Thermostable Branched-Chain Amino Acid Transaminases From the Archaea Geoglobus acetivorans and Archaeoglobus fulgidus: Biochemical and Structural Characterization

This is the final version. Available on open access from Frontiers Media via the DOI in this recordTwo new thermophilic branched chain amino acid transaminases have been identified within the genomes of different hyper-thermophilic archaea, Geoglobus acetivorans, and Archaeoglobus fulgidus. These enzymes belong to the class IV of transaminases as defined by their structural fold. The enzymes have been cloned and over-expressed in Escherichia coli and the recombinant enzymes have been characterized both biochemically and structurally. Both enzymes showed high thermostability with optimal temperature for activity at 80 and 85°C, respectively. They retain good activity after exposure to 50% of the organic solvents, ethanol, methanol, DMSO and acetonitrile. The enzymes show a low activity to (R)-methylbenzylamine but no activity to (S)-methylbenzylamine. Both enzymes have been crystallized and their structures solved in the internal aldimine form, to 1.9 Å resolution for the Geoglobus enzyme and 2.0 Å for the Archaeoglobus enzyme. Also the Geoglobus enzyme structure has been determined in complex with the amino acceptor α-ketoglutarate and the Archaeoglobus enzyme in complex with the inhibitor gabaculine. These two complexes have helped to determine the conformation of the enzymes during enzymatic turnover and have increased understanding of their substrate specificity. A comparison has been made with another (R) selective class IV transaminase from the fungus Nectria haematococca which was previously studied in complex with gabaculine. The subtle structural differences between these enzymes has provided insight regarding their different substrate specificities.Biotechnology & Biological Sciences Research Council (BBSRC

A study of prognostic factors in Chinese patients with diabetic foot ulcers

Objective: Few studies have identified factors as predictors of clinical prognosis of patients with diabetic foot ulcers (DFUs), especially of Chinese patients. In this study, we assessed the prognostic factors of Chinese patients with DFUs. Methods and materials: This was a retrospective study (January 2009–January 2011) of 194 DFUs conducted in an inpatient population at PLA 454 Hospital in Nanjing, China, to determine the prognostic influential factors of DFUs in Chinese patients. All of the studied patients were grouped into an amputation group, a non-healing group, and a cured group, according to the clinical prognosis. Patient parameters, including gender, age, smoking habits, education level, family history of diabetes mellitus, medical history, duration of foot lesions and complications, ankle-brachial index (ABI), transcutaneous oxygen pressure (TcPO2), urinary albumin/creatinine ratio (Alb/Cr), fundus oculi, electrocardiogram, DFU characteristics, bacterial nature, and neuropathy, were cross-studied among the three groups. Results: Compared with the other two groups, the amputation group showed a higher number of males, older in age, lower ABI and TcPO2 levels, higher Wagner wound grading and size, and significantly higher urinary Alb/Cr ratio, blood urea nitrogen, serum creatinine, white blood cell count, and erythrocyte sedimentation rate. Compared to the cured group (162 patients), more patients with an older age, smoking, family history of diabetes mellitus, medical history of foot ulcerations, lower ABI and TcPO2 levels, higher urine Alb/Cr ratio, and serum creatinine were found in the non-healing group. Regression analysis was used to study the correlation between various factors and clinical prognosis, and the results were as follows: age, Wagner wound classification, and heel ulcerations were negatively correlated to the DFU prognosis, whereas the female population, ABI, and TcPO2 were positively correlated with DFU prognosis. Conclusion: In this retrospective study, we conclude that the DFU prognosis may be related to age, gender, wound location (heel), Wagner wound classification, ABI, and TcPO2 levels in the Chinese population

Extensive Copy-Number Variation of Young Genes across Stickleback Populations

MM received funding from the Max Planck innovation funds for this project. PGDF was supported by a Marie Curie European Reintegration Grant (proposal nr 270891). CE was supported by German Science Foundation grants (DFG, EI 841/4-1 and EI 841/6-1). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

