Effect of spirulina on risk of hospitalization among patients with COVID-19: the TOGETHER randomized trial.

BACKGROUND: Algae-derived nutraceuticals, such as spirulina, have been reported to have biological activities that may minimize clinical consequences to COVID-19 infections. OBJECTIVES: This study aimed to determine whether spirulina is an effective treatment for high-risk patients with early COVID-19 in an outpatient setting. METHODS: The TOGETHER trial is a placebo-controlled, randomized, platform trial conducted in Brazil. Eligible participants were symptomatic adults with a positive rapid test for SARS-CoV-2 older than 50 y or with a known risk factor for disease severity. Patients were randomly assigned to receive placebo or spirulina (1 g twice daily for 14 d). The primary end point was hospitalization defined as either retention in a COVID-19 emergency setting for >6 h or transfer to tertiary hospital owing to COVID-19 at 28 d. Secondary outcomes included time-to-hospitalization, mortality, and adverse drug reactions. We used a Bayesian framework to compare spirulina with placebo. RESULTS: We recruited 1126 participants, 569 randomly assigned to spirulina and 557 to placebo. The median age was 49.0 y, and 65.3% were female. The primary outcome occurred in 11.2% in the spirulina group and 8.1% in the placebo group (odds ratio [OR]: 1.24; 95% credible interval: 0.84, 1.86). There were no differences in emergency department visit (OR: 1.21; 95% credible interval: 0.81, 1.83), nor time to symptom relief (hazard ratio: 0.90; 95% credible interval: 0.79, 1.03). Spirulina also not demonstrate important treatment effects in the prespecified subgroups defined by age, sex, BMI, days since symptom onset, or vaccination status. CONCLUSIONS: Spirulina has no any clinical benefits as an outpatient therapy for COVID-19 compared with placebo with respect to reducing the retention in an emergency setting or COVID-19-related hospitalization. There are no differences between spirulina and placebo for other secondary outcomes. This trial was registered at clinicaltrials.gov as NCT04727424.

The impact of cholesteryl ester transfer protein on the progression of cutaneous leishmaniasis.

Introduction: Pathogenesis of cutaneous leishmaniases involves parasite growth, persistent inflammation, and likely participation of lipoproteins (LP). The cholesteryl ester transfer protein (CETP), involved in LP remodeling, has been shown to participate in the inflammatory response and the evolution of infectious conditions. Methods: We evaluated the impact of the presence of CETP on infection by Leishmania (L.) amazonensis in an experimental model of cutaneous leishmaniasis using C57BL6/J mice transgenic for human CETP (CETP), having as control their littermates that do not express the protein, wild-type (WT) mice. The progression of the lesion after infection in the footpad was monitored for 12 weeks. Two groups of animals were formed to collect the plantar pad in the 4th and 12th week post-infection. Results: The lesion increased from the 3rd week onwards, in both groups, with a gradual decrease from the 10th week onwards in the CETP group compared to the WT group, showing a reduction in parasitism and an improvement in the healing process, a reduction in CD68+ cells, and an increase in CD163+ and CD206, characterizing a population of M2 macrophages. A reduction in ARG1+ cells and an increase in INOS+ cells were observed. During infection, the LP profile showed an increase in triglycerides in the VLDL fraction in the CETP group at 12 weeks. Gene expression revealed a decrease in the CD36 receptor in the CETP group at 12 weeks, correlating with healing and parasite reduction. In vitro, macrophages derived from bone marrow cells from CETP mice showed lower parasite load at 48 h and, a reduction in arginase activity at 4 h accompanied by increased NO production at 4 and 24 h compared to WT macrophages, corroborating the in vivo findings. Discussion: The data indicate that the presence of CETP plays an important role in resolving Leishmania (L.) amazonensis infection, reducing parasitism, and modulating the inflammatory response in controlling infection and tissue repair.

Matched vs Nonmatched Placebos in a Randomized Trial of COVID-19 Treatments.

