Inheritance of ambient ozone insensitivity in common bean (Phaseolus vulgaris L.)

The inheritance of ozone (O(3)) insensitivity in common bean (Phaseolus vulgaris L.) was evaluated using F(2) and F(3) populations under ambient conditions. This study was conducted over two growing seasons (1987, 1988) at Virginia State University, Randolph Research Farm, Petersburg, Virginia. Two populations were obtained by crossing insensitive plant introductions with sensitive commercial cultivars. Ratings on the scale of 1 to 5 (1 = 0 to 20% leaf injury, 2 = 21 to 40%, 3 = 41 to 60%, 4 = 61 to 80%, and 5 > 80%) were made on 160 F(2), F(3) progenies, and parental lines. Population mean injury ratings were recorded and estimates of genotypic, environmental, and phenotypic variances were computed. Estimates of heritability in the broadsense and of genetic advance were calculated for each population using F(2) and family component variance methods. Population means of the F(2) and F(3) progenies were not significantly different from their mid-parent values, suggesting that genetic variance was primarily additive. Broad-sense heritability estimates using F(2) variance method ranged from 51.4 to 70.5% and using family component variance method ranged from 62.1 to 75.6%. In this study, the computed genetic advance values closely parallel those of heritability estimated values. The high heritable nature of insensitivity would indicate that effective levels of insensitivity could be transferred to agronomically superior cultivars in a relatively short time.     

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.     

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.     

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 nitrogen dioxide on growth and yield of black turtle bean (Phaseolus vulgaris L.) cv. 'Domino'

Twenty-six-day-old black turtle bean cv. 'Domino' plants were exposed to nitrogen dioxide (0.0, 0.025, 0.05 and 0.10 microl liter(-1)), 7 h per day for 5 days per week for 3 weeks, under controlled environment. Data were collected on net photosynthesis rate (PN), stomatal resistance (SR), and dark respiration rate (DR), immediately after exposure, 24 h after the termination of exposure and at maturity (when the leaves had just started turning yellow), using a LICOR 6000 Portable Photosynthesis System. Chlorophyll-a (Ch-a), chlorophyll-b (Ch-b), total chlorophyll (tot-Ch) and leaf nitrogen were measured immediately after exposure and at maturity. Growth characteristics-relative growth rate (RGR), net assimilation rate (NAR), leaf area ratio (LAR) and root: shoot ratio (RSR)-were computed for treated plants. Net photosynthesis rate increased by 53% in 0.10 microl liter(-1) NO2 treated plants immediately after exposure compared to control plants. Dark respiration rates were also higher in treated plants. Ch-a, Ch-b and tot-Ch showed significant increases with 0.1 microl liter(-1) NO2 treatment immediately after exposure. Foliar nitrogen content showed an increase in treated plants both immediately after exposure and at maturity. Increases were also seen in RGR and NAR. Plant yield increased by 86% (number of pods), 29% (number of seeds) and 46% (weight of seeds), respectively. Nitrogen dioxide stimulated the overall plant growth and crop yield.     

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.     

Injury response of Phaseolus vulgaris to ozone flux density

This study describes a quantitative relationship between mean O₃ flux density and the length of exposure needed for the occurrence of visual injury to Phaseolus vulgaris L. Similar relationships were found for 14 day old and 6 week old plants using a whole leaf gas exchange cuvette system. Cultivars Seafarer (O₃ sensitive) and Gold Crop (O₃ resistant) exhibited similar responses at flux densities > 3 mg m⁻² h⁻¹ but only Seafarer was injured below this flux density. O₃ concentration and length of exposure period alone did not contain sufficient information to describe the onset of visual foliar injury. The use of O₃ concentrations in excess of normal ambient conditions compensated for low leaf conductances so that flux densities in the cuvette were similar to those found in the field.     

Analysis of the uptake of atmospheric ammonia by leaves of Phaseolus vulgaris L.

Individual leaves of Phaseolus vulgaris L. were exposed for 9 h in a leaf chamber to different NH₃ concentrations at different light intensities. The rates of NH₃-uptake, transpiration and photosynthesis were measured simultaneously. The flux density of NH₃ increased linearly with concentration in the range of 4–400μg m⁻³. Flux densities also increased with light intensity. Resistance analysis indicated that NH₃ transport into the leaf is via the stomata: transport via the cuticle is negligible under the experimental conditions. There is no internal resistance against NH₃ transport. The NH₃ flux was found not to influence the photosynthesis.     

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.     

