Improved Nutrient Use Efficiency Increases Plant Growth of Rice with the Use of IAA-Overproducing Strains of Endophytic Burkholderia cepacia Strain RRE25

Effect of indole acetic acid (IAA)-overproducing mutants of Burkholderia cepacia (RRE25), a member of β-subclass of Proteobacteria and naturally occurring rice endophyte, was observed on the growth of rice (Oryza sativa L.) plants grown under greenhouse conditions. Nine mutants were characterized for altered biosynthesis of IAA after nitrous acid mutagenesis. These mutants were grouped into two classes: class I mutants have reduced production of IAA as compared to the wild type, while class II mutants showed overproduction of IAA. Mutants of both classes and RRE25, the parent (wild type), were inoculated on rice seedlings of two cultivars (Sarjoo-52 and NDR-97). Uptake of nitrogen, phosphorous, and potassium was estimated in these plants. Significant increase in the amount of uptake of all three elements was observed when inoculated with the IAA-overproducing mutants over control as well as in the plants inoculated with the wild type (RRE25). Effect of inoculation of IAA-overproducing mutants was more pronounced on the uptake of phosphorous in cultivar NDR-97 than Sarjoo-52, while it was opposite with respect to potassium uptake. Any significant difference was not observed in nitrogen uptake among the two cultivars. It shows that the host also plays an important role in the beneficial endophytic association. It was concluded from these results that one of the possible mechanisms of growth promotion of rice plants inoculated with bacterial endophytes is their effects on an increase in the capability of nutritional uptake possible through the effect of IAA production which results in proliferation of root system that could mine more nutrients from the soil.     

Characterization of G-quadruplex antibody reveals differential specificity for G4 DNA forms

Accumulating evidence suggests that human genome can fold into non-B DNA structures, when appropriate sequence and favourable conditions are present. Among these, G-quadruplexes (G4-DNA) are associated with gene regulation, chromosome fragility and telomere maintenance. Although several techniques are used in detecting such structures in vitro, understanding their intracellular existence has been challenging. Recently, an antibody, BG4, was described to study G4 structures within cells. Here, we characterize BG4 for its affinity towards G4-DNA, using several biochemical and biophysical tools. BG4 bound to G-rich DNA derived from multiple genes that form G-quadruplexes, unlike complementary C-rich or random sequences. BLI studies revealed robust binding affinity (K_d = 17.4 nM). Gel shift assays show BG4 binds to inter- and intramolecular G4-DNA, when it is in parallel orientation. Mere presence of G4-motif in duplex DNA is insufficient for antibody recognition. Importantly, BG4 can bind to G4-DNA within telomere sequence in a supercoiled plasmid. Finally, we show that BG4 binds to form efficient foci in four cell lines, irrespective of their lineage, demonstrating presence of G4-DNA in genome. Importantly, number of BG4 foci within the cells can be modulated, upon knockdown of G4-resolvase, WRN. Thus, we establish specificity of BG4 towards G4-DNA and discuss its potential applications.     

In Vitro Effects of Cognitives and Nootropics on Mitochondrial Respiration and Monoamine Oxidase Activity

