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O valido foi uma personagem polêmica durante o século XVII no âmbito da cultura política do Antigo Regime. Este artigo pretende oferecer uma análise de como o valido foi tratado dentro do gênero de dramaturgia denominado Comedia de la Privanza, atendo-se à comédia ¿Cómo ha de ser el privado?, de Francisco de Quevedo, que coloca questões relevantes acerca do fenômeno do valimento, bem como da controversa relação de Quevedo com o regime do conde-duque de Olivares, valido de Felipe IV

Background: Diagnosing and predicting neonatal sepsis is challenging because of its nonspecific symptoms, lack of diagnostic criteria consensus, and absence of early, sensitive, and specific diagnostic laboratory tests., Purpose: To evaluate the diagnostic and prognostic potential of adrenomedullin (ADM), interleukin-6 (IL-6), and C-reactive protein (CRP) in late-onset neonatal sepsis (LOS)., Methods: We studied 53 neonates with culture-proven LOS by sampling at admission and on antibiotic treatment days 3 and 7. These data were compared with those of 22 healthy full-term controls sampled on day 3 before hospital discharge. Survivors and non-survivors in the sepsis group were analyzed separately., Results: Coagulase-negative Staphylococcus was the most commonly detected pathogen. ADM (cutoff, 0.5 ng/mL) and CRP (cutoff, <5 mg/L) values aligned with manufacturer recommendations, while IL-6 levels (cutoff, 10 pg/mL) were higher than expected, likely due to labor stress. The median biomarker levels significantly distinguished neonates with sepsis from controls (p < 0.0001) at all time points with ADM and IL-6 levels elevated at admission, indicating their potential as early diagnostic markers. CRP level was diagnostically useful starting on day 3. Prognostically, IL-6 (p < 0.001) and ADM (p < 0.05) differentiated survivors from non-survivors; however, only IL-6 consistently predicted mortality at all time points (area under the curve [AUC] > 0.90). ADM and CRP levels showed poor prognostic value (AUC < 0.70). ADM and IL-6 demonstrated strong diagnostic utility in early LOS, whereas CRP became relevant later. IL-6 was the only reliable biomarker for predicting mortality, supporting its integration into clinical protocols. Combining IL-6 with CRP may enhance early detection and management, potentially improving neonatal outcomes., Conclusion: IL-6 is a robust biomarker for the early diagnosis and prognosis of LOS. Incorporating IL-6 into clinical practice with CRP could improve early neonatal LOS diagnosis and patient outcomes.

The continuous increase in bacterial resistance requires the study and development of new biocompatible antibacterial materials. Galactomannan gum was extracted from endosperms of Dimorphandra gardneriana seeds, modified through quaternization reactions (with NaOH/CHPTAC), and evaluated for antimicrobial activity. Four quaternized derivatives were obtained (QG1-QG4). The derivatives were characterized by FTIR, NMR, and rheology, which confirmed the cationization. The galactomannan structure was theoretically studied, and Zeta potential exhibited positive increase in the charge of quaternized derivatives, changing from -3.2 mV (unmodified galactomannan) to +59.35 mV (QG4). Elemental analysis revealed an increase in degree of substitution (DS) between the derivatives, of which QG4 exhibited the highest DS (0.51). QG4 exhibited a bactericidal effect in the antimicrobial test for determination of minimal inhibitory and bactericidal concentrations, against Gram-positive bacteria. Atomic force microscopy showed an increase in bacterial cell surface roughness and decrease in cell height after treatment with QG4. QG3 and QG4 demonstrated antibiofilm potential, showing 50 % and 40 % of inhibition, respectively. After MTT assay, cell viability was maintained approximately 100 % for all derivatives at concentrations of 20 mg/mL on RAW 264.7 macrophages and 86 % with 250 μg/mL on L929 cells. Galactomannan was successfully modified, demonstrating strong antibacterial activity, concomitant to the increase in DS, Zeta potential and reduction of molar mass, with good biocompatibility. Therefore, this study presents a new material with potential for development of formulations for biomedical applications, from industrial waste, strengthening of the D. gardneriana production chain., 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 © 2024. Published by Elsevier B.V.)

Rare B cells can have special pathogen-recognition features giving them the potential to make outsized contributions to protective immunity. However, rare naive B cells infrequently participate in immune responses. We investigated how germline-targeting vaccine antigen delivery and adjuvant selection affect priming of exceptionally rare BG18-like HIV broadly neutralizing antibody-precursor B cells (~1 in 50 million) in non-human primates. Only escalating dose (ED) priming immunization using the saponin adjuvant SMNP elicited detectable BG18-like cells in germinal centers (GCs). All groups had strong GC responses, but only ED+SMNP and bolus+SMNP induced BG18-like memory B cells in >50% of animals. One group had vaccine-specific GC responses equivalent to ED+SMNP, but BG18-like memory B cells were rarely detected. Following homologous boosting, BG18-like memory B cells were more frequent in a bolus priming group, but had lower somatic hypermutation and affinities. This outcome was inversely associated with post-prime antibody titers, suggesting antibody feedback can significantly influence rare precursor B cell responses.

Prolonging exposure to subunit vaccines during the primary immune response enhances humoral immunity. Escalating-dose immunization (EDI), administering vaccines every other day in an increasing pattern over 2 weeks, is particularly effective but challenging to implement clinically. Here, using an HIV Env trimer/saponin adjuvant vaccine, we explored simplified EDI regimens and found that a two-shot regimen administering 20% of the vaccine followed by the remaining 80% of the dose 7 days later increased T FH responses 6-fold, antigen-specific germinal center (GC) B cells 10-fold, and serum antibody titers 10-fold compared with bolus immunization. Computational modeling of T FH priming and the GC response suggested that enhanced activation/antigen loading on dendritic cells and increased capture of antigen delivered in the second dose by follicular dendritic cells contribute to these effects, predictions we verified experimentally. These results suggest that a two-shot priming approach can be used to substantially enhance responses to subunit vaccines.

There is growing interest in using cannabinoids across various clinical scenarios, including pain medicine, leading to the disregard of regulatory protocols in some countries. Legislation has been implemented in Brazil, specifically in the state of São Paulo, permitting the distribution of cannabinoid products by health authorities for clinical purposes, free of charge for patients, upon professional prescription. Thus, it is imperative to assess the existing evidence regarding the efficacy and safety of these products in pain management. In light of this, the São Paulo State Society of Anesthesiology (SAESP) established a task force to conduct a narrative review on the topic using the Delphi method, requiring a minimum agreement of 60% among panelists. The study concluded that cannabinoid products could potentially serve as adjuncts in pain management but stressed the importance of judicious prescription. Nevertheless, this review advises against their use for acute pain and cancer-related pain. In other clinical scenarios, established treatments should take precedence, particularly when clinical protocols are available, such as in neuropathic pain. Only patients exhibiting poor therapeutic responses to established protocols or demonstrating intolerance to recommended management may be considered as potential candidates for cannabinoids, which should be prescribed by physicians experienced in handling these substances. Special attention should be given to individual patient characteristics and the likelihood of drug interactions., Competing Interests: Conflicts of Interest The authors declare no have conflicts of interest., (Copyright © 2024 Sociedade Brasileira de Anestesiologia. Published by Elsevier España S.L.U. All rights reserved.)

