Environmental metagenomics: An innovative resource for industrial biocatalysis

The current existing enzymes have been identified from cultivable micro-organisms, most frequently from bacteria. These bacterial biocatalytic capabilities have been widely used for biotransformations, resulting in the development of profitable industrial bioprocesses in the fields of feed and food processing, textiles, agro-chemistry, cosmetics, pharmaceuticals and fine chemistry. However, the originality of this bioresource is progressively drying up, while requests from industry for novel biocatalytic activities are increasing in the face of economic and environmental pressure. Metagenomics, through access to the huge reservoir of uncultivated bacteria which represents the majority of the present biodiversity, opens the door to new industrial sources of enzymes. Surmounting hurdles encountered with this technology (e.g. DNA extraction to obtain high quality DNA libraries with proper statistical representativity, setting up of relevant high throughput screenings assays, combining functional and genome-based identifications), gives unique opportunities to access novel biocatalysts that better fit with the required industrial specifications, thus providing new biocatalysis tool boxes.     

CC2D1A is a regulator of ESCRT-III CHMP4B

Endosomal sorting complexes required for transport (ESCRTs) regulate diverse processes ranging from receptor sorting at endosomes to distinct steps in cell division and budding of some enveloped viruses. Common to all processes is the membrane recruitment of ESCRT-III that leads to membrane fission. Here, we show that CC2D1A is a novel regulator of ESCRT-III CHMP4B function. We demonstrate that CHMP4B interacts directly with CC2D1A and CC2D1B with nanomolar affinity by forming a 1:1 complex. Deletion mapping revealed a minimal CC2D1A-CHMP4B binding construct, which includes a short linear sequence within the third DM14 domain of CC2D1A. The CC2D1A binding site on CHMP4B was mapped to the N-terminal helical hairpin. Based on a crystal structure of the CHMP4B helical hairpin, two surface patches were identified that interfere with CC2D1A interaction as determined by surface plasmon resonance. Introducing these mutations into a C-terminal truncation of CHMP4B that exerts a potent dominant negative effect on human immunodeficiency virus type 1 budding revealed that one of the mutants lost this effect completely. This suggests that the identified CC2D1A binding surface might be required for CHMP4B polymerization, which is consistent with the finding that CC2D1A binding to CHMP4B prevents CHMP4B polymerization in vitro. Thus, CC2D1A might act as a negative regulator of CHMP4B function.     

Improving Microstructured TiO2 Photoanodes for Dye Sensitized Solar Cells by Simple Surface Treatment

TiCl₄ surface treatment studies of porous electrode structure of TiO₂ aggregates synthesized using an acidic precursor and CTAB as a templating agent are carried out in order to understand and improve upon recombination kinetics in the photonanode film matrix, together with enhancing the intrinsic light scattering. The key beneficial features of the photoanode included high surface roughness, necessary for superior dye adsorption, nanocrystallite aggregates leading to diffuse light scattering within the film matrix, and a hierarchical macro‐ and mesopore structure allowing good access of electrolyte to the dye, thereby assisting in dye regeneration (enhanced charge transfer). Pre‐treatment of the TiO₂ electrodes reduced recombination at the fluorine‐doped tin oxide (FTO)/electrolyte interface. The post‐treatment study showed enhanced surface roughness through the deposition of a thin overlayer of amorphous TiO₂ on the film structure. This led to a notable improvement in both dye adsorption and inherent light scattering effects by the TiO₂ aggregates, resulting in enhanced energy harvesting. The thin TiO₂ overlayer also acted as a barrier in a core‐shell configuration within the porous TiO₂ matrix, and thereby reduced recombination. This allowed the hierarchical macro‐ and mesoporosity of the film matrix to be utilized more effectively for enhanced charge transfer during dye regeneration. Post‐treatment of the aggregated TiO₂ matrix resulted in a 36% enhancement in power conversion efficiency from 4.41% of untreated cells to 6.01%.     

Evaluation of Elevated Mean Pulmonary Arterial Pressure Based on Magnetic Resonance 4D Velocity Mapping: Comparison of Visualization Techniques

Purpose

Three-dimensional (3D) magnetic resonance phase contrast imaging (PC-MRI) allows non-invasive diagnosis of pulmonary hypertension (PH) and estimation of elevated mean pulmonary arterial pressure (mPAP) based on vortical motion of blood in the main pulmonary artery. The purpose of the present study was to compare the presence and duration of PH-associated vortices derived from different flow visualization techniques with special respect to their performance for non-invasive assessment of elevated mPAP and diagnosis of PH.

Methods

Fifty patients with suspected PH (23 patients with and 27 without PH) were investigated by right heart catheterization and time-resolved PC-MRI of the main pulmonary artery. PC-MRI data were visualized with dedicated prototype software, providing 3D vector, multi-planar reformatted (MPR) 2D vector, streamline, and particle trace representation of flow patterns. Persistence of PH-associated vortical blood flow (t_vortex) was evaluated with all visualization techniques. Dependencies of t_vortex on visualization techniques were analyzed by means of correlation and receiver operating characteristic (ROC) curve analysis.

