Genetic Diversity Among Iranian Isolates of Rhizoctonia solani Kühn Anastomosis Group1 Subgroups Based on Isozyme Analysis and Total Soluble Protein Pattern

Rhizoctonia solani is a destructive fungal pathogen with a wide host range. The R. solani complex species includes several divergent groups delimited by affinities for hyphal anastomosis. In this study, genetic variation among 20 isolates of R. solani anastomosis group 1 (AG1) subgroups (AG1‐IA and AG1‐IB) collected from Mâzandaran province, Iran, and standard isolates of these subgroups, was determined by isozyme analysis and total soluble protein profile. Mycelial protein pattern and isozyme analysis were studied using denaturing and non‐denaturing polyacrylamide gel electrophoresis, respectively. A total of 15 enzyme systems were tested, among which six enzymes including esterase, alkaline phosphatase, superoxide dismutase, octanol dehydrogenase, lactate dehydrogenase and mannitol dehydrogenase generated distinct and reproducible results. The soluble protein patterns were similar among the R. solani isolates examined; however, minor differences in banding pattern were observed between the two subgroups. In isozyme analysis, a total of 64 electrophoretic phenotypes were detected for all six enzymes used. Based on cluster analysis and similarity matrix, the fungal isolates were divided into two genetically distinct groups of I and II consistent with the previously reported AG1‐IA and AG1‐IB subgroups in AG1. Group I represented all isolates belonging to AG1‐IA subgroup, whereas group II represented all isolates belonging to AG1‐IB subgroup. Results from isozyme analysis suggest that the subgrouping concept within AGs is genetically based.     

Physiological and Biochemical Characteristics of Iranian Strains of Xanthomonas axonopodis pv. citri, the Causal Agent of Citrus bacterial Canker Disease

Twenty-four strains of Xanthomonas axonopodis pv. citri ( Xac ), the causal agent of bacterial canker of citrus, isolated from Mexican lime ( Citrus aurantifolia ) and lemon ( Citrus limon ) in southern Iran, were characterized phenotypically. Strains were all pathogenic on C. aurantifolia . Sodium dodecyl sulphate-polyacrylamide gel electrophoresis analysis revealed slight differences in soluble protein profiles among the strains. Based on host range specificity and phenotypic characteristics, representative strains were differentiated into two groups of Asiatic (A) and atypical Asiatic (aA) forms. DNA fingerprinting analysis using Eco RI as the restriction endonuclease showed a negligible difference in restriction pattern between the two groups. On the basis of isozymic analysis, the two groups were distinct with respect to superoxide dismutase (SOD) and esterase (EST) banding patterns. Plasmid DNA profile analysis showed that the bacterial strains were different from each other in terms of plasmid number and molecular weight. Phage typing study revealed that most of group A strains were susceptible to Cp1 and/or Cp2 and some were resistant to both phage types including the strain in aA group. Bacteriocin production test indicated that there was a variation among Xac strains using different indicators for each bacteriocin producer. It is concluded that the Iranian strains of Xac are heterogeneous and constitute a subgroup(s) within the pathotype A.     

Seaweed Liquid Extract as an Alternative Biostimulant for the Amelioration of Salt-stress Effects in Calotropis procera (Aiton) W.T

Scientists consider saltwater one of the effective environmental stress that negatively affects the growth and establishment of trees and shrubs worldwide. Utilizing the potential of Bio-stimulant compounds present in the brown seaweed extract is an alternative strategy to improve crop tolerance to salinity. This study focused on the application of seaweed extract as a Bio-stimulant agent to counteract the salt stress on the growth and some physiochemical aspects of milkweed seedlings. In this experiment, the seedlings were treated with seaweed extract (SWE) of Sargassum angustifolium at four concentrations (non-SWE or control, 0.5, 1.0, and 1.5%) and then exposed to salt stress at four levels (0, 7.5, 15, and 30 dS m^?1 of diluted seawater) in a completely randomized design (four replications per treatment) over a time-span of 3 months. The results indicated that SWE-treated seedlings could tolerate salinity up to 15 dS m^?1 and also increase the survival rate by 69%. The growth parameters like height, specific leaf area, root length and volume, root and shoot dry weight were considerably enhanced by SWE (1%) from 7.5 to 30 dS m^?1. Moreover, gas exchanges and chlorophyll pigments were markedly increased using SWE (0.5%) under salt lower 15 dS m^?1 than control. Also, both SWE and salt stress significantly enhanced antioxidant enzymes over control, but SWE more increased the parameters. SWEs agent at different dosages significantly decreased electrolyte leakage at all salinity levels (except in 7.5 and 15 dS m^?1) compared to control. SWEs (1%) resulted in increasing K⁺ uptake but decreasing Na⁺ uptake and markedly enhancing K^+/Na⁺ ratio in stressed-milkweed versus free-salt stress. Totally, this research illustrates the potential of SWEs (at lower dosages) for elevating milkweed tolerance to moderate salinity stress and highlights the possibility of applying it as Bio-stimulant fertilizer.

