'PACLIMS': A component LIM system for high-throughput functional genomic analysis

BACKGROUND: Recent advances in sequencing techniques leading to cost reduction have resulted in the generation of a growing number of sequenced eukaryotic genomes. Computational tools greatly assist in defining open reading frames and assigning tentative annotations. However, gene functions cannot be asserted without biological support through, among other things, mutational analysis. In taking a genome-wide approach to functionally annotate an entire organism, in this application the ~11,000 predicted genes in the rice blast fungus (Magnaporthe grisea), an effective platform for tracking and storing both the biological materials created and the data produced across several participating institutions was required. RESULTS: The platform designed, named PACLIMS, was built to support our high throughput pipeline for generating 50,000 random insertion mutants of Magnaporthe grisea. To be a useful tool for materials and data tracking and storage, PACLIMS was designed to be simple to use, modifiable to accommodate refinement of research protocols, and cost-efficient. Data entry into PACLIMS was simplified through the use of barcodes and scanners, thus reducing the potential human error, time constraints, and labor. This platform was designed in concert with our experimental protocol so that it leads the researchers through each step of the process from mutant generation through phenotypic assays, thus ensuring that every mutant produced is handled in an identical manner and all necessary data is captured. CONCLUSION: Many sequenced eukaryotes have reached the point where computational analyses are no longer sufficient and require biological support for their predicted genes. Consequently, there is an increasing need for platforms that support high throughput genome-wide mutational analyses. While PACLIMS was designed specifically for this project, the source and ideas present in its implementation can be used as a model for other high throughput mutational endeavors

\u27PACLIMS\u27: a component LIM system for high-throughput functional genomic analysis

BACKGROUND: Recent advances in sequencing techniques leading to cost reduction have resulted in the generation of a growing number of sequenced eukaryotic genomes. Computational tools greatly assist in defining open reading frames and assigning tentative annotations. However, gene functions cannot be asserted without biological support through, among other things, mutational analysis. In taking a genome-wide approach to functionally annotate an entire organism, in this application the approximately 11,000 predicted genes in the rice blast fungus (Magnaporthe grisea), an effective platform for tracking and storing both the biological materials created and the data produced across several participating institutions was required. RESULTS: The platform designed, named PACLIMS, was built to support our high throughput pipeline for generating 50,000 random insertion mutants of Magnaporthe grisea. To be a useful tool for materials and data tracking and storage, PACLIMS was designed to be simple to use, modifiable to accommodate refinement of research protocols, and cost-efficient. Data entry into PACLIMS was simplified through the use of barcodes and scanners, thus reducing the potential human error, time constraints, and labor. This platform was designed in concert with our experimental protocol so that it leads the researchers through each step of the process from mutant generation through phenotypic assays, thus ensuring that every mutant produced is handled in an identical manner and all necessary data is captured. CONCLUSION: Many sequenced eukaryotes have reached the point where computational analyses are no longer sufficient and require biological support for their predicted genes. Consequently, there is an increasing need for platforms that support high throughput genome-wide mutational analyses. While PACLIMS was designed specifically for this project, the source and ideas present in its implementation can be used as a model for other high throughput mutational endeavors

High risk behavior for HIV transmission among former injecting drug users: a survey from Indonesia

Contains fulltext : 88347.pdf (publisher's version ) (Open Access)BACKGROUND: Injecting drug use is an increasingly important cause of HIV transmission in most countries worldwide, especially in eastern Europe, South America, and east and southeast Asia. Among people actively injecting drugs, provision of clean needles and opioid substitution reduce HIV-transmission. However, former injecting drug users (fIDUs) are often overlooked as a high risk group for HIV transmission. We compared HIV risk behavior among current and former injecting drug users (IDUs) in Indonesia, which has a rapidly growing HIV-epidemic largely driven by injecting drug use. METHODS: Current and former IDUs were recruited by respondent driven sampling in an urban setting in Java, and interviewed regarding drug use and HIV risk behavior using the European Addiction Severity Index and the Blood Borne Virus Transmission Questionnaire. Drug use and HIV transmission risk behavior were compared between current IDUs and former IDUs, using the Mann-Whitney and Pearson Chi-square test. RESULTS: Ninety-two out of 210 participants (44%) were self reported former IDUs. Risk behavior related to sex, tattooing or piercing was common among current as well as former IDUs, 13% of former IDUs were still exposed to contaminated injecting equipment. HIV-infection was high among former (66%) and current (60%) IDUs. CONCLUSION: Former IDUs may contribute significantly to the HIV-epidemic in Indonesia, and HIV-prevention should therefore also target this group, addressing sexual and other risk behavior

