Fluorescence imaging may serve as a convenient tool for early detection of such stress induced damage. The main difference between the conventional fluorometer and the imaging fluorometer is the possibility of parallel assessment of several samples under identical conditions.
For example we treated plants of Phaseolus vulgaris cv. It is know as in plants exposed to chilling stress, photosynthetic enzymes may be inactivated or degraded and photodamage to PSII may happen, reducing photosynthesis Dai et al. The individual leaves were divided into a shaded zone and two illuminated, chilled zones. Chilling up to 7 d in the dark did not influence PSII efficiency whereas chilling in the light caused severe photoinhibition.
Data obtained from Chl fluorescence imaging were confirmed by visual appearance of symptoms which were evident in the portion of leaves chilled and illuminated. Obtained results showed that photoinhibition was due to a secondary effect in the unchilled leaf tip sink limitation as revealed by starch accumulation data. Instead it was a direct effect of chilling and irradiance in the chilled illuminated zones.
All images are normalised to the false colour bar provided. The pixel value display is based on a false-colour scale ranging from black 0. Calatayud et al. The obtained results showed as the nutrient solution temperatures of 10 C induced an increase in PSII parameters indicating that the majority of photons absorbed by PSII were used in photochemistry and that PSII centers were maintained in an oxidized state.
Water stress is another important abiotic stress that induces reduction of growth and yield of plants. For this reason the development of drought-tolerance is an important target of the researchers. Masacci et al. They found that under low and moderate light intensity, the onset of drought stress caused an increase in the operating quantum efficiency of PSII PSII which indicated increased photorespiration since photosynthesis was hardly affected by water shortage.
Chl fluorescence imaging showed a low spatial heterogeneity of PSII. The authors concluded that the increase in photorespiration rate in plants during the water stress can be seen as an acclimation process to avoid an over-excitation of PSII under more severe drought conditions.
Qing-Ming et al. They found that electron transport rate and the light saturation level declined significantly with drought stress aggravation in both CO 2 concentrations.
Drought stress decreased maximal photosynthetic ETR and subsequently decreased the capacity of preventing photodamage. At the same time, elevated CO 2 concentration increased the light saturation level significantly, irrespective of the water conditions. Elevated CO 2 concentration can alleviate drought stress-induced photoinhibitory damage by improving saturating photosynthetically active radiation.
Sommerville et al. The authors hypothesized that areole regions near primary nerves would show a smaller decline in the maximum efficiency of PSII photochemistry with drought compared with regions between secondary nerves and that the difference between areole regions would be smaller in phyllodes with higher primary nerve density.
Indeed, the phyllodes of Acacia floribunda were found to have both greater primary nerve density and show greater spatial homogeneity in photosynthetic function with drought compared with the phyllodes of Acacia pycnantha. Drought is a type of stress which can induce heterogeneity in leaf photosynthesis that probably occurs when dehydration is rapid as in the case of drought experiments performed on potted plants by withholding water.
Using Chl fluorescence imaging, Flexas et al. Even the quality and quantity of light intensity notable influence the photosynthetic apparatus and functioning. Generally, sun- and shade leaves differ in the composition of leaf pigment, electron carriers on thylakoids membranes, structure of the chloroplast and photosynthetic rate Anderson et al. Lichtenthaler et al. This tool not only provided the possibility to screen the differences in photosynthetic CO 2 assimilation rate between sun and shade leaves, but in addition permitted detection and quantification of the large gradient in photosynthetic rate across the leaf area existing in sun and shade leaves.
Chl fluorescence analysis is used also to characterized photosynthetic process in transgenic plants such as tomato Lycopersicon esculentum cv.
This gene encodes for a well known transcription factor, which is involved in anthocyanin production and is modulated by light and sucrose. The presence of a higher constitutive level of anthocyanin pigments in transgenic plants could give them some advantage, in terms of adaptation and defence against environmental stresses. To test this hypothesis, a high light experiment was carried out exposing wild type and transgenic tomato plants to a strong light irradiance for about ten days and monitoring the respective phenotypic and physiological changes.
The light intensity used was very high and likely not similar to normal environmental conditions at least for such a prolonged period. Chlorophyll fluorescence imaging on control and stressed leaves from both genotypes suggest that, in transgenic leaves, the apparent tolerance to photoinhibition was probably not due to an increased capacity for PSII to repair, but reflected instead the ability of these leaves to protect their photosynthetic apparatus.
