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The nucleocapsid (N) protein of SARS-CoV-2 is a multifunctional protein involved in nucleocapsid assembly and various regulatory functions. It is the most abundant protein during viral infection. Its functionality is closely related to its structure, which comprises two globular domains, the N-terminal domain (NTD) and the C-terminal domain (CTD), flanked by intrinsically disordered regions. The linker between the NTD and CTD includes a Serine-Arginine rich (SR) region, which is crucial for the regulation of the N protein's function. Here, we report the near-complete assignment of the construct containing the NTD followed by the SR region (NTD-SR). Additionally, we describe the dynamic nature of the SR region and compare it with all other available chemical shift assignments reported for the SR region., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

The applications of fluorinated molecules in chemical biology are rapidly expanding driven by the unique properties of C-F bonds, leading to increased interest in methodologies for controlled introduction of this atom. In this study, we present the first method for late-stage fluorination of tyrosine residues in proteins. Our results demonstrate that electrophilic fluorination using Selectfluor, a stable and non-toxic reagent, offers a straightforward and cost-effective method for labeling Cyanovirin-N (CVN), a 101-amino-acid lectin with effective antiviral activity. Uni- and bidimensional 1 H, 13 C and 19 F NMR analyses, along with mass spectrometry, revealed chemoselective fluorination of the three tyrosine residues in CVN without affecting its overall structure or mannose-binding affinity. Additionally, we observed that other aromatic amino acids, such as tryptophan, phenylalanine, and histidine, are not fluorinated using this method. These findings advance our understanding of protein fluorination and its applications in studying structure, dynamics, and interactions, as well as other biological utilities., (© 2024 Wiley-VCH GmbH.)

Disintegrins are a family of cysteine-rich small proteins that were first identified in snake venom. The high divergence of disintegrins gave rise to a plethora of functions, all related to the interaction with integrins. Disintegrins evolved to interact selectively with different integrins, eliciting many physiological outcomes and being promising candidates for the therapy of many pathologies. We used NMR to determine the structure and dynamics of the recombinant disintegrin jarastatin (rJast) and its interaction with the cancer-related integrin αVβ3. rJast displayed the canonical fold of a medium-sized disintegrin and showed complex dynamic in multiple timescales. We used NMR experiments to map the interaction of rJast with αVβ3, and molecular docking followed by molecular dynamics (MD) simulation to describe the first structural model of a disintegrin/integrin complex. We showed that not only the RGD loop participates in the interaction, but also the N-terminal domain. rJast plasticity was essential for the interaction with αVβ3 and correlated with the main modes of motion depicted in the MD trajectories. In summary, our study provides novel structural insights that enhance our comprehension of the mechanisms underlying disintegrin functionality., Competing Interests: Declaration of competing interest The authors declare no competing interests., (Copyright © 2023 Elsevier B.V. All rights reserved.)

The maize yield, nutritional status, and grain fumonisins concentration were evaluated in different genotypes, doses, and nitrogen sources (N) in two years and three locations. Two experiments were carried out in each area and year in an experimental design of a subdivided plot with four replications. One experiment involved a 4x2 factorial treatment: four nitrogen (N) doses (0, 80, 160, and 240 kg ha-1) in coverage and having urea as a source of N and two genotypes. Another experiment involved a 4x2 factorial treatment: four N sources: urea, urea covered with polymer, ammonium nitrate, and ammonium nitrate + urea (UAN), at a dose of 160 kg ha-1, in two genotypes. The genotype generally influenced maize yield more than N doses and sources, mainly due to the bushy stunt/corn stunt tolerance of AG7098 PRO2 and AG8677 PRO2. The N doses linearly increased the N leaf content. However, the N sources did not affect the N leaf content. The N doses and sources had no significant effect on the content of fumonisins, which was affected only by the genotypes in Sete Lagoas in 2016 (N doses experiment) and 2017 (N sources experiment). The hybrids, P3630H and AG8677PRO2 (Sete Lagoas, 2016, N doses experiment and 2017, N sources experiment, respectively) exceeded the Brazilian legislation for Maximum Tolerance Limit for fumonisins in corn grains, which is 5,000 µg kg-1. The best result was obtained with AG7098 PRO2, with yields (above 10,000 kg ha-1) and fumonisins consistently below 5,000 µg kg-1. Therefore, the selection of corn hybrids is a strategy to reduce the occurrence of fumonisins in the grains.

