Angeles Calatayud

Frequently asked questions about in vivo chlorophyll fluorescence: practical issues

The aim of this educational review is to provide practical information on the hardware, methodology, and the hands on application of chlorophyll (Chl) a fluorescence technology. We present the paper in a question and answer format like frequently asked questions. Although nearly all information on the application of Chl a fluorescence can be found in the literature, it is not always easily accessible. This paper is primarily aimed at scientists who have some experience with the application of Chl a fluorescence but are still in the process of discovering what it all means and how it can be used. Topics discussed are (among other things) the kind of information that can be obtained using different fluorescence techniques, the interpretation of Chl a fluorescence signals, specific applications of these techniques, and practical advice on different subjects, such as on the length of dark adaptation before measurement of the Chl a fluorescence transient. The paper also provides the physiological background for some of the applied procedures. It also serves as a source of reference for experienced scientists.

Effects of 2-month ozone exposure in spinach leaves on photosynthesis, antioxidant systems and lipid peroxidation

The photosynthesis response, antioxidant systems and lipid peroxidation were studied in leaves from spinach plants ( Spinacia oleraceaL.) in response to ozone fumigation, ambient air and charcoal filtered air treatments. The photosynthetic activity was tested through gas exchange and chlorophyll a fluorescence measurements. Ambient air and ozone fumigation caused a decrease in the photosynthetic rate (25% and 63%, respectively) mainly due to a reduced mesophyll activity, as evidenced by the increased intercellular CO 2 concentration. These data agree with a large reduction in the non-cyclic electron flow (7% and 16%), a lower capacity to reduce the quinone pool and a higher development of non-photochemical quenching upon high O3 concentration. The results suggest that the oxidative stress produced, together with the stimulation of superoxide dismutase (SOD, EC 1.15.1.1) and ascorbate peroxidase (APX, EC 1.11.1.11) activities and the increase in lipid peroxidation (20% and 36%, respectively), generated an alteration of the membrane properties.

Changes in chlorophyll a fluorescence, photosynthetic CO2 assimilation and xanthophyll cycle interconversions during dehydration in desiccation-tolerant and intolerant liverworts

Abstract. The interactions among water content, chlorophyll a fluorescence emission, xanthophyll interconversions and net photosynthesis were analyzed during dehydration in desiccation-tolerant Frullania dilatata (L.) Dum. and desiccation-intolerant Pellia endiviifolia (Dicks) Dum. Water loss led to a progressive suppression of photosynthetic carbon assimilation in both species. Their chlorophyll fluorescence characteristics at low water content were: low photosynthetic quantum conversion efficiency, high excitation pressure on photosystem II and strong non-photochemical quenching. However, dissipation activity was lower in P. endiviifolia and was not accompanied by a rise in the concentration of de-epoxidised xanthophylls as F. dilatata. The photosynthetic apparatus of F. dilatata remained fully and speedily recuperable after desiccation in as indicated by the restoration of chlorophyll fluorescence parameters to pre-desiccation values upon rehydration. A lack of recovery upon remoistening of P. endiviifolia indicated permanent and irreversible damage to photosystem II. The results suggest that F. dilatata possesses a desiccation-induced zeaxanthin-mediated photoprotective mechanism which might aid photosynthesis recovery when favourable conditions are restored by alleviating photoinhibitory damage during desiccation. This avoidance mechanism might have evolved as an adaptative response to repeated cycles of desiccation and rehydration that represent a real threat to photosynthetic viability.

Response of Spinach Leaves (Spinacia oleracea L.) to Ozone Measured by Gas Exchange, Chlorophyll a Fluorescence, Antioxidant Systems, and Lipid Peroxidation

Spinach (Spinacia oleracea L. cv. Clermont) leaves grown in open-top chambers and exposed to three different concentrations of ozone were measured for gas exchange, chlorophyll a fluorescence, antioxidant systems, and lipid peroxidation at the end of growing season. High O3 concentration reduced Fv/Fm, indicating that the efficiency in the energy conversion of photosystem 2 (PS2) was altered. The rate of non-cyclic electron transport rate and the capacity to reduce the quinone pool were also affected. The development of non-photochemical quenching was not high enough to decrease the photon excess in the PS2. The limitation of photosynthetic activity was probably correlated with stomata closure and with an increase in intercellular CO2 concentration. Under oxidative stress, superoxide dismutase (SOD) activity was stimulated in parallel with lipid peroxidation. We did not find any differences in the ascorbate (AsA) pool and ascorbate peroxidase (APX) or glutathione reductase (GR) activities between air qualities. Small, but similar responses were observed in spinach leaves exposed to ambient ozone concentration.