Certainly among abiotic stress the pollutants can alter the physiology and biochemistry of plants. Ozone is an air pollutant that induces reduction in growth and yield of plants species. The major target of the O 3 effects is represented by photosynthetic process and many works have been reported as this pollutant can impair CO 2 assimilation rate.
Plant response depends also on the dose concentration x time. In fact, it can distinguish chronic exposure to O 3 from acute one. Chen et al. Although both acute and chronic O 3 treatment resulted in a similar overall photosynthetic impairment compared to the controls, the fluorescence imaging analysis revealed that the physiological mechanisms underlying the decreases differed.
In the acute O 3 treatments over the chronic one there was a greater spatial heterogeneity related to several bases. The higher O 3 concentration typically induced oxidative stress and the hypersensitive response within a matter of hours leading to programmed cellular death PCD.
By the end of chronic O 3 treatment, control leaves showed an increase in spatial heterogeneity of photosynthesis linked to the process of natural senescence. Clearly, in this study it has been demonstrated as Chl fluorescence imaging represents a useful tool to study also mechanisms on the basis of plants responses to abiotic stress such as O 3 pollution.
Guidi et al. The aim of the work was to compare the perturbations in photosynthesis induced by an abiotic or biotic stress. In addition to, in the work were compared results obtained by conventional Chl fluorescence analysis and the technique of Chl fluorescence imaging. Indeed, in ozonated leaves fluorescence yield was lower in leaf veins than in the mesophyll with the exception of the necrotic areas where no fluorescence signals could be detected.
This suggests that the leaf area close to the veins were more sensitive to ozone. The parameter PSII decreased significantly in both infected and ozonated leaves, but image analysis provides more information than the conventional fluorometer. Plants were more affected by the mixture of salts when subjected to the first four values of electrical conductivity and, from these levels on, they were more sensitive to NaCl. Abrir menu Brasil. Abrir menu. Hidelblandi F. Recife, PE. E-mail: hidelfarias gmail.
ABSTRACT Chlorophyll a fluorescence is a very useful tool in ecophysiological studies to analyze the photosynthetic performance of plants under biotic and abiotic stresses.
Key words: photosynthesis; water and salt stress; halophyte. Introduction The use of chlorophyll a and photosynthetic pigments is an important tool to evaluate the performance in the light energy harvest by plants under abiotic stress Shu et al. Table 1 Chemical characterization of the Fluvic Neosol. Table 2 Physical characterization of the Fluvic Neosol. Literature Cited Ahmed, C. Saline water irrigation effects on soil salinity distribution and some physiological responses of field grown Chemlali olive Journal of Environmental Management, v.
Azevedo Neto, A. Baker, N. Belkheiri, O. Bjorkman, O. Photon yield of O 2 evolution and chrolophyll fluorescence characteristics at 77K among vascular plants of diverse origins. Bouchenak, F. Correia, K. Fernandez, R. Glenn, E. Hasegawa, P. Li, G. Lichtenthaler, H. Ma, Q. Sodium chloride improves photosynthesis and water status in the succulent xerophyte Zygophyte xanthoxylum Tree Physilology, v.
Mehta, P. Melo, H. Nedjimi, B. Ozfidan, C. Paridas, A. Shu, S. Silveira, J. Roots and leaves contrasting osmotic adjustment mechanisms in responses to NaCl - salinity in Atriplex nummularia Environmental and Experimental Botany, v. Souza, E. Thomas, G. Vieira, D.
Publication Dates Publication in this collection Apr History Received 22 Apr Accepted 30 Dec This is an Open Access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
Edivan R. Jailson C. Petrolina, PE. E-mail: jailson. Figures 3 Tables 3. These articles are good sources for experimental DF phenomena though. Illustration of the difference between F O and DF. Following absorption of a photon the excitation energy may be lost in the antenna and re-emitted as fluorescence F O. The excitation energy may induce a charge separation, which may be followed by electron transfer to Q A or a recombination reaction.
In the latter case the energy may again be lost as fluorescence F O. Emission occurs at subnanosecond times following excitation. As noted above, each charge pair has its own particular recombination time. Thermoluminescence TL , light emission as a result of temperature-dependent recombination of charge pairs, is a special application of DF, although it has to be noted that not all DF phenomena have a TL counterpart Vass and Govindjee As the temperature is lowered, charge recombinations become gradually inhibited.
For TL, there are different illumination strategies. This does not induce Chl or Car radicals as suggested by Stirbet and Govindjee ; to induce these radicals the temperature has to be lowered further e. Inversely, upon warming the sample, the different charge pairs can gradually recombine; each charge pair with its own typical peak temperature Vass Longer recombination times go together with higher TL peak temperatures.
