Acquired changes in stomatal characteristics in response to ozone during plant growth and leaf development of bush beans (Phaseolus vulgaris L.) indicate phenotypic plasticity

Bush bean (Phaseolus vulgaris L.) lines 'S156' (O3-sensitive)/'R123' (O3-tolerant) and cultivars 'BBL 290' (O3-sensitive)/'BBL 274' (O3-tolerant) were used to study the effects of O3 on stomatal conductance (gs), density, and aperture size on leaf and pod surfaces with the objective of establishing links between the degree of plant sensitivity to O3 and plasticity of stomatal properties in response to O3. Studies in open-top chambers (OTCs) and in continuously stirred tank reactors (CSTRs) established a clear relationship between plant developmental stages, degrees of O3 sensitivity and gs: while 'S156' had higher gs rates than 'R123' earlier in development, similar differences between 'BBL 290' and 'BBL 274' were observed at later stages. Gs rates on the abaxial leaf surfaces of 'S156' and 'BBL 290', accompanied by low leaf temperatures, were significantly higher than their O3-tolerant counterparts. Exposure to O3 in CSTRs had greater and more consistent impacts on both stomatal densities and aperture sizes of O3-sensitive cultivars. Stomatal densities were highest on the abaxial leaf surfaces of 'S156' and 'BBL 290' at higher O3 concentrations (60 ppb), but the largest aperture sizes were recorded on the adaxial leaf surfaces at moderate O3 concentrations (30 ppb). Exposure to O3 eliminated aperture size differences on the adaxial leaf surfaces between sensitive and tolerant cultivars. Regardless of sensitivity to O3 and treatment regimes, the smallest aperture sizes and highest stomatal densities were found on the abaxial leaf surface. Our studies showed that O3 has the potential to affect stomatal plasticity and confirmed the presence of different control mechanisms for stomatal development on each leaf surface. This appeared to be more evident in O3-sensitive cultivars.     

Influence of ozone and ethylenediurea (EDU) on growth and yield of bean (Phaseolus vulgaris L.) in open-top field chambers

Greenhouse and ambient air experiments have shown ethylene diurea (EDU) to be a strong and specific protective suppressant of ozone injury in plants. To examine how EDU affects plant responses to various ozone (O(3)) levels under controlled field conditions, Phaseolus vulgaris L. cv. Lit was treated with 150 ppm EDU every 14 days and exposed in open-top chambers to charcoal-filtered air (CF), nonfiltered air (NF) or two cf treatments with ozone added. The ozone treatments were proportional additions of one (CF1) and two (CF2) times ambient ozone levels. The mean ozone concentrations in the CF, NF, CF1 and CF2 treatments were 0.98, 14.1, 14.98 and 31.56 nl litre(-1). A two-way split plot ANOVA revealed that shoot dry weight was significantly reduced by ozone. EDU treatment was highly significant for leaf dry weight, root dry weight and shoot dry weight, but not for pod dry weight; leading to a higher biomass of EDU-treated plants. Ozone/EDU interactions were significant for root weight only, indicating that EDU reduced growth suppression by ozone. These results show that EDU action on plant biomass could be interpreted as a delay in senescence since EDU-treated plants showed a significant decreased biomass loss even in the CF treatment.     

Effects of ethylenediurea and ozone on the antioxidative systems in beans (Phaseolus vulgaris L.)

To study the biochemical mechanism of EDU protection against ozone injury, peroxidase, ascorbate-dependent peroxidase, and catalase activities, and the contents of ascorbic acid, dehydroascorbic acid, malondialdehyde and soluble protein were measured in Phaseolus vulgaris L. cv. Lit exposed to ozone and ethylenediurea (EDU) in open-top chambers. Plants not treated with EDU showed foliar bronzing due to ozone, while EDU-treated plants were not affected. EDU application modified the reaction of biochemical parameters to ozone. Soluble protein content was elevated by EDU. Peroxidase activity increased with ozone exposure in untreated plants only, while ascorbate-dependent peroxidase activity was lower in EDU treated plants. Catalase activity decreased in EDU-untreated plants. The ratio of ascorbic acid to dehydroascorbic acid was significantly increased in EDU treated plants. These results suggest that EDU might induce ascorbic acid synthesis and therefore provide the plant with a very potent antioxidant. Or the content of hydrogen peroxide was reduced due to other unknown processes and caused a delay in foliar senescence, regardless of whether these processes were ozone-induced or due to natural aging processes.     