Importance: Matched placebo interventions are complex and resource intensive. Recent evidence suggests matched placebos may not always be necessary. Previous studies have predominantly evaluated potential bias of nonmatched placebos (ie, differing on dose, frequency of administration, or formulation) in pain and mental health, but to date no systematic examination has been conducted in infectious disease. Objective: To test for differences between nonmatched and matched placebo arms with respect to clinical outcome measures across multiple therapeutics for COVID-19. Design, Setting, and Participants: In a comparative effectiveness research study, a post hoc analysis was conducted of data on individual patients enrolled in a large, multiarm, platform randomized clinical trial in symptomatic adult outpatients with COVID-19 between January 15, 2021, to September 28, 2023, in which the outcomes of both matched and nonmatched placebo groups were reported. Bayesian and frequentist covariate-adjusted techniques were compared with 7 intervention-placebo pairs. Exposures: Seven matched and nonmatched placebo pairs (for a total of 7 comparisons) were evaluated throughout the primary platform trial. Comparisons were made between treatment and its associated matched (concurrent) placebo, as well as with nonmatched placebo (alone and in combination) assessed at a similar time point. Main Outcomes and Measures: Outcomes assessed included hospitalizations, EuroQol 5-Dimension 5-level scores, and PROMIS Global-10 scores. Results: A total of 7 intervention-control pairs (N = 2684) were assessed, including 1620 (60.4%) women, with mean (SD) age, 47 (15.2) years; the most common comorbidities were obesity (41.9%) and hypertension (37.9%). In a meta-analysis with decoupled SEs, accounting for overlapping placebo patients, the overall odds ratio (OR) of nonmatched compared with matched placebo was 1.01 (95% credible interval, 0.77-1.32), with posterior probability of equivalence, defined as 0.8 ≤ OR ≤ 1.2 (a deviation from perfect equivalence ie, OR = 1, by no more than 0.2) of 85.4%, implying no significant difference. Unadjusted analysis of the event rate difference between all nonmatched and matched placebo groups did not identify any notable differences across all 7 treatment-placebo combinations assessed. Similar analysis that was conducted for patient-reported quality of life outcomes did not yield statistically significant differences. Conclusions and Relevance: In this post hoc study of a randomized clinical platform trial, pooling matched and nonmatched placebo patient data did not lead to inconsistencies in treatment effect estimation for any of the investigational drugs. These findings may have significant implications for future platform trials, as the use of nonmatched placebo may improve statistical power, or reduce barriers to placebo implementation.

Replacing screen time, with physical activity and sleep time: influence on cardiovascular indicators and inflammatory markers in Brazilian children

Abstract Objective: To examine if the substitution of different screen time intervals with light physical activity (LPA), moderate to vigorous physical activity (MVPA) and sleep is associated with cardiovascular indicators and inflammatory markers in children. Methods: This is a cross-sectional study developed with 186 children aged between six and 11 years old from public schools in southern Brazil. CRF was measured with the 6-minute running and walking test, following the Brazil Sports Project procedures. The percentage of fat was evaluated through DXA. LPA and MVPA were measured using accelerometers. Sleep and screen time were assessed by questionnaires answered by parents. Leptin and C-reactive protein were measured by fasting blood collection. Systolic and diastolic blood pressure were determined through a digital sphygmomanometer. Isotemporal substitution models were used for statistical analysis. Results: Replacing 1 h of screen time with MVPA was associated with lower BMI, systolic and diastolic blood pressure, fat percentage, leptin, and C-reactive protein. When screen time was substituted for sleep time, lower waist circumference was observed. Regarding the substitution of 1 h of screen time with LPA, significant values were found only for leptin. Conclusion: The replacement of screen time with physical activities of different intensities and sleep time was associated with benefits in cardiovascular indicators and inflammatory markers in childhood.

Humid and cold forest connections in South America between the eastern Andes and the southern Atlantic coast during the LGM.