Epstein-Barr virus (EBV) is associated with infectious mononucleosis, cancer, and multiple sclerosis. A vaccine that prevents infection and/or EBV-associated morbidity is an unmet need. The viral gH/gL glycoprotein complex is essential for infectivity, making it an attractive vaccine target. Here, we evaluate the immunogenicity of a gH/gL nanoparticle vaccine adjuvanted with the Sigma Adjuvant System (SAS) or a saponin/monophosphoryl lipid A nanoparticle (SMNP) in rhesus macaques. Formulation with SMNP elicits higher titers of neutralizing antibodies and more vaccine-specific CD4 + T cells. All but one animal in the SMNP group were infected after oral challenge with the EBV ortholog rhesus lymphocryptovirus (rhLCV). Their immune plasma had a 10- to 100-fold lower reactivity against rhLCV gH/gL compared to EBV gH/gL. Anti-EBV neutralizing monoclonal antibodies showed reduced binding to rhLCV gH/gL, demonstrating that EBV gH/gL neutralizing epitopes are poorly conserved on rhLCV gH/gL. Prevention of rhLCV infection despite antigenic disparity supports clinical development of gH/gL nanoparticle vaccines against EBV., Competing Interests: Declaration of interests A.T.M. holds a patent (US11116835B2) on the AMMO1 monoclonal antibody. D.J.I. is an inventor on a patent related to SMNP adjuvant (US11,547,672)., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

"Extended priming" immunization regimens that prolong exposure of the immune system to vaccines during the primary immune response have shown promise in enhancing humoral immune responses to a variety of subunit vaccines in preclinical models. We previously showed that escalating-dosing immunization (EDI), where a vaccine is dosed every other day in an increasing pattern over 2 weeks dramatically amplifies humoral immune responses. But such a dosing regimen is impractical for prophylactic vaccines. We hypothesized that simpler dosing regimens might replicate key elements of the immune response triggered by EDI. Here we explored "reduced ED" immunization regimens, assessing the impact of varying the number of injections, dose levels, and dosing intervals during EDI. Using a stabilized HIV Env trimer as a model antigen combined with a potent saponin adjuvant, we found that a two-shot extended-prime regimen consisting of immunization with 20% of a given vaccine dose followed by a second shot with the remaining 80% of the dose 7 days later resulted in increased total GC B cells, 5-10-fold increased frequencies of antigen-specific GC B cells, and 10-fold increases in serum antibody titers compared to single bolus immunization. Computational modeling of the GC response suggested that this enhanced response is mediated by antigen delivered in the second dose being captured more efficiently as immune complexes in follicles, predictions we verified experimentally. Our computational and experimental results also highlight how properly designed reduced ED protocols enhance activation and antigen loading of dendritic cells and activation of T helper cells to amplify humoral responses. These results suggest that a two-shot priming approach can be used to substantially enhance responses to subunit vaccines., Competing Interests: Competing interests S.H.B, D.J.I, A.K.C, and L.Y are inventors on a patent filing related to the extended-priming regimens described in this manuscript. For completeness, it is also noted that A.K.C. is a consultant (titled “Academic Partner”) for Flagship Pioneering, consultant and Strategic Oversight Board Member of its affiliated company, Apriori Bio, and is a consultant and Scientific Advisory Board Member of another affiliated company, Metaphore Bio.

Adjuvants and antigen delivery kinetics can profoundly influence B cell responses and should be critically considered in rational vaccine design, particularly for difficult neutralizing antibody targets such as human immunodeficiency virus (HIV). Antigen kinetics can change depending on the delivery method. To promote extended immunogen bioavailability and to present antigen in a multivalent form, native-HIV Env trimers are modified with short phosphoserine peptide linkers that promote tight binding to aluminum hydroxide (pSer:alum). Here we explore the use of a combined adjuvant approach that incorporates pSer:alum-mediated antigen delivery with potent adjuvants (SMNP, 3M-052) in an extensive head-to-head comparison study with conventional alum to assess germinal center (GC) and humoral immune responses. Priming with pSer:alum plus SMNP induces additive effects that enhance the magnitude and persistence of GCs, which correlate with better GC-T FH cell help. Autologous HIV-neutralizing antibody titers are improved in SMNP-immunized animals after two immunizations. Over 9 months after priming immunization of pSer:alum with either SMNP or 3M-052, robust Env-specific bone marrow plasma cells (BM B PC ) are observed. Furthermore, pSer-modification of Env trimer reduce targeting towards immunodominant non-neutralizing epitopes. The study shows that a combined adjuvant approach can augment humoral immunity by modulating immunodominance and shows promise for clinical translation., (© 2023. The Author(s).)

This study aimed to detect and differentiate Toxoplasma gondii by the allele typing of its polymorphic rop18 gene. For this purpose, a novel genotyping system using allele-specific oligonucleotides (ASOs) was designed, consisting of three ASO pairs. The first and third pairs specifically amplify rop18 allele I and allele III, while the second pair amplify both allele I and II. Genomic DNA from 86 congenital infections was analyzed by ASO-PCRs, successfully typing 82 (95.35%) samples. The remaining 4 samples (4.65%) required sequencing and single nucleotide polymorphism (SNP) analysis of the amplification products. The distribution of samples according to rop18 alleles was: 39.5% of allele III, 38.4% of allele II, 19.8% of mixed rop18 alleles (I/III or II/III), and 2.3% of allele I. The six severely compromised infants exhibited the highest parasite load levels and were infected during the first and early second trimesters of pregnancy. Among these cases, two were associated with rop18 allele I parasites, two with mixed rop18 alleles (I/III), one with allele II, and one with allele III parasites. In conclusion, all severe cases of congenital toxoplasmosis were infected during early pregnancy, but they were not exclusively associated with rop18 allele I parasites, as observed in murine toxoplasmosis. Furthermore, nearly one-fifth of parasites were non-archetypal, exhibiting more than one rop18 allele, indicating a higher genetic diversity of Toxoplasma gondii in this South American sample. Overall, a robust T. gondii rop18 allele typing was developed and suggested that congenital toxoplasmosis in humans involves complex mechanisms beyond the parasite genotype., 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 manuscript., (Copyright © 2023 Elsevier B.V. All rights reserved.)

In the ongoing effort to develop a vaccine against HIV, vaccine approaches that promote strong germinal center (GC) responses may be critical to enable the selection and affinity maturation of rare B cell clones capable of evolving to produce broadly neutralizing antibodies. We previously demonstrated an approach for enhancing GC responses and overall humoral immunity elicited by alum-adjuvanted protein immunization via the use of phosphoserine (pSer) peptide-tagged immunogens that stably anchor to alum particles via ligand exchange with the alum particle surface. Here, using a clinically relevant stabilized HIV Env trimer termed MD39, we systematically evaluated the impact of several parameters relevant to pSer tag composition and trimer immunogen design to optimize this approach, including phosphate valency, amino acid sequence of the trimer C-terminus used for pSer tag conjugation, and structure of the pSer tag. We also tested the impact of co-administering a potent saponin/monophosphoryl lipid A (MPLA) nanoparticle co-adjuvant with alum-bound trimers. We identified MD39 trimer sequences bearing an optimized positively-charged C-terminal amino acid sequence, which, when conjugated to a pSer tag with four phosphates and a polypeptide spacer, bound very tightly to alum particles while retaining a native Env-like antigenicity profile. This optimized pSer-trimer design elicited robust antigen-specific GC B cell and serum IgG responses in mice. Through this optimization, we present a favorable MD39-pSer immunogen construct for clinical translation., (© 2023. Springer Nature Limited.)

Immune stimulating complexes (ISCOMs) are safe and effective saponin-based adjuvants formed by the self-assembly of saponin, cholesterol, and phospholipids in water to form cage-like 30-40 nm diameter particles. Inclusion of the Toll-like receptor 4 agonist monophosphoryl lipid A (MPLA) in ISCOM particles yields a promising next-generation adjuvant termed Saponin-MPLA NanoParticles (SMNP). In this work, we detail protocols to produce ISCOMs or SMNP via a tangential flow filtration (TFF) process suitable for scalable synthesis and Good Manufacturing Practice (GMP) production of clinical-grade adjuvants. SMNP or ISCOM components were solubilized in micelles of the surfactant MEGA-10, then diluted below the critical micelle concentration (CMC) of the surfactant to drive ISCOM self-assembly. Assembly of ISCOM/SMNP particles using the purified saponin QS-21 used in clinical-grade saponin adjuvants was found to require controlled stepwise dilution of the initial micellar solution, to prevent formation of undesirable kinetically-trapped aggregate species. An optimized protocol gave yields of ~77% based on the initial feed of QS-21 and the final SMNP particle composition mirrored the feed ratios of the components. Further, samples were highly homogeneous with comparable quality to that of material prepared at lab scale by dialysis and purified via size-exclusion chromatography. This protocol may be useful for clinical preparation of ISCOM-based vaccine adjuvants and therapeutics.

