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Detection of generic differential RNA processing events from RNA-seq data. - RNA biology
RNA-seq data analysis has revealed abundant alternative splicing in eukaryotic mRNAs. However, splicing is only one of many processing events that transcripts may undergo during their lifetime. We present here RNAprof (RNA profile analysis), a program for the detection of differential processing events from the comparison of RNA-seq experiments. RNAprof implements a specific gene-level normalization procedure and compares RNA-seq coverage profiles at nucleotide resolution to detect regions of significant coverage differences, independently of splice sites or other gene features. We used RNAprof to analyze the effect of alternative-splicing regulators NSRa and NSRb on the Arabidopsis thaliana transcriptome. A number of intron retention events and alternative transcript structures were specifically detected by RNAprof and confirmed by qRT-PCR. Further tests using a public Mus musculus RNA-seq dataset and comparisons with other RNA isoform predictors showed that RNAprof uniquely identified sets of highly significant processing events as well as other relevant library-specific differences in RNA-seq profiles. This highlights an important layer of variation that remains undetected by current protocols for RNA-seq analysis.
Ex-Vivo percutaneous absorption of enrofloxacin: Comparison of LMOG organogel vs. pentravan cream. - International journal of pharmaceutics
The objective of this study was to investigate the percutaneous absorption of enrofloxacin from two base formulations, Pentravan(Â®) cream and LMOG organogel. Ex-vivo experiments were carried out on pig ear skin. The percutaneous permeation through pig skin of two formulations containing 5wt% of enrofloxacin was measured and compared using Franz diffusion cells. At appropriate intervals up to 120h, diffusion samples were taken and analyzed using HPLC assays. Permeation profiles were established and the parameters Tlag and flux values were calculated. In this ex-vivo study, the flux values were 0.35Î¼gcm(-2)h(-1) for Pentravan(Â®) and 1.22Î¼gcm(-2)h(-1) for LMOG organogel, corresponding respectively to 7.9 % and 29.3 % of enrofloxacin absorbed after 120h by these formulations. The lag time (Tlag) of Pentravan(Â®) and organogel were 6.32 and 0.015h respectively. The absorption time to reach the antibiotic concentration of enrofloxacin (2Î¼gmL(-1)) in the receptor was 60h with Pentravan(Â®) and 30h with the organogel, suggesting more effective treatment by the latter. Enrofloxacin contained in organogel could be absorbed through pig ear skin 3.7 times greater than that in Pentravan(Â®) (commercial formulation). This study demonstrates the perspective of organogel formulations as potential drug delivery systems.Copyright Â© 2015 Elsevier B.V. All rights reserved.
Micronutrient Deficiency Control in Vietnam from Policy and Research to Implementation: Keys for Success, Challenges and Lessons Learned. - Journal of nutritional science and vitaminology
Micronutrient deficiencies are still a public health problem in Vietnam. The Government of Vietnam has taken several steps to improve the situation through issuing supportive policy documents over the last several decades. Food fortification is an important complementary strategy to help bridge the nutrient gap in the population. Currently technical regulations are in place and food fortification is taking place on a voluntary basis, along with other complementary targeted programs including home fortification of complementary foods with micronutrient powders and a communications campaign to reach adolescent girls. These have been built on innotative partnerships with industries on a voluntary, market basis. Other innovative targeted nutrition programs are also being piloted, including a micronutrient supplement project in four provinces and a campain to reach adolescent girls through sports. High level political commitment and resources is a crucial element to scale up in Vietnam. A micronutrient survey planned in 2015 will help provide the evidence to support a possible mandatory decree on food fortification. Vietnam has built a solid foundation in order to scale up its national food fortification program in the future to reach the majority of the population with improved intakes of iron, vitamin A, zinc, and iodine.
Transcription Factor SomA Is Required for Adhesion, Development and Virulence of the Human Pathogen Aspergillus fumigatus. - PLoS pathogens
The transcription factor Flo8/Som1 controls filamentous growth in Saccharomyces cerevisiae and virulence in the plant pathogen Magnaporthe oryzae. Flo8/Som1 includes a characteristic N-terminal LUG/LUH-Flo8-single-stranded DNA binding (LUFS) domain and is activated by the cAMP dependent protein kinase A signaling pathway. Heterologous SomA from Aspergillus fumigatus rescued in yeast flo8 mutant strains several phenotypes including adhesion or flocculation in haploids and pseudohyphal growth in diploids, respectively. A. fumigatus SomA acts similarly to yeast Flo8 on the promoter of FLO11 fused with reporter gene (LacZ) in S. cerevisiae. FLO11 expression in yeast requires an activator complex including Flo8 and Mfg1. Furthermore, SomA physically interacts with PtaB, which is related to yeast Mfg1. Loss of the somA gene in A. fumigatus resulted in a slow growth phenotype and a block in asexual development. Only aerial hyphae without further differentiation could be formed. The deletion phenotype was verified by a conditional expression of somA using the inducible Tet-on system. A adherence assay with the conditional somA expression strain indicated that SomA is required for biofilm formation. A ptaB deletion strain showed a similar phenotype supporting that the SomA/PtaB complex controls A. fumigatus biofilm formation. Transcriptional analysis showed that SomA regulates expression of genes for several transcription factors which control conidiation or adhesion of A. fumigatus. Infection assays with fertilized chicken eggs as well as with mice revealed that SomA is required for pathogenicity. These data corroborate a complex control function of SomA acting as a central factor of the transcriptional network, which connects adhesion, spore formation and virulence in the opportunistic human pathogen A. fumigatus.