The presence of Andean plant genera in moist forests of the Brazilian Atlantic Coast has been historically hypothesized as the result of cross-continental migrations starting at the eastern Andean flanks. Here we test hypotheses of former connections between the Atlantic and Andean forests by examining distribution patterns of selected cool and moist-adapted plant arboreal taxa present in 54 South American pollen records of the Last Glacial Maximum (LGM), ca. 19-23 cal ka, known to occur in both plant domains. Pollen taxa studied include Araucaria, Drimys, Hedyosmum, Ilex, Myrsine, Podocarpus, Symplocos, Weinmannia, Myrtaceae, Ericaceae and Arecaceae. Past connectivity patterns between these two neotropical regions as well as individual ecological niches during the LGM were explored by cluster analysis of fossil assemblages and modern plant distributions. Additionally, we examined the ecological niche of 137 plant species with shared distributions between the Andes and coastal Brazil. Our results revealed five complex connectivity patterns for South American vegetation linking Andean, Amazonian and Atlantic Forests and one disjunction distribution in southern Chile. This study also provides a better understanding of vegetation cover on the large and shallow South American continental shelf that was exposed due to a global sea level drop.

Effects of a physical education intervention on children's physical activity and fitness: the PROFIT pilot study.

BACKGROUND: Physical education classes are widely accepted as one of the most effective settings for promoting physical activity and health and have often been used to implement physical activity interventions. The aim of this pilot study was to test a physical education intervention program on physical activity levels and physical fitness in a sample of school-age children. METHODS: Participants were a convenience sample of 50 children (34 experimental group and 16 in the comparative group) aged between 6 and 11 years old (Mean = 8.28 years). A 21-week intervention was implemented, consisting of high-intensity and physical fitness-focused exercises, in addition to a once-a-month extra class nutritional education. The following variables were evaluated before and post-intervention: physical fitness, sedentary behavior (SB), light physical activity (LPA), moderate physical activity (MVA), and vigorous physical activity (VPA). Propensity score analyses calculated the average treatment effect on the treated (ATET) within a quasi-experimental framework. RESULTS: Physical fitness variables showed improvements after the intervention, specifically for agility (ATET = -0.67 s; p < 0.001), cardiorespiratory fitness (ATET = 89.27 m; p = 0.045), lower limbs power (ATET = 4.47 centimeters; p = 0.025), and speed (ATET = -1.06 s; p < 0.001). For physical activity and SB levels, there were no improvements after intervention implementation. CONCLUSION: The intervention program showed preliminary effectiveness to improve physical fitness of children, but not SB nor physical activity.

Replacing screen time, with physical activity and sleep time: influence on cardiovascular indicators and inflammatory markers in Brazilian children.

OBJECTIVE: To examine if the substitution of different screen time intervals with light physical activity (LPA), moderate to vigorous physical activity (MVPA) and sleep is associated with cardiovascular indicators and inflammatory markers in children. METHODS: This is a cross-sectional study developed with 186 children aged between six and 11 years old from public schools in southern Brazil. CRF was measured with the 6-minute running and walking test, following the Brazil Sports Project procedures. The percentage of fat was evaluated through DXA. LPA and MVPA were measured using accelerometers. Sleep and screen time were assessed by questionnaires answered by parents. Leptin and C-reactive protein were measured by fasting blood collection. Systolic and diastolic blood pressure were determined through a digital sphygmomanometer. Isotemporal substitution models were used for statistical analysis. RESULTS: Replacing 1 h of screen time with MVPA was associated with lower BMI, systolic and diastolic blood pressure, fat percentage, leptin, and C-reactive protein. When screen time was substituted for sleep time, lower waist circumference was observed. Regarding the substitution of 1 h of screen time with LPA, significant values were found only for leptin. CONCLUSION: The replacement of screen time with physical activities of different intensities and sleep time was associated with benefits in cardiovascular indicators and inflammatory markers in childhood.

Historical Hg accumulation (∼65 cal kyr BP) in upland lakes of the Southeastern Brazilian Amazonia: New evidence of the extent of geogenic and diagenetic control.