Vermicomposting is the bio-oxidation and stabilization of organic matter involving relationships between the action of earthworms and microorganisms and the activation and dynamics of several enzyme activities. Semi-arid farmers to make (extra) money and organic production, produce their vermicompost using plant residues and animal manure, but there is no information about the final product generated. Thus, this study aimed to analyze the potential of vermicomposting with mixtures of animal manure and vegetable leaves in the development of Eisenia foetida, microbial biomass, and enzymatic activity in the semi-arid region, Brazil. The experimental design applied was randomized block in a 6 × 4 factorial scheme with four replicates, with six treatments (mixtures of cattle manure, goat manure, cashew leaves, and catanduva leaves) and evaluated at four-time intervals (30, 60, 90, and 120 days of vermicomposting). The treatments were placed in polyethylene pots in the same site, environmental conditions, and residues proportions as used by farmers. The characteristics analyzed were the number of earthworms (NE), total earthworm biomass (TEB) and earthworm multiplication index (MI), microbial biomass carbon (MBC), and activities of enzymes β-glucosidase, dehydrogenase, alkaline and acid phosphatases. The cattle manure vermicomposted shows the highest average values observed for NE, MI, TEB, MBC, and enzymatic activity, regardless of the plant leaves mix. In general, the enzymes activities were found in the descending order of β-glucosidase > alkaline phosphatase > dehydrogenase > acid phosphatase. The maturation dynamics of vermicompost were characterized by a decline in the microbial population and number and biomass of earthworms in the substrate and consequently a decrease in new enzyme synthesis and degradation of the remaining enzyme pool. Microbial biomass and enzymatic activity were indicators for changes in the quality of vermicompost., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

D-xylose utilization by yeasts is an essential feature for improving second-generation ethanol production. However, industrial yeast strains are incapable of consuming D-xylose. Previous analyzes of D-xylose-consuming or fermenting yeast species reveal that the genomic features associated with this phenotype are complex and still not fully understood. Here we present a previously neglected yeast enzyme related to D-xylose metabolism, D-xylose dehydrogenase (XylDH), which is found in at least 105 yeast genomes. By analyzing the XylDH gene family, we brought evidence of gene evolution marked by purifying selection on codons and positive selection evidence in D-xylose-consuming and fermenting species, suggesting the importance of XylDH for D-xylose-related phenotypes in yeasts. Furthermore, although we found no putative metabolic pathway for XylDH in yeast genomes, namely the absence of three bacterial known pathways for this enzyme, we also provide its expression profile on D-xylose media following D-xylose reductase for two yeasts with publicly available transcriptomes. Based on these results, we suggest that XylDH plays an important role in D-xylose usage by yeasts, likely being involved in a cofactor regeneration system by reducing cofactor imbalance in the D-xylose reductase pathway., (© The Author(s) 2023. Published by Oxford University Press on behalf of FEMS.)

This study investigated the chemical composition and evaluated the antibacterial and antibiofilm activities of essential oils (EOs) extracted from Ruellia asperula (EORA) and Ruellia paniculata (EORP) against oral streptococci. The EO constituents were analyzed by gas chromatography/mass spectrometry. The antimicrobial potential of EOs was evaluated using the minimum inhibitory concentration, minimum bactericidal concentration, and time-kill determination. Furthermore, the quantification of total biomass and the number of viable cells in the biofilms were evaluated. The major constituents of EORA were cariophylla-4(12)-8-(13)-dien-5β-ol (14.1%), (β)-caryophyllene (22.7%), and caryophyllene oxide (29.4%). For EORP, the major constituents were (β)-caryophyllene (11.0%), spathulenol (13.1%), and δ-amorphene (14.9%). The tested EOs exhibited antibacterial activity against planktonic growth and biofilm formation. Thus, the EOs from R. asperula and R. paniculata prove to be promising alternatives for bacterial growth control and biofilm formation prevention of oral streptococci.

Sponges have co-evolved with microbes for over 400 myr. Previous studies have demonstrated that sponges can be classified according to the abundance of microbes in their tissues as Low Microbial Abundance (LMA) and High Microbial Abundance (HMA). While LMA sponges rely mainly on water column microbes, HMA appear to rely much more on symbiotic fermentative and autotrophic microbes maintained in their tissues. However, it is unclear if this pattern holds when comparing different species of tropical sponges under extreme nutrient conditions and sediment loads in the water column, such as the Great Amazon Reef System (GARS), which covers an area of ~56,000 km 2 off the Amazon River mouth. Sponges are the major GARS benthic components. However, these sponges' microbiome across the GARS is still unknown. Here, we investigated water quality, isotopic values (δ 13 C and δ 15 N), metagenomic and lipidomic profiles of sponges obtained from different sectors throughout the GARS. >180 million shotgun metagenomic reads were annotated, covering 22 sponge species. Isotopic and lipidomic analyses suggested LMA sponges rely on the Amazon River Plume for nutrition. HMA sponges (N = 15) had higher Roseiflexus and Nitrospira abundance, whereas LMA sponges (N = 7) had higher Prochlorococcus and Pelagibacter abundance. Functional data revealed that the LMA sponge microbiomes had greater number of sequences related to phages and prophages as well as electron transport and photophosphorylation which may be related to photosynthetic processes associated with the Prochlorococcus and Synechococcus found in the LMA. The higher phages abundance in LMA sponges could be related to these holobionts' reduced defense towards phage infection. Meanwhile, HMA sponge microbiomes had higher Clustered Regularly Interspaced Short Palindromic Repeats-CRISPR abundance, which may be involved in defense against phage infection. This study sheds light on the nutrient fluxes and microbes from the Amazon River plume into the sponge holobionts., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022. Published by Elsevier B.V.)