Differences in ozone sensitivity in three varieties of cabbage (Brassica oleracea L.) in the rural Mediterranean area

Summary The effects of air quality with three levels of ozone (O 3 ) were studied on three cabbage varieties during a one month exposure period in the Valencia area by means of modulated chlorophyll (Chl) a fluorescence, lipid peroxidation and solute leakage. Increasing O 3 exposure reduced the maximum quantum yield of PSII photochemistry (F v /F m ) in line with a reduction in non-cyclic electron flow (ϕ PSII ), lower capacity to reduce the quinone pool (q P ) and a decrease in the potential phothosynthetic quantum conversion, Rfd ratio. These reductions were more evident in Caramba and Sentinel varieties but lower in Othelo. Ozone and its oxidative derivates weakened the plasmalemma, which caused a leakage of cellular liquids into intercellular spaces. In our results, the ion leakage increased with O 3 concentration. Moreover, data from thiobarbituric acid reactive (TBARS) analysis confirmed the hypothesis that O 3 modified the membrane structure by increasing the lipid peroxidation more in Caramba and Sentinel varieties. These alterations in the membranes reduced the ability to develop non-photochemical quenching (NPQ). The O 3 effects on solute leakage and lipid peroxidation in the Othelo variety were lower than in the other two varieties, which allowed non-photochemical quenching to develop in this variety. These results suggest the possibility adverse effects of ozone on winter crops grown in the Valencia region.

Salt-tolerant rootstock increases yield of pepper under salinity through maintenance of photosynthetic performance and sinks strength.

The performance of a salt-tolerant pepper (Capsicum annuum L.) accession (A25) utilized as a rootstock was assessed in two experiments. In a first field experiment under natural salinity conditions, we observed a larger amount of marketable fruit (+75%) and lower Blossom-end Root incidence (-31%) in commercial pepper cultivar Adige (A) grafted onto A25 (A/A25) when compared with ungrafted plants. In order to understand this behavior a second greenhouse experiment was conducted to determine growth, mineral partitioning, gas exchange and chlorophyll a fluorescence parameters, antioxidant systems and proline content in A and A/A25 plants under salinity conditions (80 mM NaCl for 14 days). Salt stress induced significantly stunted growth of A plants (-40.6% of leaf dry weight) compared to the control conditions, while no alterations were observed in A/A25 at the end of the experiment. Accumulation of Na(+) and Cl(-) in leaves and roots was similar in either grafted or ungrafted plants. Despite the activation of protective mechanisms (increment of superoxide dismutase, catalase, ascorbate peroxidase activity and non-photochemical quenching), A plants showed severely reduced photosynthetic CO2 assimilation (-45.6% of AN390) and substantial buildup of malondialdehyde (MDA) by-product, suggesting the inability to counteract salt-triggered damage. In contrast, A/A25 plants, which had a constitutive enhanced root apparatus, were able to maintain the shoot and root growth under salinity conditions by supporting the maintained photosynthetic performance. No increases in catalase and ascorbate peroxidase activities were observed in response to salinity, and MDA levels increased only slightly; indicating that alleviation of oxidative stress did not occur in A/A25 plants. In these plants the increased proline levels could protect enzymatic stability from salt-triggered damage, preserving the photosynthetic performance. The results could indicate that salt stress was vanished by the lack of negative effects on photosynthesis that support the maintained plant growth and increased marketable yield of the grafted plants.