The rapid inhibition of root respiration after exposure of bean (Phaseolus vulgaris L.) plants to ozone

The metabolic activity of the roots was very sensitive to the changes induced in the leaves by O₃. Respiratory activity began to decrease well before visible injury appeared on the leaves, and the per cent reduction of respiration was much greater than the per cent leaf injury. The triphenyl tetrazolium chloride (TTC) staining technique revealed changes in root tips very quickly, was generally more sensitive to changes in respiratory activity and was a convenient technique for handling large numbers of samples. Reducing foliar injury from O₃ with low levels of SO₂ reduced the effects on the roots indicating the effect of O₃ is on processes in the leaf.     

A flux-based assessment of the effects of ozone on foliar injury, photosynthesis, and yield of bean (Phaseolus vulgaris L. cv. Borlotto Nano Lingua di Fuoco) in open-top chambers

Stomatal ozone uptake, determined with the Jarvis' approach, was related to photosynthetic efficiency assessed by chlorophyll fluorescence and reflectance measurements in open-top chamber experiments on Phaseolus vulgaris. The effects of O₃ exposure were also evaluated in terms of visible and microscopical leaf injury and plant productivity. Results showed that microscopical leaf symptoms, assessed as cell death and H₂O₂ accumulation, preceded by 3-4 days the appearance of visible symptoms. An effective dose of ozone stomatal flux for visible leaf damages was found around 1.33 mmol O₃ m⁻². Significant linear dose-response relationships were obtained between accumulated fluxes and optical indices (PRI, NDI, ΔF/F_m^′). The negative effects on photosynthesis reduced plant productivity, affecting the number of pods and seeds, but not seed weight. These results, besides contributing to the development of a flux-based ozone risk assessment for crops in Europe, highlight the potentiality of reflectance measurements for the early detection of ozone stress.     

Effects of ozone on 14C translocation velocity and growth of spring wheat (Triticum aestivum L.) exposed in open-top chambers

Growth and yield were reduced but (14)C translocation velocity was not affected by increasing levels of ozone in spring wheat exposed in open top chambers to the following treatments: charcoal filtered air (CF), non-filtered ambient air (NF), or NF with addition of 30 microl litre(-1) ozone, 8 h daily (NFO). Destructive harvests were performed at anthesis and at maturity. Parts of the flag leaf or the second leaf were exposed to (14)CO(2) in small cuvettes for 5 min before, during and after anthesis. The translocation velocity was followed by autoradiography and scintillation counting of the plants frozen and lyophilized at different times after labelling. The label was transported at the same velocity in all the treatments. Ozone induced changes in carbon allocation or partitioning should probably be explained as amounts of carbon transported (mg s(-1)), rather than as transportation velocity (mm s(-1)). The amount translocated may be governed by source conditions under O(3) stress: reduced healthy green biomass and photosynthesis, but perhaps also by impairment of phloem loading because of membrane damage.     

Effects of various ozone exposures on the susceptibility of bean leaves (Phaseolus vulgaris L.) to Botrytis cinerea

The effects of various ozone exposures in predisposing bean leaves (Phaseolus vulgaris L.) to Botrytis cinerea have been investigated under laboratory conditions. Seedlings of two bean cultivars were exposed to incremental ozone concentrations (120, 180 and 270 microg m(-3) for 8-h day(-1)) for five days and primary leaves were subsequently inoculated with conidia suspended in water or in an inorganic phosphate solution (Pi), and with mycelium. Ozone injury increased with increasing ozone concentration and was much higher in the ozone-sensitive cultivar 'Pros' than in the ozone-insensitive 'Groffy'. Ozone only increased the number of lesions on leaves of Pros after inoculation with either of the conidial suspensions. The Pi-stimulated infection in Groffy was reduced by the lower ozone concentrations. Ozone decreased lesion expansion after inoculation with mycelium. In a chronic fumigation experiment, plants of the two cultivars were exposed to 90 microg m(-3) (7-h day(-1)) and the primary and the oldest tree trifoliate leaves were inoculated after five and seven weeks of exposure. Ozone enhanced the senescence-related injury only in Pros. The number of lesions was not influenced by ozone for either cultivar, conidial suspension or inoculation date. Lesion expansion after inoculation with mycelium was generally reduced in exposed plants. Thus, contrasting effects of ozone on the susceptibility of bean leaves to B. cinerea were observed depending on the cultivar, the conidial suspension, the disease parameter and the ozone exposure pattern. In extrapolating the laboratory results to the field, it is suggested that episodic and chronic exposures to ambient ozone are of minor importance in increasing the susceptibility of bean leaves to B. cinerea.     