Bioprospecting and synthesis of strategically designed molecules have been used in the search for drugs that can be in leishmaniasis. Hydrazones (HDZ) are promising compounds with extensive biological activities. The objective of this work was to perform in silico studies of hydrazones 1-5 and to evaluate their antileishmanial, cytotoxic and macrophage immunomodulatory potential in vitro. Hydrazones were subjected to prediction and molecular docking studies. Antileishmanial protocols on promastigotes and amastigotes of Leishmania amazonensis, cytotoxicity and macrophage immunomodulatory activity were performed. Hydrazones showed a good pharmacokinetic profile and hydrazone 3 and hydrazone 5 were classified as non-carcinogenic. Hydrazone 5 obtained the best conformation with trypanothione reductase. Hydrazone 1 and hydrazone 3 obtained the best mean inhibitory concentration (IC 50 ) values for promastigotes, 4.4-61.96 μM and 8.0-58.75 μM, respectively. It also showed good activity on intramacrophagic amastigotes, with hydrazone 1 being the most active (IC 50  = 6.79 μM) with selectivity index of 56. In cytotoxicity to macrophages hydrazone 3 was the most cytotoxic (CC 50  = 256.3 ± 0,04 μM), while hydrazone 4 the least (CC 50  = 1055.9 ± 0.03 μM). It can be concluded that the hydrazones revealed important pharmacokinetic and toxicological properties, in addition to antileishmania potential in reducing infection and infectivity in parasitized macrophages., Competing Interests: Declaration of Competing Interest The authors declare that they have no conflict of interest., (Copyright © 2023. Published by Elsevier Ltd.)

Objectives: To assess the prevalence of burnout, anxiety and depression symptoms, and posttraumatic stress disorder (PTSD) in PICU workers in Brazil during the first peak of the COVID-19 pandemic. To compare the results of subgroups stratified by age, gender, professional category, health system, and previous mental health disorders., Design: Multicenter, cross-sectional study using an electronic survey., Setting: Twenty-nine public and private Brazilian PICUs., Subjects: Multidisciplinary PICU workers., Interventions: None., Measurements and Main Results: Self-reported questionnaires were used to measure burnout (Maslach Burnout Inventory), anxiety and depression (Hospital Anxiety and Depression Scale), and PTSD (Posttraumatic Stress Disorder Checklist for Diagnostic and Statistical Manual of Mental Disorders, Fifth Edition [PCL-5]) in 1,084 respondents. Subjects were mainly young (37.1 ± 8.4 yr old) and females (85%), with a median workload of 50 hours per week. The prevalence of anxiety and depression was 33% and 19%, respectively, whereas PTSD was 13%. The overall median burnout scores were high in the emotional exhaustion and personal accomplishment dimensions (16 [interquartile range (IQR), 8-24] and 40 [IQR, 33-44], respectively) whereas low in the depersonalization one (2 [IQR, 0-5]), suggesting a profile of overextended professionals, with a burnout prevalence of 24%. Professionals reporting prior mental health disorders had higher prevalence of burnout (30% vs 22%; p = 0.02), anxiety (51% vs 29%; p < 0.001), and depression symptoms (32.5% vs 15%; p < 0.001), with superior PCL-5 scores for PTSD ( p < 0.001). Public hospital workers presented more burnout (29% vs 18.6%, p < 0.001) and more PTSD levels (14.8% vs 10%, p = 0.03). Younger professionals were also more burned out ( p < 0.05 in all three dimensions)., Conclusions: The prevalence of mental health disorders in Brazilian PICU workers during the first 2020 peak of COVID-19 was as high as those described in adult ICU workers. Some subgroups, particularly those reporting previous mental disorders and younger professionals, should receive special attention to prevent future crises., Competing Interests: The authors have disclosed that they do not have any potential conflicts of interest., (Copyright © 2022 by the Society of Critical Care Medicine and the World Federation of Pediatric Intensive and Critical Care Societies.)

Background: To translate and cross-culturally adapt into Brazilian-Portuguese, and to test the measurement properties of the following items of implementation outcome measures: Acceptability of Intervention Measure (AIM), Intervention Appropriateness Measure (IAM) and Feasibility of Intervention Measure (FIM)., Methods: This was a measurement properties study in accordance with the Consensus-based Standards for the selection of health status Measurement Instruments (COSMIN). We conducted a translation and cross-cultural adaptation of three implementation measures according to guidelines for translation and cross-cultural adaptation, then we collected information from patients who had participated in remotely delivered physical therapy treatment for musculoskeletal condition. The patients answered the translated versions of the implementation outcome measures. The measurement properties of the three implementation outcome measures were collected in a test-retest assessment, with an interval of 7 to 14 days.. The measurement properties evaluated in this study were interpretability, measured using Ceiling and Floor Effects, reliability in test-retest evaluation, measured using Cronbach's Alpha Coefficient, internal consistency, measured using Intraclass Correlation Coefficient and construct validity, measured using Pearson Correlation., Results: We included 104 participants (76 female). The average age of the sample was 56.8 (SD 14.8) years old. The items of implementation outcome measures (AIM, IAM, and FIM) showed 66.39%, 63.11%, and 63.93% of ceiling effects. The items of implementation outcome measures showed adequate internal consistency measured using Cronbach's Alpha Coefficient (AIM: 0.89, IAM: 0.91, FIM: 0.93) and values of Standard Error of Measurement between 5 and 10%, showing good measurement error. The results of AIM and IAM was classified as moderate reliability and the FIM as substantial reliability. In a total 96 correlations, > 75% of correlations met our prior hypothesis., Conclusion: The three Brazilian-Portuguese versions of items of implementation outcome measures had adequate internal consistency, measurement error and construct validity. The three implementation outcome measures showed moderate to substantial reliability values. The Ceiling Effect was observed in the three measures, showing maximum values ​​in more than 15% of the evaluations., (© 2023. The Author(s).)

Amniotic fluid DNA samples were genotyped by multilocus-nested-PCR-RFLP, but only three of 11 markers amplified 113 of 122 (92.6%) samples, resulting in 12 untyped and 101 partial non-archetypal genotypes. The 101 typed samples were subdivided into four groups: G1 with 73 samples (5'and 3' SAG2 allele I + SAG3 allele III + GRA6 allele III), 53 had parasite load ≤ 102 parasites/mL (43 asymptomatic, 10 mild infections), 17 had load > 102 and ≤ 103 (one mild, 13 moderate and three severe), and three had load > 103 parasites/mL (three severe); G2 with 22 samples (5'and 3' SAG2 allele I + SAG3 allele III), all parasite load levels ≤ 102 parasites/mL (18 asymptomatic and four mild); G3 with five samples (5' and 3' SAG2 allele I + SAG3 allele II), parasite load ≤ 102 parasites/mL (three asymptomatic and two mild); G4 with one sample (5' and 3' SAG2 allele II + SAG3 allele II + GRA6 allele I), a parasite load < 102 parasites/mL in an asymptomatic infant. After DNA sequencing, restriction sites confirmed SAG2, SAG3 and GRA6 alleles in 98.7%, 100% and 100% of the cases, respectively, while single nucleotide polymorphisms confirmed 90% of 5'-SAG2 allele I; 98.7% of 3'-SAG2 allele I; 98% of SAG-3 allele III, but only 40% of GRA6 allele III results. For the moment, partial non-archetypal genotypes of parasites did not show any relationship with either parasite load in amniotic fluid samples or clinical outcome of infants at the age of 12 months.

The World Health Organization classifies Leishmania as one of the 17 “neglected diseases” that burden tropical and sub-tropical climate regions with over half a million diagnosed cases each year. Despite this, currently available anti-leishmania drugs have high toxicity and the potential to be made obsolete by parasite drug resistance. We chose to analyze organoselenides for leishmanicidal potential given the reduced toxicity inherent to selenium and the displayed biological activity of organoselenides against Leishmania. Thus, the biological activities of 77 selenoesters and their N-aryl-propanamide derivatives were predicted using robust in silico models of Leishmania infantum, Leishmania amazonensis, Leishmania major, and Leishmania (Viannia) braziliensis. The models identified 28 compounds with >60% probability of demonstrating leishmanicidal activity against L. infantum, and likewise, 26 for L. amazonesis, 25 for L. braziliensis, and 23 for L. major. The in silico prediction of ADMET properties suggests high rates of oral absorption and good bioavailability for these compounds. In the in silico toxicity evaluation, only seven compounds showed signs of toxicity in up to one or two parameters. The methodology was corroborated with the ensuing experimental validation, which evaluated the inhibition of the Promastigote form of the Leishmania species under study. The activity of the molecules was determined by the IC50 value (µM); IC50 values < 20 µM indicated better inhibition profiles. Sixteen compounds were synthesized and tested for their activity. Eight molecules presented IC50 values < 20 µM for at least one of the Leishmania species under study, with compound NC34 presenting the strongest parasite inhibition profile. Furthermore, the methodology used was effective, as many of the compounds with the highest probability of activity were confirmed by the in vitro tests performed.

