Salt stress of two rice varieties: root border cell response and multi-logistic quantification.

How to capture the rice varieties salt stress sensitivity? Here, we measure responses of root border cells (1 day, ± 60 mM NaCl) and apply multi-logistic quantification of growth variables (21 days, ± 60 mM NaCl) to two rice varieties, salt-sensitive IR29 and tolerant Pokkali. Thus, logistic models determine the maximum response velocities (Vmax) and times of half-maximum (T0) for root border cell (RBC) and growth parameters. Thereof, seven variables show logistic models (0.58 < R ≤ 1) and monotonous responses in both Pokkali and IR29: root to shoot ratio by water content, primary root length, shoot water, adventitious root number, shoot dry and fresh weight, and root dry weight. Moreover, the regression to lognormal distribution (R = 0.99) of these seven Vmax fractionated by T0 represents the rice variety's comprehensive response. Its quotient IR29/Pokkali is peaking at 98-fold higher velocity of IR29, thus capturing the variety's sensitivity. Consequently, our finding of 66-fold higher Vmax of primary root length response of IR29 indicates an essential salt sensor in the root, including RBC. Finally, the effects of salt stress on RBC confirm multi-logistic quantification, showing 36% decrease of RBC mucilage layer in IR29, without change in Pokkali. Inversely, RBC number of Pokkali increases 43% without change in IR29. Briefly, this suggests both RBC and multi-logistic quantification for the screening for salt tolerance in two thousand rice varieties.

Salt stress in rice: multivariate analysis separates four components of beneficial silicon action.

How many subcellular targets of the beneficial silicon effect do exist in salt-stressed rice? Here, we investigate the effects of silicon on the different components of salt stress, i.e., osmotic stress, sodium, and chloride toxicity. These components are separated by multivariate analysis of 18 variables measured in rice seedlings (Oryza sativa L.). Multivariate analysis can dissect vectors and extract targets as principal components, given the regressions between all variables are known. Consequently, the exploration of 153 correlations and 306 regression models between all variables is essential, and regression parameters for variables of shoot (silicon, sodium, chloride, carotenoids, chlorophylls a and b, and relative growth rate) and variables of shoot and root (hydrogen peroxide, ascorbate peroxidase (APX), catalase (CAT), fresh weight, dry weight, root-to-shoot ratio) are determined. The regression models [log (y) = y0 + a × log (x)] are confirmed by variance analysis of global goodness of fits (p < 0.0001). Thereby, logarithmic transformation yields linearization for multivariate analysis by Pearson's correlation. Four principal components are extracted: two targets of osmotic stress, one target of sodium toxicity, and one target of chloride toxicity. Thereby, silicon improves salt tolerance by increasing APX and CAT activities and decreasing hydrogen peroxide, salt ion accumulation, photosynthetic pigment losses, and growth inhibition. Salt stress increases silicon uptake pointing to a physiological regulation of plant salt stress in the presence of silicon. This mechanism and its four components are promising targets for further agricultural application.

"Singing in the Tube"--audiovisual assay of plant oil repellent activity against mosquitoes (Culex pipiens).

Plant essential oils have been suggested as a promising alternative to the established mosquito repellent DEET (N,N-diethyl-meta-toluamide). Searching for an assay with generally available equipment, we designed a new audiovisual assay of repellent activity against mosquitoes "Singing in the Tube," testing single mosquitoes in Drosophila cultivation tubes. Statistics with regression analysis should compensate for limitations of simple hardware. The assay was established with female Culex pipiens mosquitoes in 60 experiments, 120-h audio recording, and 2580 estimations of the distance between mosquito sitting position and the chemical. Correlations between parameters of sitting position, flight activity pattern, and flight tone spectrum were analyzed. Regression analysis of psycho-acoustic data of audio files (dB[A]) used a squared and modified sinus function determining wing beat frequency WBF ± SD (357 ± 47 Hz). Application of logistic regression defined the repelling velocity constant. The repelling velocity constant showed a decreasing order of efficiency of plant essential oils: rosemary (Rosmarinus officinalis), eucalyptus (Eucalyptus globulus), lavender (Lavandula angustifolia), citronella (Cymbopogon nardus), tea tree (Melaleuca alternifolia), clove (Syzygium aromaticum), lemon (Citrus limon), patchouli (Pogostemon cablin), DEET, cedar wood (Cedrus atlantica). In conclusion, we suggest (1) disease vector control (e.g., impregnation of bed nets) by eight plant essential oils with repelling velocity superior to DEET, (2) simple mosquito repellency testing in Drosophila cultivation tubes, (3) automated approaches and room surveillance by generally available audio equipment (dB[A]: ISO standard 226), and (4) quantification of repellent activity by parameters of the audiovisual assay defined by correlation and regression analyses.