Responses of bean (Phaseolus vulgaris L. cv. Pros) to chronic ozone exposure at two levels of atmospheric ammonia

Plants of bean (Phaseolus vulgaris cv. Pros) were exposed to a range of O3 concentrations up to 70 nl litre(-1) for 9 h day(-1) in the presence (45 nl litre(-1)) and absence (21 nl litre(-1)) of enhanced NH3 in 12 open-top chambers. Treatment effects on visible injury, growth and yield were assessed after 49 (intermediate harvest) and 62 days of exposure (final harvest). The proportion of leaves with visible injury at final harvest increased with increasing concentrations of O3. Enhanced NH3 did not cause any symptoms and did not affect injury by O3. The estimated seasonal mean concentration corresponding with 5% injury was circa 23 nl litre(-1) O3. Biomass production and green pod yield decreased with increasing concentrations of O3 and were generally stimulated by enhanced NH3 at both harvests. Adverse effects of O3 on biomass and pod yield did not depend on the NH3 level. Relative yield response to increasing 9-h daily mean O3 concentrations was nonlinear and yield losses of 5 and 10% were calculated to occur at seasonal daytime mean concentrations of 27 and 33 nl litre(-1) O3, respectively. Linear regression showed that the Accumulated exposures Over a Threshold of 30 (AOT30) and 40 nl litre(-1) (AOT40) O3 performed equally well. The estimated accumulated O3 exposures corresponding with a yield loss of 5% were 1600 nl litre(-1) h for AOT30 and 400 nl litre(-1) h for AOT40. The results are discussed in relation to the long-term critical level that is used as a guideline to protect crops against adverse effects by O3.     

Residues and dissipation kinetic of abamectin,chlorfenapyr and pyridaben acaricides in green beans (Phaseolus vulgaris L.) under field conditions using QuEChERS method and HPLC

Abstract

The current study estimated the dissipation rates of abamectin, chlorfenapyr and pyridaben acaricides in pods of green beans (Phaseolus vulgaris L.) under field conditions in Egypt. Pesticides were extracted and cleaned-up by QuEChERS method and were analyzed by HPLC. The dissipation of these acaricides followed the first order kinetics model with half-life (t_1/2) values 1.00, 3.50 and 1.50?days for abamectin, chlorfenapyr and pyridaben, respectively. The lowest residues, at different time intervals of field application rate of each pesticide, were observed with abamectin followed by pyridaben and then chlorfenapyr. Pre-harvest intervals (PHIs) were 10.00, 13.50 and 6.00?days for abamectin, chlorfenapyr and pyridaben, respectively and were below the established European maximum residue limits (EU MRLs) 10–14, 14–21 and 7–10?days after application, respectively. If the fresh pods will be consumed after harvest, it is expected that the presence of these pesticides in the food will have a negative impact on human health. Therefore, the elimination of the residues of these harmful pesticides must be carried out.     

