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Physical activity (PA) built environments may support PA among rural youth and families. In the United States (U.S.), differences between rural and urban PA built environments are assessed using coarse scale, county-level methods. However, this method insufficiently examines environmental differences within rural counties. The present study uses rural-specific geospatial mapping techniques and a fine scale, within-rural grouping strategy to identify differing levels of access to the PA built environment among a rural sample. First, PA infrastructure variables (parks, sidewalks) within a rural region of the Midwest U.S. were mapped. Then, households (N = 112) of participants in the NU-HOME study, a childhood obesity prevention trial, were categorized to community-level and neighborhood-level PA built environment groups using two access indicators; Rural-Urban Commuting Area (RUCA) codes and Walk Scores®, respectively. Finally, households were categorized to new groups that combined community-level RUCA codes and neighborhood-level Walk Scores® to indicate the diverse ways in which rural families might access PA built environments, including by vehicle travel and pedestrian commuting. Household access to PA infrastructure (per geospatial proximity and density analyses), parent perceptions of the PA environment, and child PA were examined across the new combined access groups. All measures of household access to PA infrastructure significantly differed by group (p

MalariaGEN is a data-sharing network that enables groups around the world to work together on the genomic epidemiology of malaria. Here we describe a new release of curated genome variation data on 7,000 Plasmodium falciparum samples from MalariaGEN partner studies in 28 malaria-endemic countries. High-quality genotype calls on 3 million single nucleotide polymorphisms (SNPs) and short indels were produced using a standardised analysis pipeline. Copy number variants associated with drug resistance and structural variants that cause failure of rapid diagnostic tests were also analysed. Almost all samples showed genetic evidence of resistance to at least one antimalarial drug, and some samples from Southeast Asia carried markers of resistance to six commonly-used drugs. Genes expressed during the mosquito stage of the parasite life-cycle are prominent among loci that show strong geographic differentiation. By continuing to enlarge this open data resource we aim to facilitate research into the evolutionary processes affecting malaria control and to accelerate development of the surveillance toolkit required for malaria elimination.

MalariaGEN is a data-sharing network that enables groups around the world to work together on the genomic epidemiology of malaria. Here we describe a new release of curated genome variation data on 7,000 Plasmodium falciparum samples from MalariaGEN partner studies in 28 malaria-endemic countries. High-quality genotype calls on 3 million single nucleotide polymorphisms (SNPs) and short indels were produced using a standardised analysis pipeline. Copy number variants associated with drug resistance and structural variants that cause failure of rapid diagnostic tests were also analysed. Almost all samples showed genetic evidence of resistance to at least one antimalarial drug, and some samples from Southeast Asia carried markers of resistance to six commonly-used drugs. Genes expressed during the mosquito stage of the parasite life-cycle are prominent among loci that show strong geographic differentiation. By continuing to enlarge this open data resource we aim to facilitate research into the evolutionary processes affecting malaria control and to accelerate development of the surveillance toolkit required for malaria elimination.

Canine Chronic Ulcerative Stomatitis is a spontaneously occurring inflammatory disease of the oral mucosa. An immune-mediated pathogenesis is suspected though not yet proven. We have recently reported on the clinical and histologic features, and identification of select leukocyte cell populations within the lesion. A clinical and histologic similarity to oral lichen planus of people was proposed. In the present study, these initial observations are extended by examining lesions from 24 dogs with clinical evidence of chronic ulcerative stomatitis. Because dogs with chronic ulcerative stomatitis often have concurrent periodontal disease, we wondered if dental plaque/biofilm may be a common instigator of inflammation in both lesions. We hypothesized that dogs with chronic ulcerative stomatitis would exhibit a spectrum of pathologic changes and phenotype of infiltrating leukocytes that would inform lesion pathogenesis and that these changes would differ from inflammatory phenotypes in periodontitis. Previously we identified chronic ulcerative stomatitis lesions to be rich in FoxP3+ and IL17+ cells. As such, we suspect that these leukocytes play an important role in lesion pathogenesis. The current study confirms the presence of moderate to large numbers of FoxP3+ T cells and IL17+ cells in all ulcerative stomatitis lesions using confocal immunofluorescence. Interestingly, the majority of IL17+ cells were determined to be non-T cells and IL17+ cell frequencies were negatively correlated with severity on the clinical scoring system. Three histologic subtypes of ulcerative stomatitis were determined; lichenoid, deep stomatitis and granulomatous. Periodontitis lesions, like stomatitis lesions, were B cell and plasma cell rich, but otherwise differed from the stomatitis lesions. Direct immunofluorescence results did not support an autoantibody-mediated autoimmune disease process. This investigation contributes to the body of literature regarding leukocyte involvement in canine idiopathic inflammatory disease pathogenesis.