Risk to human health related to the presence of perfluoroalkyl substances in food

Acknowledgements: The Panel wishes to thank the following for their support provided to this scientific output as Hearing experts: Klaus Abraham, Esben Budtz-Jørgensen, Tony Fletcher, Philippe Grandjean, Hans Mielke and Hans Rumke and EFSA staff members: Davide Arcella, Marco Binaglia, Petra Gergelova, Elena Rovesti and Marijke Schutte. The Panel wishes to acknowledge all European competent institutions, Member State bodies and other organisations that provided data for this scientific output. The Panel would also like to thank the following authors and co-authors for providing additional information in relation to their respective studies: Berit Granum, Margie M Peden-Adams, Thomas Webster.Peer reviewedPublisher PD

Psychosocial impact of undergoing prostate cancer screening for men with BRCA1 or BRCA2 mutations.

OBJECTIVES: To report the baseline results of a longitudinal psychosocial study that forms part of the IMPACT study, a multi-national investigation of targeted prostate cancer (PCa) screening among men with a known pathogenic germline mutation in the BRCA1 or BRCA2 genes. PARTICPANTS AND METHODS: Men enrolled in the IMPACT study were invited to complete a questionnaire at collaborating sites prior to each annual screening visit. The questionnaire included sociodemographic characteristics and the following measures: the Hospital Anxiety and Depression Scale (HADS), Impact of Event Scale (IES), 36-item short-form health survey (SF-36), Memorial Anxiety Scale for Prostate Cancer, Cancer Worry Scale-Revised, risk perception and knowledge. The results of the baseline questionnaire are presented. RESULTS: A total of 432 men completed questionnaires: 98 and 160 had mutations in BRCA1 and BRCA2 genes, respectively, and 174 were controls (familial mutation negative). Participants' perception of PCa risk was influenced by genetic status. Knowledge levels were high and unrelated to genetic status. Mean scores for the HADS and SF-36 were within reported general population norms and mean IES scores were within normal range. IES mean intrusion and avoidance scores were significantly higher in BRCA1/BRCA2 carriers than in controls and were higher in men with increased PCa risk perception. At the multivariate level, risk perception contributed more significantly to variance in IES scores than genetic status. CONCLUSION: This is the first study to report the psychosocial profile of men with BRCA1/BRCA2 mutations undergoing PCa screening. No clinically concerning levels of general or cancer-specific distress or poor quality of life were detected in the cohort as a whole. A small subset of participants reported higher levels of distress, suggesting the need for healthcare professionals offering PCa screening to identify these risk factors and offer additional information and support to men seeking PCa screening

Development and Validation of a 3-Dimensional Flexible Laryngoscopy Training Simulator

The objective of this study was to validate the use of a 3-Dimensional Flexible Laryngoscopy Training Simulator. This is a simulation device development and validation study. Anonymized CT scan data from a head/neck CT of a patient with normal anatomy was imported and a head/neck digital model was created. A 3D simulation model was printed using a stiff (Stratasys Vero) and flexible (Stratasys Agilus) material combination with a ShoreA hardness value of 60. Novices and experts were instructed and provided 5 trials to pass the laryngoscope. The videos of the first and the last trial were recorded and evaluated by three different evaluators. Performances were measured by the amount of time spent and precision of the task. Repeated measures of ANOVA and generalized linear model with binomial proportion was used were utilized to analyze the data. The post training scores were statistically significantly higher than pre training scores (Mean: 15.57 vs. 13.01, p  <0.0001) controlling for trainee experience. The time taken to complete a successful pass post training was statistically significantly lesser than pre training (Mean: 62.55 secs vs. 36.36 secs, p-value = 0.0007) controlling for individual’s experience. The odds of becoming skilled at the task was 4 times higher post training in comparison to pre training, controlling for individual’s experience (OR: 4.05, p-value: 0.0026). The 3-Dimensional Flexible Laryngoscopy Training Simulator is a valid trainer for both novice and experienced individuals. The simulator can improve technical skill performance and is critical for medical training

