The emergence and spread of Zika virus (ZIKV) presented a challenge to the diagnosis of ZIKV infections in areas with transmission of dengue (DENV) and chikungunya (CHIKV) viruses. To facilitate ...detection of ZIKV infections, and differentiate these infections from DENV and CHIKV, we developed the Trioplex real-time RT-PCR assay (Trioplex assay). Here, we describe the optimization of multiplex and singleplex formats of the assay for a variety of chemistries and instruments to facilitate global standardization and implementation. We evaluated the analytical performance of all Trioplex modalities for detection of these three pathogens in serum and whole blood, and for ZIKV in urine. The limit of detection for the three viruses and in different RNA-extraction modalities is near 10
genome copy equivalents per milliliter (GCE/mL). Simultaneous testing of more than one specimen type from each patient provides a 6.4% additional diagnostic sensitivity. Overall, the high sensitivity of the Trioplex assay demonstrates the utility of this assay ascertaining Zika cases.
OBJECTIVES/GOALS: To introduce the new Team Science Community Toolkit, co-created by community and academic partners, and showcase its potential to empower Community Organizations (COs) in achieving ...equity in community-engaged research (CER). METHODS/STUDY POPULATION: In response to the challenges faced by COs in CER collaborations, qualitative interviews were conducted with CO staff from historically marginalized communities. These interviews informed the development of the Team Science Community Toolkit, a collaborative effort involving a Community Advisory Board (CAB) and Team Science experts from Northwestern University. The toolkit, designed using a community-based participatory research approach, incorporates the Science of Team Science and User-Centered Design principles. Integrated into the NIH-sponsored COALESCE website, it includes templates, checklists, and interactive tools, along with a real-world simulation, to support COs in all stages of the research process. RESULTS/ANTICIPATED RESULTS: Focus groups and usability testing involving external community experts validated the toolkit’s content and usability. Participants expressed enthusiasm and a sense of empowerment, indicating that the toolkit allows them to actively shape research processes and infuse their specific voices and needs into their partnerships. The toolkit is designed to support breaking down barriers like jargon and cultural adaptability to improve accessibility and open conversation. The impact of this Team Science focused toolkit is under evaluation. This presentation will showcase the toolkit, detail its collaborative development, and explore potential applications, ultimately offering a path to more equitable and valuable community-based research. DISCUSSION/SIGNIFICANCE: By providing COs with the resources and knowledge to participate as equal partners in research collaborations, it enhances self-advocacy, transparency, and equity. The toolkit has the potential to utilize Team Science to foster productive communication in community-academic research partnerships.
Megalencephaly-capillary malformation (MCAP) and megalencephaly-polymicrogyria-polydactyly-hydrocephalus (MPPH) syndromes are sporadic overgrowth disorders associated with markedly enlarged brain ...size and other recognizable features. We performed exome sequencing in 3 families with MCAP or MPPH, and our initial observations were confirmed in exomes from 7 individuals with MCAP and 174 control individuals, as well as in 40 additional subjects with megalencephaly, using a combination of Sanger sequencing, restriction enzyme assays and targeted deep sequencing. We identified de novo germline or postzygotic mutations in three core components of the phosphatidylinositol 3-kinase (PI3K)-AKT pathway. These include 2 mutations in AKT3, 1 recurrent mutation in PIK3R2 in 11 unrelated families with MPPH and 15 mostly postzygotic mutations in PIK3CA in 23 individuals with MCAP and 1 with MPPH. Our data highlight the central role of PI3K-AKT signaling in vascular, limb and brain development and emphasize the power of massively parallel sequencing in a challenging context of phenotypic and genetic heterogeneity combined with postzygotic mosaicism.