Positive parent-child attachment can be determined by opportunities for the child to interact with his/her parent and can influence a child's physical activity (PA) behavior. Therefore, an intervention that provides children and their parent more time to interact positively could impact children's PA. We examined the efficacy of a 12-week mother-daughter intervention on African-American girls' PA levels. In Spring of 2013 and 2014, mother-daughter dyads (n = 76) from Springfield, MA, were randomly assigned to one of three groups [child-mother (CH-M, n = 28), child alone (CH, n = 25), or control (CON, n = 23)] that participated in an afterschool culturally-tailored dance intervention (60 min/day, 3 days/week, 12 weeks). Girls in the CH-M group participated in the intervention with their maternal figure, while girls in the CH group participated in the intervention alone. CON group participants received weekly health-related newsletters. PA was assessed with accelerometers for seven days at baseline, 6-weeks, and 12-weeks. Hierarchical linear modeling was used to examine rates of change in PA. During the afterschool intervention time, girls in the CH-M group displayed a significantly steeper rate of increase in their percent time spent in vigorous PA compared to both the CON (γ = 0.80, p

BACKGROUND:Peroxisome proliferator-activated receptor gamma (PPAR-γ) is a nuclear receptor that modulates inflammation in models of asthma. To determine whether pioglitazone improves measures of asthma control and airway inflammation, we performed a single-center randomized, double-blind, placebo-controlled, parallel-group trial. METHODS:Sixty-eight participants with mild asthma were randomized to 12 weeks pioglitazone (30 mg for 4 weeks, then 45 mg for 8 weeks) or placebo. The primary outcome was the adjusted mean forced expiratory volume in one second (FEV1) at 12 weeks. The secondary outcomes were mean peak expiratory flow (PEF), scores on the Juniper Asthma Control Questionnaire (ACQ) and Asthma Quality of Life Questionnaire (AQLQ), fractional exhaled nitric oxide (FeNO), bronchial hyperresponsiveness (PD20), induced sputum counts, and sputum supernatant interferon gamma-inducible protein-10 (IP-10), vascular endothelial growth factor (VEGF), monocyte chemotactic protein-1 (MCP-1), and eosinophil cationic protein (ECP) levels. Study recruitment was closed early after considering the European Medicines Agency's reports of a potential increased risk of bladder cancer with pioglitazone treatment. Fifty-five cases were included in the full analysis (FA) and 52 in the per-protocol (PP) analysis. RESULTS:There was no difference in the adjusted FEV1 at 12 weeks (-0.014 L, 95% confidence interval [CI] -0.15 to 0.12, p = 0.84) or in any of the secondary outcomes in the FA. The PP analysis replicated the FA, with the exception of a lower evening PEF in the pioglitazone group (-21 L/min, 95% CI -39 to -4, p = 0.02). CONCLUSIONS:We found no evidence that treatment with 12 weeks of pioglitazone improved asthma control or airway inflammation in mild asthma. TRIAL REGISTRATION:ClinicalTrials.gov NCT01134835.

Our ability to engineer organisms with new biosynthetic pathways and genetic circuits is limited by the availability of protein characterization data and the cost of synthetic DNA. With new tools for reading and writing DNA, there are opportunities for scalable assays that more efficiently and cost effectively mine for biochemical protein characteristics. To that end, we have developed the Multiplex Library Synthesis and Expression Correction (MuLSEC) method for rapid assembly, error correction, and expression characterization of many genes as a pooled library. This methodology enables gene synthesis from microarray-synthesized oligonucleotide pools with a one-pot technique, eliminating the need for robotic liquid handling. Post assembly, the gene library is subjected to an ampicillin based quality control selection, which serves as both an error correction step and a selection for proteins that are properly expressed and folded in E. coli. Next generation sequencing of post selection DNA enables quantitative analysis of gene expression characteristics. We demonstrate the feasibility of this approach by building and testing over 90 genes for empirical evidence of soluble expression. This technique reduces the problem of part characterization to multiplex oligonucleotide synthesis and deep sequencing, two technologies under extensive development with projected cost reduction.