Results

t_vortex values from 3D vector visualization correlated strongly with those from other visualization techniques (r = 0.98, 0.98 and 0.97 for MPR, streamline and particle trace visualization, respectively). Areas under ROC curves for diagnosis of PH based on t_vortex did not differ significantly and were 0.998 for 3D vector, MPR vector and particle trace visualization and 0.999 for streamline visualization. Correlations between elevated mPAP and t_vortex in patients with PH were r = 0.96, 0.93, 0.95 and 0.92 for 3D vector, MPR vector, streamline and particle trace visualization, respectively. Corresponding standard deviations from the linear regression lines ranged between 3 and 4 mmHg.

Conclusion

3D vector, MPR vector, streamline as well as particle trace visualization of time-resolved 3D PC-MRI data of the main pulmonary artery can be employed for accurate vortex-based diagnosis of PH and estimation of elevated mPAP.     

Slow life history and rapid extreme flood: demographic mechanisms and their consequences on population viability in a threatened amphibian

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The Third Replicon of Members of the Burkholderia cepacia Complex,Plasmid pC3, Plays a Role in Stress Tolerance

The metabolically versatile Burkholderia cepacia complex (Bcc) occupies a variety of niches, including the plant rhizosphere and the cystic fibrosis lung (where it is often fatal to the patient). Bcc members have multipartite genomes, of which the third replicon, pC3 (previously chromosome 3), has been shown to be a nonessential megaplasmid which confers virulence and both antifungal and proteolytic activity on several strains. In this study, pC3 curing was extended to cover strains of 16 of the 17 members of the Bcc, and the phenotypes conferred by pC3 were determined. B. cenocepacia strains H111, MCO-3, and HI2424 were previously cured of pC3; however, this had not proved possible in the epidemic strain K56-2. Here, we investigated the mechanism of this unexpected stability and found that efficient toxin-antitoxin systems are responsible for maintaining pC3 of strain K56-2. Identification of these systems allowed neutralization of the toxins and the subsequent deletion of K56-2pC3. The cured strain was found to exhibit reduced antifungal activity and was attenuated in both the zebrafish and the Caenorhabditis elegans model of infection. We used a PCR screening method to examine the prevalence of pC3 within 110 Bcc isolates and found that this replicon was absent in only four cases, suggesting evolutionary fixation. It is shown that plasmid pC3 increases the resistance of B. cenocepacia H111 to various stresses (oxidative, osmotic, high-temperature, and chlorhexidine-induced stresses), explaining the prevalence of this replicon within the Bcc.     

The Detection of the Methylated Wif-1 Gene Is More Accurate than a Fecal Occult Blood Test for Colorectal Cancer Screening

Background

The clinical benefit of guaiac fecal occult blood tests (FOBT) is now well established for colorectal cancer screening. Growing evidence has demonstrated that epigenetic modifications and fecal microbiota changes, also known as dysbiosis, are associated with CRC pathogenesis and might be used as surrogate markers of CRC.

Patients and Methods

We performed a cross-sectional study that included all consecutive subjects that were referred (from 2003 to 2007) for screening colonoscopies. Prior to colonoscopy, effluents (fresh stools, sera-S and urine-U) were harvested and FOBTs performed. Methylation levels were measured in stools, S and U for 3 genes (Wif1, ALX-4, and Vimentin) selected from a panel of 63 genes; Kras mutations and seven dominant and subdominant bacterial populations in stools were quantified. Calibration was assessed with the Hosmer-Lemeshow chi-square, and discrimination was determined by calculating the C-statistic (Area Under Curve) and Net Reclassification Improvement index.

Results

There were 247 individuals (mean age 60.8±12.4 years, 52% of males) in the study group, and 90 (36%) of these individuals were patients with advanced polyps or invasive adenocarcinomas. A multivariate model adjusted for age and FOBT led to a C-statistic of 0.83 [0.77–0.88]. After supplementary sequential (one-by-one) adjustment, Wif-1 methylation (S or U) and fecal microbiota dysbiosis led to increases of the C-statistic to 0.90 [0.84–0.94] (p = 0.02) and 0.81 [0.74–0.86] (p = 0.49), respectively. When adjusted jointly for FOBT and Wif-1 methylation or fecal microbiota dysbiosis, the increase of the C-statistic was even more significant (0.91 and 0.85, p<0.001 and p = 0.10, respectively).

Conclusion

The detection of methylated Wif-1 in either S or U has a higher performance accuracy compared to guaiac FOBT for advanced colorectal neoplasia screening. Conversely, fecal microbiota dysbiosis detection was not more accurate. Blood and urine testing could be used in those individuals reluctant to undergo stool testing.