Identification and characterization of a novel Iraqi isolate of Fusarium pseudograminearum causing crown rot in wheat

Crown rot is one of the main important fungal diseases affecting wheat in many areas of the world, including Australia, USA, and Iran. Until now, there had been no report of this pathogen in Iraq. Plants displaying crown rot symptoms were observed in Shaat Alarab (Basra, Iraq); we investigated the causal agent of the disease. Samples were surface-sterilized in bleach (1% available chlorine) and cultured on quarter-strength potato dextrose agar plates. DNA was extracted from fungal mycelia, using a modified CTAB protocol. The ITS/5.8S regions were amplified using primer pair ITS1 and ITS4. PCR products purified using a gel extraction kit were sequenced. The sequence that was detected was used to BLAST against NCBI data. The most similar sequence was the ITS/5.8S rDNA region of Fusarium pseudograminearum (strain NRRL28062), showing 97.95% identity. This species normally causes crown rot, resulting in severe damage under dry spring conditions. A pathogenicity test employed to assess the disease-causing ability of the strain showed significant disease symptoms up to 57% infected spikelets. The results confirmed the presence of F. pseudograminearum as a causal agent of wheat crown rot in Iraq. The presence of this pathogen demands further investigations to develop resistant cultivars and/or mechanical control.     

Genetic characterization of Chikungunya virus from New Delhi reveal emergence of a new molecular signature in Indian isolates

ABSTRACT: BACKGROUND: Chikungunya (CHIK) is currently endemic in South and Central India and exist as co-infections with dengue in Northern India. In 2010, New Delhi witnessed an outbreak of CHIK in the months October-December. This was the first incidence of a dominant CHIK outbreak in Delhi and prompted us to characterize the Delhi virus strains. We have also investigated the evolution of CHIK spread in India. FINDINGS: Clinical samples were subjected to RT-PCR to detect CHIK viral RNA. The PCR amplified products were sequenced and the resulting sequences were genetically analyzed. Phylogenetic analysis based on partial sequences of the structural proteins E1 and E2 revealed that the viruses in the latest outbreak exhibited ECSA lineage. Two novel mutations, E1 K211E and E2 V264A were observed in all Delhi isolates. In addition, CHIKV sequences from eight states in India were analyzed along with Delhi sequences to map the genetic diversity of CHIKV within the country. Estimates of average evolutionary divergence within states showed varying divergence among the sequences both within the states and between the states. We identified distinct molecular signatures of the different genotypes of CHIKV revealing emergence of a new signature in the New Delhi clade. Statistical analyses and construction of evolutionary path of the virus within the country revealed gradual spread of one specific strain all over the country. CONCLUSION: This study has identified unique mutations in the E1 and E2 genes and has revealed the presence of ancestral CHIKV population with maximum diversity circulating in Maharashtra. The study has further revealed the trend of CHIK spread in India since its first report in 1963 and its subsequent reappearance in 2005.     

Prostaglandin E2 upregulates EGF-stimulated signaling in mitogenic pathways involving Akt and ERK in hepatocytes

Prostaglandins (PGs) such as PGE2 enhance proliferation in many cells, apparently through several distinct mechanisms, including transactivation of the epidermal growth factor (EGF) receptor (EGFR) as well as EGFR-independent pathways. In this study we found that in primary cultures of rat hepatocytes PGE2 did not induce phosphorylation of the EGFR, and the EGFR tyrosine kinase blockers gefitinib and AG1478 did not affect PGE2-stimulated phosphorylation of ERK1/2. In contrast, PGE2 elicited EGFR phosphorylation and EGFR tyrosine kinase inhibitor-sensitive ERK phosphorylation in MH1C1 hepatoma cells. These findings suggest that PGE2 elicits EGFR transactivation in MH1C1 cells but not in hepatocytes. Treatment of the hepatocytes with PGE2 at 3 h after plating amplified the stimulatory effect on DNA synthesis of EGF administered at 24 h and advanced and augmented the cyclin D1 expression in response to EGF in hepatocytes. The pretreatment of the hepatocytes with PGE2 resulted in an increase in the magnitude of EGF-stimulated Akt phosphorylation and ERK1/2 phosphorylation and kinase activity, including an extended duration of the responses, particularly of ERK, to EGF in PGE2-treated cells. Pertussis toxin abolished the ability of PGE2 to enhance the Akt and ERK responses to EGF. The results suggest that in hepatocytes, unlike MH1C1 hepatoma cells, PGE2 does not transactivate the EGFR, but instead acts in synergism with EGF by modulating mitogenic mechanisms downstream of the EGFR. These effects seem to be at least in part G(i) protein-mediated and include upregulation of signaling in the PI3K/Akt and the Ras/ERK pathways.     