The historical upland lake sediments in the Brazilian Amazon witnessed significant enrichment of total mercury (Hg). However, its spatio-temporal relationships between lakes and the main factors responsible for this enrichment are still poorly constrained. Given this, we geochemically investigated 12 radiometrically dated (extending back to ∼65 cal kyr BP) sediment cores from the Carajás plateau, Brazil. The Hg level in historical sediments presented a large temporal variability (from 1 to 3200 µg/kg), with maximum accumulation peaks observed between 30 and 45 cal kyr BP in core R2, LB3, and R1. However, the lack of the Hg peak in other cores (LV2 and LTI3) during the same period despite being proximity and non-correlation of these Hg peaks with the onset of major volcanic events indicates that this source has little bearing. Hg enrichment is highly dependent on the type of sedimentary facies, with higher values were associated with detritic facies (MI) and detritic+organic facies (P/M). Principal component analysis shows that aluminosilicate minerals and organic matter are essential hosts of Hg in sediments. The positive correlation between Al, Ti, and Hg in detritic facies and their strong coherence with Hg/TOC in R1, R5, LSL, ST02, and LB3 cores indicate that Hg is primarily of lithogenic origin. This can be substantiated by the higher background threshold value of Hg (574 µg/kg) in historical lake sediments compared to those in recent lake sediments (340 µg/kg). However, the most pronounced Hg peak (3200 µg/kg) in R2 around 45 cal kyr BP, which correlates positively with TOC, S, Se, As, and Mo indicates their diagenetic enrichment in organic-rich sediments under anoxic conditions. Thus, in addition to the lithogenic effect, it can be argued that diagenesis can play a significant role in prompting Hg enrichment in the Carajás lake sediments in Amazonia.

A comprehensive procedure for pollen extraction from bat guano deposits in organic and detrital matrices.

Although bat guano deposits have been proven to be excellent environmental archives for paleoecological and paleoclimate studies, the development of a standardized method specially catering to pollen extraction has received no attention so far. In general, the processing procedure is quite similar among published studies, but adjustments must be made regarding the proportion of organic and particulate matter in the guano deposit. In this study, we present step-by-step optimized sample processing methods for pollen analysis. These procedures first apply a chemical treatment for the removal of siliceous and organic material, followed by a sieving step to remove the remaining inorganic matter from those samples with high detrital content. Overall, our methods can efficiently remove particulate matter and improve the quality of the final residue, resulting in cleaner slides and better visualization of pollen and spores.•Remove humic acid and organic material with Potassium hydroxide.•Remove inorganic matter with hydrofluoric acid and sieving.•Concentrate and store the pollen residues in glycerin.

Unconventional Sites for Diagnosis of Leptospirosis in Bovine Anicteric Fetuses.

BACKGROUND: Bovine leptospirosis is an important reproductive disease and abortion is a major sign, leading to economic impacts. Due to its multifactorial etiology, the proper diagnosis of the cause of the abortion is crucial. Necropsy of the fetuses followed by molecular analysis is recommended for diagnosis, and the investigation mainly occurs in the kidneys and liver. This study aimed to analyze unconventional sites for the presence of leptospiral DNA in bovine anicteric aborted fetuses. METHODS: Five fetuses of the same herd were received for necropsy and diagnosis. Conventional lipL32-PCR was performed in the fetuses' kidneys, livers, lungs, hearts, spleens, subcapsular kidney content, abomasal fluid, and in the cavity's hemorrhagic contents. To complete the investigation, the sera of 30 cows of the herd were collected to perform the serologic screening by Microscopic Agglutination Test. In addition, six subfertile non-pregnant cows from the same herd were selected due to their low reproductive performance, and genital samples (uterine fragment and cervicovaginal mucus) and urine were collected for lipL32-PCR. PCR-positive samples were submitted to a nested PCR of the secY gene and intended for sequencing. RESULTS: The herd presented seroreactive animals (11/30, 36.6%), all against the Sejroe serogroup, with titers between 200 and 1600. In necropsy, four fetuses showed hemorrhagic and anicteric lesions, while one fetus had no macroscopic lesions. Regarding molecular analysis, all the fetuses were positive in lipL32-PCR and the positive sites were the heart, lungs, subcapsular kidney content, thymus, kidneys, liver, and abomasal fluid. Only one fetus presented positive results in the kidney and liver, while three fetuses were positive in the abomasal fluid. Five of six cows were positive for lipL32-PCR, all being positive only in genital samples. Of the fetuses and the cows, seven sequences were obtained and all were identified as Leptospira interrogans serogroup Sejroe serovar Hardjoprajitno. CONCLUSIONS: In order to improve the diagnosis of leptospirosis in cows, it is recommended to perform a comprehensive analysis of the samples, beyond the kidneys and liver. Thus, we highly encourage testing multiple organs by PCR to investigate abortions suspected of bovine leptospirosis, particularly in anicteric fetuses.