Cell-specific association of heat shock-induced proton flux with actin ring formation in Chenopodium cells: comparison of auto- and heterotroph cultures.

A comparison of the responses of extracellular pH, buffering capacity and actin cytoskeleton in autotroph and heterotroph Chenopodium rubrum cells to heat shock revealed cell-specific reactions: alkalinization caused by the heat shock at 25-35 degrees C was higher in heterotroph cells and characterized by heat shock-induced changes in the actin cytoskeleton and ring formation at 35-37 degrees C. Rings (diameter up to 3 mum) disappeared and extracellular pH recovered after the heat-shocked cells were transferred into control medium. At 41 degrees C, no rings but a network of coarse actin filaments were induced; at higher temperatures, fragmentation of the actin cytoskeleton and release of buffering compounds occurred, indicating sudden membrane leakage at 45-47 degrees C. The calcium chelator EGTA [ethylene-glycol-bis(beta-aminoethyl-ether)-N,N,N',N'-tetraacetic-acid] increased the frequency of heat shock-induced rings. Ionophore (10 microM nigericin) and the sodium/proton antiport blocker [100 microM 5-(N-ethyl-N-isopropyl)-amiloride] mimicked the effect of the 37 degrees C heat shock. The cytoskeleton inhibitors latrunculin B, cytochalasin D and 2,3-butanedione monoxime inhibited ring formation but not alkalinization. In autotroph cells, the treatment with nigericin (10 microM) produced rings, although the actin cytoskeleton was not affected by temperatures up to 45 degrees C. We conclude that Chenopodium cells express a specific temperature sensor that has ascendancy over the organization of the actin cytoskeleton; this is probably a temperature- and potential-sensitive proton-transporting mechanism that is dependent on the culture conditions of the heterotroph cells.

Extracellular alkaline phosphatase is a sensitive marker for cellular stimulation and exocytosis in heterotroph cell cultures of Chenopodium rubrum.

We investigated the response of extracellular phosphatase to heat shock in heterotrophic Chenopodium rubrum L. cell cultures. Surprisingly, in contrast to the generally used acid phosphatase, an extracellular alkaline phosphatase showed the most sensitive response. This phosphatase was characterized as a marker for cellular stimulation by its high correlations with induced changes of extracellular pH: 10microM nigericin (correlation coefficient r=0.91), 100microM salicylic acid (r=0.84), heat shock 5min 37 degrees C (r=0.79), and heat shock after pre-treatment with 5microM fusicoccin (r=0.92) or 0.5% ethanol (r=0.90). Cellular stimulation was estimated with concentrations of acids and bases, yielding similar levels of pH change (0.5 pH) in cell-free supernatant: salicylic acid (200microM), benzoic acid (600microM), HCl (140microM), NaOH (100microM), and KOH (100microM). The Golgi apparatus inhibitor Brefeldin A (200microM) reduced the heat-shock-induced phosphatase (-33%). The pH optimum of heat-shock-induced phosphatase was 3; however, there the proportion of constitutive phosphatase was higher than at pH 8-9.5, indicating different pH dependence of constitutive and induced activity. Thus, heat-shock-induced phosphatase was characterized by alkaline activity with inhibitors (10microM molybdate: -52%, 2.5mM phosphate: -64%, 10microM ZnCl(2): -82%), substrates (2.5mM, tyrosine phosphate: 255pkat g(-1), p-nitrophenyl phosphate: 92pkat g(-1), serine phosphate: 0, threonine phosphate: 0), Hill coefficient (nH=1.4) indicating two binding sites, and the extent of heat-shock stimulation (p-nitrophenyl phosphate: +190%, tyrosine phosphate: +180%). SDS-PAGE showed a correlation of alkaline phosphatase with the heat-shock-induced release of highly N-glycosylated 53kDa protein, detected by peroxidase-labeled concanavalin A affinoblotting after endoglycosidase H treatment. The 53kDa protein showed no in-gel phosphatase activity after SDS-PAGE and regeneration treatment, in contrast to a putative dimer (105kDa).