Therapeutic strategies capable of inducing and enhancing prosthesis embodiment are a key point for better adaptation to and acceptance of prosthetic limbs. In this study, we developed a training protocol using an EMG-based human-machine interface (HMI) that was applied in the preprosthetic rehabilitation phase of people with amputation. This is a case series with the objective of evaluating the induction and enhancement of the embodiment of a virtual prosthesis. Six men and a woman with unilateral transfemoral traumatic amputation without previous use of prostheses participated in the study. Participants performed a training protocol with the EMG-based HMI, composed of six sessions held twice a week, each lasting 30 mins. This system consisted of myoelectric control of the movements of a virtual prosthesis immersed in a 3D virtual environment. Additionally, vibrotactile stimuli were provided on the participant's back corresponding to the movements performed. Embodiment was investigated from the following set of measurements: skin conductance response (affective measurement), crossmodal congruency effect (spatial perception measurement), ability to control the virtual prosthesis (motor measurement), and reports before and after the training. The increase in the skin conductance response in conditions where the virtual prosthesis was threatened, recalibration of the peripersonal space perception identified by the crossmodal congruency effect, ability to control the virtual prosthesis, and participant reports consistently showed the induction and enhancement of virtual prosthesis embodiment. Therefore, this protocol using EMG-based HMI was shown to be a viable option to achieve and enhance the embodiment of a virtual prosthetic limb., 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 © 2022 Rodrigues, Moreira, Pinheiro, Dantas, Santos, Nepomuceno, Nogueira, Cavalheiro and Faber.)

Chagas disease, a neglected tropical disease, is endemic in 21 Latin American countries and particularly prevalent in Brazil. Chagas disease has drawn more attention in recent years due to its expansion into non-endemic areas. The aim of this work was to computationally identify and experimentally validate the natural products from an Annonaceae family as antichagasic agents. Through the ligand-based virtual screening, we identified 57 molecules with potential activity against the epimastigote form of T. cruzi. Then, 16 molecules were analyzed in the in vitro study, of which, six molecules displayed previously unknown antiepimastigote activity. We also evaluated these six molecules for trypanocidal activity. We observed that all six molecules have potential activity against the amastigote form, but no molecules were active against the trypomastigote form. 13-Epicupressic acid seems to be the most promising, as it was predicted as an active compound in the in silico study against the amastigote form of T. cruzi, in addition to having in vitro activity against the epimastigote form., (© 2022 Wiley-VCH GmbH.)

To combat the HIV epidemic and emerging threats such as SARS-CoV-2, immunization strategies are needed that elicit protection at mucosal portals of pathogen entry. Immunization directly through airway surfaces is effective in driving mucosal immunity, but poor vaccine uptake across the mucus and epithelial lining is a limitation. The major blood protein albumin is constitutively transcytosed bidirectionally across the airway epithelium through interactions with neonatal Fc receptors (FcRn). Exploiting this biology, here, we demonstrate a strategy of "albumin hitchhiking" to promote mucosal immunity using an intranasal vaccine consisting of protein immunogens modified with an amphiphilic albumin-binding polymer-lipid tail, forming amph-proteins. Amph-proteins persisted in the nasal mucosa of mice and nonhuman primates and exhibited increased uptake into the tissue in an FcRn-dependent manner, leading to enhanced germinal center responses in nasal-associated lymphoid tissue. Intranasal immunization with amph-conjugated HIV Env gp120 or SARS-CoV-2 receptor binding domain (RBD) proteins elicited 100- to 1000-fold higher antigen-specific IgG and IgA titers in the serum, upper and lower respiratory mucosa, and distal genitourinary mucosae of mice compared to unmodified protein. Amph-RBD immunization induced high titers of SARS-CoV-2-neutralizing antibodies in serum, nasal washes, and bronchoalveolar lavage. Furthermore, intranasal amph-protein immunization in rhesus macaques elicited 10-fold higher antigen-specific IgG and IgA responses in the serum and nasal mucosa compared to unmodified protein, supporting the translational potential of this approach. These results suggest that using amph-protein vaccines to deliver antigen across mucosal epithelia is a promising strategy to promote mucosal immunity against HIV, SARS-CoV-2, and other infectious diseases.

This study assessed the technical performance of a rapid lateral flow immunochromatographic assay (LFIA) for the detection of anti-SARS-CoV-2 IgG and compared LFIA results with chemiluminescent immunoassay (CLIA) results and an in-house enzyme immunoassay (EIA). To this end, a total of 216 whole blood or serum samples from three groups were analyzed: the first group was composed of 68 true negative cases corresponding to blood bank donors, healthy young volunteers, and eight pediatric patients diagnosed with other coronavirus infections. The serum samples from these participants were obtained and stored in a pre-COVID-19 period, thus they were not expected to have COVID-19. In the second group of true positive cases, we chose to replace natural cases of COVID-19 by 96 participants who were expected to have produced anti-SARS-CoV-2 IgG antibodies 30-60 days after the vaccine booster dose. The serum samples were collected on the same day that LFIA were tested either by EIA or CLIA. The third study group was composed of 52 participants (12 adults and 40 children) who did or did not have anti-SARS-CoV-2 IgG antibodies due to specific clinical scenarios. The 12 adults had been vaccinated more than seven months before LFIA testing, and the 40 children had non-severe COVID-19 diagnosed using RT-PCR during the acute phase of infection. They were referred for outpatient follow-up and during this period the serum samples were collected and tested by CLIA and LFIA. All tests were performed by the same healthcare operator and there was no variation of LFIA results when tests were performed on finger prick whole blood or serum samples, so that results were grouped for analysis. LFIA's sensitivity in detecting anti-SARS-CoV-2 IgG antibodies was 90%, specificity 97.6%, efficiency 93%, PPV 98.3%, NPV 86.6%, and likelihood ratio for a positive or a negative result were 37.5 and 0.01 respectively. There was a good agreement (Kappa index of 0.677) between LFIA results and serological (EIA or CLIA) results. In conclusion, LFIA analyzed in this study showed a good technical performance and agreement with reference serological assays (EIA or CLIA), therefore it can be recommended for use in the outpatient follow-up of non-severe cases of COVID-19 and to assess anti-SARS-CoV-2 IgG antibody production induced by vaccination and the antibodies decrease over time. However, LFIAs should be confirmed by using reference serological assays whenever possible.

Protein antigens are often combined with aluminum hydroxide (alum), the most commonly used adjuvant in licensed vaccines; yet the immunogenicity of alum-adjuvanted vaccines leaves much room for improvement. Here, the authors demonstrate a strategy for codelivering an immunostimulatory cytokine, the interleukin IL-21, with an engineered outer domain (eOD) human immunodeficiency virus gp120 Env immunogen eOD, bound together to alum to bolster the humoral immune response. In this approach, the immunogen and cytokine are co-anchored to alum particles via a short phosphoserine (pSer) peptide linker, promoting stable binding to alum and sustained bioavailability following injection. pSer-modified eOD and IL-21 promote enhanced lymphatic drainage and lead to accumulation of the vaccine in B cell follicles in the draining lymph nodes. This in turn promotes enhanced T follicular helper cell priming and robust germinal center responses as well as increased antigen-specific serum IgG titers. This is a general strategy for codelivery of immunostimulatory cytokine with immunogens providing a facile approach to modulate T cell priming and GC reactions toward enhanced protective immunity using the most common clinical vaccine adjuvant., Competing Interests: D.J.I., Y.A., and T.J.M. are inventors on patent applications related to the technology described in the manuscript., (© 2022 The Authors. Advanced Therapeutics published by Wiley‐VCH GmbH.)

