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The legume crops such as chickpea and lentils are mainly cultivated in semi-arid tropical lands. Chickpea chlorotic dwarf virus (CpCDV) causes major losses to legumes throughout the world. Producing of specific antibody against this virus is crucial for surveys of disease in the fields and assessment of vial resistance in plant cultivars. Present article describes developing of specific antibody against the CpCDV virus by applying recombinant protein. In this study, coat protein of CpCDV was selected as a target for detection and preparation of polyclonal antibody. To achieve this aim CP gene encoding coat protein of CpCDV was initially PCR-amplified and inserted into bacterial expression vector. Expression of recombinant protein was performed in Bl21 strain of Escherichia coli. Purification was carried out under native conditions and the accuracy of recombinant protein production was confirmed by electrophoresis. The purified recombinant coat protein of CpCDV was used for immunization of rabbit. Purification of immunoglobulin molecules was performed by affinity chromatography using protein A column followed by conjugating of IgG to alkaline phosphatase enzyme. The capability of purified antibodies and conjugates for efficient detection of infected plants was assessed by double antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA), western blotting and dot immunosorbent assay (DIBA). These results proved that prepared IgG and conjugate are able to distinguish with high efficiency CpCDV infected plants. To the best of our knowledge, this is the first report for production of anti-CpCDV antibodies raised through recombinant protein technology.

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The Xanthomonas citri pv. citri (Xcc) is causal agent of bacterial citrus canker which is major disease of citrus throughout the world. The pthA bacterial effector protein is presented within the infected plants and indispensable of canker. The scFv antibodies are valuable tools for diagnosis and suppression of pathogens within plants. The present article describes developing and characterization of specific recombinant monoclonal scFv antibodies against pthA effector protein. For this aim, the gene encoding pthA protein was heterologously expressed in Escherichia coli and used for screening of Tomlinson phage display antibody library to pinpoint specific single chain variable fragment (scFv). In each round of panning, the affinity of phage towards pthA was checked by enzyme linked immunosorbent assay (ELISA). The data was indicative of about 50% of the monoclonal phages to be reactive strongly against pthA protein. Among the positive clones, 5 samples (A12, B8, C1, H8 and G8) were capable of detecting Xcc-infected plant samples and recombinant pthA protein. Restriction fragment length polymorphism showed similar banding pattern for all 5 scFvs as renamed to pthA-scFG8. HB2151 E. coli cells were infected by the phage bearing pthA-scFG8, and the expression of the peptide was induced by IPTG to produce a 30 kDa recombinant molecule. I-TASSER was used for homology modeling of both scFv and pthA and docking was carried out by Hex program. The latter demonstrated binding energy of −784 kcal/mol in scFv-pthA. 

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Grapevine viruses cause significant losses in the yield of grape. This study describes applying silver nanoparticles (AgNPs) to produce virus-free grapevine plants and compares it with chemo and thermotherapy. Preliminary molecular analysis proved the presence of Grapevine fanleaf virus (GFLV) and grapevine leafroll-associated virus-1 (GLRaV-1) in the ʻAsgariʼ, ʻPeykaniʼ, and ʻShahaniʼ cultivar samples, then single node explants were cultivated in the MS medium. Thermotherapy at 35 ± 1 ºC and cycles of 35/38 ± 1 ºC, chemotherapy with ribavirin 0, 20, 25, and 30 μg.ml^-1 and using AgNPs at 0, 10, 15, and 20 ppm in medium and 40, 50, and 60 ppm sprayed during acclimatization stage were applied to obtain virus-free explants. The results indicated that using 20 ppm AgNPs in medium and AgNPs combined treatment (15 ppm AgNPs in medium and sprayed with 50 ppm AgNPs in the acclimatization stage) were the most effective treatments for the elimination of viruses. The best treatment led to 100% eradication of GLRaV-1 and 67% of GFLV in ʻAsgariʼ, 100% eradication of GLRaV-1 and GFLV in ʻPeykaniʼ and 100% eradication of GLRaV-1 and 67% of GFLV in ʻShahaniʼ. Furthermore, applying of AgNPs improved plant growth parameters, including plant height, which in infected plantlets was (18.06, 12.36, and 14.92 cm in ʻAsgariʼ, ʻPeykaniʼ, and ʻShahaniʼ, respectively) less than virus-free plantlets. Leaf number was 45, 34, and 27 in virus-free plantlets of ʻAsgariʼ, ʻPeykaniʼ, and ʻShahaniʼ, respectively, but in infected plantlets, it was 24.40, 19.80, and 12. Leaf area increased from 5.34, 5.50, and 5.94 cm² in infected plantlets to 9.56, 11.43, and 12.33 cm² in virus-free plantlets of ʻAsgariʼ, ʻPeykaniʼ, and ʻShahaniʼ, respectively. Complementary results proved that chlorophyll content in virus-free is significantly higher than in virus-infected plantlets, which explains and confirms the change in growth parameters after virus removal.
 