The effects of foliar fertilization with iron sulfate in chlorotic leaves are limited to the treated area. A study with peach trees (Prunus persica L. Batsch) grown in the field and sugar beet (Beta vulgaris L.) grown in hydroponics

Crop Fe deficiency is a worldwide problem. The aim of this study was to assess the effects of foliar Fe applications in two species grown in different environments: peach (Prunus persica L. Batsch) trees grown in the field and sugar beet (Beta vulgaris L. cv. “Orbis”) grown in hydroponics. The distal half of Fe-deficient, chlorotic leaves was treated with Fe sulfate by dipping and using a brush in peach trees and sugar beet plants, respectively. The re-greening of the distal (Fe-treated) and basal (untreated) leaf areas was monitored, and the nutrient and photosynthetic pigment composition of the two areas were also determined. Leaves were also studied using chlorophyll fluorescence imaging, low temperature-scanning electron microscopy microanalysis, scanning transmission ion microscopy-particle induced X-ray emission and Perls Fe staining. The distal, Fe-treated leaf parts of both species showed a significant increase in Fe concentrations (across the whole leaf volume) and marked re-greening, with significant increases in the concentrations of all photosynthetic pigments, as well as decreases in de-epoxidation of xanthophyll cycle carotenoids and increases in photochemical efficiency. In the basal, untreated leaf parts, Fe concentrations increased slightly, but little re-greening occurred. No changes in the concentrations of other nutrients were found. Foliar Fe fertilization was effective in re-greening treated leaf areas both in peach trees and sugar beet plants. Results indicate that the effects of foliar Fe-sulfate fertilization in Fe-deficient, chlorotic leaves were minor outside the leaf surface treated, indicating that Fe mobility within the leaf is a major constraint for full fertilizer effectiveness in crops where Fe-deficiency is established and leaf chlorosis occurs.

Chlorophyll a Fluorescence Emission, Xanthophyll Cycle Activity, and Net Photosynthetic Rate Responses to Ozone in Some Foliose and Fruticose Lichen Species

The lichens Parmelia quercina, Parmelia sulcata, Evernia prunastri, Hypogymnia physodes, and Anaptychia ciliaris were exposed to ozone (O3) in controlled environment cuvettes designed to maintain the lichens at optimal physiological activity during exposure. Measurements of gas exchange, modulated chlorophyll (Chl) fluorescence, and pigment analysis were conducted before and after exposure to 300 mm3 (O3) m−3, 4 h per d for 14 d. No changes in the efficiency of photosystem 2 (PS2) photochemistry, the reduction state of QA, or the electron flow through PS2, measured by Chl fluorescence, were detected in any of the five lichen species studied. Additionally, neither photosynthetic CO2 assimilation nor xanthophyll cycle activity or photosynthetic pigment concentration were affected by high O3 concentrations. Thus the studied lichen species have significant capacities to withstand oxidative stresses induced by high concentration of O3.

Similar Effects of Ozone on Four Cultivars of Lettuce in Open Top Chambers During Winter

Ozone is the major phytotoxic air pollutant that reduces the yield of several agricultural crops in the Spanish Mediterranean area. We studied four lettuce cultivars (Lactuca sativa L.) for the effects of different O3 concentrations during the winter on chlorophyll (Chl) a fluorescence, lipid peroxidation, and root length in outdoor open-top chambers. Under O3 the photosynthetic quantum conversion declined while heat emissions increased in all cultivars; these results provide more evidence of non-filtered air with additional ozone (NFA+O3) treatment compared with non-filtered air (NFA) and charcoal filtered ozone-free air (CFA). Changes in the Chl a fluorescence may be associated with an increase in membrane lipid peroxidation as well as with observed reduction of root length under O3 stress.

Foliar Spraying with Zineb Increases Fruit Productivity and Alleviates Oxidative Stress in Two Tomato Cultivars

The effects of foliar spraying of the dithiocarbamate zineb on two cultivars of tomato grown in the field in a site with high ozone concentrations were studied by means of biomass assessment, antioxidant enzyme assays, lipid peroxidation, and chlorophyll fluorescence measurements. Zineb prevented the peroxidation of membrane lipids and decreased the activity of scavenging enzymes, which suggests that plants sprayed with zineb are subjected to lower oxidative stress than controls. The beneficial effects of zineb protection is the utilization of a larger fraction of absorbed radiant energy in photosynthesis and a larger fruit yield in plants of both cultivars.