Schistosomiasis is one of the most important tropical diseases. A fundamental strategy to control its spread is the use of natural products against its vectors, which are snails of the genus Biomphalaria. The present study evaluated the chemical composition, the molluscicidal and cercaricidal effects, and the ecotoxicity of the essential oil from the aerial parts of Dysphania ambrosioides (L.) Mosyakin & Clemants (DAEO). The essential oil was obtained by hydrodistillation and analyzed by gas chromatography-mass spectrometry (GC-MS). Molluscicidal and cercaricidal activities were determined by the immersion method. Environmental toxicity was assessed from bioassays using Artemia salina larvae and Danio rerio fish. DAEO presented a 0.8% yield. The GC-MS analysis revealed the predominance of hydrocarbon monoterpenes in the oil. A total of 32 constituents was identified, with α-terpinene (50.69%) being the major compound, followed by p-cymene (13.27%) and ascaridole (10.26%). DAEO was active against adult Biomphalaria glabrata snails and demonstrated lethal effect against Schistosoma mansoni cercariae, with LC 50 values of 25.2 (22.7-27.8) and 62.4 (61.8-62.9) μg/mL, respectively. Regarding toxicity to non-target aquatic organisms, the oil showed LC 50 values of 86.9 (84.7-87.6) and 18.6 μg/mL (15.5-22.8) for A. salina and D. rerio, respectively. DAEO proved to be a promising natural product for the control of schistosomiasis, acting on both the vectors and the etiological agent of the disease. However, the use of the oil is safer in transmission sites where there are no non-target organisms, as it has showed toxicity to D. rerio fish., (Copyright © 2022. Published by Elsevier B.V.)

Anti-tumour inflammatory cytokines are highly toxic when administered systemically. Here, in multiple syngeneic mouse models, we show that the intratumoural injection of recombinantly expressed cytokines bound tightly to the common vaccine adjuvant aluminium hydroxide (alum) (via ligand exchange between hydroxyls on the surface of alum and phosphoserine residues tagged to the cytokine by an alum-binding peptide) leads to weeks-long retention of the cytokines in the tumours, with minimal side effects. Specifically, a single dose of alum-tethered interleukin-12 induced substantial interferon-γ-mediated T-cell and natural-killer-cell activities in murine melanoma tumours, increased tumour antigen accumulation in draining lymph nodes and elicited robust tumour-specific T-cell priming. Moreover, intratumoural injection of alum-anchored cytokines enhanced responses to checkpoint blockade, promoting cures in distinct poorly immunogenic syngeneic tumour models and eliciting control over metastases and distant untreated lesions. Intratumoural treatment with alum-anchored cytokines represents a safer and tumour-agnostic strategy to improving local and systemic anticancer immunity., (© 2022. The Author(s), under exclusive licence to Springer Nature Limited.)

There is a need for additional rapidly scalable, low-cost vaccines against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) to achieve global vaccination. Aluminum hydroxide (alum) adjuvant is the most widely available vaccine adjuvant but elicits modest humoral responses. We hypothesized that phosphate-mediated coanchoring of the receptor binding domain (RBD) of SARS-CoV-2 together with molecular adjuvants on alum particles could potentiate humoral immunity by promoting extended vaccine kinetics and codelivery of vaccine components to lymph nodes. Modification of RBD immunogens with phosphoserine (pSer) peptides enabled efficient alum binding and slowed antigen clearance, leading to notable increases in germinal center responses and neutralizing antibody titers in mice. Adding phosphate-containing CpG or saponin adjuvants to pSer-RBD:alum immunizations synergistically enhanced vaccine immunogenicity in mice and rhesus macaques, inducing neutralizing responses against SARS-CoV-2 variants. Thus, phosphate-mediated coanchoring of RBD and molecular adjuvants to alum is an effective strategy to enhance the efficacy of SARS-CoV-2 subunit vaccines.

Saponins are potent and safe vaccine adjuvants, but their mechanisms of action remain incompletely understood. Here, we explored the properties of several saponin formulations, including immune-stimulatory complexes (ISCOMs) formed by the self-assembly of saponin and phospholipids in the absence or presence of the Toll-like receptor 4 agonist monophosphoryl lipid A (MPLA). We found that MPLA self-assembles with saponins to form particles physically resembling ISCOMs, which we termed saponin/MPLA nanoparticles (SMNP). Saponin-containing adjuvants exhibited distinctive mechanisms of action, altering lymph flow in a mast cell–dependent manner and promoting antigen entry into draining lymph nodes. SMNP was particularly effective, exhibiting even greater potency than the compositionally related adjuvant AS01 B in mice, and primed robust germinal center B cell, T FH , and HIV tier 2 neutralizing antibodies in nonhuman primates. Together, these findings shed new light on mechanisms by which saponin adjuvants act to promote the immune response and suggest that SMNP may be a promising adjuvant in the setting of HIV, SARS-CoV-2, and other pathogens.

Global containment of COVID-19 still requires accessible and affordable vaccines for low- and middle-income countries (LMICs). Recently approved vaccines provide needed interventions, albeit at prices that may limit their global access. Subunit vaccines based on recombinant proteins are suited for large-volume microbial manufacturing to yield billions of doses annually, minimizing their manufacturing cost. These types of vaccines are well-established, proven interventions with multiple safe and efficacious commercial examples. Many vaccine candidates of this type for SARS-CoV-2 rely on sequences containing the receptor-binding domain (RBD), which mediates viral entry to cells via ACE2. Here we report an engineered sequence variant of RBD that exhibits high-yield manufacturability, high-affinity binding to ACE2, and enhanced immunogenicity after a single dose in mice compared to the Wuhan-Hu-1 variant used in current vaccines. Antibodies raised against the engineered protein exhibited heterotypic binding to the RBD from two recently reported SARS-CoV-2 variants of concern (501Y.V1/V2). Presentation of the engineered RBD on a designed virus-like particle (VLP) also reduced weight loss in hamsters upon viral challenge., Competing Interests: Competing interest statement: L.E.C., K.R.L., and J.C.L. have filed patents related to the InSCyT system and methods. N.C.D., S.R.A., and J.C.L. have filed a patent related to the RBD-L452K-F490W sequence. K.R.L., L.E.C., and M.K.T. are current employees at Sunflower Therapeutics PBC. N.P.K., J.Y.W., and C.W. are named as inventors on patent applications filed by the University of Washington related to the I3-01 nanoparticle. N.P.K. is a co-founder, shareholder, paid consultant, and chair of the scientific advisory board of Icosavax, Inc. and has received an unrelated sponsored research agreement from Pfizer. J.C.L. has interests in Sunflower Therapeutics PBC, Pfizer, Honeycomb Biotechnologies, OneCyte Biotechnologies, QuantumCyte, Amgen, and Repligen. J.C.L.’s interests are reviewed and managed under MIT’s policies for potential conflicts of interest. J.M.S. is an employee of the Bill & Melinda Gates Medical Research Institute. H.K. is an employee of the Bill & Melinda Gates Foundation., (Copyright © 2021 the Author(s). Published by PNAS.)

Hyaluronic acid (HA) is a biopolymer formed by UDP-glucuronic acid and UDP-N-acetyl-glucosamine disaccharide units linked by β-1,4 and β-1,3 glycosidic bonds. It is widely employed in medical and cosmetic procedures. HA is synthesized by hyaluronan synthase (HAS), which catalyzes the precursors' ligation in the cytosol, elongates the polymer chain, and exports it to the extracellular space. Here, we engineer Ogataea ( Hansenula ) polymorpha for HA production by inserting the genes encoding UDP-glucose 6-dehydrogenase, for UDP-glucuronic acid production, and HAS. Two microbial HAS, from Streptococcus zooepidemicus ( hasAs ) and Pasteurella multocida ( hasAp ), were evaluated separately. Additionally, we assessed a genetic switch using integrases in O. polymorpha to uncouple HA production from growth. Four strains were constructed containing both has genes under the control of different promoters. In the strain containing the genetic switch, HA production was verified by a capsule-like layer around the cells by scanning electron microscopy in the first 24 h of cultivation. For the other strains, the HA was quantified only after 48 h and in an optimized medium, indicating that HA production in O. polymorpha is limited by cultivation conditions. Nevertheless, these results provide a proof-of-principle that O. polymorpha is a suitable host for HA production.

The present study aimed to evaluate the nutritional composition and production of mycelial biomass of Pleurotus eryngii in different culture media, formulated with infusion of purple-skinned sweet potato, glucose concentrations, and the presence or absence of yeast extract. The mushroom was grown for 15 days at 25°C and 150 rpm. The best production of mycelial biomass occurred in the culture medium with 40 g/L of glucose plus yeast extract. At the second cultivation, the fungus entered the exponential growth phase from days 6 until 14. These results indicate that the insertion of nutritional sources in purple-skinned sweet potato media positively influences mycelial biomass production and its nutritional characteristics make it useful for food industry applications.