Disintegrins are a group of cysteine-rich proteins found in a wide variety of snake venoms. These proteins selectively bind to integrins, which play a fundamental role in the regulation of many physiological and pathological processes. Here, we report the NMR chemical shift assignments for 1 H, 15 N, and 13 C nuclei in the backbone and side chains of recombinant disintegrin Jarastatin (rJast), which was further validated by secondary structure prediction using the TALOS-N server. Taken together, these data are essential to perform NMR-based experiments, including structure determination, backbone dynamics, mapping ligand sites and enabling a deeper understanding of the effect of hydrophobic surface clusters, which are important elements to stabilize some 3D proteins structure/folding., (© 2021. The Author(s), under exclusive licence to Springer Nature B.V.)

The SARS-CoV-2 nucleocapsid protein (N) is a multifunctional promiscuous nucleic acid-binding protein, which plays a major role in nucleocapsid assembly and discontinuous RNA transcription, facilitating the template switch of transcriptional regulatory sequences (TRS). Here, we dissect the structural features of the N protein N-terminal domain (N-NTD) and N-NTD plus the SR-rich motif (N-NTD-SR) upon binding to single and double-stranded TRS DNA, as well as their activities for dsTRS melting and TRS-induced liquid-liquid phase separation (LLPS). Our study gives insights on the specificity for N-NTD(-SR) interaction with TRS. We observed an approximation of the triple-thymidine (TTT) motif of the TRS to β-sheet II, giving rise to an orientation difference of ~25° between dsTRS and non-specific sequence (dsNS). It led to a local unfavorable energetic contribution that might trigger the melting activity. The thermodynamic parameters of binding of ssTRSs and dsTRS suggested that the duplex dissociation of the dsTRS in the binding cleft is entropically favorable. We showed a preference for TRS in the formation of liquid condensates when compared to NS. Moreover, our results on DNA binding may serve as a starting point for the design of inhibitors, including aptamers, against N, a possible therapeutic target essential for the virus infectivity., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Nanotechnology is a cutting-edge area with numerous industrial applications. Nanoparticles are structures that have dimensions ranging from 1 - 100 nm, which significantly exhibit different mechanical, optical, electrical, and chemical properties when compared with their larger counterparts. Synthetic routes that use natural sources, such as plant extracts, honey, and microorganisms, are environmentally friendly and low-cost methods that can be used to obtain nanoparticles. These methods of synthesis generate products that are more stable and less toxic than those obtained using conventional methods. Nanoparticles formed by titanium dioxide, zinc oxide, silver, gold, and copper, as well as cellulose nanocrystals, are among the nanostructures obtained by green synthesis that have shown interesting applications in several technological industries. Several analytical techniques have also been used to analyze the size, morphology, hydrodynamics, diameter, and chemical functional groups involved in the stabilization of the nanoparticles as well as to quantify and evaluate their formation. Despite their pharmaceutical, biotechnological, cosmetic, and food applications, studies have detected their harmful effects on human health and the environment, and thus, caution must be taken in uses involving living organisms. The present review aims to present an overview of the applications, the structural properties, and the green synthesis methods that are used to obtain nanoparticles, and special attention is given to those obtained from metal ions. The review also presents the analytical methods used to analyze, quantify, and characterize these nanostructures., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

The routes of uptake and efflux should be considered when developing new drugs so that they can effectively address their intracellular targets. As a general rule, drugs appear to enter cells via protein carriers that normally carry nutrients or metabolites. A previously developed pipeline that searched for drug transporters using Saccharomyces cerevisiae mutants carrying single-gene deletions identified import routes for most compounds tested. However, due to the redundancy of transporter functions, we propose that this methodology can be improved by utilizing double mutant strains in both low- and high-throughput screens. We constructed a library of over 14,000 strains harboring double deletions of genes encoding 122 nonessential plasma membrane transporters and performed low- and high-throughput screens identifying possible drug import routes for 23 compounds. In addition, the high-throughput assay enabled the identification of putative efflux routes for 21 compounds. Focusing on azole antifungals, we were able to identify the involvement of the myo- inositol transporter, Itr1p, in the uptake of these molecules and to confirm the role of Pdr5p in their export. IMPORTANCE Our library of double transporter deletion strains is a powerful tool for rapid identification of potential drug import and export routes, which can aid in determining the chemical groups necessary for transport via specific carriers. This information may be translated into a better design of drugs for optimal absorption by target tissues and the development of drugs whose utility is less likely to be compromised by the selection of resistant mutants.