Diagnostic accuracy of existing methods for identifying diabetic foot ulcers from inpatient and outpatient datasets

<p>Abstract</p> <p>Background</p> <p>As the number of persons with diabetes is projected to double in the next 25 years in the US, an accurate method of identifying diabetic foot ulcers in population-based data sources are ever more important for disease surveillance and public health purposes. The objectives of this study are to evaluate the accuracy of existing methods and to propose a new method.</p> <p>Methods</p> <p>Four existing methods were used to identify all patients diagnosed with a foot ulcer in a Department of Veterans Affairs (VA) hospital from the inpatient and outpatient datasets for 2003. Their electronic medical records were reviewed to verify whether the medical records positively indicate presence of a diabetic foot ulcer in diagnoses, medical assessments, or consults. For each method, five measures of accuracy and agreement were evaluated using data from medical records as the gold standard.</p> <p>Results</p> <p>Our medical record reviews show that all methods had sensitivity > 92% but their specificity varied substantially between 74% and 91%. A method used in Harrington et al. (2004) was the most accurate with 94% sensitivity and 91% specificity and produced an annual prevalence of 3.3% among VA users with diabetes nationwide. A new and simpler method consisting of two codes (707.1× and 707.9) shows an equally good accuracy with 93% sensitivity and 91% specificity and 3.1% prevalence.</p> <p>Conclusions</p> <p>Our results indicate that the Harrington and New methods are highly comparable and accurate. We recommend the Harrington method for its accuracy and the New method for its simplicity and comparable accuracy.</p

Transgenerational Stress Memory Is Not a General Response in Arabidopsis

Adverse conditions can trigger DNA damage as well as DNA repair responses in plants. A variety of stress factors are known to stimulate homologous recombination, the most accurate repair pathway, by increasing the concentration of necessary enzymatic components and the frequency of events. This effect has been reported to last into subsequent generations not exposed to the stress. To establish a basis for a genetic analysis of this transgenerational stress memory, a broad range of treatments was tested for quantitative effects on homologous recombination in the progeny. Several Arabidopsis lines, transgenic for well-established recombination traps, were exposed to 10 different physical and chemical stress treatments, and scored for the number of somatic homologous recombination (SHR) events in the treated generation as well as in the two subsequent generations that were not treated. These numbers were related to the expression level of genes involved in homologous recombination and repair. SHR was enhanced after the majority of treatments, confirming previous data and adding new effective stress types, especially interference with chromatin. Compounds that directly modify DNA stimulated SHR to values exceeding previously described induction rates, concomitant with an induction of genes involved in SHR. In spite of the significant stimulation in the stressed generations, the two subsequent non-treated generations only showed a low and stochastic increase in SHR that did not correlate with the degree of stimulation in the parental plants. Transcripts coding for SHR enzymes generally returned to pre-treatment levels in the progeny. Thus, transgenerational effects on SHR frequency are not a general response to abiotic stress in Arabidopsis and may require special conditions

Reliability of a tool for measuring theory of planned behaviour constructs for use in evaluating research use in policymaking