Staphylococcus aureus-Cure-Associated Antigens Elicit Type 3 Immune Memory T Cells.

BACKGROUND: Staphylococcus aureus is one of the most frequently major mastitis pathogens that cause clinical and subclinical mastitis worldwide. Current antimicrobial treatments are usually ineffective, and the commercially available vaccines lack proven effectiveness. The immunological response elicited by the recombinant S. aureus-cure-associated proteins phosphoglycerate kinase (PGK), enolase (ENO), and elongation factor-G (EF-G) in combination with the granulocyte-macrophage colony-stimulating factor (GM-CSF) DNA vaccination was studied in this work. METHODS: Here, twenty-three C57BL/6 mice were divided into four groups and vaccinated with: G1: none (control); G2: GM-CSF DNA plasmid DNA vaccine; G3: the combination of EF-G+ENO+PGK; and G4: the combinations of EF-G+ENO+PGK proteins plus GM-CSF plasmid DNA vaccine. After 44 days, spleen cells were collected for immunophenotyping and lymphocyte proliferation evaluation by flow cytometry upon S. aureus stimulus. RESULTS: Immunization with the three S. aureus recombinant proteins alone resulted in a higher percentage of IL-17A+ cells among CD8+ T central memory cells, as well as the highest intensity of IL-17A production by overall lymphocytes indicating that the contribution of the combined lymphocyte populations is crucial to sustaining a type 3 cell immunity environment. CONCLUSION: The immunization with three S. aureus-cure-associated recombinant proteins triggered type 3 immunity, which is a highly interesting path to pursue an effective bovine S. aureus mastitis vaccine.

Memory CD4+ and CD8+ T lymphocyte proliferation in vaccinated dairy cows with different histories of Staphylococcus aureus mastitis.

Staphylococcus aureus mastitis constitutes a serious threat to dairy cows. The reasons why available vaccines are not fully effective remain poorly understood; thus, in the present study, we investigated CD4+ and CD8+ T lymphocyte proliferation in dairy cows vaccinated with a polyvalent mastitis vaccine that had distinct precedent Staphylococcus aureus mastitis. We studied 17 S. aureus-infected dairy cows (11 vaccinated and six unvaccinated) and eight vaccinated healthy dairy cows with no previous S. aureus mastitis infections. Flow cytometry was used to assess lymphocyte proliferation using an anti-Ki67 antibody, and monoclonal antibodies were used to identify T cell subsets. S. aureus-infected cows exhibited reduced overall lymphocyte proliferation, including CD4+ T lymphocyte proliferation, and memory lymphocyte proliferation in response to S. aureus isolate stimulus. Immunization did not influence the expansion of blood lymphocyte populations. Furthermore, CD8+ T cells, memory CD8+ T lymphocytes, and effector memory CD8+ T lymphocytes displayed reduced proliferation 21 days after the third vaccine dose compared with before vaccination at time zero. The present data demonstrates an overall negative regulation of the T-cell response suggesting its detrimental impact leading to the persistence of S. aureus intramammary infections. Furthermore, the lack of vaccination effect on T-cell mediated immunity (e.g., proliferation) may be related to poor vaccine efficacy.