Disintegrins are small cysteine-rich proteins found in a variety of snake venom. These proteins selectively modulate integrin function, heterodimeric receptors involved in cell-cell and cell-matrix interaction that are widely studied as therapeutic targets. Snake venom disintegrins emerged from the snake venom metalloproteinase and are classified according to the sequence size and number of disulfide bonds. Evolutive structure and function diversification of disintegrin family involves a stepwise decrease in the polypeptide chain, loss of cysteine residues, and selectivity. Since the structure elucidation of echistatin, the description of the structural properties of disintegrins has allowed the investigation of the mechanisms involved in integrin-cell-extracellular matrix interaction. This review provides an analysis of the structures of all family groups enabling the description of an expanded classification of the disintegrin family in seven groups. Each group presents a particular disulfide pattern and sequence signatures, facilitating the identification of new disintegrins. The classification was based on the disintegrin-like domain of the human metalloproteinase (ADAM-10). We also present the sequence and structural signatures important for disintegrin-integrin interaction, unveiling the relationship between the structure and function of these proteins., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Vasconcelos, Estrada, David, Wermelinger, Almeida and Zingali.)

Schinus terebinthifolia is a species native to different ecoregions in the Brazilian Atlantic Forest. The plant is listed on the National Relation of Medicinal Plants and recommended as phytomedicine, however while extractive exploitation prevails as the main route of raw material a significant variation of compounds will be detected. To assure the expansion of productive chain it is important to start by studying population diversity and chemical variations. We used SSR markers for studies of genetic structure among populations from dense ombrophilous forest (ES); the deciduous seasonal forest (SM); the savanna (DOU) and the sandbanks (ITA and MSP), and compared the results to their chemical profiles of essential oil. Genetic structure revealed differences among populations and significant fixation rates. Pairwise studies and Bayesian analysis showed similarities between ITA and SM and between DOU and MSP, proving that the patterns of distribution for the species do not follow the isolation by distance or similarity by environmental conditions. The comparison between PCA of genotypes and chemodiversity reinforces the unique profile for each population despite the environmental similarity observed and genetic analysis. The most divergent genotype and chemical group was found at the ombrophilous forest, strong evidence that we should undertake conservation efforts to prevent losses of biodiversity in that area., (© 2021. The Author(s).)

Background: Plant pathogenesis related-1 (PR-1) proteins belong to the CAP superfamily and have been characterized as markers of induced defense against pathogens. Moniliophthora perniciosa and Moniliophthora roreri are hemibiotrophic fungi that respectively cause the witches' broom disease and frosty pod rot in Theobroma cacao. Interestingly, a large number of plant PR-1-like genes are present in the genomes of both species and many are up-regulated during the biotrophic interaction. In this study, we investigated the evolution of PR-1 proteins from 22 genomes of Moniliophthora isolates and 16 other Agaricales species, performing genomic investigation, phylogenetic reconstruction, positive selection search and gene expression analysis., Results: Phylogenetic analysis revealed conserved PR-1 genes (PR-1a, b, d, j), shared by many Agaricales saprotrophic species, that have diversified in new PR-1 genes putatively related to pathogenicity in Moniliophthora (PR-1f, g, h, i), as well as in recent specialization cases within M. perniciosa biotypes (PR-1c, k, l) and M. roreri (PR-1n). PR-1 families in Moniliophthora with higher evolutionary rates exhibit induced expression in the biotrophic interaction and positive selection clues, supporting the hypothesis that these proteins accumulated adaptive changes in response to host-pathogen arms race. Furthermore, although previous work showed that MpPR-1 can detoxify plant antifungal compounds in yeast, we found that in the presence of eugenol M. perniciosa differentially expresses only MpPR-1e, k, d, of which two are not linked to pathogenicity, suggesting that detoxification might not be the main function of most MpPR-1., Conclusions: Based on analyses of genomic and expression data, we provided evidence that the evolution of PR-1 in Moniliophthora was adaptive and potentially related to the emergence of the parasitic lifestyle in this genus. Additionally, we also discuss how fungal PR-1 proteins could have adapted from basal conserved functions to possible roles in fungal pathogenesis.

Antimalarial drugs with novel modes of action and wide therapeutic potential are needed to pave the way for malaria eradication. Violacein is a natural compound known for its biological activity against cancer cells and several pathogens, including the malaria parasite, Plasmodium falciparum (Pf). Herein, using chemical genomic profiling (CGP), we found that violacein affects protein homeostasis. Mechanistically, violacein binds Pf chaperones, Pf Hsp90 and Pf Hsp70-1, compromising the latter's ATPase and chaperone activities. Additionally, violacein-treated parasites exhibited increased protein unfolding and proteasomal degradation. The uncoupling of the parasite stress response reflects the multistage growth inhibitory effect promoted by violacein. Despite evidence of proteotoxic stress, violacein did not inhibit global protein synthesis via UPR activation-a process that is highly dependent on chaperones, in agreement with the notion of a violacein-induced proteostasis collapse. Our data highlight the importance of a functioning chaperone-proteasome system for parasite development and differentiation. Thus, a violacein-like small molecule might provide a good scaffold for development of a novel probe for examining the molecular chaperone network and/or antiplasmodial drug design.