<p>Abstract</p> <p>Background</p> <p>Although measures of knowledge translation and exchange (KTE) effectiveness based on the theory of planned behavior (TPB) have been used among patients and providers, no measure has been developed for use among health system policymakers and stakeholders. A tool that measures the intention to use research evidence in policymaking could assist researchers in evaluating the effectiveness of KTE strategies that aim to support evidence-informed health system decision-making. Therefore, we developed a 15-item tool to measure four TPB constructs (intention, attitude, subjective norm and perceived control) and assessed its face validity through key informant interviews.</p> <p>Methods</p> <p>We carried out a reliability study to assess the tool's internal consistency and test-retest reliability. Our study sample consisted of 62 policymakers and stakeholders that participated in deliberative dialogues. We assessed internal consistency using Cronbach's alpha and generalizability (G) coefficients, and we assessed test-retest reliability by calculating Pearson correlation coefficients (<it>r</it>) and G coefficients for each construct and the tool overall.</p> <p>Results</p> <p>The internal consistency of items within each construct was good with alpha ranging from 0.68 to alpha = 0.89. G-coefficients were lower for a single administration (G = 0.34 to G = 0.73) than for the average of two administrations (G = 0.79 to G = 0.89). Test-retest reliability coefficients for the constructs ranged from <it>r </it>= 0.26 to <it>r </it>= 0.77 and from G = 0.31 to G = 0.62 for a single administration, and from G = 0.47 to G = 0.86 for the average of two administrations. Test-retest reliability of the tool using G theory was moderate (G = 0.5) when we generalized across a single observation, but became strong (G = 0.9) when we averaged across both administrations.</p> <p>Conclusion</p> <p>This study provides preliminary evidence for the reliability of a tool that can be used to measure TPB constructs in relation to research use in policymaking. Our findings suggest that the tool should be administered on more than one occasion when the intervention promotes an initial 'spike' in enthusiasm for using research evidence (as it seemed to do in this case with deliberative dialogues). The findings from this study will be used to modify the tool and inform further psychometric testing following different KTE interventions.</p

Polyploidization as a Retraction Force in Plant Genome Evolution: Sequence Rearrangements in Triticale

BACKGROUND: Polyploidization is a major evolutionary process in plants where hybridization and chromosome doubling induce enormous genomic stress and can generate genetic and epigenetic modifications. However, proper evaluation of DNA sequence restructuring events and the precise characterization of sequences involved are still sparse. METHODOLOGY/PRINCIPAL FINDINGS: Inter Retrotransposons Amplified Polymorphism (IRAP), Retrotransposons Microsatellite Amplified Polymorphism (REMAP) and Inter Simple Sequence Repeat (ISSR) largely confirmed the absence of any intraspecific variation in wheat, rye and triticale. The comparative analysis of banding profiles between wheat and rye inbred lines revealed 34% of monomorphic (common to both parental species) bands for the ten different primer combinations used. The analysis of triticale plants uncovered nearly 51% of rearranged bands in the polyploid, being the majority of these modifications, due to the loss of rye bands (83%). Sequence analysis of rye fragments absent in triticale revealed for instance homology with hydroxyproline-rich glycoproteins (HRGP), a protein that belongs to a major family of inducible defence response proteins. Conversely, a wheat-specific band absent in triticale comprises a nested structure of copia-like retrotransposons elements, namely Claudia and Barbara. Sequencing of a polyploid-specific band (absent in both parents) revealed a microsatellite related sequence. Cytological studies using Fluorescent In Situ Hybridization (FISH) with REMAP products revealed a widespread distribution of retrotransposon and/or microsatellite flanking sequences on rye chromosomes, with a preferential accumulation in heterochromatic sub-telomeric domains. CONCLUSIONS/SIGNIFICANCE: Here, we used PCR-based molecular marker techniques involving retrotransposons and microsatellites to uncover polyploidization induced genetic restructuring in triticale. Sequence analysis of rearranged genomic fragments either from rye or wheat origin showed these to be retrotransposon-related as well as coding sequences. Further FISH analysis revealed possible chromosome hotspots for sequence rearrangements. The role of chromatin condensation on the origin of genomic rearrangements mediated by polyploidization in triticale is also discussed

Challenges of self-reported medical conditions and electronic medical records among members of a large military cohort

