Detection and quantitation of colored deposit-forming <Emphasis Type="Italic">Meiothermus</Emphasis> spp. in paper industry processes and end products

Colored biofilms cause problems in paper industry. In this work we used real-time PCR to detect and to quantitate members of the genus Meiothermus from the process samples and end products from 24 machines manufacturing pulp, paper and board in four countries. The results obtained from 200 samples showed the importance of members of the genus Meiothermus as ubiquitous biofoulers in paper machines. This genus was the dominant biofouler in some mills. From ≤10⁴ to 10¹¹ copies of Meiothermus 16S rRNA genes were found per gram of process deposit (wet weight). Meiothermus spp. were found in paper and board products with colored defects and connection between deposit-forming microbes and end-product spots was shown. 16S rRNA gene sequences of 29 biofilm producing bacterial isolates from different mills were determined. Based on sequence data, 25 of the isolates were assigned to the genus Meiothermus, with Meiothermus silvanus and M. ruber as the most frequent species.     

Exercise enhancement by RGS14 disruption is mediated by brown adipose tissue

Enhanced exercise capacity is not only a feature of healthful aging, but also a therapy for aging patients and patients with cardiovascular disease. Disruption of the Regulator of G Protein Signaling 14 (RGS14) in mice extends healthful lifespan, mediated by increased brown adipose tissue (BAT). Accordingly, we determined whether RGS14 knockout (KO) mice exhibit enhanced exercise capacity and the role of BAT in mediating exercise capacity. Exercise was performed on a treadmill and exercise capacity was assessed by maximal running distance and work to exhaustion. Exercise capacity was measured in RGS14 KO mice and their wild types (WT), and also in WT mice with BAT transplantation from RGS14 KO mice or from other WT mice. RGS14 KO mice demonstrated 160 ± 9% increased maximal running distance and 154 ± 6% increased work to exhaustion, compared to WT mice. RGS14 KO BAT transplantation to WT mice, resulted in a reversal of phenotype, with the WT mice receiving the BAT transplant from RGS14 KO mice demonstrating 151 ± 5% increased maximal running distance and 158 ± 7% increased work to exhaustion, at three days after BAT transplantation, compared to RGS14 KO donors. BAT transplantation from WT to WT mice also resulted in increased exercise performance, but not at 3 days, but only at 8 weeks after transplantation. The BAT induced enhanced exercise capacity was mediated by (1) mitochondrial biogenesis and SIRT3; (2) antioxidant defense and the MEK/ERK pathway, and increased hindlimb perfusion. Thus, BAT mediates enhanced exercise capacity, a mechanism more powerful with RGS14 disruption.     

Fluctuations of Serum Neuron Specific Enolase and Protein S-100B Concentrations in Relation to the Use of Shunt during Carotid Endarterectomy

Objective

To evaluate the changes in serum neuron specific enolase and protein S-100B, after carotid endarterectomy performed using the conventional technique with routine shunting and patch closure, or eversion technique without the use of shunt.

Materials and Methods

Prospective non-randomized study included 43 patients with severe (>80%) carotid stenosis undergoing carotid endarterectomy in regional anesthesia. Patients were divided into two groups: conventional endarterectomy with routine use of shunt and Dacron patch (csCEA group) and eversion endarterectomy without the use of shunt (eCEA group). Protein S-100B and NSE concentrations were measured from peripheral blood before carotid clamping, after declamping and 24 hours after surgery.

Results

Neurologic examination and brain CT findings on the first postoperative day did not differ from preoperative controls in any patients. In csCEA group, NSE concentrations decreased after declamping (P<0.01), and 24 hours after surgery (P<0.01), while in the eCEA group NSE values slightly increased (P=ns), accounting for a significant difference between groups on the first postoperative day (P=0.006). In both groups S-100B concentrations significantly increased after declamping (P<0.05), returning to near pre-clamp values 24 hours after surgery (P=ns). Sub-group analysis revealed significant decline of serum NSE concentrations in asymptomatic patients shunted during surgery after declamping (P<0.05) and 24 hours after surgery (P<0.01), while no significant changes were noted in non-shunted patients (P=ns). Decrease of NSE serum levels was also found in symptomatic patients operated with the use of shunt on the first postoperative day (P<0.05). Significant increase in NSE serum levels was recorded in non-shunted symptomatic patients 24 hours after surgery (P<0.05).