Filamentous fungi are well known for producing secondary metabolites applied in various industrial segments. Among these, lovastatin and itaconic acid, produced by Aspergillus terreus, have applications in the pharmaceutical and chemical industries. Lovastatin is primarily used for the control of hypercholesterolemia, while itaconic acid is a building block for the production of synthetic fibers, coating adhesives, among others. In this study, for the first time, 35 strains of Aspergillus sp. from four Brazilian culture collections were evaluated for lovastatin and itaconic acid production and compared to a reference strain, ATCC 20542. From an initial screening, the strains ATCC 20542, URM 224, URM1876, URM 5061, URM 5254, URM 5256, URM 5650, and URM 5961 were selected for genomic comparison. Among tested strains, the locus corresponding to the lovastatin genomic cluster was assembled, showing that all genes essential for lovastatin biosynthesis were present in producing URM 5961 and URM 5650 strains, with 100% and 98.5% similarity to ATCC 20542, respectively. However, in the no producing URM 1876, URM 224, URM 5254, URM 5061, and URM 5256 strains, this cluster was either fragmented or missing. Among the 35 strains evaluated for itaconic acid production in this study, only three strains had titers above 0.5 g/L, 16 strains had production below 0.5 g/L, and the remaining 18 strains had no production, with the highest production of itaconic acid observed in the URM 5254 strain with 2.2 g/L. The essential genes for itaconic acid production, mttA, cadA msfA were also mapped, where all three genes linked to itaconic acid production were found in a single contig in the assembly of each strain. In contrast to lovastatin loci, there is no correlation between the level of itaconic acid production and genetic polymorphisms in the genes associated with its biosynthesis., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Background: Chemoinformatics has several applications in the field of drug design, helping to identify new compounds against a range of ailments. Among these are Leishmaniasis, effective treatments for which are currently limited., Objective: To construct new indole 2-aminothiophene molecules using computational tools and to test their effectiveness against Leishmania amazonensis (sp.)., Methods: Based on the chemical structure of thiophene-indol hybrids, we built regression models and performed molecular docking, and used these data as bases for design of 92 new molecules with predicted pIC50 and molecular docking. Among these, six compounds were selected for the synthesis and to perform biological assays (leishmanicidal activity and cytotoxicity)., Results: The prediction models and docking allowed inference of characteristics that could have positive influences on the leishmanicidal activity of the planned compounds. Six compounds were synthesized, one-third of which showed promising antileishmanial activities, with IC50 ranging from 2.16 and 2.97 μM (against promastigote forms) and 0.9 and 1.71 μM (against amastigote forms), with selectivity indexes (SI) of 52 and 75., Conclusion: These results demonstrate the ability of Quantitative Structure-Activity Relationship (QSAR)-based rational drug design to predict molecules with promising leishmanicidal potential, and confirming the potential of thiophene-indole hybrids as potential new leishmanial agents., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

This study is a report on the anti-Leishmania activity of Morita-Baylis-Hillman (MBH) homodimers adducts against the promastigote and axenic amastigote forms of Leishmania (Leishmania) infantum and Leishmania (Leishmania) amazonensis and on the cytotoxicity of these adducts to human blood cells. Both studied homodimers, MBH 1 and MBH 2, showed activity against the promastigote forms of L. infantum and L. amazonensis, which are responsible for visceral and cutaneous leishmaniasis, respectively. Additionally, the homodimers presented biological activity against the axenic amastigote forms of these two Leishmania species. The adducts exhibited no hemolytic activity to human peripheral blood mononuclear cells or erythrocytes at the tested concentrations and achieved higher selectivity indices than amphotericin B. Evaluation of cell death by apoptosis revealed that the homodimers had better apoptosis/necrosis profiles than amphotericin B in the promastigote forms of both L. infantum and L. amazonensis. In conclusion, these Morita-Baylis-Hillman adducts had anti-Leishmania activity in an in vitro model and may thus be promising molecules in the search for new drugs to treat leishmaniasis.

Ankle sprain in plantarflexion and inversion is one of the most common injuries occurring in daily activities and sports. Although acute symptoms may resolve quickly, many individuals have reported persistent pain and instability. Thus, understanding the factors that contribute to the occurrence of this type of injury is extremely important. Although sprains are multifactorial, a relationship can be established between sprain and fatigue. Therefore, the present study examined the latency and intensity of activation of the peroneus longus and brevis muscles under conditions of fatigue. Twenty-three women participated in the study, including 12 with functional instability of the ankle and 11 without a history of sprain. To induce fatigue, the volunteers maintained the force of eversion and plantarflexion at 70% of the maximum voluntary isometric contraction for as long as possible until a 10% decline in the rated force occurred. Ten simulations of ankle sprains were performed before and after fatigue at random for each side, with simultaneous recordings of the electromyographic signals, using a simulator platform for inversion sprain and plantarflexion. As a result, after fatigue, no change in latency was observed. However, a reduction in the intensity of contraction of the muscles analyzed in both groups was observed. Neuromuscular control was concluded to be compromised in situations of fatigue, while differences in muscle behavior were not observed between stable and unstable ankles., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Myracrodruon urundeuva (Engl.) Fr. All., commonly known as "aroeira-do-sertão", is a medicinal plant from Anacardiaceae family. In this study, the chemical composition of M. urundeuva essential oil (MuEO) was evaluated by gas chromatography-mass spectrometry (GC-MS), as well as its anti-Leishmania potential, cytotoxicity, and macrophage activation capability as possible antiprotozoal mechanism of action were assessed. Fourteen compounds were identified, which constituted 94.87% of total oil composition. The most abundant components were monoterpenes (80.35%), with β-myrcene (42.46%), α-myrcene (37.23%), and caryophyllene (4.28%) as the major constituents. The MuEO inhibited the growth of promastigotes (IC 50 205 ± 13.4 μg mL -1 ), axenic amastigotes (IC 50 104.5 ± 11.82 μg mL -1 ) and decreased percentage of macrophage infection and number of amastigotes per macrophage (IC 50 of 44.5 ± 4.37 μg⋅mL -1 ), suggesting significant anti-Leishmania activity. The cytotoxicity of MuEO was assessed by MTT test in Balb/c murine macrophages and by human erythrocytes lysis assay and low cytotoxicity for these cells was observed. The CC 50 value against macrophages were 550 ± 29.21 μg mL -1 , while cytotoxicity for erythrocytes was around 20% at the highest concentration assessed, with HC 50  > 800 μg mL -1 . While MuEO-induced anti-Leishmania activity is not mediated by increases in both lysosomal activity and nitric oxide production in macrophages, the results suggest the antiamastigote activity is associated with an immunomodulatory activity of macrophages due to an increase of phagocytic capability induced by MuEO. Thus, MuEO presented significant activity against Leishmania amazonensis, probably modulating the activation of macrophages, with low cytotoxicity to murine macrophages and human erythrocytes., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Leishmaniasis is an infectious disease complex caused by protozoa from the Leishmania genus, which presents a broad spectrum of clinical manifestations: cutaneous, mucocutaneous and visceral forms. The current treatments are unsatisfactory considering that few drugs are available and present some level of toxicity. Many lignans and neolignans have been used for the development of new antileishmania drugs. The capability in vitro of the neolignan 2,3-dihydrobenzofuran (2,3-DBF), a commonly found constituent of propolis and other plants, to inhibit the growth of promastigote and macrophage-internalized amastigote forms of Leishmania amazonensis was investigated. The cytotoxicity of this compound was assessed by MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) test in BALB/c murine macrophages and human erythrocyte lysis assay. The 2,3-DBF was active against promastigote (IC 50 =1.042 μM) and amastigote (IC 50 =1.43 μM) forms, indicating a potent antileishmanial effect. There was no evidence of cytotoxicity to macrophages or erythrocytes at concentrations ranging from 13 to 0.5 μM, after 48 hr of exposure. The antileishmanial activity is probably mediated by the activation of macrophages, because treatment with 2,3-DBF increases both phagocytic and lysosomal activities, as well as the nitrite (NO 2 - ) levels. These results suggest that 2,3-DBF may be a potential candidate for the development of a new promising antileishmanial drug. Further studies are needed to determine its potential in vivo effect as well as additional mechanisms underlying the antileishmanial and immunomodulatory activities., (© 2016 Nordic Association for the Publication of BCPT (former Nordic Pharmacological Society).)