Tissue inhibitor of metalloproteinase (TIMP)-4 binds pro-matrix metalloproteinase (MMP)-2 and efficiently inhibits MT1-MMP, but unlike TIMP-2 neither forms a trimolecular complex nor supports ...pro-MMP-2 activation. To investigate the structural and functional differences between these two TIMPs, the C-terminal domains (C-TIMP-4 and C-TIMP-2) were expressed independently from their N domains and mutations were introduced into the C-terminal tails. Myoglobin was used as a novel recombinant fusion protein partner because spectroscopic measurement of the heme Soret absorbance at 408 nm readily enabled calculation of the molar equivalent of the red-colored recombinant protein, even in complex protein mixtures. Both C-TIMP-4 and C-TIMP-2 bound pro-MMP-2 and blocked concanavalin A-induced cellular activation of the enzyme. Measurement of k(on) rates revealed that the inhibition of MMP-2 by TIMP-4 is preceded by a C domain docking interaction, but in contrast to TIMP-2, this is not enhanced by a C-terminal tail interaction and so occurs at a slower rate. Indeed, the binding stability of C-TIMP-4 was unaltered by deletion of the C-terminal tail, but replacement with the tail of TIMP-2 increased its affinity for pro-MMP-2 by approximately 2-fold, as did substitution with the TIMP-2 C-terminal tail acidic residues in the tail of C-TIMP-4 (V193E/Q194D). Conversely, substitution of the C-terminal tail of C-TIMP-2 with that of TIMP-4 reduced pro-MMP-2 binding by approximately 75%, as did reduction of its acidic character by mutation to the corresponding TIMP-4 amino acid residues (E192V/D193Q). Together, this shows the importance of Glu(192) and Asp(193) in TIMP-2 binding to pro-MMP-2; the lack of these acidic residues in the TIMP-4 C-terminal tail, which reduces the stability of complex formation with the MMP-2 hemopexin C domain, probably precludes TIMP-4 from supporting the activation of pro-MMP-2.
Thesis (M.S.P.H.)--University of South Florida, 2004.
Title from PDF of title page. Document formatted into pages; contains 185 pages. Includes bibliographical references.
West Nile is an important novel virus in the United States, having spread rapidly since it was first detected in New York in 1999. The Centers for Disease Control and Prevention as well as many State ...Health Departments, have mandated programs for surveillance of West Nile Virus activity. These programs incorporate many different aspects including existing arboserology programs with additional testing for West Nile Virus and new plans that incorporate active and passive surveillance methods.
The objective of this study was to examine all aspects of the Florida West Nile surveillance program to determine if there was transmission in the animal systems prior to human cases. The predictive analyses were done using regional data graphs, spatial information, correlations and regression models.
Data for sentinel chickens, bird necropsy and mosquito pool surveillance from participating counties in Florida were obtained from the State of Florida surveillance database. The human data was obtained from the State of Florida reportable disease database for each county whether participating in the state surveillance programs or not. Clinical cases were examined by demographics (gender and age) and an incidence rate was calculated to demonstrate the effects of disease. Specific statistical methods used included Pearson's coefficient correlation, Poisson distribution regression modeling to show if any of the surveillance systems were predictors for human disease.
The incidence rate analysis for clinical cases showed clustering of cases in adjacent counties within a region where Florida's panhandle and adjacent counties northeast had the highest incidence. Florida's central and southern regions had moderate human incidence. This provides useful information in transmission geography for prevention and control measures. Demographic analysis showed that there were twice as many males than females diagnosed with West Nile in Florida, this was true across the groups as well. The highest number of cases was seen within the age group over 55 years of age for West Nile Neuroinvasive Disease and for West Nile Fever the highest number of cases was within the 36-54 age range.
The temporal distribution was determined using graphical representations of all of the surveillance types and clinical cases. In order to include all relevant data, the temporality was set from week 20 to week 52. This study found that all of the surveillance types (dead birds, mosquitoes and sentinels) offered a specialized strength for predicting clinical cases. However, mosquitoes proved to be the least efficient out of the three surveillance systems. The regional and spatial analysis showed that positive dead birds and sentinels provided the coverage for the surveillance systems in the state. However, Pearson's correlation coefficient was low for sentinel surveillance; this may be due to higher participation showing West Nile Virus activity in areas (especially rural) that have no reported human cases. This analysis did show that West Nile is detected in mosquito pool samples before it is detected in the dead bird or sentinel surveillance systems which provides an earlier warning for human cases. The Poisson distribution regression model was only useful for the pooled years and 2003. These showed that mosquitoes, positive dead birds and sentinels were good predictors for clinical cases for the combined years and dead birds and sentinels were significant for 2003 as well. The recommendations based on the results from this study would be to continue all the current surveillance efforts but with the following enhancements: 1. Increase the coverage and consistency of submissions for all surveillance types. 2. Set standard levels of participation for all counties based on the regional analyses and populations at risk. 3. Create standardized approaches for sampling, shipping and submitting samples (especially for mosquito pool submissions) and require that participating counties adhere to these standards. 4. Only submit specific birds known to be especially susceptible to West Nile Virus (e.g. corvids). 5. Targeted prevention and education strategies for higher risk groups based on their potential levels of exposure.