Growth and yield responses of spring wheat (Triticum aestivum L. CV. Turbo) grown in open-top chambers to ozone and water stress

Spring wheat (Triticum aestivum L.) cv. Turbo was exposed to different levels of ozone and water supply in open-top chambers in 1991. The plants were grown either in charcoal filtered air (CF), not filtered air (NF), in charcoal filtered air with proportional addition of ambient ozone (CF1), or in charcoal filtered air with twice proportional addition of ambient ozone (CF2). The mean seasonal ozone concentrations (24 h mean) were 2.3, 20.6, 17.3, and 24.5 nl litre(-1) for CF, NF, CF1, and CF2 treatments, respectively. Ozone enhanced senescence and reduced growth and yield of the wheat plants. At final harvest, dry weight reductions were mainly due to reductions in ear weight. Grain yield loss by ozone mainly resulted from depressions of 1000 grain weight, whereas numbers of ears per plant and of grains per ear remained unchanged. Pollutants other than ozone did not alter the response to ozone, as was obvious from comparisons between CF1 and NF responses. Water stress alone did not enhance senescence, but also reduced growth and yield. However, yield loss mainly resulted from reductions in the number of ears per plant; 1000 grain weight was not influenced by water stress. No water supply by ozone treatment interactions were detected for any of the estimated parameters.     

Comparison of a diurnal vs steady-state ozone exposure profile on growth and yield of oilseed rape (Brassica napus L.) in open-top chambers in the Yangtze Delta, China

Most available exposure-response relationships for assessing crop loss due to elevated ozone (O₃) have been established using data from chamber and open-top chamber experiments, using a simulated constant O₃ concentration exposure (square wave), which is not consistent with the diurnal variation of O₃ concentration that occurs in nature. We investigated the response of oilseed rape (Brassica napus L.) to O₃ as affected by two exposure regimes: one with a diurnal variation (CF100D) and another with a constant concentration (CF100). Although the two exposure regimes have the same mean O₃ concentration and accumulated O₃ concentration above 40 ppb (AOT40), our results show that O₃ at CF100D reduced biomass and number of pods/plant more than O₃ at CF100. Both O₃ exposures resulted in larger seed weights/100 pods compared to CF. Numbers of seeds/100 pods were reduced by CF100, while numbers of seeds/100 pods in the CF100D chambers were comparable to those in CF. Our results suggest that chamber experiments that use a constant O₃ exposure may underestimate O₃ effects on biomass and yields.     

Yield and grain quality of spring wheat (Triticum aestivum L., cv. Drabant) exposed to different concentrations of ozone in open-top chambers

Spring wheat (Triticum aestivum L., cv. Drabant) was exposed to different concentrations of ozone in open-top chambers for two growing seasons, 1987 and 1988, at a site located in south-west Sweden. The chambers were placed in a field of commercially grown spring wheat. The treatments were charcoal-filtered air (CF), non-filtered air (NF) and non-filtered air plus extra ozone (NF(+)). In 1988, one additional ozone concentration (NF(++)) was used. Grain yield was affected by the ozone concentration of the air. Air filtration resulted in an increase in grain yield of about 7% in both years, compared to NF. The addition of ozone (NF(+), NF(++)) reduced grain yield and increased the content of crude protein of the grain in both years. Filtration of the air had no significant effect on the content of crude protein, compared to NF. The results showed a strong positive chamber effect on grain yield in the cold and wet summer of 1987. In 1988, there was no net chamber effect on grain yield. The relative differences between the CF, NF and NF(+) treatments with respect to grain yield were of the same magnitude in the two years, despite the very different weather conditions.     

Effects of ozone, acid mist and soil characteristics on clonal Norway spruce (Picea abies (L.) Karst.)--an introduction to the joint 14 month tree exposure experiment in closed chambers

This paper introduces a series of publications referring to a single 14-month laboratory study testing the hypothesis that the recent decline of Norway spruce (Picea abies (L.) Karst.) at higher elevations of the Bavarian Forest and comparable forests in medium-range mountains and in the calcareous Alps is caused by an interaction of elevated ozone concentrations, acid mist and site-specific soil (nutritional) characteristics. The effect of climatic extremes, a further important factor, was not included as an experimental variable but was considered by testing of the frost resistance of the experimental plants. Results of these individual studies are presented and discussed in the following 14 papers. Plants from six pre-selected clones of 3-year-old Norway spruce (Picea abies (L.) Karst.) were planted in April 1985 in an acidic soil from the Bavarian Forest, or a calcareous soil from the Bavarian Alps. After a transition period, plants were transferred, in July 1986, into four large environmental chambers and exposed for 14 months to an artificial climate and air pollutant regime based on long-term monitoring in the Inner Bavarian Forest. The climatic exposure protocol followed realistic seasonal and diurnal cycles (summer maximum temperature, 26 degrees C; total mean temperature, 9.8 degrees C; winter minimum, -14 degrees C; mean relative humidity, 70%; maximum irradiance, 500 W m(-2); daylength summer maximum, 17 h; winter minimum, 8 h). Plants were fumigated with ozone, generated from pure oxygen (control: annual mean of 50 microg m(-3); pollution treatment: annual mean of 100 microg m(-3) with 68 episodes of 130-360 microg m(-3) lasting 4-24 h), and background concentrations of SO(2) (22 microg m(-3)) and NO(2) (20 microg m(-3)); windspeed was set at a constant 0.6 m s(-1). Plants were additionally exposed to prolonged episodes of misting at pH 5.6 (control) and pH 3.0 (treatment). Simulation of the target climatic and fumigation conditions was highly reliable and reproducible (temperature +/-0.5 degrees C; rh+/-10%; ozone+/-10 microg m(-3);SO(2) and NO(2)+/-15 microg m(-3)).     