Deep Brain Stimulation (DBS) is increasingly used to treat a variety of brain diseases by sending electrical impulses to deep brain nuclei through long, electrically conductive leads. Magnetic resonance imaging (MRI) of patients pre- and post-implantation is desirable to target and position the implant, to evaluate possible side-effects and to examine DBS patients who have other health conditions. Although MRI is the preferred modality for pre-operative planning, MRI post-implantation is limited due to the risk of high local power deposition, and therefore tissue heating, at the tip of the lead. The localized power deposition arises from currents induced in the leads caused by coupling with the radiofrequency (RF) transmission field during imaging. In the present work, parallel RF transmission (pTx) is used to tailor the RF electric field to suppress coupling effects. Electromagnetic simulations were performed for three pTx coil configurations with 2, 4, and 8-elements, respectively. Optimal input voltages to minimize coupling, while maintaining RF magnetic field homogeneity, were determined for all configurations using a Nelder-Mead optimization algorithm. Resulting electric and magnetic fields were compared to that of a 16-rung birdcage coil. Experimental validation was performed with a custom-built 4-element pTx coil. In simulation, 95-99% reduction of the electric field at the tip of the lead was observed between the various pTx coil configurations and the birdcage coil. Maximal reduction in E-field was obtained with the 8-element pTx coil. Magnetic field homogeneity was comparable to the birdcage coil for the 4- and 8-element pTx configurations. In experiment, a temperature increase of 2±0.15°C was observed at the tip of the wire using the birdcage coil, whereas negligible increase (0.2±0.15°C) was observed with the optimized pTx system. Although further research is required, these initial results suggest that the concept of optimizing pTx to reduce DBS heating effects holds considerable promise.

The analysis of in vitro anti-malarial drug susceptibility testing is vulnerable to the effects of different statistical approaches and selection biases. These confounding factors were assessed with respect to pfmdr1 gene mutation and amplification in 490 clinical isolates. Two statistical approaches for estimating the drug concentration associated with 50% effect (EC50 ) were compared: the commonly used standard two-stage (STS) method, and nonlinear mixed-effects modelling. The in vitro concentration-effect relationships for, chloroquine, mefloquine, lumefantrine and artesunate, were derived from clinical isolates obtained from patients on the western border of Thailand. All isolates were genotyped for polymorphisms in the pfmdr1 gene. The EC50 estimates were similar for the two statistical approaches but 15-28% of isolates in the STS method had a high coefficient of variation (>15%) for individual estimates of EC50 and these isolates had EC50 values that were 32 to 66% higher than isolates derived with more precision. In total 41% (202/490) of isolates had amplification of pfmdr1 and single nucleotide polymorphisms were found in 50 (10%). Pfmdr1 amplification was associated with an increase in EC50 for mefloquine (139% relative increase in EC50 for 2 copies, 188% for 3+ copies), lumefantrine (82% and 75% for 2 and 3+ copies respectively) and artesunate (63% and 127% for 2 and 3+ copies respectively). In contrast pfmdr1 mutation at codons 86 or 1042 were associated with an increase in chloroquine EC50 (44-48%). Sample size calculations showed that to demonstrate an EC50 shift of 50% or more with 80% power if the prevalence was 10% would require 430 isolates and 245 isolates if the prevalence was 20%. In conclusion, although nonlinear mixed-effects modelling did not demonstrate any major advantage for determining estimates of anti-malarial drug susceptibility, the method includes all isolates, thereby, potentially improving confirmation of candidate molecular markers of anti-malarial drug susceptibility.

BackgroundSynthetic biological systems are currently created by an ad-hoc, iterative process of specification, design, and assembly. These systems would greatly benefit from a more formalized and rigorous specification of the desired system components as well as constraints on their composition. Therefore, the creation of robust and efficient design flows and tools is imperative. We present a human readable language (Eugene) that allows for the specification of synthetic biological designs based on biological parts, as well as provides a very expressive constraint system to drive the automatic creation of composite Parts (Devices) from a collection of individual Parts.ResultsWe illustrate Eugene's capabilities in three different areas: Device specification, design space exploration, and assembly and simulation integration. These results highlight Eugene's ability to create combinatorial design spaces and prune these spaces for simulation or physical assembly. Eugene creates functional designs quickly and cost-effectively.ConclusionsEugene is intended for forward engineering of DNA-based devices, and through its data types and execution semantics, reflects the desired abstraction hierarchy in synthetic biology. Eugene provides a powerful constraint system which can be used to drive the creation of new devices at runtime. It accomplishes all of this while being part of a larger tool chain which includes support for design, simulation, and physical device assembly.

Deflazacort slows the progress of Duchenne muscular dystrophy (DMD) with fewer side effects than prednisone. In mdx mice, deflazacort treatment augments repair and growth of new muscle fibers. We tested the hypothesis that deflazacort improves muscle function and promotes repair by increasing myogenic cell proliferation and fiber differentiation. mdx mice (3.5 weeks old) were treated with deflazacort (1.2 mg/kg) or vehicle for 4 weeks. Forelimb grip strength was measured. After 4 weeks, the right tibialis anterior muscle (TA) was crush injured to induce synchronous regeneration. DNA was labeled using different markers 24 and 2 h before collecting tissues 4 days after injury. The expression of creatine kinase (CK) isoforms, laminin-2 (merosin) mRNA and protein, and proliferation by myogenic cells were measured and satellite cells were identified by immunolocalization of c-met receptor. Peak grip strength increased 15% within 10 days of treatment, and was maintained up to 6 weeks after the end of treatment in a second experiment. Expression of CK MM in the regenerating TA rose from 46% to 55% of total CK activity after deflazacort treatment. Satellite cells were more numerous and appeared earlier on new fibers, in concert with a threefold increase in proliferation by myogenin+ (but not MyoD+) myoblasts. α2-Laminin mRNA expression and protein increased 1.3–5.5-fold relative to MM CK in regenerating and dystrophic TA, respectively. These studies support the hypothesis that deflazacort promotes functional gains, myogenic differentiation, myoblast fusion, and laminin expression in regenerating dystrophic muscle. The potential to augment precursor specification, strength, and possible membrane stability may be useful in directing long-term benefits for DMD patients and short-term amplification of precursors prior to myoblast transfer.