Análise da qualidade de vida em pacientes pediátricos com alergia alimentar / Quality of life assessment in pediatric patients with food allergy

Introdução: A alergia alimentar pode afetar o bem-estar dos pacientes e de seus familiares. Esse trabalho busca, por meio de questionário validado, investigar a qualidade de vida desses pacientes, acompanhados em um centro de tratamento multidisciplinar. Métodos: Pacientes entre 0 e 18 anos, monitorados no Ambulatório de Alergia Alimentar do Hospital Infantil João Paulo II entre 2012 e 2017, foram selecionados para responder a um questionário de avaliação de qualidade de vida com coleta de informações acerca do tipo de alergia, sua apresentação clínica, presença de dermatite atópica, prescrição ou não de kit de Adrenalina®, tempo de acompanhamento no serviço e tempo de acompanhamento por nutricionista. Resultados: Foram incluídos 77 pacientes, com idade média de 3,38 anos, em sua maioria revelando qualidade de vida regular (43%) e com acompanhamento no Serviço inferior a seis meses (52%). Daqueles acompanhados por nutricionista, 52,4% o faziam há menos de seis meses. Alergia IgE mediada foi identificada em 51% dos sujeitos da pesquisa, com 66,66% dos mesmos sob prescrição de kit de Adrenalina®. Não houve associação estatisticamente significativa entre qualidade de vida e as variáveis analisadas. Conclusão: O questionário de qualidade de vida é um importante instrumento de avaliação de pacientes com alergia alimentar, permitindo traçar o perfil dos mesmos e atuar individualmente nos quesitos que impactam negativamente o seu dia a dia.

Background: Food allergy can affect the well-being of patients and their families. Objective: To investigate the quality of life of patients with food allergy followed up at a multidisciplinary treatment center using a validated questionnaire. Methods: Patients aged 0 to 18 years followed up at the Food Allergy Outpatient Clinic of João Paulo II Pediatric Hospital between 2012 and 2017 were invited to answer a quality-of-life assessment questionnaire for information on type of allergy, clinical presentation, presence of atopic dermatitis, prescription of an epinephrine kit, duration of follow-up at the clinic, and duration of follow-up with a dietitian. Results: A total of 77 patients were included, with a mean age of 3.38 years. Most participants rated their quality of life as fair (43%) and had less than 6 months of outpatient follow-up (52%). From those meeting with a dietitian, 52.4% had less than 6 months of follow-up. Immunoglobulin E (IgE)-mediated allergy was identified in 51% of participants, and 66.66% of them required an epinephrine kit. There was no statistically significant association between quality of life and the study variables. Conclusion: A quality-of-life assessment questionnaire is an important tool for evaluating patients with food allergy, allowing us to profile these patients and to act individually on issues that might negatively impact their daily lives.

Corrigendum: miR-548d-3p Is Up-regulated in Human Visceral Leishmaniasis and Suppresses Parasite Growth in Macrophages.

[This corrects the article DOI: 10.3389/fcimb.2022.826039.].

miR-548d-3p Is Up-Regulated in Human Visceral Leishmaniasis and Suppresses Parasite Growth in Macrophages.

Visceral leishmaniasis caused by Leishmania (Leishmania) infantum in Latin America progress with hepatosplenomegaly, pancytopenia, hypergammaglobulinemia, and weight loss and maybe lethal mainly in untreated cases. miRNAs are important regulators of immune and inflammatory gene expression, but their mechanisms of action and their relationship to pathogenesis in leishmaniasis are not well understood. In the present study, we sought to quantify changes in miRNAs associated with immune and inflammatory pathways using the L. (L.) infantum promastigote infected- human monocytic THP-1 cell model and plasma from patients with visceral leishmaniasis. We identified differentially expressed miRNAs in infected THP-1 cells compared with non-infected cells using qPCR arrays. These miRNAs were submitted to in silico analysis, revealing targets within functional pathways associated with TGF-ß, chemokines, glucose metabolism, inflammation, apoptosis, and cell signaling. In parallel, we identified differentially expressed miRNAs in active visceral leishmaniasis patient plasma compared with endemic healthy controls. In silico analysis of these data indicated different predicted targets within the TGF-ß, TLR4, IGF-I, chemokine, and HIF1α pathways. Only a small number of miRNAs were commonly identified in these two datasets, notably with miR-548d-3p being up-regulated in both conditions. To evaluate the potential biological role of miR-548d-3p, we transiently transfected a miR-548d-3p inhibitor into L. (L.) infantum infected-THP-1 cells, finding that inhibition of miR-548d-3p enhanced parasite growth, likely mediated through reduced levels of MCP-1/CCL2 and nitric oxide production. Further work will be required to determine how miR-548d-3p plays a role in vivo and whether it serves as a potential biomarker of progressive leishmaniasis.

