The Agrobacterium tumefaciens virulence protein VirE3 is a transcriptional activator of the F-box gene VBF.

During Agrobacterium tumefaciens-mediated transformation of plant cells a part of the tumour-inducing plasmid, T-DNA, is integrated into the host genome. In addition, a number of virulence proteins are translocated into the host cell. The virulence protein VirE3 binds to the Arabidopsis thaliana pBrp protein, a plant-specific general transcription factor of the TFIIB family. To study a possible role for VirE3 in transcriptional regulation, we stably expressed virE3 in A. thaliana under control of a tamoxifen-inducible promoter. By RNA sequencing we showed that upon expression of virE3 the RNA levels of 607 genes were increased more than three-fold and those of 132 genes decreased more than three-fold. One of the strongly activated genes was that encoding VBF (At1G56250), an F-box protein that may affect the levels of the VirE2 and VIP1 proteins. Using Arabidopsis cell suspension protoplasts we showed that VirE3 stimulates the VBF promoter, especially when co-expressed with pBrp. Although pBrp is localized at the external surface of plastids, co-expression of VirE3 and pBrp in Arabidopsis cell suspension protoplasts resulted in the accumulation of pBrp in the nucleus. Our results suggest that VirE3 affects the transcriptional machinery of the host cell to favour the transformation process.

Antiviral activity of the "Virus Blocking Factor" (VBF) derived i.a. from Pelargonium extract and Sambucus juice against different human-pathogenic cold viruses in vitro.

THE AIM: The in-vitro antiviral activity of the "Virus Blocking Factor" (VBF), a combination of Pelargonium extract and Sambucus juice with addition of Betaglucan 1,3 / 1,6, Zincum gluconium, Acidum ascorbicum, was studied against human pathogenic viruses: Influenza A H1N1 (FluA H1N1), Rhinovirus B subtype 14 (HRV14), Respiratory Syncytial Virus (RSV), Parainfluenzavirus subtype 3 (Para 3), and Adenovirus C subtype 5 (Adeno 5). METHOD: Antiviral activity was assessed using plaque-reduction assays after adding the test substance post infection of the MDCK, HeLa and HEp-2 cells with the viruses. Ribavirin Virazol and - in case of Adenovirus an internal laboratory standard - were used as positive controls. Cytotoxic effects of VBF and VBF Control onto the virus permissive MDCK, HeLa and HEp-2 cells were examined. Non-toxic concentrations of VBF were determined by the Methylthiazoletetrazolium test (MTT-Test). RESULTS AND CONCLUSIONS: In all antiviral studies VBF showed (2.1%) a dose-dependent antiviral activity against FluA H1N1 and HRV14 at non-toxic concentrations. A very strong effect was demonstrated in concentrations of 2.5% and 1.25% where replication of H1N1 and HRV14 was nearly completely blocked. Dose-dependent antiviral activity was detectable against RSV in a concentration range of 1.25% to 0.63% of the test item. Due to toxic side effects of a 2.5% concentration at least a "minor effect" of about 30% (1.25% solution) against Para 3 infected HEp-2 cells could be determined. Concerning Adeno 5 not any antiviral activity could be demonstrated in all studies with all tested substance concentrations of VBF. VBF Control did not show any cytotoxicity and antiviral effects. Further research is needed to elucidate clinical effect of VBF.

Antiviral activity of the "Virus Blocking Factor" (VBF) derived i.a. from Pelargonium extract and Sambucus juice against different human-pathogenic cold viruses in vitro.

THE AIM: The in-vitro antiviral activity of the "Virus Blocking Factor" (VBF), a combination of Pelargonium extract and Sambucus juice with addition of Betaglucan 1,3 / 1,6, Zincum gluconium, Acidum ascorbicum, was studied against human pathogenic viruses: Influenza A H1N1 (FluA H1N1), Rhinovirus B subtype 14 (HRV14), Respiratory Syncytial Virus (RSV), Parainfluenzavirus subtype 3 (Para 3), and Adenovirus C subtype 5 (Adeno 5). METHOD: Antiviral activity was assessed using plaque-reduction assays after adding the test substance post infection of the MDCK, HeLa and HEp-2 cells with the viruses. Ribavirin Virazol and - in case of Adenovirus an internal laboratory standard - were used as positive controls. Cytotoxic effects of VBF and VBF Control onto the virus permissive MDCK, HeLa and HEp-2 cells were examined. Non-toxic concentrations of VBF were determined by the Methylthiazoletetrazolium test (MTT-Test). RESULTS AND CONCLUSIONS: In all antiviral studies VBF showed (2.1%) a dose-dependent antiviral activity against FluA H1N1 and HRV14 at non-toxic concentrations. A very strong effect was demonstrated in concentrations of 2.5% and 1.25% where replication of H1N1 and HRV14 was nearly completely blocked. Dose-dependent antiviral activity was detectable against RSV in a concentration range of 1.25% to 0.63% of the test item. Due to toxic side effects of a 2.5% concentration at least a "minor effect" of about 30% (1.25% solution) against Para 3 infected HEp-2 cells could be determined. Concerning Adeno 5 not any antiviral activity could be demonstrated in all studies with all tested substance concentrations of VBF. VBF Control did not show any cytotoxicity and antiviral effects. Further research is needed to elucidate clinical effect of VBF.