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Rigid Body-Spring Models (RBSM) are a kind of discrete models which are developed mainly for the simulation of quasi-brittle materials ranging from ceramic, concrete, and masonry, to rock and soil. In this approach, material domain is discretized to a set of rigid cells interconnected through a set of translational and rotational springs located at cell interfaces. These cells are constructed over a set of points (seeds) distributed regularly or randomly over the domain. When it comes to heterogeneous materials, the seeds may be located in accord to the geometry and distribution of inclusions. For two-dimensional problems, each rigid cell has normally two translational and one rotational degrees of freedom (DOFs). The springs may be distributed along the interface or lumped at a point called contact/computational point (CP) and activated by the relative movement of connecting cells. As a fundamental issues, before being applicable for the simulation of inelastic behavior of materials, the kinematics of an RBSM and also the force-displacement relations of its springs should be defined in such a way that the model can adequately predict the elastic behavior of continuum at both macro and micro scales. Our review of the literature shows that except one of the RBSMs, used in the current paper for comparison, others suffer from some shortcomings which result in their inaccurate elastic predictions. In the aforementioned model, cells are convex polygons generated by the Voronoi diagram of seeds (cell nucleus) and the spring set of an interface is comprised of two translational (normal and tangential) and one rotational springs located at the midpoint of the interface. Our study shows that, although this RBSM presents generally a reliable predictions, however, there exists some kind of scattering in the predicted micro strain and stress distributions. Accordingly, with the aim of eliminating the observed scatters, this paper borrows the interpolation functions of the conventional finite element method and presents a new kinematic formulation for the RBSM. In the new model, called FE-RBSM, a Delaunay tessellation is constructed over cell nuclei. This results in a network of triangular elements which can be considered as 3-node constant strain triangular finite elements. Two translational DOFs at each nucleus and two CPs per interface with normal and tangential springs are assumed. Next the triangles including the CPs are determined. Finally, the normal, tangential, and lateral strains of each CP are calculated by projecting the constant strain tensor of the associated triangle on the corresponding interface. In order to examine the efficiency and accuracy of the proposed FE-RBSM formulation, two kinds of numerical analyses including constant and variable stress fields are employed. For the case of constant stress field, a 100mm square sample is analyzed in uniaxial tension and pure shear. Besides, for the case of variable stress field, a 300mm square sample including a 10mm diameter hole at its centroid is analyzed in uniaxial and biaxial tension. Also, a 300mm diameter circle sample is analyzed under splitting compression. The results are compared with those of the selected RBSM and also the analytical solutions. They show that, compared to the RBSM, the FE-RBSM can better predict the macro elastic properties and also gives scatter-free microstress fields.

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Production of virus-free stocks is crucial for efficient management of plant viruses in cultivation of pome fruits. Regarding the importance of producing the pre-basic stocks of valuable fruit trees, pear cultivar ʽNatanzʼ, an important local pear cultivar in Iran, was selected for virus eradication. In the present study, tissue culture combined with in vitro thermotherapy and thermo-chemotherapy techniques were used for elimination of Apple Stem Pitting Virus (ASPV) and Apple Mosaic Virus (ApMV). In thermotherapy approach, in vitro shoots were initially incubated for 55, 60, 65, and 70 days in alternating temperatures (32/38°C), then, meristems were cultivated on meristem medium. In thermo-chemotherapy approach, in vitro shoots were incubated for 50 days at 32/38°C, and then meristems were cultivated on a medium containing ribavirin. Virus detection by RT-PCR using specific primers was carried out after rooting and adaptation of the regenerated shoots. The percentage of survived shoots and meristem establishment were depended on thermo-duration. After 55 days, 83.33% of shoots survived, while it decreased to 33.33% after 70 days. Both ASPV and ApMV were eliminated after 60 days of thermotherapy. Ribavirin at 10 and 20 mg L^-1 reduced the percentage of meristem establishment to 50 and 37%, respectively, compared to the control (88.88%). Thermo-chemothery was also effective for ASPV and ApMV eradication from pear shoots.

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Aspergillus flavus is a major fungal phytopathogen and an opportunistic pathogen to humans and livestock. The fungus produces immunosuppressive and carcinogenic aflatoxins that acts as a burden in food and feed industries. Phylogenetic analysis indicated that the cell wall protein, A. flavus mannoprotein 1 (AFLMP1), is exclusively present in Aspergillus section Flavi such as A. flavus and A. parasiticus. This makes AFLMP1 an excellent candidate for siRNA-based control of aflatoxigenic fungi in farms and processing units, and fungal therapy in hospitals. Here and for the first time, mode of action of a chemically synthesized RNA interference (siRNA) was investigated for the control of AFLMP1 synthesis. The efficacy of direct uptake of different concentration of siRNA on spore germination of A. flavus was monitored via Opera High Content Screening confocal microscope. siRNA caused growth inhibition at lower concentrations (0.65 nM) and germination failure (more than 90%) at higher concentrations (5 nM), most likely by interfering in mannoprotein biosynthesis. It is assumed that siRNA technology can be implemented as a promising suppressive agent in treatment of target genes for inactivation. It can be considered as an intervention in food/feed industries to control the development and reproduction of fungi to keep the fungal population below hazard critical points.