Early reduction in PD-L1 expression predicts faster treatment response in human cutaneous leishmaniasis.

Cutaneous leishmaniasis (CL) is caused by Leishmania donovani in Sri Lanka. Pentavalent antimonials (e.g., sodium stibogluconate [SSG]) remain first-line drugs for CL with no new effective treatments emerging. We studied whole blood and lesion transcriptomes from Sri Lankan patients with CL at presentation and during SSG treatment. From lesions but not whole blood, we identified differential expression of immune-related genes, including immune checkpoint molecules, after onset of treatment. Using spatial profiling and RNA-FISH, we confirmed reduced expression of programmed death-ligand 1 (PD-L1) and indoleamine 2,3-dioxygenase 1 (IDO1) proteins on treatment in lesions of a second validation cohort and further demonstrated significantly higher expression of these checkpoint molecules on parasite-infected compared with noninfected lesional CD68+ monocytes and macrophages. Crucially, early reduction in PD-L1 but not IDO1 expression was predictive of rate of clinical cure (HR = 4.88) and occurred in parallel with reduction in parasite load. Our data support a model whereby the initial anti-leishmanial activity of antimonial drugs alleviates checkpoint inhibition on T cells, facilitating immune-drug synergism and clinical cure. Our findings demonstrate that PD-L1 expression can be used as a predictor of rapidity of clinical response to SSG treatment in Sri Lanka and support further evaluation of PD-L1 as a host-directed therapeutic in leishmaniasis.

Staphylococcus aureus Protection-Related Type 3 Cell-Mediated Immune Response Elicited by Recombinant Proteins and GM-CSF DNA Vaccine.

Staphylococcus aureus mastitis remains a major challenge for dairy farming. Here, 24 mice were immunized and divided into four groups: G1: control; G2: Granulocyte Macrophage Colony-Stimulating Factor (GM-CSF) DNA vaccine; G3: F0F1 ATP synthase subunit α (SAS), succinyl-diaminopimelate (SDD), and cysteinyl-tRNA synthetase (CTS) recombinant proteins; and G4: SAS+SDD+CTS plus GM-CSF DNA vaccine. The lymphocyte subpopulations, and the intracellular interleukin-17A (IL-17A) and interferon-γ production in the draining lymph node cells were immunophenotyped by flow cytometry. The immunophenotyping and lymphocyte proliferation was determined in spleen cells cultured with and without S. aureus stimulus. Immunization with S. aureus recombinant proteins generated memory cells in draining lymph nodes. Immunization with the three recombinant proteins plus GM-CSF DNA led to an increase in the percentage of IL-17A+ cells among overall CD44+ (memory), T CD4+, CD4+ T CD44+ CD27-, γδ TCR, γδ TCR+ CD44+ CD27+, and TCRVγ4+ cells. Vaccination with S. aureus recombinant proteins associated with GM-CSF DNA vaccine downregulated TH2 immunity. Immunization with the three recombinant proteins plus the GM-CSF DNA led to a proliferation of overall memory T, CD4+, and CD4+ TEM cells upon S. aureus stimulus. This approach fostered type 3 immunity, suggesting the development of a protective immune response against S. aureus.

miR-548d-3p Alters Parasite Growth and Inflammation in Leishmania (Viannia) braziliensis Infection.