RNA Enrichment Method for Quantitative Transcriptional Analysis of Pathogens In Vivo Applied to the Fungus Candida albicans. - mBio
In vivo transcriptional analyses of microbial pathogens are often hampered by low proportions of pathogen biomass in host organs, hindering the coverage of full pathogen transcriptome. We aimed to address the transcriptome profiles of Candida albicans, the most prevalent fungal pathogen in systemically infected immunocompromised patients, during systemic infection in different hosts. We developed a strategy for high-resolution quantitative analysis of the C.Â albicans transcriptome directly from early and late stages of systemic infection in two different host models, mouse and the insect Galleria mellonella. Our results show that transcriptome sequencing (RNA-seq) libraries were enriched for fungal transcripts up to 1,600-fold using biotinylated bait probes to capture C.Â albicans sequences. This enrichment biased the read counts of only ~3% of the genes, which can be identified and removed based on a priori criteria. This allowed an unprecedented resolution of C.Â albicans transcriptome in vivo, with detection of over 86% of its genes. The transcriptional response of the fungus was surprisingly similar during infection of the two hosts and at the two time points, although some host- and time point-specific genes could be identified. Genes that were highly induced during infection were involved, for instance, in stress response, adhesion, iron acquisition, and biofilm formation. Of the in vivo-regulated genes, 10% are still of unknown function, and their future study will be of great interest. The fungal RNA enrichment procedure used here will help a better characterization of the C.Â albicans response in infected hosts and may be applied to other microbial pathogens.Understanding the mechanisms utilized by pathogens to infect and cause disease in their hosts is crucial for rational drug development. Transcriptomic studies may help investigations of these mechanisms by determining which genes are expressed specifically during infection. This task has been difficult so far, since the proportion of microbial biomass in infected tissues is often extremely low, thus limiting the depth of sequencing and comprehensive transcriptome analysis. Here, we adapted a technology to capture and enrich C.Â albicans RNA, which was next used for deep RNA sequencing directly from infected tissues from two different host organisms. The high-resolution transcriptome revealed a large number of genes that were so far unknown to participate in infection, which will likely constitute a focus of study in the future. More importantly, this method may be adapted to perform transcript profiling of any other microbes during host infection or colonization.Copyright Â© 2015 Amorim-Vaz et al.
Has the employment status of people living with HIV changed since the early 2000s? - AIDS (London, England)
In a context marked by major changes in the field of HIV and in the general socioeconomic context, this study aimed at investigating changes in the employment situation of people living with HIV (PLWHIV) in France since the early 2000s.Repeated cross-sectional survey among two nationally representative samples of PLWHIV followed at hospital in France in 2003 (Nâ€Š=â€Š2932) and 2011 (Nâ€Š=â€Š3022).Differences between 2003 and 2011 in (1) rates of employment, unemployment, and inactivity and (2) rates of work cessation and access to work following HIV diagnosis were measured using two-step multivariate Poisson regression models adjusted for individual sociodemographic determinants of position on the labor market, and then additionally for health status characteristics.Overall, among working-age PLWHIV 60.9% in 2003 and 59.6% in 2011 were employed; 12.6 and 15.8%, respectively, were unemployed; and 26.5 and 24.6%, respectively, were inactive. Adjusting for sociodemographic determinants of position on the labor market, while employment rate was not different in 2011 compared with 2003, inactivity rate significantly decreased (adjusted prevalence rate ratio: 0.83, 95% confidence interval: [0.72-0.96]) and unemployment rate significantly increased (1.28 [1.04-1.57]). After additional adjustment for health status characteristics, the difference was no longer significant for inactivity (0.89 [0.77-1.03]) but remained significant for unemployment (1.55 [1.24-1.93]). Compared with 2003, in 2011 transitions to unemployment following HIV diagnosis tended to be more frequent (1.58 [0.92-2.68]) and access to work was significantly less frequent (0.57 [0.33-0.99]).Improvements in HIV care have not translated into improvements in PLWHIV's situation regarding employment.