Conclusion

Variations of NSE concentrations seemed to be influenced by cerebral perfusion alterations, while protein S-100B values were unaffected by shunting strategy. Routine shunting during surgery for symptomatic carotid stenosis may have the potential to prevent postoperative increase of serum NSE levels, a potential marker of brain injury.     

DNA2 drives processing and restart of reversed replication forks in human cells

Accurate processing of stalled or damaged DNA replication forks is paramount to genomic integrity and recent work points to replication fork reversal and restart as a central mechanism to ensuring high-fidelity DNA replication. Here, we identify a novel DNA2- and WRN-dependent mechanism of reversed replication fork processing and restart after prolonged genotoxic stress. The human DNA2 nuclease and WRN ATPase activities functionally interact to degrade reversed replication forks with a 5′-to-3′ polarity and promote replication restart, thus preventing aberrant processing of unresolved replication intermediates. Unexpectedly, EXO1, MRE11, and CtIP are not involved in the same mechanism of reversed fork processing, whereas human RECQ1 limits DNA2 activity by preventing extensive nascent strand degradation. RAD51 depletion antagonizes this mechanism, presumably by preventing reversed fork formation. These studies define a new mechanism for maintaining genome integrity tightly controlled by specific nucleolytic activities and central homologous recombination factors.     

Stochastic search variable selection based on two mixture components and continuous‐scale weighting

Stochastic search variable selection (SSVS) is a Bayesian variable selection method that employs covariate‐specific discrete indicator variables to select which covariates (e.g., molecular markers) are included in or excluded from the model. We present a new variant of SSVS where, instead of discrete indicator variables, we use continuous‐scale weighting variables (which take also values between zero and one) to select covariates into the model. The improved model performance is shown and compared to standard SSVS using simulated and real quantitative trait locus mapping datasets. The decision making to decide phenotype‐genotype associations in our SSVS variant is based on median of posterior distribution or using Bayes factors. We also show here that by using continuous‐scale weighting variables it is possible to improve mixing properties of Markov chain Monte Carlo sampling substantially compared to standard SSVS. Also, the separation of association signals and nonsignals (control of noise level) seems to be more efficient compared to the standard SSVS. Thus, the novel method provides efficient new framework for SSVS analysis that additionally provides whole posterior distribution for pseudo‐indicators which means more information and may help in decision making.     

Multi-step analysis as a tool for kinetic parameter estimation and mechanism discrimination in the reaction between tight-binding fasciculin 2 and electric eel acetylcholinesterase

The mechanism of action of a potent peptidic inhibitor fasciculin 2 (Fas2) on electric eel acetylcholinesterase (eleelAChE) has been examined in a three-level analysis. Classical steps included equilibration experiments for the evaluation of high affinity binding constant and the existence of residual hydrolytic activity in a solution of completely Fas2 saturated enzyme. The two rate constants for the association (k(on)) and the dissociation (k(off)) of Fas2 with free enzyme were determined by the time course of residual enzyme activity measurements. In the third step, with a nonclassical progress curve analysis, we found that the Fas2-enzyme complex exhibited hydrolytic activity in a butyrylcholinesterase-like kinetics. The switch appears to be a consequence of steric obstruction, but also the consequence of subtle rapid conformational changes around catalytic site, upon slow single-step binding of large Fas2 molecule at the peripheral site. An unusual unilateral effect of bound Fas2 is reflected by acylation-independent association and dissociation rates and might indeed be due to inability of small acylation agent to influence the binding of a large opponent.     

Improving Education in Medical Statistics: Implementing a Blended Learning Model in the Existing Curriculum

Background

Although recent studies report on the benefits of blended learning in improving medical student education, there is still no empirical evidence on the relative effectiveness of blended over traditional learning approaches in medical statistics. We implemented blended along with on-site (i.e. face-to-face) learning to further assess the potential value of web-based learning in medical statistics.

Methods

This was a prospective study conducted with third year medical undergraduate students attending the Faculty of Medicine, University of Belgrade, who passed (440 of 545) the final exam of the obligatory introductory statistics course during 2013–14. Student statistics achievements were stratified based on the two methods of education delivery: blended learning and on-site learning. Blended learning included a combination of face-to-face and distance learning methodologies integrated into a single course.