TL measurements become distorted during the transition from the frozen to the thawed state Vass et al. See Ducruet and Vass for a practical guide to the application of TL. TL measurements are destructive. DF measurements, on the other hand, are non-destructive. Even so, TL measurements have been more popular than DF measurements due to the absence of commercially available instruments for the measurement of DF.
It may also be mentioned that the analysis of TL measurements is better developed DeVault et al. It is also a convenient technique for the study of environmental stresses that affect PSII e. PF intensity is linearly related to the actinic light intensity see, e. In the case of DF only absorbed light that induces a stable charge separation can lead to DF and especially at high light intensities, this is only a small fraction of the absorbed light.
In addition, DF emission is much more spread out in time because the DF-inducing charge recombinations can be due to different charge pairs, with different recombination times.
A third factor that reduces the DF yield is the existence of several recombination pathways of which only one leads to DF see, e. Pathways 2 and 3 are called indirect charge recombination pathways. Krieger-Liszkay and Rutherford showed that DCMU bound to the Q B site increases the midpoint potential of Q A , stabilizes the charge separation shift of TL bands to higher temperatures and increases the probability that recombination occurs via tunneling pathway 1.
Bromoxynil, on the other hand, decreases the midpoint potential of Q A , destabilizes the charge separation shift TL bands to lower temperature and increases the probability that recombination occurs via pathways 2 and 3. Cser and Vass confirmed this interpretation using mutants in which amino acids near Pheo and P were modified using site-directed mutagenesis.
In addition, the emission spectra of both types of fluorescence emission are identical Strehler and Arnold ; Arnold and Thompson ; Sonneveld et al. As a consequence, both cannot be detected at the same time and thus a separation strategy is needed. Two approaches for the simultaneous monitoring of PF and DF signals have been developed: 1 quasi-continuous illumination of dark-adapted samples and 2 illumination of a dark-adapted sample by single pulse usually by a laser [see, e.
The results of these two methods differ. The first approach has been used for the evaluation of the induction kinetics of the two fluorescence signals over short time periods seconds Wraight and Crofts ; Zaharieva and Goltsev ; Strasser et al. This type of illumination is often called quasi-continuous.
Prompt fluorescence is measured during the illumination periods, and the decaying DF signal is recorded during the dark intervals. It should be noted that the dark intervals reduce the effective light intensity by one-third. In addition, during the dark intervals the dark reactions continue electron flow , whereas the light reactions charge separations stop, changing the relationship between dark and light reactions. Using quasi-continuous illumination, DF induction transients complementing the PF induction transients can be constructed.
To do this, the DF intensities measured during a particular dark interval for all dark periods are selected and averaged, and then used as single points for the DF induction curve. By selecting different decay intervals, induction curves can be constructed that show DF kinetic components with different lifetimes Kalaji et al. Using the second approach, Chl a fluorescence decay is recorded after a short ns laser pulse or after continuous illumination. The strong fluorescence signal registered during the first few nanoseconds following illumination is due to PF, while at longer times after illumination the emitted light is the result of charge recombinations i.
Alternatively, single light pulses of variable length e. The M-PEA also allows this type of measurement. This approach is much more time-consuming than method 1 , because each time point is a separate measurement, but has the advantage that the intensity of the actinic light is also the effective light intensity and, therefore, the results can be related to OJIP transients on a one-to-one basis.
The DF induction curve is often compared to the simultaneously measured PF induction curve. To allow a direct comparison of the maxima for both fluorescence types, they are superimposed on each other on the same timescale Govindjee and Papageorgiou ; Krause and Weis ; Malkin et al.
For the construction of DF induction transients, see Question During induction, different reactions occur in parallel in different RCs. Upon turning off the light, different reoxidation and recombination reactions all occur separately with their own particular rate constants for PF see, e. To understand DF measurements during the first few ms of induction, it is important to understand what happens on the donor side of PSII.
These authors also showed for the ms DF component that only the state induced by three flashes on going from S 3 via S 4 to S 0 still yields a high DF intensity. And as mentioned above, the peak DF intensity S 4 state occurs after 3—4 ms. This reaction sequence accounts for the DF rise to the I1 peak.
Using single turnover flashes, several S state cycles can be detected see Grabolle and Dau for DF. OJIP transients, on the other hand, are induced by much lower light intensities that are much less concentrated in time.