Premise: A novel set of nuclear microsatellite markers was developed and characterized for Campomanesia adamantium (Myrtaceae) and tested for cross-amplification in the related species C. sessiliflora ., Methods and Results: Forty-one primer pairs were designed for simple sequence repeat loci, of which 36 successfully amplified and were polymorphic. The number of alleles ranged from two to 14, with an average of 8.14 alleles per locus. Additionally, cross-amplification was tested in C. sessiliflora ; more than 55.5% of the microsatellite loci amplified, confirming the use of these microsatellite markers in a related species., Conclusions: We developed a set of microsatellite markers that will be useful for future studies of genetic diversity and population structure of C. adamantium and a closely related species, which will aid in future conservation efforts., (© 2019 Crispim et al. Applications in Plant Sciences is published by Wiley Periodicals, Inc. on behalf of the Botanical Society of America.)

Galactosaminoglycans (GalAGs) are sulfated glycans composed of alternating N -acetylgalactosamine and uronic acid units. Uronic acid epimerization, sulfation patterns and fucosylation are modifications observed on these molecules. GalAGs have been extensively studied and exploited because of their multiple biomedical functions. Chondroitin sulfates (CSs), the main representative family of GalAGs, have been used in alternative therapy of joint pain/inflammation and osteoarthritis. The relatively novel fucosylated chondroitin sulfate (FCS), commonly found in sea cucumbers, has been screened in multiple systems in addition to its widely studied anticoagulant action. Biomedical properties of GalAGs are directly dependent on the sugar composition, presence or lack of fucose branches, as well as sulfation patterns. Although research interest in GalAGs has increased considerably over the three last decades, perhaps motivated by the parallel progress of glycomics, serious questions concerning the effectiveness and potential side effects of GalAGs have recently been raised. Doubts have centered particularly on the beneficial functions of CS-based therapeutic supplements and the potential harmful effects of FCS as similarly observed for oversulfated chondroitin sulfate, as a contaminant of heparin. Unexpected components were also detected in CS-based pharmaceutical preparations. This review therefore aims to offer a discussion on (1) the current and potential therapeutic applications of GalAGs, including those of unique features extracted from marine sources, and (2) the potential drawbacks of this class of molecules when applied to medicine.

The anticoagulant and antithrombotic properties of three structurally correlated sea urchin-derived 3-linked sulfated α-glycans and their low molecular-weight derivatives were screened comparatively through various in vitro and in vivo methods. These methods include activated partial thromboplastin time, the inhibitory activity of antithrombin over thrombin and factor Xa, venous antithrombosis, the inhibition of platelet aggregation, the activation of factor XII, and bleeding. While the 2-sulfated fucan from Strongylocentrotus franciscanus was observed to be poorly active in most assays, the 4-sulfated fucan from Lytechinus variegatus , the 2-sulfated galactan from Echinometra lucunter and their derivatives showed multiple effects. All marine compounds showed no capacity to activate factor XII and similar low bleeding tendencies regardless of the dose concentrations used to achieve the highest antithrombotic effect observed. The 2-sulfated galactan showed the best combination of results. Our work improves the background about the structure-function relationship of the marine sulfated glycans in anticoagulation and antithrombosis. Besides confirming the negative effect of the 2-sulfated fucose and the positive effect of the 2-sulfated galactose on anticoagulation in vitro, our results also demonstrate the importance of this set of structural requirements on antithrombosis in vivo, and further support the involvement of high-molecular weight and 4-sulfated fucose in both activities.

The oceans harbor a great diversity of organisms, and have been recognized as an important source of new compounds with nutritional and therapeutic potential. Among these compounds, carbohydrate-based compounds are of particular interest because they exhibit numerous biological functions associated with their chemical diversity. This gives rise to new substances for the development of bioactive products. Many are the known applications of substances with glycosidic domains obtained from marine species. This review covers the structural properties and the current findings on the antioxidant, anti-inflammatory, anticoagulant, antitumor and antimicrobial activities of medium and high molecular-weight carbohydrates or glycosylated compounds extracted from various marine organisms.

Glycosaminoglycans (GAGs) are sulfated glycans capable of regulating various biological and medical functions. Heparin, heparan sulfate, chondroitin sulfate, dermatan sulfate, keratan sulfate and hyaluronan are the principal classes of GAGs found in animals. Although GAGs are all composed of disaccharide repeating building blocks, the sulfation patterns and the composing alternating monosaccharides vary among classes. Interestingly, GAGs from marine organisms can present structures clearly distinct from terrestrial animals even considering the same class of GAG. The holothurian fucosylated chondroitin sulfate, the dermatan sulfates with distinct sulfation patterns extracted from ascidian species, the sulfated glucuronic acid-containing heparan sulfate isolated from the gastropode Nodipecten nodosum , and the hybrid heparin/heparan sulfate molecule obtained from the shrimp Litopenaeus vannamei are some typical examples. Besides being a rich source of structurally unique GAGs, the sea is also a wealthy environment of GAG-resembling sulfated glycans. Examples of these mimetics are the sulfated fucans and sulfated galactans found in brown, red and green algae, sea urchins and sea cucumbers. For adequate visualization, representations of all discussed molecules are given in both Haworth projections and 3D models., Competing Interests: The authors declare no conflict of interest.