Disease-Specific Survival with Spindle Cell Carcinoma of the Head and Neck. - Otolaryngology--head and neck surgery : official journal of American Academy of Otolaryngology-Head and Neck Surgery
(1) Determine factors influencing survival in patients diagnosed with spindle cell carcinoma (SpCC), a rare variant of head and neck squamous cell carcinoma (SCC). (2) Compare survival of patients with SpCC to those with conventional SCC.Retrospective cohort study.Surveillance, Epidemiology, and End Results 18 database (years 2004-2009).Among patients receiving treatment for a single primary in the oral cavity, oropharynx, hypopharynx, or larynx, 118 subjects with SpCC and 18,298 subjects with SCC were identified with complete data for the variables of age, sex, grade, tumor size, stage group, and TNM stage. Disease-specific survival curves were compared. Univariate and multivariate analyses were used to examine the effects of each factor on survival over all sites and within each of 3 sites.Univariate analysis of the combination of the 3 anatomic subsites showed survival with SpCC was worse than with conventional SCC (P < .001). Three-year disease-specific survival with SpCC was 49.5%, and 5-year disease-specific survival was 40.2%. Compared with conventional SCC, survival was worse for SpCC of the oral cavity (P < .001) and oropharynx (P < .001) but no different for the larynx and hypopharynx site (P = .15). Multivariate analysis identified age (P = .02), tumor size (P = .006), and M stage (P < .001) as the only variables significantly affecting survival with SpCC. All variables significantly affected survival with conventional SCC.Spindle cell carcinoma carries a worse prognosis than SCC. Larger tumor size, older age, and metastatic disease portend worse survival with SpCC of the head and neck.Â© American Academy of Otolaryngologyâ€”Head and Neck Surgery Foundation 2015.
Antimicrobial resistance in Helicobacter pylori: current situation and management strategy in Vietnam. - Journal of infection in developing countries
Increasing antimicrobial resistance to key antibiotics in Helicobacter pylori has become a main cause of treatment failures in many countries, including Vietnam. For this reason it is advisable to perform antimicrobial sensitivity tests to provide more focused regimens for H. pylori eradication. However, this approach is generally unavailable for H. pylori in Vietnam and the selection of treatment regimens is mainly based on the trend of antibiotic use in the population, resistance development in the region, and history of H. pylori eradication of patients. The aim of this review is to examine the current situation of antimicrobial resistance in Vietnam and suggest management strategies for treatment selection.
Validation and utilization of an internally controlled multiplex Real-time RT-PCR assay for simultaneous detection of enteroviruses and enterovirus A71 associated with hand foot and mouth disease. - Virology journal
Hand foot and mouth disease (HFMD) is a disease of public health importance across the Asia-Pacific region. The disease is caused by enteroviruses (EVs), in particular enterovirus A71 (EV-A71). In EV-A71-associated HFMD, the infection is sometimes associated with severe manifestations including neurological involvement and fatal outcome. The availability of a robust diagnostic assay to distinguish EV-A71 from other EVs is important for patient management and outbreak response.We developed and validated an internally controlled one-step single-tube real-time RT-PCR in terms of sensitivity, linearity, precision, and specificity for simultaneous detection of EVs and EV-A71. Subsequently, the assay was then applied on throat and rectal swabs sampled from 434 HFMD patients.The assay was evaluated using both plasmid DNA and viral RNA and has shown to be reproducible with a maximum assay variation of 4.41Â % and sensitive with a limit of detection less than 10 copies of target template per reaction, while cross-reactivity with other EV serotypes was not observed. When compared against a published VP1 nested RT-PCR using 112 diagnostic throat and rectal swabs from 112 children with a clinical diagnosis of HFMD during 2014, the multiplex assay had a higher sensitivity and 100Â % concordance with sequencing results which showed EVs in 77/112 (68.8Â %) and EV-A71 in 7/112 (6.3Â %). When applied to clinical diagnostics for 322 children, the assay detected EVs in throat swabs of 257/322 (79.8Â %) of which EV-A71 was detected in 36/322 (11.2Â %) children. The detection rate increased to 93.5Â % (301/322) and 13.4Â % (43/322) for EVs and EV-A71, respectively, when rectal swabs from 65 throat-negative children were further analyzed.We have successfully developed and validated a sensitive internally controlled multiplex assay for rapid detection of EVs and EV-A71, which is useful for clinical management and outbreak control of HFMD.
Geometric and Electronic Structures for MnS2(-/0) Clusters by Interpreting the Anion Photoelectron Spectrum with Quantum Chemical Calculations. - The journal of physical chemistry. A
Geometric and electronic structures of linear SMnS, cyclic Î·(2)-MnS2, and linear Î·(1)-MnS2 isomers of MnS2(-) clusters have been investigated with B3LYP, CCSD(T), and NEVPT2 methods. The ground state of the anionic cluster is determined as (5)Î g of the linear SMnS(-) isomer, while the ground state of the neutral cluster may be either the (4)Î£g(-) of the same isomer or the (6)A1 of the Î·(2)-MnS2 cluster. The experimental photoelectron spectrum of the MnS2(-) cluster is interpreted by contributions of these two isomers. The high-intensity band at a binding energy of 2.94 eV is attributed to the (5)Î g â†’ (4)Î£g(-) transition between the linear SMnS(-/0) clusters. The lower energy feature in the spectrum at binding energies between 1.9 and 2.8 eV and exhibiting a low intensity, is ascribed to electron detachments within the less stable Î·(2)-MnS2(-/0) clusters. Ionizations from the lowest energy (7)A1 state of this isomer to the neutral (6)A1, (6)A2, (8)A2, and (6)B2 states are responsible for this part of the spectrum. The extreme low intensity part between 1.3 and 1.9 eV can be due to excited states of either SMnS(-) or Î·(2)-MnS2(-).
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