The putative Agrobacterium transcriptional activator-like virulence protein VirD5 may target T-complex to prevent the degradation of coat proteins in the plant cell nucleus.

Agrobacterium exports at least five virulence proteins (VirE2, VirE3, VirF, VirD2, VirD5) into host cells and hijacks some host plant factors to facilitate its transformation process. Random DNA binding selection assays (RDSAs), electrophoretic mobility shift assays (EMSAs) and yeast one-hybrid systems were used to identify protein-bound DNA elements. Bimolecular fluorescence complementation, glutathione S-transferase pull-down and yeast two-hybrid assays were used to detect protein interactions. Protoplast transformation, coprecipitation, competitive binding and cell-free degradation assays were used to analyze the relationships among proteins. We found that Agrobacterium VirD5 exhibits transcriptional activation activity in yeast, is located in the plant cell nucleus, and forms homodimers. A specific VirD5-bound DNA element designated D5RE (VirD5 response element) was identified. VirD5 interacted directly with Arabidopsis VirE2 Interacting Protein 1 (AtVIP1). However, the ternary complex of VirD5-AtVIP1-VirE2 could be detected, whereas that of VirD5-AtVIP1-VBF (AtVIP1 Binding F-box protein) could not. We demonstrated that VirD5 competes with VBF for binding to AtVIP1 and stabilizes AtVIP1 and VirE2 in the cell-free degradation system. Our results indicated that VirD5 may act as both a transcriptional activator-like effector to regulate host gene expression and a protector preventing the coat proteins of the T-complex from being quickly degraded by the host's ubiquitin proteasome system (UPS).

Bioelectrochemically-assisted vermibiofilter process enhancing stabilization of sewage sludge with synchronous electricity generation.

Bioelectrochemically-assisted vermifilter (VBFBE) with sewage sludge as the anode fuel was constructed to accelerate composting of sewage sludge, which could increase the quality of the compost and harvest electric energy in comparison with vermicomposting and electrochemical only. Results revealed that the sludge stabilization with a higher soluble chemical oxygen demand (SCOD) and lower NH4+-H during 40 days of composting. At the composting, pH, C/N, electrical conductivity (EC) and germination index (GI) results demonstrated that the maturity degree of VBFBE4 was higher than that of other VBFBE. The VBFBE4 yielded a voltage of 1.024 V and maximum power density of 105.28 mW/m2 on 3th day. The bacteria in VBFBE4 were richer and higher in terms of diversity than those in other VBFBE, that was demonstrated that combination vermicomposting and electrochemistry could improve the sludge stabilization degree, accelerate sludge composting process and enhance composting maturity.

Contribution to the development of low frequency terahertz coherent Raman micro-spectroscopy and microscopy.

We report the construction and characterization of a coherent Raman tabletop system utilizing a novel astigmatic optical focusing geometry, a broadband nanosecond optical parametric oscillator and volumetric Bragg filters assisting 3CBCRS measuring system for the first time. In order to illustrate the versatility of the measurements and reveal the molecular information obtainable, two well-characterized chemicals were selected. Polarization sensitive epi-detected 3CBCRS spectra of liquid CCl4 and calcite crystal were recorded and analyzed. An unexpected polarization dependence of the signals of the lowest frequency modes of CCl4 was observed. The 1122 third order susceptibility component was phase flipped. The non-resonant susceptibility normalized 1122 component was found to be larger than the 1111 component for the lowest vibrational modes. This anomalous comportment was attributable to the anisotropy Raman tensor invariant in the third order nonlinear susceptibility tensor.