<p>Abstract</p> <p>Background</p> <p>Self-reported medical history data are frequently used in epidemiological studies. Self-reported diagnoses may differ from medical record diagnoses due to poor patient-clinician communication, self-diagnosis in the absence of a satisfactory explanation for symptoms, or the "health literacy" of the patient.</p> <p>Methods</p> <p>The US Department of Defense military health system offers a unique opportunity to evaluate electronic medical records with near complete ascertainment while on active duty. This study compared 38 self-reported medical conditions to electronic medical record data in a large population-based US military cohort. The objective of this study was to better understand challenges and strengths in self-reporting of medical conditions.</p> <p>Results</p> <p>Using positive and negative agreement statistics for less-prevalent conditions, near-perfect negative agreement and moderate positive agreement were found for the 38 diagnoses.</p> <p>Conclusion</p> <p>This report highlights the challenges of using self-reported medical data and electronic medical records data, but illustrates that agreement between the two data sources increases with increased surveillance period of medical records. Self-reported medical data may be sufficient for ruling out history of a particular condition whereas prevalence studies may be best served by using an objective measure of medical conditions found in electronic healthcare records. Defining medical conditions from multiple sources in large, long-term prospective cohorts will reinforce the value of the study, particularly during the initial years when prevalence for many conditions may still be low.</p

Cost-of-Illness Analysis of Type 2 Diabetes Mellitus in Iran

Diabetes is a worldwide high prevalence chronic progressive disease that poses a significant challenge to healthcare systems. The aim of this study is to provide a detailed economic burden of diagnosed type 2 diabetes mellitus (T2DM) and its complications in Iran in 2009 year.This is a prevalence-based cost-of-illness study focusing on quantifying direct health care costs by bottom-up approach. Data on inpatient hospital services, outpatient clinic visits, physician services, drugs, laboratory test, education and non-medical cost were collected from two national registries. The human capital approach was used to calculate indirect costs separately in male and female and also among different age groups.The total national cost of diagnosed T2DM in 2009 is estimated at 3.78 billion USA dollars (USD) including 2.04±0.28 billion direct (medical and non-medical) costs and indirect costs of 1.73 million. Average direct and indirect cost per capita was 842.6±102 and 864.8 USD respectively. Complications (48.9%) and drugs (23.8%) were main components of direct cost. The largest components of medical expenditures attributed to diabetes's complications are cardiovascular disease (42.3% of total Complications cost), nephropathy (23%) and ophthalmic complications (14%). Indirect costs include temporarily disability (335.7 million), permanent disability (452.4 million) and reduced productivity due to premature mortality (950.3 million).T2DM is a costly disease in the Iran healthcare system and consume more than 8.69% of total health expenditure. In addition to these quantified costs, T2DM imposes high intangible costs on society in terms of reduced quality of life. Identification of effective new strategies for the control of diabetes and its complications is a public health priority

Tubule-Guided Cell-to-Cell Movement of a Plant Virus Requires Class XI Myosin Motors

Cell-to-cell movement of plant viruses occurs via plasmodesmata (PD), organelles that evolved to facilitate intercellular communications. Viral movement proteins (MP) modify PD to allow passage of the virus particles or nucleoproteins. This passage occurs via several distinct mechanisms one of which is MP-dependent formation of the tubules that traverse PD and provide a conduit for virion translocation. The MP of tubule-forming viruses including Grapevine fanleaf virus (GFLV) recruit the plant PD receptors called Plasmodesmata Located Proteins (PDLP) to mediate tubule assembly and virus movement. Here we show that PDLP1 is transported to PD through a specific route within the secretory pathway in a myosin-dependent manner. This transport relies primarily on the class XI myosins XI-K and XI-2. Inactivation of these myosins using dominant negative inhibition results in mislocalization of PDLP and MP and suppression of GFLV movement. We also found that the proper targeting of specific markers of the Golgi apparatus, the plasma membrane, PD, lipid raft subdomains within the plasma membrane, and the tonoplast was not affected by myosin XI-K inhibition. However, the normal tonoplast dynamics required myosin XI-K activity. These results reveal a new pathway of the myosin-dependent protein trafficking to PD that is hijacked by GFLV to promote tubule-guided transport of this virus between plant cells