More than 60 zoonoses are linked to small mammals, including some of the most devastating pathogens in human history. Millions of museum-archived tissues are available to understand natural history of those pathogens. Our goal was to maximize the value of museum collections for pathogen-based research by using targeted sequence capture. We generated a probe panel that includes 39,916 80-bp RNA probes targeting 32 pathogen groups, including bacteria, helminths, fungi, and protozoans. Laboratory-generated, mock-control samples showed that we are capable of enriching targeted loci from pathogen DNA 2,882‒6,746-fold. We identified bacterial species in museum-archived samples, including Bartonella, a known human zoonosis. These results showed that probe-based enrichment of pathogens is a highly customizable and efficient method for identifying pathogens from museum-archived tissues.

Abstract Infrasound may be used to detect the approach of hazardous volcanic mudflows, known as lahars, tens of minutes before their flow fronts arrive. We have analyzed signals from more than 20 secondary lahars caused by precipitation events at Fuego Volcano during Guatemala’s rainy season in May through October of 2022. We are able to quantify the capabilities of infrasound monitoring through comparison with seismic data, time lapse camera imagery, and high-resolution video of a well-recorded event on August 17. We determine that infrasound sensors, deployed adjacent to the lahar path and in small-aperture (10 s of meters) arrays, are particularly sensitive to remote detection of lahars, including small-sized events, at distances of at least 5 km. At Fuego Volcano these detections could be used to provide timely alerts of up to 30 min before lahars arrive at a downstream monitoring site, such as in the frequently impacted Ceniza drainage. We propose that continuous infrasound monitoring, from locations adjacent to a drainage, may complement seismic monitoring and serve as a valuable tool to help identify approaching hazards. On the other hand, infrasound arrays located a kilometer or more from the lahar path can be effectively used to track a lahar’s progression.

The Construction File (CF) specification establishes a standardized interface for molecular biology operations, laying a foundation for automation and enhanced efficiency in experiment design. It is implemented across three distinct software projects: PyDNA_CF_Simulator, a Python project featuring a ChatGPT plugin for interactive parsing and simulating experiments; ConstructionFileSimulator, a field-tested Java project that showcases 'Experiment' objects expressed as flat files; and C6-Tools, a JavaScript project integrated with Google Sheets via Apps Script, providing a user-friendly interface for authoring and simulation of CF. The CF specification not only standardizes and modularizes molecular biology operations but also promotes collaboration, automation, and reuse, significantly reducing potential errors. The potential integration of CF with artificial intelligence, particularly GPT-4, suggests innovative automation strategies for synthetic biology. While challenges such as token limits, data storage, and biosecurity remain, proposed solutions promise a way forward in harnessing AI for experiment design. This shift from human-driven design to AI-assisted workflows, steered by high-level objectives, charts a potential future path in synthetic biology, envisioning an environment where complexities are managed more effectively.

OBJECTIVES/SPECIFIC AIMS: Inflammatory bowel disease (IBD) patients are at an increased risk of Clostridium difficile infection (CDI) but the impact of CDI on disease severity is unclear. The aim of this study was to determine the effect of CDI on long-term disease outcome in a cohort of IBD patients. METHODS/STUDY POPULATION: We analyzed patients enrolled in a prospective IBD natural history registry. Patients who tested positive at least once formed the CDI positive group. We generated a 2:1 propensity matched control cohort based on risk factors of CDI in the year before infection. Healthcare utilization data (emergency department use, subsequent hospitalizations, telephone encounters), medications, labs, disease activity, and quality of life metrics were temporally organized. RESULTS/ANTICIPATED RESULTS: A total of 198 patients (66 CDI, 132 matched controls) were included [56.6% female; 60.1% Crohn’s disease (CD), 39.9% ulcerative colitis (UC)]. Groups were not significantly different in the year before infection in all metrics but in the year of infection, having CDI was significantly associated with more steroid and antibiotic exposure, elevated C-reactive protein or erythrocyte sedimentation rate, and low vitamin D (all p