Cell-specific localization of Na+ in roots of durum wheat and possible control points for salt exclusion

Sodium exclusion from leaves is an important mechanism for salt tolerance in durum wheat. To characterize possible control points for Na(+) exclusion, quantitative cryo-analytical scanning electron microscopy was used to determine cell-specific ion profiles across roots of two durum wheat genotypes with contrasting rates of Na(+) transport from root to shoot grown in 50 mm NaCl. The Na(+) concentration in Line 149 (low transport genotype) declined across the cortex, being highest in the epidermal and sub-epidermal cells (48 mm) and lowest in the inner cortical cells (22 mm). Na(+) was high in the pericycle (85 mm) and low in the xylem parenchyma (34 mm). The Na(+) profile in Tamaroi (high transport genotype) had a similar trend but with a high concentration (130 mm) in the xylem parenchyma. The K(+) profiles were generally inverse to those of Na(+). Chloride was only detected in the epidermis. These data suggest that the epidermal and cortical cells removed most of the Na(+) and Cl(-) from the transpiration stream before it reached the endodermis, and that the endodermis is not the control point for salt uptake by the plant. The pericycle as well as the xylem parenchyma may be important in the control of net Na(+) loading of the xylem.     

Zinc binding ligands and cellular zinc trafficking: apo-metallothionein, glutathione, TPEN, proteomic zinc, and Zn-Sp1

Many cell types contain metal-ion unsaturated metallothionein (MT). Considering the Zn(2+) binding affinity of metallothionein, the existence of this species in the intracellular environment constitutes a substantial "thermodynamic sink". Indeed, the mM concentration of glutathione may be thought of in the same way. In order to understand how apo-MT and the rest of the Zn-proteome manage to co-exist, experiments examined the in vitro reactivity of Zn-proteome with apo-MT, glutathione (GSH), and a series of common Zn(2+) chelating agents including N,N,N',N'-(2-pyridylethyl)ethylenediammine (TPEN), EDTA, and [(2,2'-oxyproplylene-dinitrilo]tetraacetic acid (EGTA). Less than 10% of Zn-proteome from U87mg cells reacted with apo-MT or GSH. In contrast, each of the synthetic chelators was 2-3 times more reactive. TPEN, a cell permeant reagent, also reacted rapidly with both Zn-proteome and Zn-MT in LLC-PK(1) cells. Taking a specific zinc finger protein for further study, apo-MT, GSH, and TPEN inhibited the binding of Zn(3)-Sp1 with its cognate DNA site (GC-1) in the sodium-glucose co-transporter promoter of mouse kidney. In contrast, preformation of Zn(3)-Sp1-(GC-1) prevented reaction with apo-MT and GSH; TPEN remained active but at a higher concentration. Whereas, Zn(3)-Sp1 is active in cells containing apo-MT and GSH, exposure of LLC-PK(1) cells to TPEN for 24h largely inactivated its DNA binding activity. The results help to rationalize the steady state presence of cellular apo-MT in the midst of the many, diverse members of the Zn-proteome. They also show that TPEN is a robust intracellular chelator of proteomic Zn(2+).     

Plasmodium falciparum: effect of radiation on levels of gene transcripts in sporozoites

Humans immunized by the bites of irradiated Plasmodium falciparum (Pf) sporozoite-infected mosquitoes are protected against malaria. Radiation attenuates the sporozoites preventing them from fully developing and replicating in hepatocytes, but the effects of radiation on gene expression in sporozoites are unknown. We used RT-PCR (35 cycles of PCR followed by densitometry) to assess the expression of ten genes in Pf sporozoites, and in sporozoites irradiated with 15,000cGy. Irradiation reduced expression substantially (>60%) of two DNA repair genes; moderately (30-60%) of PfUIS3, the Pf orthologue of PbUIS3, a gene up-regulated in Plasmodium berghei sporozoites and of a third DNA repair gene; and minimally (<30%) of the Pf18S ribosomal RNA, PfCSP, PfSSP2/TRAP, and PfCELTOS genes. Irradiation increased expression of PfSPATR minimally. PfLSA1 RNA was not detectable in sporozoites. These results establish that radiation of sporozoites affects gene expression levels and provide the foundation for studies to identify specific genes involved in attenuation and protective immunity.     

