Munc18a is an SM protein required for SNARE-mediated fusion. The molecular details of how Munc18a acts to enhance neurosecretion have remained elusive. Here, we use in vitro fusion assays to characterize how specific interactions between Munc18a and the neuronal SNAREs enhance the rate and extent of fusion. We show that Munc18a interacts directly and functionally with the preassembled t-SNARE complex. Analysis of Munc18a point mutations indicates that Munc18a interacts with helix C of the Syntaxin1a NRD in the t-SNARE complex. Replacement of the t-SNARE SNAP25b with yeast Sec9c had little effect, suggesting that Munc18a has minimal contact with SNAP25b within the t-SNARE complex. A chimeric Syntaxin built of the Syntaxin1a NRD and the H3 domain of yeast Sso1p and paired with Sec9c eliminated stimulation of fusion, suggesting that Munc18a/Syntaxin1a H3 domain contacts are important. Additionally, a Syntaxin1A mutant lacking a flexible linker region that allows NRD movement abolished stimulation of fusion. These experiments suggest that Munc18a binds to the Syntaxin1a NRD and H3 domain within the assembled t-SNARE complex, positioning them for productive VAMP2 binding. In this capacity, Munc18a serves as a platform for trans-SNARE complex assembly that facilitates efficient SNARE-mediated membrane fusion.

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Chagas disease, caused by Trypanosoma cruzi, results in an acute febrile illness that progresses to chronic chagasic cardiomyopathy in 30% of patients. Current treatments have significant side effects and poor efficacy during the chronic phase; therefore, there is an urgent need for new treatment modalities. A robust T H 1-mediated immune response correlates with favorable clinical outcomes. A therapeutic vaccine administered to infected individuals could bolster the immune response, thereby slowing or stopping the progression of chagasic cardiomyopathy. Prior work in mice has identified an efficacious T. cruzi DNA vaccine encoding Tc24. To elicit a similar protective cell-mediated immune response to a Tc24 recombinant protein, we utilized a poly(lactic-co-glycolic acid) nanoparticle delivery system in conjunction with CpG motif-containing oligodeoxynucleotides as an immunomodulatory adjuvant. In a BALB/c mouse model, the vaccine produced a T H 1-biased immune response, as demonstrated by a significant increase in antigen-specific IFNg-producing splenocytes, IgG2a titers, and proliferative capacity of CD8 C T cells. When tested for therapeutic efficacy, significantly reduced systemic parasitemia was seen during peak parasitemia. Additionally, there was a significant reduction in cardiac parasite burden and inflammatory cell infiltrate. This is the first study demonstrating immunogenicity and efficacy of a therapeutic Chagas vaccine using a nanoparticle delivery system.

Chagas disease, caused by the parasite Trypanosoma cruzi , develops into chronic Chagas’ cardiomyopathy in ~30% of infected individuals, characterized by conduction disorders, arrhythmias, heart failure, and even sudden cardiac death. Current anti-parasitic treatments are plagued by significant side effects and poor efficacy in the chronic phase of disease; thus, there is a pressing need for new treatment options. A therapeutic vaccine could bolster the protective T H 1-mediated immune response, thereby slowing or halting the progression of chronic Chagas’ cardiomyopathy. Prior work in mice has demonstrated therapeutic efficacy of a Tc24 recombinant protein vaccine in the acute phase of Chagas disease. However, it is anticipated that humans will be vaccinated therapeutically when in the chronic phase of disease. This study investigates the therapeutic efficacy of a vaccine prototype containing recombinant protein Tc24, formulated with an emulsion containing the Toll-like receptor 4 agonist E6020 as an immunomodulatory adjuvant in a mouse model of chronic T . cruzi infection. Among outbred ICR mice vaccinated during chronic T . cruzi infection, there is a significant increase in the number of animals with undetectable systemic parasitemia (60% of vaccinated mice compared to 0% in the sham vaccine control group), and a two-fold reduction in cardiac fibrosis over the control group. The vaccinated mice produce a robust protective T H 1-biased immune response to the vaccine, as demonstrated by a significant increase in antigen-specific IFNγ-production, the number of antigen-specific IFNγ-producing cells, and IgG2a antibody titers. Importantly, therapeutic vaccination significantly reduced cardiac fibrosis in chronically infected mice. This is a first study demonstrating therapeutic efficacy of the prototype Tc24 recombinant protein and E6020 stable emulsion vaccine against cardiac fibrosis in a mouse model of chronic T . cruzi infection.

