Cristina Nali

Effect of chronic ozone fumigation on the photosynthetic process of poplar clones showing different sensitivity.

Rooted cuttings from two poplar clones (Populus x euramericana, I-214, and Populus deltoides x maximowiczii, Eridano) were exposed for 15 days to diurnal square-wave treatment with ozone (60 nL L-1 for 5 h day-1). Completely fully expanded leaves exposed to ozone showed a reduction in net CO2 assimilation rate as compared to the control leaves during whole exposure period in both the clones. The reduction was related to a strong stomatal closure in clone I-214, but also to an altered mesophyll activity ascribed to limitation of the dark reactions of photosynthetic process. The results obtained in leaves of I-214 subjected to long-term fumigation seem to support the view that the decrease in quantum yield of electron transport may be a mechanism to down-regulate photosynthetic electron transport so that production of ATP and NADPH would be in equilibrium with the decreased demand in the Calvin cycle. In Eridano the CO2 assimilation was reduced because of the exposure and any alteration in stomatal conductance was observed. Thus, chlorophyll fluorescence parameters showed that an inhibition of photosystem II had occurred (reduction in Fv/Fm ratio), while no alterations in quenching parameters were observed upon illumination. The results seem to indicate that an alternative sink for reducing equivalent, other than carbon metabolism is present.

Leaf level early assessment of ozone injuries by passive fluorescence and photochemical reflectance index

Excess energy dissipation pathways (heat and fluorescence) were monitored, at leaf level as indicators of plant physiological status, with field spectroscopy techniques on poplar clones subjected to ozone fumigation. Measurements of spectral radiance emerging from a leaf provide a fast, non‐destructive method for the assessment of excess energy dissipation: xanthophyll‐related heat dissipation was estimated with the photochemical reflectance index (PRI) calculated from a traditional field spectrometer, and steady‐state fluorescence (Fs) under natural illumination conditions was estimated by exploiting a variation of the Fraunhofer line‐depth principle, where the radiance collected with very high resolution spectrometers (FWHM = 0.13 nm) was spectrally modelled. Both remotely‐sensed dissipation pathways responded to fumigation. During a 26‐day fumigation experiment, four diurnal cycles of spectral measurements were collected in parallel to meteorological and key physiological variables (active fluorescence, net photosynthesis) and leaf sample collection for pigment extraction. We outline evidence of a link between the remotely‐sensed Fs and PRI and leaf physiological status. These results open up new possibilities for assessment of plant stress by means of hyperspectral remote sensing.

Surface ozone in Pisa (Italy): A Six-year study

Abstract The paper reports the results of ground level distribution of ozone in Pisa (Italy) in the periods May to September, 1986–1991. The recurrence of relevant levels of photochemically produced ozone is described. The maximum hourly peak (M 1) proved to be 128 ppb and the average of the M 1 peaks turned out to be 60 ppb. The threshold of 100 ppb was surpassed in 17 d. The average value of the “daily highest consecutive 7-h mean concentrations” (M7) was 53 ppb. M1 and M7 are closely correlated and both show approximately a log-normal distribution. Monthly variations are relatively low, with September showing the lowest values. The years 1986 and 1991 show values that are clearly higher than the other years, and no temporal trend has been verified. Ozone levels were positively correlated with temperature and solar radiation.

Ozone distribution in central Italy and its effect on crop productivity

The focus of the present study was to assess the ozone levels in a typical area of the Mediterranean basin, viz. Tuscany (central Italy). Eighty-thousand hourly mean ozone concentrations were recorded by 10 automatic analysers in the districts of Florence, Pisa, Lucca and Prato, from May to September 1995 to 1997. The highest daily mean concentrations were reached in Florence, with a maximum hourly average of 197 ppb. In Lucca and Pisa, the peaks were close to 100 ppb. Data from Prato were much lower. Long-term critical levels for vegetation, as set by United Nations Economic Commission for Europe (UNECE), were constantly exceeded in Florence and Pisa, occasionally in Lucca, never in Prato. The results were used to fit exposure/yield response relationships proposed by UNECE and US National Crop Loss Assessment Network for some important crops. The estimated yield losses varied in Florence from 8% for corn and alfalfa to 27% for soybean, in Pisa from 5% for corn to 24% for soybean, in Lucca from 3% for corn to 17% for soybean. A preliminary economic estimate for corn, wheat, barley, soybean, tomato and alfalfa, calculated annual damage to be 4.6 M Euro in Florence, 0.5 M Euro in Lucca and 3 M Euro in Pisa. The picture must be regarded as only partial, as exposure/yield response relationships for important Italian crops (such as grapevine and vegetables) are not available.