Leishmaniasis represents a series of severe neglected tropical diseases caused by protozoa of the genus Leishmania and is widely distributed around the world. Here, we present the syntheses of Morita-Baylis-Hillman adducts (MBHAs) prepared from eugenol, thymol and carvacrol, and their bioevaluation against promastigotes of Leishmania amazonensis . The new MBHAs are prepared in two steps from essential oils in moderate to good yields and present IC 50 values in the range of 22.30-4.71 μM. Moreover, the selectivity index to the most potent compound is very high (SIrb > 84.92), far better than that of Glucantime ® (SIrb 1.39) and amphotericin B (SIrb = 22.34). Conformational analysis were carried out at the M062X//6-31+G(d,p) level of theory to corroborate a hypothesis about the nitroaromatic bioreduction mechanism.

Neglected tropical diseases (NTD) are treated with toxic therapy of limited efficacy. Previously, we studied the antimicrobial effect of Dinoponera quadriceps venom (DqV) against bacteria. To continue the study, we report in this short communication the antimicrobial effect of DqV against Leishmania amazonensis and Trypanosoma cruzi. DqV inhibits the promastigote forms of L. amazonensis and all T. cruzi developmental forms, with low toxicity in host cells. DqV causes cell death in T. cruzi through necrotic and apoptotic mechanisms observed by staining the cells with annexin V-FITC (AX) and propidium iodide (PI), loss of mitochondrial membrane potential by flow cytometry analyses and confocal microscopy and morphological alterations, such as loss of membrane integrity and cell shrinkage by scanning electron microscopy (SEM). In conclusion, we suggest there is an antimicrobial effect also on parasites., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

The present study aimed to evaluate the effects of Trypanosoma vivax infection on the shape of the lactation curve and the milk quality of dairy goats experimentally infected with T. vivax. In total, twenty Saanen goats, aged 26-30 months and the same number of calving (two calvings), were divided into two experimental groups: an infected group, consisting of ten goats intravenously infected with 0.5 ml of blood containing approximately 1.25 × 10(5) trypomastigotes of T. vivax and ten uninfected animals as the control group. Clinical tests and hematocrit, parasitemia, and serum biochemistry evaluations were performed on all of the goats. Milk production was measured daily for 152 days by hand milking the goats and weighing the milk. Every seven days, physiochemical analyses were performed to evaluate the milk. Wood's nonlinear model was used to analyze the lactation curve parameters. The infected goats had high levels of parasitemia and hyperthermia, significantly reduced hematocrit, serum total protein, albumin, and glucose levels and increased cholesterol and urea concentrations. Wood's model indicated that the milk production of goats in the infected group declined sharply over a short period of time and produced a flattened yield curve and significant difference (P < 0.05) in the rate of increase of peak milk production, rate of decrease of milk production after the peak, day of peak milk production, and maximum peak milk production compared with that of the control group. Trypanosomiasis also affected the persistency of lactation, which was significantly reduced in goats in the infected group. In addition, the physico-chemical properties of the milk, including the fat content, defatted dry extracts (DDE) and protein content, decreased significantly (P < 0.05) in the goats in the infected group compared with those in the control group. The T. vivax-infected goats showed reduction in milk production, persistence of lactation, and fat levels, the defatted dry extract (DDE) content, and protein, changing the quality of milk., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Zanthoxylum rhoifolium Lam. (Rutaceae) has been traditionally used in the treatment of microbial infections and parasitic diseases. In the present study, the antileishmanial effect induced by the ethanol extract of stem barks from Z. rhoifolium (ZR-EEtOH) and its n-hexane fraction (ZR-FHEX) on infection and infectivity of murine macrophages by promastigote forms of Leishmania amazonensis were investigated. In different set of experiments, macrophages or promastigotes were pretreated with ZR-EEtOH or ZR-FHEX at non-lethal concentrations for 24 hours, and then macrophages were submitted to infection by promastigotes. Moreover, their effects on activation of macrophages, as well as on the DNA content, size and number of promastigotes by flow cytometry were also evaluated. The infection rate and the number of internalized amastigote forms were markedly decreased after pretreatment of macrophages or promastigotes when compared with non-treated cells. The increase in phagocytic capability and nitrite content was also observed. Furthermore, the decrease of DNA content, size and number of promastigotes was also observed. In conclusion, ZR-EEtOH and ZR-FHEX promoted a markedly significant antileishmanial effect and reduction of infection of macrophages, probably underlying defense mechanisms activation in macrophages. These findings reinforce the potential application of Z. rhoifolium in the treatment of leishmaniasis.

This study evaluated the effects of 2-amino-thiophene derivatives on the promastigote and amastigote forms of Leishmania (Leishmania) amazonensis and their possible mechanisms of action. Initially, we evaluated the antileishmanial activity of ten 2-amino-thiophene derivatives on promastigote and axenic amastigote forms of Leishmania amazonensis and their cytotoxicity against murine macrophages and human red blood cells. Three promising compounds were selected for studies of the cell death process using flow cytometry analysis and a DNA fragmentation assay. The effects of the compounds were assessed on intramacrophagic amastigotes, and the modulation of cytokine and NO production was investigated. All thiophene derivatives showed antileishmanial activity against promastigotes and axenic amastigotes with less toxicity for murine macrophages and human red blood cells. The best values were obtained for compounds containing a lateral indole ring. Docking studies suggested that these compounds played an important role in inhibiting trypanothione reductase (TryR) activity. The selected compounds SB-200, SB-44, and SB-83 induced apoptosis in promastigotes involving phosphatidylserine externalization and DNA fragmentation in a pattern similar to that observed for the positive control. Additionally, SB-200, SB-44, and SB-83 significantly reduced the infection index of macrophages by the parasites; for compounds SB-200 and SB-83 this reduction was associated with increased TNF-α, IL-12, and NO levels. This study demonstrated the effective and selective action of 2-amino-thiophene derivatives against L. amazonensis, resulting in apoptosis-like cell death and immunomodulation in vitro. The results suggest that they are promising compounds for the development of new leishmanicidal drugs., (Copyright © 2015 Elsevier Masson SAS. All rights reserved.)

Ethnopharmacological Relevance: Syzygium cumini (L.) Skeels (Myrtaceae), commonly known as "jambolão" in Brazil is widely used in folk medicine against leishmaniasis, inflammation, chronic diarrhea, and ulcers. It is one of the most commonly used plants for the treatment of diabetes worldwide. In previous studies, Syzygium cumini was shown to possess antihyperlipidemic and anti-allergic properties, and to exhibit good performance as an antimicrobial agent against bacteria, fungi, and protozoa parasites of the genus Leishmania and Trypanosoma. This study was aimed at evaluating the effects of S. cumini essential oil (ScEO) and its major component α-pinene on Leishmania (Leishmania) amazonensis, as well as their cytotoxicity and possible mechanisms of action., Materials and Methods: To evaluate the anti-proliferative effect on Leishmania, effects on promastigote and axenic amastigote forms were assessed using tetrazolium salt (MTT) assay. The intramacrophagic amastigotes were exposed to ScEO and α-pinene to determine the survival index. To gain insight into the mechanism of action involved in the effect on the samples, we evaluated the modulation of macrophage activation state by observing structural (phagocytic and lysosomal activities) and cellular (nitric oxide increase) changes. To assess the safety profile of ScEO and α-pinene, murine macrophages and human red blood cells were treated with ScEO and α-pinene and the selectivity index was calculated for each treatment., Results: α-Pinene was effective against Leishmania amazonensis promastigote forms, with a half-maximal inhibitory concentration (IC50) value of 19.7µg/mL. α-Pinene was more active (IC50 values of 16.1 and 15.6µg/mL against axenic and intracellular amastigotes, respectively) than ScEO (IC50 values of 43.9 and 38.1µg/mL against axenic and intracellular amastigotes, respectively). Our results showed that the anti-Leishmania effects were mediated by immunomodulatory activity, as evidenced by the observed increases in both phagocytic and lysosomal activity, and the elevated NO levels. ScEO and α-pinene exhibited low cytotoxicity against murine macrophages and human erythrocytes. The 50% cytotoxicity concentration (CC50) values for the macrophages in the MTT assay were 614.1 and 425.2µg/mL for ScEO and α-pinene, respectively, while the corresponding half-maximal hemolytic concentration (HC50) values were 874.3 and 233.3µg/mL., Conclusions: Taken together, the results demonstrate that ScEO and its major constituent α-pinene have significant anti-Leishmania activity, modulated by macrophage activation, with acceptable levels of cytotoxicity in murine macrophages and human erythrocytes. Further work is warranted, involving more in-depth mechanistic studies and in vivo investigations., (Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.)

