Isolation,identification, and biochemical characterization of a brown rot fungus capable of textile dye decolorization

Twenty-two local brown rot fungal isolates were obtained from 5 different environmental sources. Fourteen isolates were presumptively identified as Aspergillus sp. and eight as Penicillium sp. using mycelium and spore morphology. All the fungal isolates were screened for their ability to decolorize Isolan Red and colored waste water and to produce oxidase activity. Aspergillus isolate 2 was chosen for further study because it could decolorize both dyes and produce oxidase. A 400 bp fragment of the 18S rRNA gene from isolate 2 was analyzed by nucleotide sequence analysis. Blastn analysis of sequence data demonstrated 100% identity to Aspergillus sp. and isolate 2 was assigned the strain designation Aspergillus sp. EL-2 (Accession number: HM140797). EL-2 could remove up to 80% of Disperse Blue in waste water effluent within 48 h in submerged shake culture. The decolorization process was energy dependent, growth related, and required viable biomass. EL-2 was able to grow and decolorize waste water over a broad pH range. Addition of inducers and inhibitors of specific enzymes or families of enzymes demonstrated the involvement of phenol oxidase (laccase), cytochrome p-450 oxygenase and hydroxyl radicals in the decolorization process. The data also suggest that it may be practical to enhance decolorizing activity of Aspergillus sp. EL-2 through the metabolic control of fungal degradative pathways by altering media composition.     

Herpes Simplex Virus Type 1 Single Strand DNA Binding Protein and Helicase/Primase Complex Disable Cellular ATR Signaling

Herpes Simplex Virus type 1 (HSV-1) has evolved to disable the cellular DNA damage response kinase, ATR. We have previously shown that HSV-1-infected cells are unable to phosphorylate the ATR substrate Chk1, even under conditions in which replication forks are stalled. Here we report that the HSV-1 single stranded DNA binding protein (ICP8), and the helicase/primase complex (UL8/UL5/UL52) form a nuclear complex in transfected cells that is necessary and sufficient to disable ATR signaling. This complex localizes to sites of DNA damage and colocalizes with ATR/ATRIP and RPA, but under these conditions, the Rad9-Rad1-Hus1 checkpoint clamp (9-1-1) do not. ATR is generally activated by substrates that contain ssDNA adjacent to dsDNA, and previous work from our laboratory has shown that ICP8 and helicase/primase also recognize this substrate. We suggest that these four viral proteins prevent ATR activation by binding to the DNA substrate and obstructing loading of the 9-1-1 checkpoint clamp. Exclusion of 9-1-1 prevents recruitment of TopBP1, the ATR kinase activator, and thus effectively disables ATR signaling. These data provide the first example of viral DNA replication proteins obscuring access to a DNA substrate that would normally trigger a DNA damage response and checkpoint signaling. This unusual mechanism used by HSV suggests that it may be possible to inhibit ATR signaling by preventing recruitment of the 9-1-1 clamp and TopBP1.     

CellProfiler: Novel Automated Image Segmentation Procedure for Super-Resolution Microscopy

Background

Super resolution (SR) microscopy enabled cell biologists to visualize subcellular details up to 20 nm in resolution. This breakthrough in spatial resolution made image analysis a challenging procedure. Direct and automated segmentation of SR images remains largely unsolved, especially when it comes to providing meaningful biological interpretations.

Results

Here, we introduce a novel automated imaging analysis routine, based on Gaussian, followed by a segmentation procedure using CellProfiler software (www.cellprofiler.org). We tested this method and succeeded to segment individual nuclear pore complexes stained with gp210 and pan-FG proteins and captured by two-color STED microscopy. Test results confirmed accuracy and robustness of the method even in noisy STED images of gp210.

Conclusions

Our pipeline and novel segmentation procedure may benefit end-users of SR microscopy to analyze their images and extract biologically significant quantitative data about them in user-friendly and fully-automated settings.

     

Lessons Learned from Crowdsourcing Complex Engineering Tasks

Crowdsourcing

Crowdsourcing is the practice of obtaining needed ideas, services, or content by requesting contributions from a large group of people. Amazon Mechanical Turk is a web marketplace for crowdsourcing microtasks, such as answering surveys and image tagging. We explored the limits of crowdsourcing by using Mechanical Turk for a more complicated task: analysis and creation of wind simulations.

Harnessing Crowdworkers for Engineering

Our investigation examined the feasibility of using crowdsourcing for complex, highly technical tasks. This was done to determine if the benefits of crowdsourcing could be harnessed to accurately and effectively contribute to solving complex real world engineering problems. Of course, untrained crowds cannot be used as a mere substitute for trained expertise. Rather, we sought to understand how crowd workers can be used as a large pool of labor for a preliminary analysis of complex data.

Virtual Wind Tunnel

We compared the skill of the anonymous crowd workers from Amazon Mechanical Turk with that of civil engineering graduate students, making a first pass at analyzing wind simulation data. For the first phase, we posted analysis questions to Amazon crowd workers and to two groups of civil engineering graduate students. A second phase of our experiment instructed crowd workers and students to create simulations on our Virtual Wind Tunnel website to solve a more complex task.

Conclusions

With a sufficiently comprehensive tutorial and compensation similar to typical crowd-sourcing wages, we were able to enlist crowd workers to effectively complete longer, more complex tasks with competence comparable to that of graduate students with more comprehensive, expert-level knowledge. Furthermore, more complex tasks require increased communication with the workers. As tasks become more complex, the employment relationship begins to become more akin to outsourcing than crowdsourcing. Through this investigation, we were able to stretch and explore the limits of crowdsourcing as a tool for solving complex problems.     

A novel layered double hydroxide-hesperidin nanoparticles exert antidiabetic,antioxidant and anti-inflammatory effects in rats with diabetes

Molecular Biology Reports - Incidence of diabetes has increased significantly worldwide over recent decades. Our objective was to prepare and characterize a novel nano-carrier of hesperidin to...     

Direct measurement of protein–protein interactions by FLIM-FRET at UV laser-induced DNA damage sites in living cells

Protein–protein interactions are essential to ensure timely and precise recruitment of chromatin remodellers and repair factors to DNA damage sites. Conventional analyses of protein–protein interactions at a population level may mask the complexity of interaction dynamics, highlighting the need for a method that enables quantification of DNA damage-dependent interactions at a single-cell level. To this end, we integrated a pulsed UV laser on a confocal fluorescence lifetime imaging (FLIM) microscope to induce localized DNA damage. To quantify protein–protein interactions in live cells, we measured Förster resonance energy transfer (FRET) between mEGFP- and mCherry-tagged proteins, based on the fluorescence lifetime reduction of the mEGFP donor protein. The UV-FLIM-FRET system offers a unique combination of real-time and single-cell quantification of DNA damage-dependent interactions, and can distinguish between direct protein–protein interactions, as opposed to those mediated by chromatin proximity. Using the UV-FLIM-FRET system, we show the dynamic changes in the interaction between poly(ADP-ribose) polymerase 1, amplified in liver cancer 1, X-ray repair cross-complementing protein 1 and tripartite motif containing 33 after DNA damage. This new set-up complements the toolset for studying DNA damage response by providing single-cell quantitative and dynamic information about protein–protein interactions at DNA damage sites.     

Assessment of In Ovo Administration of Bifidobacterium bifidum and Bifidobacterium longum on Performance,Ileal Histomorphometry,Blood Hematological,and Biochemical Parameters of Broilers

Probiotics and Antimicrobial Proteins - Bifidobacterium is one of the most promising probiotics which was recently used as an alternative growth promoter in poultry. This trial was considered to...