Development and Validation of a 3-Dimensional Flexible Laryngoscopy Training Simulator

The objective of this study was to validate the use of a 3-Dimensional Flexible Laryngoscopy Training Simulator. This is a simulation device development and validation study. Anonymized CT scan data from a head/neck CT of a patient with normal anatomy was imported and a head/neck digital model was created. A 3D simulation model was printed using a stiff (Stratasys Vero) and flexible (Stratasys Agilus) material combination with a ShoreA hardness value of 60. Novices and experts were instructed and provided 5 trials to pass the laryngoscope. The videos of the first and the last trial were recorded and evaluated by three different evaluators. Performances were measured by the amount of time spent and precision of the task. Repeated measures of ANOVA and generalized linear model with binomial proportion was used were utilized to analyze the data. The post training scores were statistically significantly higher than pre training scores (Mean: 15.57 vs. 13.01, p  <0.0001) controlling for trainee experience. The time taken to complete a successful pass post training was statistically significantly lesser than pre training (Mean: 62.55 secs vs. 36.36 secs, p-value = 0.0007) controlling for individual’s experience. The odds of becoming skilled at the task was 4 times higher post training in comparison to pre training, controlling for individual’s experience (OR: 4.05, p-value: 0.0026). The 3-Dimensional Flexible Laryngoscopy Training Simulator is a valid trainer for both novice and experienced individuals. The simulator can improve technical skill performance and is critical for medical training

Development and Validation of a 3-Dimensional Flexible Laryngoscopy Training Simulator

The objective of this study was to validate the use of a 3-Dimensional Flexible Laryngoscopy Training Simulator. This is a simulation device development and validation study. Anonymized CT scan data from a head/neck CT of a patient with normal anatomy was imported and a head/neck digital model was created. A 3D simulation model was printed using a stiff (Stratasys Vero) and flexible (Stratasys Agilus) material combination with a ShoreA hardness value of 60. Novices and experts were instructed and provided 5 trials to pass the laryngoscope. The videos of the first and the last trial were recorded and evaluated by three different evaluators. Performances were measured by the amount of time spent and precision of the task. Repeated measures of ANOVA and generalized linear model with binomial proportion was used were utilized to analyze the data. The post training scores were statistically significantly higher than pre training scores (Mean: 15.57 vs. 13.01, p  <0.0001) controlling for trainee experience. The time taken to complete a successful pass post training was statistically significantly lesser than pre training (Mean: 62.55 secs vs. 36.36 secs, p-value = 0.0007) controlling for individual’s experience. The odds of becoming skilled at the task was 4 times higher post training in comparison to pre training, controlling for individual’s experience (OR: 4.05, p-value: 0.0026). The 3-Dimensional Flexible Laryngoscopy Training Simulator is a valid trainer for both novice and experienced individuals. The simulator can improve technical skill performance and is critical for medical training

A Role for Mycobacterium tuberculosis Sigma Factor C in Copper Nutritional Immunity

Sigma factor C (SigC) contributes to Mycobacterium tuberculosis virulence in various animal models, but the stress response coordinated by this transcription factor was undefined. The results presented here indicate that SigC prevents copper starvation. Whole genome expression studies demonstrate short-term (4-h) induction of sigC, controlled from a tetracycline-inducible promoter, upregulates ctpB and genes in the nonribosomal peptide synthase (nrp) operon. These genes are expressed at higher levels after 48-h sigC induction, but also elevated are genes encoding copper-responsive regulator RicR and RicR-regulated copper toxicity response operon genes rv0846–rv0850, suggesting prolonged sigC induction results in excessive copper uptake. No growth and global transcriptional differences are observed between a sigC null mutant relative to its parent strain in 7H9 medium. In a copper-deficient medium, however, growth of the sigC deletion strain lags the parent, and 40 genes (including those in the nrp operon) are differentially expressed. Copper supplementation reverses the growth defect and silences most transcriptional differences. Together, these data support SigC as a transcriptional regulator of copper acquisition when the metal is scarce. Attenuation of sigC mutants in severe combined immunodeficient mice is consistent with an inability to overcome innate host defenses that sequester copper ions to deprive invading microbes of this essential micronutrient