Responses of sensitive and tolerant bush beans (Phaseolus vulgaris L.) to ozone in open-top chambers are influenced by phenotypic differences, morphological characteristics, and the chamber environment

Responses of bush bean (Phaseolus vulgaris L.) lines 'S156' (O(3)-sensitive) and 'R123' (O(3)-tolerant), and cultivars 'BBL 290' (O(3)-sensitive) and 'BBL 274' (O(3)-tolerant) to ambient ozone (O(3)) were investigated during the 2001 and 2002 growing seasons. Seedlings were grown in pots inside open-top chambers (OTCs), with charcoal filtered (CF) and non-filtered (NF) ambient air, and in non-chambered ambient air (AA) plots. Growth parameters from individual plants were evaluated after harvests at the end of vegetative (V(4)) and reproductive (R(10)) growth phases. Results at V(4) indicated that CF did not provide additional benefits over NF in 'S156' in 2001 and 2002. In contrast, exposure to CF significantly impaired the growth of 'R123'. At the end of R(10), 'S156' produced more pods, most of which remained immature, and contained fewer seeds or were more frequently aborted, whereas pods produced in 'R123' reached pod maturation and senescence more consistently. Despite increased seed weights inside the OTCs, as observed in 'S156', differences between the two lines were insignificant when grown outside OTCs. Results from the 'BBL 290'/'BBL 274' pair, especially at V(4) phase, remained inconclusive. Plant morphological characteristics, variabilities in environmental conditions, and 'chamber effects' inside OTCs were influential in determining plant response to ambient O(3).     

Field controlled experiments on the physiological responses of maize (Zea mays L.) leaves to low-level air and soil mercury exposures

Thousands of tons of mercury (Hg) are released from anthropogenic and natural sources to the atmosphere in a gaseous elemental form per year, yet little is known regarding the influence of airborne Hg on the physiological activities of plant leaves. In the present study, the effects of low-level air and soil Hg exposures on the gas exchange parameters of maize (Zea mays L.) leaves and their accumulation of Hg, proline, and malondialdehyde (MDA) were examined via field open-top chamber and Hg-enriched soil experiments, respectively. Low-level air Hg exposures (<50 ng m^?3) had little effects on the gas exchange parameters of maize leaves during most of the daytime (p?>?0.05). However, both the net photosynthesis rate and carboxylation efficiency of maize leaves exposed to 50 ng m^?3 air Hg were significantly lower than those exposed to 2 ng m^?3 air Hg in late morning (p?<?0.05). Additionally, the Hg, proline, and MDA concentrations in maize leaves exposed to 20 and 50 ng m^?3 air Hg were significantly higher than those exposed to 2 ng m^?3 air Hg (p?<?0.05). These results indicated that the increase in airborne Hg potentially damaged functional photosynthetic apparatus in plant leaves, inducing free proline accumulation and membrane lipid peroxidation. Due to minor translocation of soil Hg to the leaves, low-level soil Hg exposures (<1,000 ng g^?1) had no significant influences on the gas exchange parameters, or the Hg, proline, and MDA concentrations in maize leaves (p?>?0.05). Compared to soil Hg, airborne Hg easily caused physiological stress to plant leaves. The effects of increasing atmospheric Hg concentration on plant physiology should be of concern.     

Effects of ozone on the foliar histology of the mastic plant (Pistacia lentiscus L.)

An open-top chamber study was conducted to investigate the tissue and cellular-level foliar effects of ozone (O3) on a Mediterranean evergreen species, the mastic plant (Pistacia lentiscus L.). Plants were exposed at three different O3 levels, and leaf samples were collected periodically from the beginning of the exposure. Although no visible foliar injury was evident, alterations of the plastids and vacuoles in the mesophyll were observed. Senescence processes were accelerated with an anomalous stacking of tannin vacuoles, and a reduction in the size and number of the chloroplasts. Overall, most of the modifications induced by O3 were consistent with previously reported observations on deciduous broadleaf species, with the exception of alterations in the cells covering the secretory channels, reported here as a new finding. Comments on the feasibility of using microscopy to validate O3 related field observations and subtle foliar injury are also given.     

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.     

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 on chlorophyll and quantum yield of tobacco (Nicotiana tabacum L.) varieties

Plants of Bel-W3 and of seven commercial tobacco varieties (Nicotiana tabacum L.) were exposed to two relatively low ozone concentrations (90 or 135 ppb) for 20 consecutive days, for 8 h per day. Ozone caused necrotic and chlorotic spots, acceleration of leaf senescence, depression of photosynthetic mechanism, chlorophyll diminution and greater destruction of chl a than of chl b. The higher sensitivity of chl a was also confirmed by exposure of segments of leaves in test tubes to high ozone concentration (>1000 ppb) as well as by bubbling of ozone in extracts of chlorophyll in vitro. The quantum yield (QY) of photosynthesis was positively correlated with the chlorophyll content and negatively correlated with the visible injury and the chl b/a ratio.     

