cDNA-RNA subtractive hybridization reveals increased expression of mycocerosic acid synthase in intracellular Mycobacterium bovis BCG.

Identifying genes that are differentially expressed by Mycobacterium bovis BCG after phagocytosis by macrophages will facilitate the understanding of the molecular mechanisms of host cell-intracellular pathogen interactions. To identify such genes a cDNA-total RNA subtractive hybridization strategy has been used that circumvents the problems both of limited availability of bacterial RNA from models of infection and the high rRNA backgrounds in total bacterial RNA. The subtraction products were used to screen a high-density gridded Mycobacterium tuberculosis genomic library. Sequence data were obtained from 19 differential clones, five of which contained overlapping sequences for the gene encoding mycocerosic acid synthase (mas). Mas is an enzyme involved in the synthesis of multi-methylated long-chain fatty acids that are part of phthiocerol dimycocerosate, a major component of the complex mycobacterial cell wall. Northern blotting and primer extension data confirmed up-regulation of mas in intracellular mycobacteria and also revealed a putative extended -10 promoter structure and a long untranslated upstream region 5' of the mas transcripts, containing predicted double-stranded structures. Furthermore, clones containing overlapping sequences for furB, groEL-2, rplE and fadD28 were identified and the up-regulation of these genes was confirmed by Northern blot analysis. The cDNA-RNA subtractive hybridization enrichment and high density gridded library screening, combined with selective extraction of bacterial mRNA represents a valuable approach to the identification of genes expressed during intra-macrophage residence for bacteria such as M. bovis BCG and the pathogenic mycobacterium, M. tuberculosis

Methods for Calibration of Prout-Tompkins Kinetics Parameters Using EZM Iteration and GLO

This document contains information regarding the standard procedures used to calibrate chemical kinetics parameters for the extended Prout-Tompkins model to match experimental data. Two methods for calibration are mentioned: EZM calibration and GLO calibration. EZM calibration matches kinetics parameters to three data points, while GLO calibration slightly adjusts kinetic parameters to match multiple points. Information is provided regarding the theoretical approach and application procedure for both of these calibration algorithms. It is recommended that for the calibration process, the user begin with EZM calibration to provide a good estimate, and then fine-tune the parameters using GLO. Two examples have been provided to guide the reader through a general calibrating process

Large-Scale First-Principles Molecular Dynamics Simulations on the BlueGene/L Platform using the Qbox Code

First-Principles Molecular Dynamics (FPMD) is an accurate, atomistic simulation approach that is routinely applied to a variety of areas including solid-state physics, chemistry, biochemistry and nanotechnology. FPMD enables one to perform predictive materials simulations, as no empirical or adjustable parameters are used to describe a given system. Instead, a quantum mechanical description of electrons is obtained by solving the Kohn-Sham equations within a pseudopotential plane-wave formalism. This rigorous first-principles treatment of electronic structure is computationally expensive and limits the size of tractable systems to a few hundred atoms on most currently available parallel computers. Developed specifically for large parallel systems at LLNL's Center for Applied Scientific Computing, the Qbox implementation of the FPMD method shows unprecedented performance and scaling on BlueGene/L

Refinement type contracts for verification of scientific investigative software

Our scientific knowledge is increasingly built on software output. User code which defines data analysis pipelines and computational models is essential for research in the natural and social sciences, but little is known about how to ensure its correctness. The structure of this code and the development process used to build it limit the utility of traditional testing methodology. Formal methods for software verification have seen great success in ensuring code correctness but generally require more specialized training, development time, and funding than is available in the natural and social sciences. Here, we present a Python library which uses lightweight formal methods to provide correctness guarantees without the need for specialized knowledge or substantial time investment. Our package provides runtime verification of function entry and exit condition contracts using refinement types. It allows checking hyperproperties within contracts and offers automated test case generation to supplement online checking. We co-developed our tool with a medium-sized ($\approx$3000 LOC) software package which simulates decision-making in cognitive neuroscience. In addition to helping us locate trivial bugs earlier on in the development cycle, our tool was able to locate four bugs which may have been difficult to find using traditional testing methods. It was also able to find bugs in user code which did not contain contracts or refinement type annotations. This demonstrates how formal methods can be used to verify the correctness of scientific software which is difficult to test with mainstream approaches