American Tegumentary Leishmaniasis (ATL) is an endemic disease in Latin America, mainly caused in Brazil by Leishmania (Viannia) braziliensis. Clinical manifestations vary from mild, localized cutaneous leishmaniasis (CL) to aggressive mucosal disease. The host immune response strongly determines the outcome of infection and pattern of disease. However, the pathogenesis of ATL is not well understood, and host microRNAs (miRNAs) may have a role in this context. In the present study, miRNAs were quantified using qPCR arrays in human monocytic THP-1 cells infected in vitro with L. (V.) braziliensis promastigotes and in plasma from patients with ATL, focusing on inflammatory response-specific miRNAs. Patients with active or self-healed cutaneous leishmaniasis patients, with confirmed parasitological or immunological diagnosis, were compared with healthy controls. Computational target prediction of significantly-altered miRNAs from in vitro L. (V.) braziliensis-infected THP-1 cells revealed predicted targets involved in diverse pathways, including chemokine signaling, inflammatory, cellular proliferation, and tissue repair processes. In plasma, we observed distinct miRNA expression in patients with self-healed and active lesions compared with healthy controls. Some miRNAs dysregulated during THP-1 in vitro infection were also found in plasma from self-healed patients, including miR-548d-3p, which was upregulated in infected THP-1 cells and in plasma from self-healed patients. As miR-548d-3p was predicted to target the chemokine pathway and inflammation is a central to the pathogenesis of ATL, we evaluated the effect of transient transfection of a miR-548d-3p inhibitor on L. (V.) braziliensis infected-THP-1 cells. Inhibition of miR-548d-3p reduced parasite growth early after infection and increased production of MCP1/CCL2, RANTES/CCL5, and IP10/CXCL10. In plasma of self-healed patients, MCP1/CCL2, RANTES/CCL5, and IL-8/CXCL8 concentrations were significantly decreased and MIG/CXCL9 and IP-10/CXCL10 increased compared to patients with active disease. These data suggest that by modulating miRNAs, L. (V.) braziliensis may interfere with chemokine production and hence the inflammatory processes underpinning lesion resolution. Our data suggest miR-548d-3p could be further evaluated as a prognostic marker for ATL and/or as a host-directed therapeutic target.

Pleiotropic Effect of Hormone Insulin-Like Growth Factor-I in Immune Response and Pathogenesis in Leishmaniases.

Leishmaniases are diseases caused by several Leishmania species, and many factors contribute to the development of the infection. Because the adaptive immune response does not fully explain the outcome of Leishmania infection and considering that the initial events are crucial in the establishment of the infection, we investigated one of the growth factors, the insulin-like growth factor-I (IGF-I), found in circulation and produced by different cells including macrophages and present in the skin where the parasite is inoculated. Here, we review the role of IGF-I in leishmaniasis experimental models and human patients. IGF-I induces the growth of different Leishmania species in vitro and alters the disease outcome increasing the parasite load and lesion size, especially in L. major- and L. amazonensis-infected mouse leishmaniasis. IGF-I affects the parasite interacting with the IGF-I receptor present on Leishmania. During Leishmania-macrophage interaction, IGF-I acts on the arginine metabolic pathway, resulting in polyamine production both in macrophages and Leishmania. IGF-I and cytokines interact with reciprocal influences on their expression. IL-4 is a hallmark of susceptibility to L. major in murine leishmaniasis, but we observed that IGF-I operates astoundingly as an effector element of the IL-4. Approaching human leishmaniasis, patients with mucosal, disseminated, and visceral diseases presented surprisingly low IGF-I serum levels, suggesting diverse effects than parasite growth. We observed that low IGF-I levels might contribute to the inflammatory response persistence and delayed lesion healing in human cutaneous leishmaniasis and the anemia development in visceral leishmaniasis. We must highlight the complexity of infection revealed depending on the Leishmania species and the parasite's developmental stages. Because IGF-I exerts pleiotropic effects on the biology of interaction and disease pathogenesis, IGF-I turns up as an attractive tool to explore biological and pathogenic processes underlying infection development. IGF-I pleiotropic effects open further the possibility of approaching IGF-I as a therapeutical target.