Background and Aims: Between 10 and 20% of all patients undergoing bariatric surgery procedures regain weight secondary to a gastrojejunostomy enlargement. The aim of this study was to validate the interobserver agreement while measuring gastric outlet diameters using a new standard guidewire., Methods: We selected thirty-five videos of consecutive endoscopic procedures on patients undergoing esophagogastroduodenoscopy after a Roux-en-Y gastric bypass procedure. All videos were evaluated by four raters: two expert endoscopists and two trainees. We excluded videos having a slipped Fobi ring or a strictured gastric outlet. Anastomosis diameter was measured using a novel device with standardized markings on a guidewire (Hydra jagwire, Boston Scientific, Natick. MA) as well as the current gold standard defined as a calibrated endoscopic measuring instrument (Olympus America, Center Valley, PA)., Results: We obtained 272 measurements of the gastric outlet. Overall agreement measured through intra-class correlation coefficients for the gold standard was 0.84 (p < 0.01) and 0.83 (p < 0.01) for the new guidewire. Agreement among experts was 0.699 (p < 0.01), while among trainees it was 0.822 (p < 0.01)., Conclusion: The new guidewire demonstrated a high degree of observer reliability, also presenting similar results between expert endoscopists and trainees.

Schinus terebinthifolius Raddi is a perennial native from Atlantic forest. It is of high ecological plasticity and is used in traditional medicine. Based on promising reports concerning its bioactivity, it was included as a species of great interest for distribution through the National Health System. A number of agronomic studies to guide its crop production are therefore underway. This study examined diversity and phylogenetic relationships among native S. terebinthifolius populations from different Brazilian ecosystems: Cerrado; sandbanks; dense rainforest; and deciduous forest. The intergenic regions rpl20-5'rps12, trnH-psbA, and trnS-trnG were sequenced from cpDNA and aligned using BLASTn. There were few fragments for comparison in GenBank and so only region trnS-trnG was informative. There were variations among and within populations with intravarietal polymorphisms and three distinct haplotypes (HpSM, HpDDO, HpNE), once populations from NE (sandbanks and rainforest) clustered together. Sequences from HpSM, HpNE, and HpDDO returned greater similarity to haplotypes A (AY928398.1), B (AY928399.1), and C (AY928400.1), respectively. A network, built by median-joining among native haplotypes and 10 available on GenBank, revealed HpSM as the origin of all other haplogroups. HpDDO showed the most mutations and was closely related to haplogroups from Argentina. While this could indicate hybridization, we believe that the polymorphisms resulted from adaptation to events such as deforestation, fire, rising temperature, and seasonal drought during the transition from Atlantic forest to Cerrado. While more detailed phylogeographical studies are needed, these results indicate eligible groups for distinct climates as an important step for pre-breeding programs before field propagation.

Background: The high-sensitivity C-reactive protein (hs-CRP) seems to be associated with the presence of atherosclerosis. The association between menopause-related changes in endogenous sex hormones and hs-CRP levels remains controversial., Objective: To compare the levels of hs-CRP in pre- and postmenopausal women and to evaluate the association of endogenous sex hormones, waist circumference (WC) and insulin resistance with the levels of hs-CRP., Methods: This cross-sectional study included 145 women (age range: 45-65 years), 56 premenopausal and 89 postmenopausal. Patients were evaluated for hormonal and lipid profile, HOMA-IR, hs-CRP and WC. Each variable was first assessed for correlation with log hs-CRP using a univariate model, and significant variables were then added to the multivariate regression model. A p value <0.05 was considered statistically significant., Results: The hs-CRP is higher in postmenopausal patients (3.6 ± 4.9 versus 2.6 ± 3.7, p = 0.004). The log hs-CRP was positively correlated with WC (r = 0.13, p = 0.005), HOMA-IR (r = 0.29, p = 0.001), and triglycerides (r = 0.50, p = 0.01). The menopausal status (p = 0.02) and WC (p = 0.00003) behaved as independent predictors of hs-CRP levels. No correlation was found between hs-CRP and the time since menopause (r = -0.1, p = 0.58)., Conclusions: The hs-CRP is higher in postmenopausal patients. The menopausal status and WC were independently associated with hs-CRP levels in this sample of pre and postmenopausal women.