     

Novel spiro[indene-1,2′-quinazolin]-4′(3′H)-one derivatives as potent anticonvulsant agents: One-pot synthesis,in vivo biological evaluation,and molecular docking studies

A new series of spiro[indene-1,2′-quinazolin]-4′(3′H)-one derivatives 4a–m were synthesized via a one-pot method and evaluated for anticonvulsant activities using pentylenetetrazole (PTZ) and maximal electroshock (MES)-induced seizures. Obtained results demonstrated that these compounds have not anticonvulsant activity in PTZ test while are active in the MES test. Among the synthesized compounds, the best anticonvulsant activity was obtained with compound 4h . This compound also was not neurotoxic. Given that the title new compounds have the pharmacophore requirement for benzodiazepine (BZD) receptor agonist, the most potent compound was assayed in vivo and in silico as BZD receptor agonist. After treatment with flumazenil as a standard BZD receptor antagonist, anticonvulsant activity of compound 4h decreased. Therefore, the involvement of BZD receptors in anticonvulsant activity of this compound confirmed. Furthermore, docking study of compound 4h in the BZD-binding site of GABA_A receptor confirmed that this compound interacted with the important residues.     

Development of a new live attenuated mumps virus vaccine in human diploid cells

A new live attenuated mumps vaccine was developed in human diploid cells. The S-12 virus was isolated from a 10-year-old girl showing typical symptoms of mumps infection, the diagnosis was confirmed by a pediatrician. The virus was isolated in green monkey kidney cells, without passage in chick embryo cavity or chick embryo fibroblasts. Attenuation of the wild virus was performed by serial passages in human diploid cells (MRC-5). The attenuated virus was characterized by identity tests, as well as by a reduction in plaque size, as marker tests. The virus was free from adventitious agents and safe for laboratory animals as well as for monkeys. The reactogenicity and immunogenicity of the S-12 virus for man was investigated by administration of a monovalent vaccine to 20 seronegative adult male volunteers and 30 children aged 1 to 5 years without history of mumps infection or vaccination. Seroconversion was obtained in 95% of the vaccinees. The new vaccine has the advantage of not requiring specific pathogen-free eggs, and being free from avian proteins and therefore can be used in sensitized patients.     

Generation of recombinant cytoplasmic domain of epidermal growth factor receptor with intrinsic protein tyrosine kinase activity

We have generated a recombinant baculovirus using the high expression vector pVL941 containing the complementary DNA encoding the intracellular domain of the human epidermal growth factor receptor (EGFR-IC). Upon infection of Spodoptera frugiperda insect cells, protein tyrosine kinase-active EGFR-IC was produced. The expressed protein has a molecular weight of 61,000 and is specifically recognized by antibodies directed against peptides representing different regions of human EGFR-IC. Upon sonication of infected cells, EGFR-IC was detected in both the soluble and insoluble fractions of the cell lysate. About 20-50% of the expressed EGFR-IC was soluble. Metabolic labeling and protein analyses showed that EGFR-IC comprised 7% of newly synthesized proteins in the cytoplasmic lysate and 0.1-0.2% of the total soluble protein. We have used a three-step purification procedure (fast-Q-Sepharose and phenyl-Sepharose column chromatographies and 30% ammonium sulfate precipitation) to purify EGFR-IC to 85% purity with 15-20% recovery from the initial soluble lysate. A yield of 3-4 mg of purified EGFR-IC has been consistently produced from 20 roller bottles with 2-4 x 10(8) infected cells/bottle. The tyrosine kinase activity was retained through purification. The enzyme demonstrated much higher autophosphorylation activity in the presence of Mn2+ than Mg2+. Phosphopeptide mapping revealed the same autophosphorylation sites utilized by EGFR-IC as those identified in wild-type EGFR. EGFR-IC-catalyzed phosphorylation of either a synthetic peptide representing the major autophosphorylation site or angiotensin II showed that the baculovirus-expressed EGFR-IC exhibits similar enzymatic kinetic characteristics to the intact activated EGFR kinase.     

Validation of EST-derived STS markers localized on Qfhs.ndsu-3BS for Fusarium head blight resistance in wheat using a 'Wangshuibai' derived population

A few EST-derived STS markers localized on Qfhs.ndsu-3BS, a major QTL for resistance to Fusarium head blight (FHB) in wheat, have been previously identified in the 'Sumai 3'/'Stoa' population. In this study, we used a 'Wangshuibai' (resistant)/'Seri82' (susceptible) derived population, linkage group, QTL, and quantitative gene expression analysis to assess the genetic background dependence and stability of the EST-derived STS markers for use in marker aided selection to improve FHB resistance in wheat. Based on our results, a QTL in the map interval of Xsts3B-138_1-Xgwm493 on chromosome 3BS was detected for FHB resistance, which accounted for up to 16% of the phenotypic variation. BLASTN analysis indicated that Xsts3B-138_1 sequence had significant similarity with the resistance gene analogue. Real-time quantitative PCR showed that the relative expression of Xsts3B-1381 in 'Wangshuibai' at 96 h after inoculation was 2.6 times higher than 'Seri82'. Our results underlined that EST-derived STS3B-138 markers could be predominantly used in marker aided selection to improve FHB resistance in wheat.