Responses of spring wheat (Triticum aestivum L.) to ozone produced by either electric discharge and dry air or by UV-lamps and ambient air

The aim of the present study was to examine if ozone produced similar effects on spring wheat growth with and without small amounts of nitrogen oxides. Two methods were used to produce ozone: the first method consisted of dry pressurized air fed to an electric discharge generator generating the byproducts, N2O5 and N2O, the second method consisted of ambient air fed to UV-lamps. Two spring wheat cultivars (Triticum aestivum L. cvs Minaret and Eridano) were exposed in small open-top chambers to charcoal-filtered air, non-filtered ambient air, and non-filtered ambient air with the addition of ozone for 8 h (0900 to 1700 h) daily, for five weeks. Plants were harvested every week. The growth of Minaret was shown to be more sensitive to O3 than that of Eridano. Leaf senescence increased with increasing ozone level in both cultivars. The total above-ground biomass dry weight decreased with increasing ozone concentration in Minaret, but not in Eridano. The Minaret plants reacted with more damaged leaf dry weight and inhibition of growth when O3 was produced by UV-lamps than when O3 was produced by air fed to an electric discharge generator. This could be explained by more nitrogen content per plant but not by increased nitrogen concentration in plant tissue in plants exposed to increased O3 and small amounts of incidental nitrogen oxides.     

Differential response of CO2 uptake parameters of soil- and sand-grown phaseolus vulgaris (L.) plants to absorbed ozone flux

Shoots of a soil- or sand-grown dwarf bean variety were exposed to O(3) concentrations in the range of 500 to 900 ppb for up to 5 h. The measured exchange rates of water vapor and CO(2) during exposures were used to calculate stomatal and mesophyll conductances averaged across all leaves. Changes in conductances were related to exposure duration and absorbed O(3) totals (AOT). Both conductances were more sensitive to AOT in sand-grown plants, which also had more visible injury under comparable AOT values. Measurements of the relationship between CO(2) exchange and internal CO(2) concentration of single leaflets of treated plants also showed greater sensitivity of CO(2)-saturated photosynthesis in sand-grown plants. Diffusional processes were not likely to have been the cause of dissimilar responses because the O(3) absorption rate was lower in sand-grown plants. A difference in the scaveninng capacities in cells is suggested to be the cause of the differences in sensitivity to acute O(3) exposure.     

Effects of ozone, acid mist and soil characteristics on clonal Norway spruce (Picea abies (L.) Karst.)--overall results and conclusions of the joint 14 month tree exposure experiment in closed chambers

This paper summarizes and evaluates the main findings of 14 preceding papers related to the joint 14-month tree-exposure experiment carried out by the 'Munich Working Party on Air Pollution' at the GSF, Munich, FRG, from July 1986 to September 1987. The experiment tested the hypothesis that an interaction of ozone/acid mist/soil/extreme climatic conditions is the cause of decline of Norway spruce (Picea abies (L.) Karst.) at higher altitudes of the Inner Bavarian Forest. The main findings of the individual studies are presented and their implications for the hypothesis are discussed. Clear effects of soil and genetic factors (differences between clones), for example on growth and frost resistance were found. Treatment with O(3)/acid mist was shown to have effects on plant biochemistry, physiology, histology/ cytology, and growth. The wide scattering of these effects, and the lack of a consistent pattern of response across all clones does not permits a firm conclusion on the validity of the experimental hypothesis. These effects were not confounded by the nutrient stresses imposed during the initial exposure period and were not found to be cumulative during repeated treatments, as was proposed by the hypothesis. It is concluded that the experimental evidence does not indicate that ozone/acid mist are major factors to explain the Norway spruce decline on acidic sites at higher altitudes of the Inner Bavarian Forest and probably similar forest areas.     