Results

Mean exam scores for the blended learning student group were higher than for the on-site student group for both final statistics score (89.36±6.60 vs. 86.06±8.48; p = 0.001) and knowledge test score (7.88±1.30 vs. 7.51±1.36; p = 0.023) with a medium effect size. There were no differences in sex or study duration between the groups. Current grade point average (GPA) was higher in the blended group. In a multivariable regression model, current GPA and knowledge test scores were associated with the final statistics score after adjusting for study duration and learning modality (p<0.001).

Conclusion

This study provides empirical evidence to support educator decisions to implement different learning environments for teaching medical statistics to undergraduate medical students. Blended and on-site training formats led to similar knowledge acquisition; however, students with higher GPA preferred the technology assisted learning format. Implementation of blended learning approaches can be considered an attractive, cost-effective, and efficient alternative to traditional classroom training in medical statistics.     

Mouse genetics identifies unique and overlapping functions of fibroblast growth factor receptors in keratinocytes

Fibroblast growth factors (FGFs) are key regulators of tissue development, homeostasis and repair, and abnormal FGF signalling is associated with various human diseases. In human and murine epidermis, FGF receptor 3 (FGFR3) activation causes benign skin tumours, but the consequences of FGFR3 deficiency in this tissue have not been determined. Here, we show that FGFR3 in keratinocytes is dispensable for mouse skin development, homeostasis and wound repair. However, the defect in the epidermal barrier and the resulting inflammatory skin disease that develops in mice lacking FGFR1 and FGFR2 in keratinocytes were further aggravated upon additional loss of FGFR3. This caused fibroblast activation and fibrosis in the FGFR1/FGFR2 double‐knockout mice and even more in mice lacking all three FGFRs, revealing functional redundancy of FGFR3 with FGFR1 and FGFR2 for maintaining the epidermal barrier. Taken together, our study demonstrates that FGFR1, FGFR2 and FGFR3 act together to maintain epidermal integrity and cutaneous homeostasis, with FGFR2 being the dominant receptor.     

Lipoproteins in Drosophila melanogaster-Assembly, Function, and Influence on Tissue Lipid Composition

Interorgan lipid transport occurs via lipoproteins, and altered lipoprotein levels correlate with metabolic disease. However, precisely how lipoproteins affect tissue lipid composition has not been comprehensively analyzed. Here, we identify the major lipoproteins of Drosophila melanogaster and use genetics and mass spectrometry to study their assembly, interorgan trafficking, and influence on tissue lipids. The apoB-family lipoprotein Lipophorin (Lpp) is the major hemolymph lipid carrier. It is produced as a phospholipid-rich particle by the fat body, and its secretion requires Microsomal Triglyceride Transfer Protein (MTP). Lpp acquires sterols and most diacylglycerol (DAG) at the gut via Lipid Transfer Particle (LTP), another fat body-derived apoB-family lipoprotein. The gut, like the fat body, is a lipogenic organ, incorporating both de novo-synthesized and dietary fatty acids into DAG for export. We identify distinct requirements for LTP and Lpp-dependent lipid mobilization in contributing to the neutral and polar lipid composition of the brain and wing imaginal disc. These studies define major routes of interorgan lipid transport in Drosophila and uncover surprising tissue-specific differences in lipoprotein lipid utilization.     

The comparative effects of metals on the hatching of earthworm cocoons

To establish the use of Metaphire posthuma as a sensitive model for ecotoxicological studies, the comparative effects of five metals on the hatching profiles of the cocoons of the earthworms, Metaphire posthuma, Eisenia foetida and Perionyx excavatus, were studied. The cocoons of the three species of earthworms were exposed to copper, chromium (III), chromium (VI), lead and zinc at 1.25, 2.5, 5.0 and 10.0 ppm. Viable cocoons were incubated at 20 +/- degrees C by using the immersion method. The results indicated that the inhibition of cocoon hatching was concentration dependent. The normal hatching, delayed hatching and non-viability of cocoons were recorded. At a concentration of 1.25 ppm, there was almost no effect on the hatching of the cocoons of all three species of earthworms, except when exposed to chromium (VI), but higher concentrations (2.5 and 5.0 ppm) caused severe effects. It was concluded that M. posthuma was more sensitive than the other two species, and that it is a suitable model for use in ecotoxicity testing.