Foliar injury response of petunia and kidney bean to simultaneous and alternate exposures to ozone and pan

Petunia at about 6 weeks old and kidney bean at two growing stages (6–7 days old and 16–18 days old) were exposed separately to O₃, (0–0.40 ppm) and PAN (0–0.25 ppm) for 4 h and to the mixture for the same time. In addition, petunia was exposed to O, (0.10–0.40 ppm) and then PAN (0.010−0.040 ppm) for 4 h, respectively. Foliar injury of petunia and kidney bean in exposures to the mixtures of O₃ and PAN was significantly smaller than that induced by each oxidant, with the exception of PAN injury on young leaves of 16–18 day-old kidney bean. The percentage of foliar injury caused by either of the mixed pollutants decreased with an increase of the concentration of the other oxidant, and was found to approximate a logarithmic function of the combined pollutant concentrations expressed as O₃, minum PAN or vice versa. Alternate exposures caused no additive or synergistic injuries.     

Impact of three different fungicides on fungal epi- and endophytic communities of common bean (Phaseolus vulgaris) and broad bean (Vicia faba)

In this study, the impacts of three different fungicides to fungal phyllosphere communities on broad bean (Vicia faba, Fabaceae) and common bean (Phaseolus vulgaris, Fabaceae) were analyzed. The fungicides included copper, sulfur, and azoxystrobin. The plants were sowed, grown, and treated under conditions occurring in conventional and organic farming. A culture-based approach was used to identify changes in the phyllosphere fungal community after the treatment. Different effects on species richness and growth index of the epiphytic and endophytic communities for common bean and broad bean could be shown. Treatments with sulfur showed the weakest effect, followed by those based on copper and the systemic azoxystrobin, which showed the strongest effect especially on endophytic communities. The epiphytic fungal community took five weeks to recover after treatment with azoxystrobin. However, the effect of azoxystrobin on the endophytic community lasted more than five weeks. Finally, the data suggest that the surface structure of the host leaves have a huge impact on the mode of action that the fungicides exert.     

Bioremediation model for atrazine contaminated agricultural soils using phytoremediation (using Phaseolus vulgaris L.) and a locally adapted microbial consortium

The objective of the present study was to examine a biological model under greenhouse conditions for the bioremediation of atrazine contaminated soils. The model consisted in a combination of phytoremediation (using Phaseolus vulgaris L.) and rhizopheric bio-augmentation using native Trichoderma sp., and Rhizobium sp. microorganisms that showed no inhibitory growth at 10,000 mg L^?1 of herbicide concentration. 33.3 mg of atrazine 50 g^?1 of soil of initial concentration was used and an initial inoculation of 1 × 10⁹ UFC mL^?1 of Rhizobium sp. and 1 × 10⁵ conidia mL^?1 of Trichoderma sp. were set. Four treatments were arranged: Bean + Trichoderma sp. (B+T); Bean + Rhizobium sp. (BR); Bean + Rhizobium sp. + Trichoderma sp. (B+R+T) and Bean (B). 25.51 mg of atrazine 50 g^?1 of soil (76.63%) was removed by the B+T treatment in 40 days (a = 0.050, Tukey). This last indicate that the proposed biological model and methodology developed is useful for atrazine contaminated bioremediation agricultural soils, which can contribute to reduce the effects of agrochemical abuse.     

Effects of ozone on chlorophyll and quantum yield of tobacco (Nicotiana tabacum L.) varieties

Plants of Bel-W3 and of seven commercial tobacco varieties (Nicotiana tabacum L.) were exposed to two relatively low ozone concentrations (90 or 135 ppb) for 20 consecutive days, for 8 h per day. Ozone caused necrotic and chlorotic spots, acceleration of leaf senescence, depression of photosynthetic mechanism, chlorophyll diminution and greater destruction of chl a than of chl b. The higher sensitivity of chl a was also confirmed by exposure of segments of leaves in test tubes to high ozone concentration (>1000 ppb) as well as by bubbling of ozone in extracts of chlorophyll in vitro. The quantum yield (QY) of photosynthesis was positively correlated with the chlorophyll content and negatively correlated with the visible injury and the chl b/a ratio.