BlueGene/L Applications: Parallelism on a Massive Scale

BlueGene/L (BG/L), developed through a partnership between IBM and Lawrence Livermore National Laboratory (LLNL), is currently the world's largest system both in terms of scale with 131,072 processors and absolute performance with a peak rate of 367 TFlop/s. BG/L has led the Top500 list the last four times with a Linpack rate of 280.6 TFlop/s for the full machine installed at LLNL and is expected to remain the fastest computer in the next few editions. However, the real value of a machine like BG/L derives from the scientific breakthroughs that real applications can produce by successfully using its unprecedented scale and computational power. In this paper, we describe our experiences with eight large scale applications on BG/L from several application domains, ranging from molecular dynamics to dislocation dynamics and turbulence simulations to searches in semantic graphs. We also discuss the challenges we faced when scaling these codes and present several successful optimization techniques. All applications show excellent scaling behavior, even at very large processor counts, with one code even achieving a sustained performance of more than 100 TFlop/s, clearly demonstrating the real success of the BG/L design

Clinical safety assessment of oral higenamine supplementation in healthy, young men

Objective: Higenamine, an herbal agent also known as norcoclaurine, is thought to stimulate β-androgenic receptors and possess lipolytic activity. It is currently making its way into the dietary supplement market. To our knowledge, no studies have been conducted to determine the safety profile of oral higenamine when used alone and in conjunction with other commonly used lipolytic agents. Methods: Forty-eight men were assigned to ingest either a placebo, higenamine, caffeine, or higenamine + caffeine + yohimbe bark extract daily for a period of 8 weeks. Before and after 4 and 8 weeks of supplementation, the following variables were measured: resting respiratory rate, heart rate, blood pressure, urinalysis, complete blood count, metabolic panel, liver enzyme activity, and lipid panel. Results: No interaction effects were noted for any variable (p \u3e 0.05), with no changes of statistical significance occurring across time for any of the four conditions (p \u3e 0.05). Conclusion: This is the first study to determine the safety profile of oral higenamine intake in human infjects. Our data indicate that 8 weeks of daily higenamine supplementation, either alone or in conjunction with caffeine and yohimbe bark extract, does not result in a statistically significant change in any of the measured outcome variables. Additional studies, inclusive of a larger sample size, are needed to extend these initial findings

Comparison of a Restricted and Unrestricted Vegan Diet Plan with a Restricted Omnivorous Diet Plan on Health-Specific Measures

Background: We have previously noted beneficial health outcomes when individuals follow a dietary restriction plan in accordance with the Daniel Fast (DF). This is true whether individuals eliminate all animal products or include small amounts of meat and dairy in their plan. The present study sought to compare anthropometric and biochemical measures of health in individuals following a traditional DF (i.e., restricted vegan) or modified DF (i.e., restricted omnivorous; inclusive of ad libitum meat and skim milk consumption), with those following an unrestricted vegan diet plan. Methods: 35 subjects (six men; 29 women; 33 ± 2 years; range: 18–67 years) completed a 21-day diet plan. Subjects reported to the lab for pre- (day 1) and post-intervention testing (day 22) in a 10 h fasted state. Blood samples were collected and assayed for complete blood count, metabolic panel, lipid panel, insulin, HOMA-IR, C-reactive protein, and oxidative stress biomarkers (malondialdehyde, advanced oxidation protein products, and nitrate/nitrite). Heart rate and blood pressure were measured and body composition was determined via dual energy X-ray absorptiometry. Subjects’ self-reported compliance, mental and physical health, and satiety in relation to the dietary modification were recorded. Results: No interaction effects were noted for our outcome measures (p > 0.05). However, subjects in the traditional DF group reported an approximate 10% increase in perceived mental and physical health, with a 25% reduction in malondialdehyde and a 33% reduction in blood insulin. Systolic BP was reduced approximately 7 mmHg in subjects assigned to the traditional DF, with an approximate 5 mmHg reduction in subjects assigned to the modified DF and the unrestricted vegan plan. A small (2 mmHg) reduction in diastolic BP was noted for subjects in both DF groups; a slight increase in diastolic BP was noted for subjects assigned to the unrestricted vegan group. An approximate 20% reduction was noted in total and LDL cholesterol for subjects in the traditional DF group, with an approximate 10% decrease for subjects in the modified DF group. No decrease in total or LDL cholesterol was noted for subjects in the unrestricted vegan group. Conclusion: These data indicate that both a traditional or modified DF may improve blood pressure and blood lipids in a clinically meaningful manner if these results are sustained over the long term. A traditional DF also results in a significant reduction in blood insulin and oxidative stress. An unrestricted vegan diet may improve systolic blood pressure, but in the absence of measures to strictly monitor adherence, it does not favorably impact other markers of health measured in the present study