Obesity increases cardiovascular risk. However, the extent to which various measures of body composition are associated with abnormalities in cardiac structure and function, independent of ...comorbidities commonly affecting obese individuals, is not clear. This study sought to examine the relationship between body mass index, waist circumference, and percent body fat with conventional and advanced measures of cardiac structure and function.
We studied 4343 participants of the ARIC study (Atherosclerosis Risk in Communities) who were aged 69 to 82 years, free of coronary heart disease and heart failure, and underwent comprehensive echocardiography. Increasing body mass index, waist circumference, and body fat were associated with greater left ventricular (LV) mass and left atrial volume indexed to height(2.7) in both men and women (P<0.001). In women, all 3 measures were associated with abnormal LV geometry, and increasing waist circumference and body fat were associated with worse global longitudinal strain, a measure of LV systolic function. In both sexes, increasing body mass index was associated with greater right ventricular end-diastolic area and worse right ventricular fractional area change (P≤0.001). We observed similar associations for both waist circumference and percent body fat.
In a large, biracial cohort of older adults free of clinically overt coronary heart disease or heart failure, obesity was associated with subclinical abnormalities in cardiac structure in both men and women and with adverse LV remodeling and impaired LV systolic function in women. These data highlight the association of obesity and subclinical abnormalities of cardiac structure and function, particularly in women.
Effector T cells exhibiting features of either T helper 1 (Th1) or T follicular helper (Tfh) populations are essential to control experimental Plasmodium infection and are believed to be critical for ...resistance to clinical malaria. To determine whether Plasmodium-specific Th1- and Tfh-like effector cells generate memory populations that contribute to protection, we developed transgenic parasites that enable high-resolution study of anti-malarial memory CD4 T cells in experimental models. We found that populations of both Th1- and Tfh-like Plasmodium-specific memory CD4 T cells persist. Unexpectedly, Th1-like memory cells exhibit phenotypic and functional features of Tfh cells during recall and provide potent B cell help and protection following transfer, characteristics that are enhanced following ligation of the T cell co-stimulatory receptor OX40. Our findings delineate critical functional attributes of Plasmodium-specific memory CD4 T cells and identify a host-specific factor that can be targeted to improve resolution of acute malaria and provide durable, long-term protection against Plasmodium parasite re-exposure.
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•Reagents generated to study Plasmodium-specific memory CD4 T cell development•Th1- and Tfh-like memory CD4 T cells exhibit functional overlap during recall•Potent B cell helper function can be recalled from Th1-like memory cells•Functions of parasite-specific memory CD4 T cells are enhanced by ligation of OX40
Th1 CD4 T cells are widely described as terminally differentiated with a relatively reduced capacity to form memory or support humoral immunity. Using experimental malaria models, Zander et al. show that potent proliferative and B cell helper activity unexpectedly resides within the Plasmodium-specific Th1-like memory CD4 T cell compartment.
A meta-analysis study design was used to analyze 7 data sets of invasive and carriage pneumococcal isolates recovered from children, to determine whether invasive disease potential differs for each ...serotype and, if so, whether it has changed over time or differs geographically. Serotype- and serogroup-specific odds ratios (ORs) were calculated for each study and as a pooled estimate, with use of serotype 14 as the reference group. ORs varied widely: the serotypes with the highest ORs (1, 5, and 7) were 60-fold more invasive than those with the lowest ORs (3, 6A, and 15). There was a significant inverse correlation between invasive disease and carriage prevalence for the serotypes that we considered, which implies that the most invasive serotypes and serogroups were the least commonly carried and that the most frequently carried were the least likely to cause invasive disease. There was no evidence of any temporal change or major geographical differences in serotype- or serogroup-specific invasive disease potential.
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