Seasonal and diurnal gas exchange differences in ozone-sensitive common milkweed (Asclepias syriaca L.) in relation to ozone uptake

Stomatal conductance and net photosynthesis of common milkweed (Asclepias syriaca L.) plants in two different soil moisture regimes were directly quantified and subsequently modeled over an entire growing season. Direct measurements captured the dynamic response of stomatal conductance to changing environmental conditions throughout the day, as well as declining gas exchange and carbon assimilation throughout the growth period beyond an early summer maximum. This phenomenon was observed in plants grown both with and without supplemental soil moisture, the latter of which should theoretically mitigate against harmful physiological effects caused by exposure to ozone. Seasonally declining rates of stomatal conductance were found to be substantial and incorporated into models, making them less susceptible to the overestimations of effective exposure that are an inherent source of error in ozone exposure indices. The species-specific evidence presented here supports the integration of dynamic physiological processes into flux-based modeling approaches for the prediction of ozone injury in vegetation.     

Response to ambient ozone of two white clover (Trifolium repens L.cv. "Regal") clones, one resistant and one sensitive, grown in a Mediterranean environment

Two clones of white clover (Trifolium repens L.) differing in ozone tolerance were grown in southern Italy during 1997 and 1998 to study the effects of ambient ozone exposure on yield, leaf morphology and water use. Ambient ozone levels were high in both years with values exceeding the threshold for leaf injury reported in the literature. In both years ozone injury was observed on the sensitive clone (NC-S) but not on the resistant one (NC-R), and leaf and stolon dry matter production was significantly lower in NC-S than in NC-R. However, it cannot be excluded that other factors, such as high temperature, interacted with the effect of ozone on biomass production. The clones differed in morphological characteristics. Lower total leaf area in NC-S plants was due to a smaller number of leaves per plant, but the average area per leaf was higher in NC-S. Specific leaf weight and net assimilation rate were higher in the more productive clone (NC-R). Cumulative plant water use was higher in NC-R in each growth period because of the larger leaf area; by contrast, water use per unit leaf area was higher in NC-S, indicating higher leaf conductance to water vapour. The results suggest that ozone significantly reduces the yield of sensitive white clover plants under well-watered conditions, and that the difference in ozone tolerance between clover clones is related to differences in leaf morphology and water use.     

Responses of photosynthesis,antioxidant enzymes,and related gene expression to nicosulfuron stress in sweet maize (Zea mays L.)

Environmental Science and Pollution Research - Weed control in maize (Zea mays L.) crops is usually undertaken using the postemergence herbicide nicosulfuron. The toxicity of nicosulfuron on maize,...     

Physiological and chemical response of the lichen, Flavoparmelia caperata (L.) Hale, to the urban environment of Kolkata, India

The present study was focused on the effect of increasing urbanization including industrial and traffic activity on the accumulation of heavy metals and possible damage of selected physiological parameters (composition of assimilation pigments, membrane lipid peroxidation, and membrane integrity) of an epiphytic foliose lichen, Flavoparmelia caperata (L.) Hale. The lichen samples were collected from three different localities in and around Kolkata, India, two sites being from the urban area and one from the relatively non-polluted sub-urban area. The results showed that thalli from the urban sites have significantly higher concentrations of Fe, Cr, Cu, Zn, and Pb compared to those collected from the sub-urban site. Physiological parameters of damage also exhibited stress symptoms in thalli from the urban sites—decreased chlorophyll a indicating less photosynthetic efficiency, and increase in lipid peroxidation and electrolyte conductivity indicating cell membrane injuries. Correlation studies among metals pinpointed vehicular traffic as the main source of pollution in this area.