This study evaluated the antimicrobial effect of 3β,6β,16β-trihydroxylup-20(29)-ene (CLF1), a triterpene isolated from Combretum leprosum Mart., in inhibiting the planktonic growth and biofilms of Gram positive bacteria Streptococcus mutans and S. mitis. The antimicrobial activity was assessed by determining the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC). The antibiofilm potential was determined by quantifying total biomass and enumerating biofilm-entrapped viable bacteria. In addition, the acute toxicity of CLF1 on Artemia sp. nauplii was also determined. The results showed that CLF1 was able in inhibiting the growth of S. mutans and S. mitis with MIC and MBC of 7.8 μg/mL and 15.6 μg/mL, respectively. CLF1 was highly effective on biofilms of both bacteria. Only 7.8 μg/mL CLF1 was enough to inhibit by 97% and 90% biomass production of S. mutans and S. mitis, respectively. On the other hand, such effects were not evident on Gram negative Pseudomonas aeruginosa and Klebsiella oxytoca. The toxicity tests showed that the LC50 of CLF1 was 98.19 μg/mL. Therefore, CLF1 isolated from C. leprosum may constitute an important natural agent for the development of new therapies for caries and other infectious diseases caused by S. mutans and S. mitis.

Objective: To analyze training and knowledge of professionals in the Family Health Team on reporting the mistreatment of children and adolescents., Methods: Cross-sectional study carried out in three municipalities of Ceará State, Northeast Brazil, from January to April 2012. The research included 51 professionals: physicians (9), nurses (26), and dentists (16) who worked in the Family Health Strategy. A questionnaire was used for data collection, which received descriptive statistical analysis with the Pearson's chi-square test, being significant p≤0.05., Results: There was a predominance of professionals who had not participated in violence against children and adolescents training (86.3%); who knew the Child and Adolescent Statute (90.2%), and how to notify mistreatment (62.7%). Most interviewees said that the health unit had the form (70.5%), and they knew where to refer victims to (82.3%). Most professionals did not have any contact with mistreatment situations (62.8%). Only 37.2% had already identified some case and, among them, 60.0% reported the occurrences. There was a significant association (p=0.035) between the act of notifying and the participation in a training on the subject., Conclusions: This study showed that the participants have difficulties in the reporting mistreatment of children and adolescents, and there are gaps in knowledge and weaknesses in training in this area.

Objectives: The aim of this in vitro study was to investigate the efficacy of in-office bleaching technique combined with the application of a casein phosphopeptide-amorphous calcium phosphate (CPP-ACP) paste (MI Paste-MI) at different moments and its influence on enamel surface properties., Methods: Eighty bovine dental crowns were randomly allocated into eight groups (n = 10), and bleached with either 35% hydrogen peroxide (HP) or 37% carbamide peroxide (CP). Four different protocols of application of MI were considered: without MI, MI applied before bleaching, MI applied after bleaching, and MI applied both before and after bleaching. Bleaching effectiveness was measured by the VITA EasyShade spectrophotometer utilizing the CIEL*a*b* system (ΔE, ΔL*, Δa*, and Δb*). Color readings were measured at baseline, 7, 14, and 21 days. Hardness and roughness were measured at baseline (T0) and immediately after bleaching (T14). Data were subjected to the two-way ANOVA for repeated measurements and Tukey's test at 5%., Results: HP groups achieved the greatest color change. The application of a CPP-ACP paste did not reduce the efficacy of bleaching peroxides. Samples bleached with CP showed decreased hardness at T14. Samples bleached with HP that received the application of MI before and before/after bleaching did not present hardness decrease at T14. Samples bleached with peroxides only and received MI after bleaching showed increased roughness at T14., Conclusions: The use of CPP-ACP was able to prevent negative changes in roughness and hardness of bovine enamel when associated to hydrogen peroxide, and might be applied before/after the bleaching protocol., (Copyright © 2012 Wiley Periodicals, Inc.)

Background: This study evaluated the impact of bleaching teeth using blends of a CPP-ACP paste (MI Paste; MI) and carbamide/hydrogen peroxides in different proportions on surface properties of bleached enamel., Methods: Ninety bovine incisors were bleached with 7.5% hydrogen peroxide (HP), 16% carbamide peroxide (CP), MI and blends of HP or CP:MI at three proportions (1:1, 2:1, 1:2). Hardness and roughness were measured at baseline and after bleaching. Enamel morphology was evaluated by Scanning Electron Microscopy (SEM). The data were analyzed by two-way ANOVA for repeated measurements and Tukey's test., Results: Most of the samples bleached with MI in combination with peroxides presented increased hardness and roughness which were associated to mineral deposition, as observed by SEM images. Blends with higher fractions of MI did not offer superior benefits., Conclusions: The use of a CPP-ACP paste mixed to carbamide/hydrogen peroxides can decrease adverse side-effects from tooth bleaching on an enamel surface.