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The neglected tropical diseases (NTDs) are a group of 17 lesser known chronic infections which predominantly affect poor and disenfranchised communities. There are a number of NTDs that cause significant global morbidity in children, including the three major soil transmitted helminth (STH) infections (ascariasis, trichuriasis and hookworm infection), schistosomiasis and trachoma. These NTDs, together with lymphatic filariasis and onchocerciasis, are currently being targeted for global control and elimination through mass drug administration (MDA) campaigns. They represent the most common NTDs and share significant geographical overlap. Additionally, many individuals are polyparasitised with more than a single NTD. Integrated NTD control and elimination MDA programmes offer safe and efficacious treatments for all seven NTDs. However, the current global level of MDA coverage for the leading childhood NTDs, that is, STH infections, schistosomiasis and trachoma, remains well under 50%. Limiting factors for global coverage include insufficient global financial support, drug donation capacity of pharmaceutical companies and targeting school age children to the exclusion of other age groups in need of treatment, such as preschool age children. There is also a need for development of novel prevention and treatment modalities, such as next-generation small molecule drugs and vaccines. Efforts are underway to harness the momentum of a 2012 London Declaration on NTDs and a 2013 World Health Assembly (WHA) resolution as a means to control or in some cases eliminate by 2020 these NTDs that affect children worldwide.

Introduction Although emergency medicine (EM) training programmes have begun to be introduced in low- and middle-income countries (LMICs), minimal data exist on their effects on patient-centered outcomes in such settings. This study evaluated the impact of EM training and associated systems implementation on mortality among patients treated at the University Teaching Hospital-Kigali (UTH-K). Methods At UTH-K an EM post-graduate diploma programme was initiated in October 2013, followed by a residency-training programme in August 2015. Prior to October 2013, care was provided exclusively by general practice physicians (GPs); subsequently, care has been provided through mutually exclusive shifts allocated between GPs and EM trainees. Patients seeking Emergency Centre (EC) care during November 2012–October 2013 (pre-training) and August 2015–July 2016 (post-training) were eligible for inclusion. Data were abstracted from a random sample of records using a structured protocol. The primary outcomes were EC and overall hospital mortality. Mortality prevalence and risk differences (RD) were compared pre- and post-training. Magnitudes of effects were quantified using regression models to yield adjusted odds ratios (aOR) with 95% confidence intervals (CI). Results From 43,213 encounters, 3609 cases were assessed. The median age was 32 years with a male predominance (60.7%). Pre-training EC mortality was 6.3% (95% CI 5.3–7.5%), while post-training EC mortality was 1.2% (95% CI 0.7–1.8%), constituting a significant decrease in adjusted analysis (aOR = 0.07, 95% CI 0.03–0.17; p < 0.001). Pre-training overall hospital mortality was 12.2% (95% CI 10.9–13.8%). Post-training overall hospital mortality was 8.2% (95% CI 6.9–9.6%), resulting in a 43% reduction in mortality likelihood (aOR = 0.57, 95% CI 0.36–0.94; p = 0.016). Discussion In the studied population, EM training and systems implementation was associated with significant mortality reductions demonstrating the potential patient-centered benefits of EM development in resource-limited settings.

Diarrheal diseases lead to an estimated 1.3 million deaths each year, with the majority of those deaths occurring in patients over five years of age. As the severity of diarrheal disease can vary widely, accurately assessing dehydration status remains the most critical step in acute diarrhea management. The objective of this study is to empirically derive clinical diagnostic models for assessing dehydration severity in patients over five years with acute diarrhea in low resource settings. We enrolled a random sample of patients over five years with acute diarrhea presenting to the icddr,b Dhaka Hospital. Two blinded nurses independently assessed patients for symptoms/signs of dehydration on arrival. Afterward, consecutive weights were obtained to determine the percent weight change with rehydration, our criterion standard for dehydration severity. Full and simplified ordinal logistic regression models were derived to predict the outcome of none (<3%), some (3–9%), or severe (>9%) dehydration. The reliability and accuracy of each model were assessed. Bootstrapping was used to correct for over-optimism and compare each model’s performance to the current World Health Organization (WHO) algorithm. 2,172 patients were enrolled, of which 2,139 (98.5%) had complete data for analysis. The Inter-Class Correlation Coefficient (reliability) was 0.90 (95% CI = 0.87, 0.91) for the full model and 0.82 (95% CI = 0.77, 0.86) for the simplified model. The area under the Receiver-Operator Characteristic curve (accuracy) for severe dehydration was 0.79 (95% CI: 0.76–0.82) for the full model and 0.73 (95% CI: 0.70, 0.76) for the simplified model. The accuracy for both the full and simplified models were significantly better than the WHO algorithm (p<0.001). This is the first study to empirically derive clinical diagnostic models for dehydration severity in patients over five years. Once prospectively validated, the models may improve management of patients with acute diarrhea in low resource settings.