

Questions What are the main features of the seed rain in a fragmented Atlantic forest landscape? Can seed rain species attributes (life form, dispersal mode, successional status) relate to the spatial arrangement (size and number of fragments, edge density and presence of corridor) of forest fragments in the landscape? How does the rain forest landscape structure affect the seed rain? Location Atlantic rainforest, Sao Paulo State, Southeastern Brazil. Methods Seed rain samples were collected monthly throughout 1yr, counted, identified and classified according to species dispersal mode, successional status and life form. Seed rain composition was compared with woody species near the seed traps. Relationships between seed rain composition and landscape spatial arrangement (fragment area, presence of corridor, number of fragments in the surroundings, proximity of fragments, and edge density) were tested using canonical correspondence analysis (CCA). Results We collected 20142 seeds belonging to 115 taxa, most of them early successional and anemochorous trees. In general, the seed rain had a species composition distinct from that of the nearby forest tree community. Small isolated fragments contained more seeds, mainly of anemochorous, epiphytic and early-successional species; large fragments showed higher association with zoochorous and late-successional species compared to small fragments. The CCA significantly distinguished the species dispersal mode according to fragment size and isolation, anemochorous species being associated to small and isolated fragments, and zoochorous species to larger areas and fragment aggregation. Nevertheless, a gradient driven by proximity (PROX) and edge density (ED) segregated lianas (in the positive extremity), early successional and epiphyte species (in the negative end); large fragments were positively associated to PROX and ED. Conclusions The results highlight the importance of the size and spatial arrangement of forest patches to promote habitat connectivity and improve the flux of animal-dispersed seeds. Landscape structure controls seed fluxes and affects plant dispersal capacity, potentially influencing the composition and structure of forest fragments. The seed rain composition may be used to assess the effects of landscape spatial structure on plant assemblages, and provide relevant information for biodiversity conservation.

Rationale, study design, and analysis plan of the Alveolar Recruitment for ARDS Trial (ART): Study protocol for a randomized controlled trial

Background: Acute respiratory distress syndrome (ARDS) is associated with high in-hospital mortality. Alveolar recruitment followed by ventilation at optimal titrated PEEP may reduce ventilator-induced lung injury and improve oxygenation in patients with ARDS, but the effects on mortality and other clinical outcomes remain unknown. This article reports the rationale, study design, and analysis plan of the Alveolar Recruitment for ARDS Trial (ART). Methods/Design: ART is a pragmatic, multicenter, randomized (concealed), controlled trial, which aims to determine if maximum stepwise alveolar recruitment associated with PEEP titration is able to increase 28-day survival in patients with ARDS compared to conventional treatment (ARDSNet strategy). We will enroll adult patients with ARDS of less than 72 h duration. The intervention group will receive an alveolar recruitment maneuver, with stepwise increases of PEEP achieving 45 cmH(2)O and peak pressure of 60 cmH2O, followed by ventilation with optimal PEEP titrated according to the static compliance of the respiratory system. In the control group, mechanical ventilation will follow a conventional protocol (ARDSNet). In both groups, we will use controlled volume mode with low tidal volumes (4 to 6 mL/kg of predicted body weight) and targeting plateau pressure <= 30 cmH2O. The primary outcome is 28-day survival, and the secondary outcomes are: length of ICU stay; length of hospital stay; pneumothorax requiring chest tube during first 7 days; barotrauma during first 7 days; mechanical ventilation-free days from days 1 to 28; ICU, in-hospital, and 6-month survival. ART is an event-guided trial planned to last until 520 events (deaths within 28 days) are observed. These events allow detection of a hazard ratio of 0.75, with 90% power and two-tailed type I error of 5%. All analysis will follow the intention-to-treat principle. Discussion: If the ART strategy with maximum recruitment and PEEP titration improves 28-day survival, this will represent a notable advance to the care of ARDS patients. Conversely, if the ART strategy is similar or inferior to the current evidence-based strategy (ARDSNet), this should also change current practice as many institutions routinely employ recruitment maneuvers and set PEEP levels according to some titration method.