Background: Different enhanced peroxide formulations have been developed to overcome mineral loss during tooth whitening. This study investigated the impact on tooth whitening from combining a CPP-ACP paste with home peroxide agents at several proportions., Methods: Ninety bovine incisors were randomly allocated into nine groups (n = 10) and exposed to 7.5% hydrogen peroxide (HP), 16% carbamide peroxide (CP), a CPP-ACP paste (Tooth Mousse/MI Paste, GC Corporation, Tokyo, Japan) and blends of HP/CP:MI at three proportions (1:1, 2:1 and 1:2). Tooth whitening was performed for 14 days and its effectiveness was measured by a spectrophotometer utilizing the CIE L*a*b* system (ΔE, ΔL*, Δa* and Δb*). Colour readings were measured at baseline (T0), 7 days of bleaching (T7), 14 days of bleaching (T14) and 7 days after the end of bleaching (T21). Data were analysed by two-way ANOVA for paired averages followed by the Tukey's test and Pearson's correlation at 5%., Results: Although CP produced the greatest colour change (ΔE), all whitening protocols were considered to be effective regardless of the CPP-ACP presence. In general, greater changes in ΔE, ΔL*, Δa* and Δb* were detected at T14, with T21 exhibiting similar results to those at T7. A greater number of strong correlations with ΔE values was found for a* (11/15)., Conclusions: Within the limitations of this study, the results indicate that the CPP-ACP paste did not affect tooth whitening efficacy., (© 2012 Australian Dental Association.)

Currently, there has been an increased frequency of fungal infections. Candida albicans and other Candida spp. have been proven to be major causes for urinary tract infection. Increased resistance to antifungals indicates the need to develop strategies in order to prevent the spread of resistance. Chromogenic medium have been proven to be useful in the detection of yeasts in clinical specimens containing mixed cultures of Candida. The aim of this study was to compare the results of antifungal susceptibility testing with fluconazole and amphotericin B on strains of Candida spp. isolated from urine, conducted on a Mueller-Hinton Agar with Glucose and Methylene Blue (MHAGMB) medium and on a Hicrome Candida® Agar with 2% Glucose (HCAG) medium. We used 40 samples of Candida spp. isolated from urine samples from inpatients and outpatients. The results showed that both media presented high rates of agreement, above 94%. The use of the HCAG medium decreases the release time of the results by 24-48 h, which may be decisive for initiating the correct drug treatment.

We compared the effect of the number of weekly repetitions of a static stretching program on the flexibility, hamstring tightness and electromyographic activity of the hamstring and of the triceps surae muscles. Thirty-one healthy subjects with hamstring tightness, defined as the inability to perform total knee extension, and shortened triceps surae, defined by a tibiotarsal angle wider than 90 degrees during trunk flexion, were divided into three groups: G1 performed the stretching exercises once a week; G2, three times a week, and G3, five times a week. The parameters were determined before and after the stretching program. Flexibility improved in all groups after intervention, from 7.65 +/- 10.38 to 3.67 +/- 12.08 in G1, from 10.73 +/- 12.07 to 0.77 +/- 10.45 in G2, and from 14.20 +/- 10.75 to 6.85 +/- 12.19 cm in G3 (P < 0.05 for all comparisons). The increase in flexibility was higher in G2 than in G1 (P = 0.018), while G2 and G3 showed no significant difference (G1: 4 +/- 2.17, G2: 10 +/- 5.27; G3: 7.5 +/- 4.77 cm). Hamstring tightness improved in all groups, from 37.90 +/- 6.44 to 29 +/- 11.65 in G1, from 39.82 +/- 9.63 to 21.91 +/- 8.40 in G2, and from 37.20 +/- 6.63 to 26.10 +/- 5.72 degrees in G3 (P < 0.05 for all comparisons). During stretching, a statistically significant difference was observed in electromyographic activity of biceps femoris muscle between G1 and G3 (P = 0.048) and G2 and G3 (P = 0.0009). No significant differences were found in electromyographic activity during maximal isometric contraction. Stretching exercises performed three times a week were sufficient to improve flexibility and range of motion compared to subjects exercising once a week, with results similar to those of subjects who exercised five times a week.

Purpose: To evaluate both the efficacy and reliability of the cytologic method in the diagnosis of conjunctival lesions and compare it with the histologic examination., Methods: A prospective study on 31 conjunctival lesions was performed. Samples were obtained with a Kimura spatula. The lesions were, then, surgically removed and sent for histologic examination. The Papanicolaou technique was used for the cytologic examination., Results: The cytologic method showed high specificity (100%) and sensitivity (92.9%). The positive predictive value was 100% and the negative was 94.4%., Conclusions: The simple cytologic method is effective and reliable for the diagnosis of tumoral lesions of the conjunctiva.

A case of lichen planus verrucosus of the legs in a 29 year old patient is presented by the authors. 18 years after the outset a squamous cell carcinoma giving rise to lymph node metastasis appeared on a lesion localized on the external side of the right leg. The chronic inflammation and the traumatism are the only carcinogenetic factors to be considered. In this case there were no antecedents of arsenic administration. The therapy with bleomicin and R.X. was effective. The review of the literature shows the extreme rarity of the malignant transformation of the lichen planus verrucose.