Impairment of mitochondrial metabolism, particularly the electron transport chain (ETC), as well as increased oxidative stress might play a significant role in pathogenesis of Alzheimer’s disease (AD). Some effects of drugs used for symptomatic AD treatment may be related to their direct action on mitochondrial function. In vitro effects of pharmacologically different cognitives (galantamine, donepezil, rivastigmine, 7-MEOTA, memantine) and nootropic drugs (latrepirdine, piracetam) were investigated on selected mitochondrial parameters: activities of ETC complexes I, II + III, and IV, citrate synthase, monoamine oxidase (MAO), oxygen consumption rate, and hydrogen peroxide production of pig brain mitochondria. Complex I activity was decreased by galantamine, donepezil, and memantine; complex II + III activity was increased by galantamine. None of the tested drugs caused significant changes in the rate of mitochondrial oxygen consumption, even at high concentrations. Except galantamine, all tested drugs were selective MAO-A inhibitors. Latrepirdine, donepezil, and 7-MEOTA were found to be the most potent MAO-A inhibitors. Succinate-induced mitochondrial hydrogen peroxide production was not significantly affected by the drugs tested. The direct effect of cognitives and nootropics used in the treatment of AD on mitochondrial respiration is relatively small. The safest drugs in terms of disturbing mitochondrial function appear to be piracetam and rivastigmine. The MAO-A inhibition by cognitives and nootropics may also participate in mitochondrial neuroprotection. The results support the future research aimed at measuring the effects of currently used drugs or newly synthesized drugs on mitochondrial functioning in order to understand their mechanism of action.     

A cross-sectional study of pandemic influenza health literacy and the effect of a public health campaign

ABSTRACT: BACKGROUND: To ascertain the understanding of 2009 pandemic (H1N1) influenza and relevant infection control measures in an emergency department population and to assess the effectiveness of education campaigns in informing the public about the pandemic. METHODS: Questionnaires were administered to patients, visitors, non-clinical staff and volunteers. Data were collected on knowledge, preventative measures, information sources, attitudes to government and media reporting, perceived seriousness, behaviour change and intended compliance with future measures. Results were used to construct an overall knowledge score. RESULTS: There were 252 participants. Traditional forms of mass media (138 [55%]) remained the principal information source. Approximately 70% (176) accurately described mode of transmission and recommended precautions and 68% (175) reported behaviour change because of the pandemic. Gaps in knowledge included failure to identify certain high risk groups. Recall of government campaigns was significantly associated with a higher knowledge score. 60% (151) thought that authorities and media had exaggerated the threat; only 40% (101) would comply with recommended measures in a future pandemic. CONCLUSIONS: The knowledge regarding pandemic influenza was high in this population and positively affected by official campaigns. Pandemic planning should address knowledge gaps and the impression that authorities had exaggerated the public-health threat.     

siRNAs containing 2′-fluorinated Northern-methanocarbacyclic (2′-F-NMC) nucleotides: in vitro and in vivo RNAi activity and inability of mitochondrial polymerases to incorporate 2′-F-NMC NTPs

We recently reported the synthesis of 2′-fluorinated Northern-methanocarbacyclic (2′-F-NMC) nucleotides, which are based on a bicyclo[3.1.0]hexane scaffold. Here, we analyzed RNAi-mediated gene silencing activity in cell culture and demonstrated that a single incorporation of 2′-F-NMC within the guide or passenger strand of the tri-N-acetylgalactosamine-conjugated siRNA targeting mouse Ttr was generally well tolerated. Exceptions were incorporation of 2′-F-NMC into the guide strand at positions 1 and 2, which resulted in a loss of the in vitro activity. Activity at position 1 was recovered when the guide strand was modified with a 5′ phosphate, suggesting that the 2′-F-NMC is a poor substrate for 5′ kinases. In mice, the 2′-F-NMC-modified siRNAs had comparable RNAi potencies to the parent siRNA. 2′-F-NMC residues in the guide seed region position 7 and at positions 10, 11 and 12 were well tolerated. Surprisingly, when the 5′-phosphate mimic 5′-(E)-vinylphosphonate was attached to the 2′-F-NMC at the position 1 of the guide strand, activity was considerably reduced. The steric constraints of the bicyclic 2′-F-NMC may impair formation of hydrogen-bonding interactions between the vinylphosphonate and the MID domain of Ago2. Molecular modeling studies explain the position- and conformation-dependent RNAi-mediated gene silencing activity of 2′-F-NMC. Finally, the 5′-triphosphate of 2′-F-NMC is not a substrate for mitochondrial RNA and DNA polymerases, indicating that metabolites should not be toxic.