Photosynthetic response of tomato plants to vascular wilt diseases

Greenhouse experiments were conducted to study the chronology of effects on gas exchange and chlorophyll-a fluorescence, visible symptoms and hyphal colonization in plants of the susceptible tomato cultivar Bonny Best inoculated with tracheomycotic fungi Fusarium oxysporum f. sp. lycopersici or Verticillium albo-atrum. The net photosynthetic rates and related parameters of healthy (uncolonized and asymptomatic) leaves of infected plants were affected by both the parasites. In the first uncolonized leaf, net photosynthesis was depressed in different ways: in Fusarium-infected individuals, the maximum detrimental effect was observed a week after inoculation, while in Verticillium-infected plants the most severe depression was detected 21 days after inoculation. The behaviour of the physiological parameters investigated, together with the data relative to chlorophyll fluorescence measurements highlighted the fact that the depression in photosynthetic activity was caused by different concomitant factors in Verticillium-infected plants and was due mainly to drought stress in plants inoculated with Fusarium.

Photosynthesis of two poplar clones contrasting in O3 sensitivity

Abstract Two clones of poplar known for their phenomenological difference in response to ozone were fumigated with 150 nl l–1 of ozone for 5 h. In both clones the treatment significantly reduced the light-saturated rate of CO2 uptake of recently mature leaves and this was accompanied by a decrease in stomatal conductance. Intercellular CO2 concentration of the resistant clone increased following the fumigation. After 20 h of recovery, photosynthesis recovered completely only in the resistant clone. Electrolyte leakage of leaf disks increased in both clones to indicate damage to membranes; after the recovery time this parameter only reached values of the control in the resistant clone. The photochemical efficiency of PSII slightly decreased in the resistant clone. In the other clone, the treatment caused a decline of all chlorophyll fluorescence parameters and only some of them returned to normal values after the recovery time. The physiological response appears to be different in the two clones. In the resistant one, the most probable mechanism involved in the photosynthetic reduction was a regulatory reduction in CO2 fixation. Also data obtained by the solute leakage indicate that in the resistant clone repair mechanisms play a role. The reduction of photosynthesis observed in the sensitive clone is related both to strong stomatal closure and to an impairment in fluorescence parameters. These alterations can indicate a general disruption at the membrane level as confirmed by the solute leakage data.

MONITORING AND BIOMONITORING OF SURFACE OZONE IN FLORENCE, ITALY

An ambient air study was conducted inthe city of Florence, Italy, in the summer 1996.Tropospheric ozone was continuously monitored withautomatic analyzers in three stations, two located inthe urban area and one in the hilly surroundings(Settignano). A biomonitoring campaign based on thetobacco cv. Bel-W3 plants was performed in the samearea. The highest values were constantly recorded inthe Settignano station. The highest 1-hour meanrecorded was 197 nl/l; the accumulated exposure overa threshold of 40 nl/l (AOT40) was well above thecritical levels standards for protection of thevegetation. A consistent temporal variation wasobserved and July proved to be the month with thehighest ozone levels. Cumulative frequencydistribution of ozone maximum daily concentrationsexhibited a good fitting to log-normality. No`week-end’ effect was observed. Biomonitoring datawere in good agreement with chemico-physical ones.

The protective effect of rust infection against ozone, sulphur dioxide and paraquat toxicity symptoms in broad bean

A localized but highly protective effect of rust infection towards injury induced by the air pollutants ozone and sulphur dioxide, either alone or in mixture and the herbicide paraquat was found. The effect was proportional to inoculum density, was non-translocatable, and dependent on temperature. Protection continued even after the infection was no longer active, and was induced by different strains of the fungus in cultivars of both broad bean and pea. Protection was not due to the exclusion of pollutants resulting from stomatal closure. Histopathological investigations indicated that the effect was present in uncolonized tissues adjacent to the site of infection. Ultrastructural studies showed normal cell structure in the green islands surrounding the infection centres, even when nearby regions had collapsed. Peroxidase activity was increased rapidly in the infected tissues both in the colonized and in the adjacent non-colonized cells. Such activity was not detectable at significant levels in non-parasitized tissues. The results are discussed in connection with the possible mechanisms involved.