Dietary fish oil associated with increased apoptosis and modulated expression of Bax and Bcl-2 during 7,12-dimethylbenz(alpha)anthracene-induced mammary carcinogenesis in rats

The present study investigated the chemopreventive effect of dietary fish oil (Maxepa), rich in eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) on induction of apoptosis in mammary carcinogenesis model. Mammary carcinogenesis was initiated by a single, tail vein injection of 7,12-dimethylbenz(alpha)anthracene (DMBA) (0.5mg/0.2ml corn oil/100g body weight) at 7 weeks of animal age. Ninety female Sprague-Dawley rats were divided into two parts: part one was used for histology and immunohistochemical study and part two for morphological analysis. Each part consists of three experimental groups having 15 animals, i.e., Group A (DMBA control), Group B (DMBA+fish oil) and Group C (DMBA+corn oil). Rats were fed either fish oil or corn oil (0.5ml/day/rat) by oral gavage, 2 weeks prior to DMBA injection. Treatment was continued 25 weeks, studying histopathology, expression of Bax and Bcl-2 proteins by immunohistochemistry and apoptosis by TUNEL assay and morphological study at 36 weeks. Results showed that the fish oil-treated group exhibited a substantial increase in Bax (p<0.05) immunolabelling and a reduction of Bcl-2 immunopositivity (p<0.05), and increased TUNEL-positive apoptotic cells (p<0.05); however, corn oil treatment did not show these beneficial effects toward mammary preneoplasia. We conclude that fish oil has the potential to play a significant role in limiting mammary tumourigenesis in vivo.     

Bringing science and pragmatism together a tiered approach for modelling toxicological impacts in LCA

Goal, Scope and Background  The EU 5^th framework project OMNIITOX will develop models calculating characterisation factors for assessing the potential toxic impacts of chemicals within the framework of LCA. These models will become accessible through a web-based information system. The key objective of the OMNIITOX project is to increase the coverage of substances by such models. In order to reach this objective, simpler models which need less but available data, will have to be developed while maintaining scientific quality. Methods. Experience within the OMNIITOX project has taught that data availability and quality are crucial issues for calculating characterisation factors. Data availability determines whether calculating characterisation factors is possible at all, whereas data quality determines to what extent the resulting characterisation factors are reliable. Today, there is insufficient knowledge and/or resources to have high data availability as well as high data quality and high model quality at the same time. Results  The OMNIITOX project is developing two inter-related models in order to be able to provide LCA impact assessment characterisation factors for toxic releases for as broad a range of chemicals as possible: 1) A base model representing a state-of-the-art multimedia model and 2) a simple model derived from the base model using statistical tools. Discussion. A preliminary decision tree for using the OMNIITOX information system (IS) is presented. The decision tree aims to illustrate how the OMNIITOX IS can assist an LCA practitioner in finding or deriving characterisation factors for use in life cycle impact assessment of toxic releases. Conclusions and Outlook  Data availability and quality are crucial issues when calculating characterisation factors for the toxicity impact categories. The OMNIITOX project is developing a tiered model approach for this. It is foreseen that a first version of the base model will be ready in late summer of 2004, whereas a first version of the simple base model is expected a few months later.     

Effect of Calotropis procera latex on isoproterenol induced myocardial infarction in albino rats.

The alcoholic extract of the latex obtained from Calotropis procera (Asclepidaceae) was evaluated for protection against isoproterenol (20 mg/100 g body wt., s.c.)-induced myocardial infarction in albino rats. The heart damage induced by isoproterenol was indicated by elevated levels of the marker enzymes such as Creatine Kinase-isoenzyme (CK-MB), Lactate dehydrogenase (LDH), Serum Glutamate Oxaloacetic Transaminase (SGOT) and Serum Glutamate Pyruvate Transaminase (SGPT) in serum with increased lipid peroxide and reduced glutathione content in heart homogenates. Microscopical examination (histopathology) was also performed on the myocardial tissue. Pretreatment with an ethanolic latex extract of Calotropis procera at a dose of 300 mg/kg body wt., administered orally thrice a day for 30 days, reduced significantly (p < 0.01) the elevated marker enzyme levels in serum and heart homogenates in isoproterenol-induced myocardial infarction. Histopathological observation revealed a marked protection by the extract in myocardial necrotic damage.     

Identification of the Axin and Frat binding region of glycogen synthase kinase-3.

Glycogen synthase kinase-3 (GSK-3) is a key component of several signaling pathways including those regulated by Wnt and insulin ligands. Specificity in GSK-3 signaling is thought to involve interactions with scaffold proteins that localize GSK-3 regulators and substrates. This report shows that GSK-3 forms a low affinity homodimer that is disrupted by binding to Axin and Frat. Based on the crystal structure of GSK-3, we have used surface-scanning mutagenesis to identify residues that differentially affect GSK-3 interactions. Mutations that disrupt Frat and Axin cluster at the dimer interface explaining their effect on homodimer formation. Loss of the Axin binding site blocks the ability of dominant negative GSK-3 to cause axis duplication in Xenopus embryos. The Axin binding site is conserved within all GSK-3 proteins, and its loss affects both cell motility and gene expression in the nonmetazoan, Dictyostelium. Surprisingly, we find no genetic interaction between a non-Axin-binding GSK-3 mutant and T-cell factor activity, arguing that Axin interactions alone cannot explain the regulation of T-cell factor-mediated gene expression.