The mosquito Aedes aegypti L. (Diptera: Culicidae) is the major vector of dengue and chikungunya fever. The lack of effective therapies and vaccines for these diseases highlights the need for alternative strategies to control the spread of virus. Therefore, this study investigated the larvicidal potential of essential oils from common plant species obtained from the Chapada das Mesas National Park, Brazil, against third instar A. aegypti larvae. The chemical composition of these oils was determined by gas chromatography coupled to mass spectrometry. The essential oils of Eugenia piauhiensis Vellaff., Myrcia erythroxylon O. Berg, Psidium myrsinites DC., and Siparuna camporum (Tul.) A. DC. were observed to be mainly composed of sesquiterpene hydrocarbons. The essential oil of Lippia gracilis Schauer was composed of oxygenated monoterpenes. Four of the five tested oils were effective against the A. aegypti larvae, with the lethal concentration (LC50) ranging from 230 to 292 mg/L after 24 h of exposure. Overall, this work demonstrated the possibility of developing larvicidal products against A. aegypti by using essential oils from the flora of the Brazilian Legal Amazon. This in turn demonstrates the potential of using natural resources for the control of disease vectors.

Objective: The aim of the present study was to assess the feasibility of the early implementation of a swallowing rehabilitation program in tracheostomized patients under mechanical ventilation with dysphagia., Methods: This prospective study was conducted in the intensive care units of a university hospital. We included hemodynamically stable patients under mechanical ventilation for at least 48 hours following 48 hours of tracheostomy and with an appropriate level of consciousness. The exclusion criteria were previous surgery in the oral cavity, pharynx, larynx and/or esophagus, the presence of degenerative diseases or a past history of oropharyngeal dysphagia. All patients were submitted to a swallowing rehabilitation program. An oropharyngeal structural score, a swallowing functional score and an otorhinolaryngological structural and functional score were determined before and after swallowing therapy., Results: We included 14 patients. The mean duration of the rehabilitation program was 12.4 ± 9.4 days, with 5.0 ± 5.2 days under mechanical ventilation. Eleven patients could receive oral feeding while still in the intensive care unit after 4 (2 - 13) days of therapy. All scores significantly improved after therapy., Conclusion: In this small group of patients, we demonstrated that the early implementation of a swallowing rehabilitation program is feasible even in patients under mechanical ventilation.

Background: The recognition and measurement of pain in cattle are important in determining the necessity for and efficacy of analgesic intervention. The aim of this study was to record behaviour and determine the validity and reliability of an instrument to assess acute pain in 40 cattle subjected to orchiectomy after sedation with xylazine and local anaesthesia. The animals were filmed before and after orchiectomy to record behaviour. The pain scale was based on previous studies, on a pilot study and on analysis of the camera footage. Three blinded observers and a local observer assessed the edited films obtained during the preoperative and postoperative periods, before and after rescue analgesia and 24 hours after surgery. Re-evaluation was performed one month after the first analysis. Criterion validity (agreement) and item-total correlation using Spearman's coefficient were employed to refine the scale. Based on factor analysis, a unidimensional scale was adopted., Results: The internal consistency of the data was excellent after refinement (Cronbach's α coefficient = 0.866). There was a high correlation (p < 0.001) between the proposed scale and the visual analogue, simple descriptive and numerical rating scales. The construct validity and responsiveness were confirmed by the increase and decrease in pain scores after surgery and rescue analgesia, respectively (p < 0.001). Inter- and intra-observer reliability ranged from moderate to very good. The optimal cut-off point for rescue analgesia was > 4, and analysis of the area under the curve (AUC = 0.963) showed excellent discriminatory ability., Conclusion: The UNESP-Botucatu unidimensional pain scale for assessing acute postoperative pain in cattle is a valid, reliable and responsive instrument with excellent internal consistency and discriminatory ability. The cut-off point for rescue analgesia provides an additional tool for guiding analgesic therapy.

Background: Mimosa caesalpiniifolia Benth. (Leguminosae) is widely found in the Brazilian Northeast region and markedly contributes to production of pollen and honey, being considered an important honey plant in this region., Objective: To investigate the chemical composition of the ethanol extract of leaves from M. caesalpiniifolia by GC-MS after derivatization (silylation), as well as to evaluate the in vitro and in vivo toxicological effects and androgenic activity in rats., Materials and Methods: The ethanol extract of leaves from Mimosa caesalpiniifolia was submitted to derivatization by silylation and analyzed by gas chromatography-mass spectrometry (GC-MS) to identification of chemical constituents. In vitro toxicological evaluation was performed by MTT assay in murine macrophages and by Artemia salina lethality assay, and the in vivo acute oral toxicity and androgenic evaluation in rats., Results: Totally, 32 components were detected: Phytol-TMS (11.66%), lactic acid-2TMS (9.16%), α-tocopherol-TMS (7.34%) and β-sitosterol-TMS (6.80%) were the major constituents. At the concentrations analyzed, the ethanol extract showed low cytotoxicity against brine shrimp (Artemia salina) and murine macrophages. In addition, the extract did not exhibit any toxicological effect or androgenic activity in rats., Conclusions: The derivatization by silylation allowed a rapid identification of chemical compounds from the M. caesalpiniifolia leaves extract. Besides, this species presents a good safety profile as observed in toxicological studies, and possess a great potential in the production of herbal medicines or as for food consumption.

Heparin is routinely administered in postoperative abdominal surgery aiming to prevent adhesions formation; however, there is no consensus indicating its effectiveness. This study evaluated the effect of heparin on peritoneal reactivity after abdominal surgery, through the association between peritoneal fluid features and ultrasonographic and laparoscopic examination. Ten adult horses were used: control group (CG) and treated group (TG). Both groups underwent laparotomy and small colon enterotomy. TG received subcutaneous heparin at 150 IU/kg every 12 hours for 5 days. The animals underwent ultrasonography and peritoneal fluid examination prior to enterotomy (M0) 12 hours (M1), 1 day (M2), 2 days (M3), 4 days (M4), 6 days (M5), 10 days (M6), and 14 days after enterotomy (M7) with laparoscopic examination being performed on the fifth postoperative day. Peritoneal inflammatory response was observed in both groups. The peritoneal fluid of TG animals showed higher echogenicity during heparin therapy. No inflammatory difference was observed between groups through peritoneal fluid features, except for the higher D-dimer concentration in CG. On laparoscopy, slightly diffuse peritoneal reactivity for both groups was observed, being higher for TG. Laparoscopy and ultrasonography association allowed detailed access to the abdominal cavity. Ultrasonography assessed the diffuse peritoneal inflammation, and laparoscopy allowed the detailed analysis of the segments. No gross beneficial reactions resulting from the use of heparin on peritoneal reactivity were observed; however, it was observed by D-dimer evaluation that the TG had less fibrin deposition, which is directly related to a lower rate of abdominal adhesions formation.

Unlabelled: CONTEXT. Cymbopogon winterianus Jowitt ex Bor (Poaceae), known as citronella grass, is an aromatic herbaceous plant and the essential oil extracted from this grass is used in cosmetics, perfumes, hygiene and cleanliness products worldwide., Objective: This study investigated the composition and molluscicidal and larvicidal activities of the essential oil of C. winterianus cultivated in North Brazil., Materials and Methods: The oil was obtained by hydrodistillation, analyzed by gas chromatography (GC) and GC-mass spectrometry and then its molluscicidal and larvicidal activities against snails (Biomphalaria glabrata) and hatched larvae of Artemia salina, respectively, were evaluated at concentrations from 10 to 1000 mg/L., Results: The main constituents of oil were citronellal (26.5%), geraniol (16.2%), elemol (14.5%) and citronellol (7.3%). The molluscicidal test revealed significant lethal concentration (LC) values (LC90=97.0 mg/L, LC50=54.0 mg/L and LC20=22.0 mg/L), indicating the presence of molluscicidal compounds in the oil. In addition, the oil showed moderate larvicidal activity (LC50=181.0 mg/L) against the larvae of A. salina, which could justify its use in the aquatic environment without affecting other living organisms., Discussion and Conclusion: The results suggest that the oil of C. winterianus could be an effective alternative to control schistosomiasis, with an average margin of safety to other living organisms that coexist with snails.