Antioxidant enzyme isoforms on gels in two poplar clones differing in sensitivity after exposure to ozone

The effect of acute ozone (O3) fumigation on isozyme patterns of superoxide dismutase (SOD), peroxidase (POD) and ascorbate peroxidase (APX) in mature (ML) and young leaves (YL) of two poplar clones, contrasting in O3-sensitivity was analysed. Untreated leaves of both the O3-sensitive (O3-S) clone Eridano of Populus deltoides×P. maximowiczii and the O3-resistant (O3-R) clone I-214 of P.×euramericana showed four distinct SOD isoforms with a relative mobility (Rf) of 0.54 (MnSOD), 0.60 (Cu/ZnSOD), 0.65 (unidentified), and 0.71 (Cu/ZnSOD). After O3-fumigation the activity of the SOD isoforms showed only quantitative variations with respect to control plants. In ML of untreated O3-R plants seven POD isoforms (Rf= 0.13, 0.19, 0.34, 0.59, 0.64, 0.70 and 0.75) were found, while in YL one isoform (Rf= 0.34) was undetected. Only three POD isoforms in both ML and YL of untreated O3-S plants were resolved. The electrophoretic pattern of POD in O3-S leaves was greatly modified by acute O3-fumigation with the appearance of new isoforms in both YL and ML and the disappearance of an isoform (Rf= 0.13) in YL. Additionally, O3-exposure induced the appearance of two APX isoforms in YL (Rf= 0.66 and 0.70), and one isoform in ML (Rf= 0.70) of the O3-S clone. By contrast, the activity of the three APX isoformes (Rf= 0.64, 0.70 and 0.76) detected in O3-R leaves showed only quantitative variation with respect to untreated plants. From these data it is concluded that: 1) in these poplar hybrids antioxidant enzyme activity is developmentally regulated and greatly affected by acute O3 stress treatments and 2) the different enzymes activity displayed by the two poplar clones, especially for POD and APX isoformes, could partly explain their distinct O3-sensitivity.

Suppression Substractive Hybridization and NGS Reveal Differential Transcriptome Expression Profiles in Wayfaring Tree (Viburnum lantana L.) Treated with Ozone.

Tropospheric ozone (O3) is a global air pollutant that causes high economic damages by decreasing plant productivity. It enters the leaves through the stomata, generates reactive oxygen species, which subsequent decrease in photosynthesis, plant growth, and biomass accumulation. In order to identify genes that are important for conferring O3 tolerance or sensitivity to plants, a suppression subtractive hybridization analysis was performed on the very sensitive woody shrub, Viburnum lantana, exposed to chronic O3 treatment (60 ppb, 5 h d−1 for 45 consecutive days). Transcript profiling and relative expression assessment were carried out in asymptomatic leaves, after 15 days of O3 exposure. At the end of the experiment symptoms were observed on all treated leaves and plants, with an injured leaf area per plant accounting for 16.7% of the total surface. Cloned genes were sequenced by 454-pyrosequencing and transcript profiling and relative expression assessment were carried out on sequenced reads. A total of 38,800 and 12,495 high quality reads obtained in control and O3-treated libraries, respectively (average length of 319 ± 156.7 and 255 ± 107.4 bp). The Ensembl transcriptome yielded a total of 1241 unigenes with a total sequence length of 389,126 bp and an average length size of 389 bp (guanine-cytosine content = 49.9%). mRNA abundance was measured by reads per kilobase per million and 41 and 37 ensembl unigenes showed up- and down-regulation respectively. Unigenes functionally associated to photosynthesis and carbon utilization were repressed, demonstrating the deleterious effect of O3 exposure. Unigenes functionally associated to heat-shock proteins and glutathione were concurrently induced, suggesting the role of thylakoid-localized proteins and antioxidant-detoxification pathways as an effective strategy for responding to O3. Gene Ontology analysis documented a differential expression of co-regulated transcripts for several functional categories, including specific transcription factors (MYB and WRKY). This study demonstrates that a complex sequence of events takes place in the cells at intracellular and membrane level following O3 exposure and elucidates the effects of this oxidative stress on the transcriptional machinery of the non-model plant species V. lantana, with the final aim to provide the molecular supportive knowledge for the use of this plant as O3-bioindicator.