As a consequence the S states will dampen due to double hits and misses; Kok et al. As the PQ pool becomes reduced the availability of oxidized PQ decreases. The position of the I1 peak is sensitive to the light intensity. Destruction of the Mn cluster changes the DF response. Two DF induction curves are shown in Fig. The characteristic points of the DF induction are indicated according to the nomenclature proposed by Goltsev and Yordanov and Goltsev et al.
Similarly to the PF induction curve, the DF induction curve can be divided into fast and slow phases Itoh et al. The slow phase lasts several minutes, reaching a stationary level after 5—10 min of illumination by quasi-continuous actinic light.
Two maxima I1 and I2 sometimes with a minimum D1 in between are observed in the fast phase, after which DF drops to a minimum labeled as D2 Goltsev and Yordanov ; Goltsev et al. I2 strongly depends on the actinic light intensity; at high light intensities it is visible only as a shoulder Schansker et al. In a 2D graph, DF values collected from different dark delay intervals are plotted as a function of relative double-normalized PF, V t Fig.
For the two plots there is a long induction interval, where the intensities of PF and DF change synchronously inversely. This appears on the diagram as an almost linear section between phases S and T of the PF transient.
During this time interval both PF and DF intensities are modified, and it has been suggested that this is mainly due to changes in fluorescence quantum yield Lavorel ; Goltsev et al. See above for a discussion of the reasons for this relationship.
Between 0. The restarted electron flow is also accompanied by a further energetization of the thylakoid membranes. External electric fields are known to stimulate recombination reactions Vos et al. In other words, both signals monitor the reduction kinetics of the photosynthetic electron transport chain in different ways, providing complementary information. To produce comparable experimental data several factors are important.
In the previous paper the variability among instruments and the extent to which parameters vary naturally was discussed Kalaji et al. In the first study it is shown that the statistical distribution of eight OJIP-derived parameters does not follow the Gaussian distribution law, and in the second study it is shown that a stress like high-temperature stress affects the statistical distribution.
Changes occurring in leaves during the day and their effect on leaf sampling form another factor. An important variable is the orientation of a leaf. Certain leaves are oriented toward the east and receive most of their daily light dose in the morning, whereas other leaves may be oriented toward the west and receive a large part of their daily light dose in the afternoon.
Losciale et al. Another important factor is the angle of the leaf relative to the plane. Leaves that are oriented almost orthogonally will intercept only a relatively small part of the sunlight, whereas leaves oriented nearly parallel to the plane will intercept a large part of the sunlight. A steeper orientation allows a better distribution of the light over different layers of leaves e. Another factor is wind. Leaves that move around in the wind will continuously change their orientation relative to the sun causing fluctuations in the intercepted light.
These fluctuations are further modulated by the passage of clouds and sunflecks. The intensity of sunlight also varies over the day, being low in the early morning and late afternoon and peaking around noon.
In this sense, photosynthetic activity is strongly dependent on the perceived average light intensity, which ultimately drives long-term light acclimation. In shade leaves, having a lower photosynthetic capacity, saturation will be reached at lower light intensities than in sun leaves. It may be noted, on the other hand, that shade leaves exhibit a higher photosynthetic activity at low light intensities and a lower compensation point. Saturation is further modulated by temperature.
With the exception of the light reactions photophysical , photosynthetic electron transport, Calvin—Benson cycle and the photorespiratory pathway have a biochemical character and, as a consequence, photosynthetic activity is quite strongly temperature dependent. A lowering of the temperature will shift the light intensity at which saturation occurs to lower light intensities.
Another important variable is the humidity of the air. Dry air is likely to lead to high levels of evaporation, to which the plant may respond by partially closing its stomata Lange et al. This lowers the uptake of CO 2 and increases photorespiratory activity at the expense of the assimilatory activity Medrano et al. Closing of the stomata will, however, increase the leaf temperature e.
All these variables should be considered when designing a leaf sampling protocol for an experiment Rousseau et al. On bright days, solar radiation is supersaturating with respect to the photosynthetic capacity during a considerable part of the day in most plant species and regions of the world Long and Humphries Dark acclimation with leaf clips for 20 min during daytime hours ensures the relaxation of the transthylakoid pH difference Quick and Stitt ; Nilkens et al.
Photosynthesis frequently remains depressed during the afternoon hours Correia et al. It is worth noting that nighttime recovery of photoinhibition may be inhibited or slowed down by low night temperatures Strand and Lundmark ; Bussotti The temperature dependence of the repair cycle and its inhibition under low temperature conditions has been described under in vitro conditions Aro et al.
In the JIP test this parameter is associated with electron transport activity Strasser et al. The water status of plants may play an important role. Desotgiu et al. The amplitude of the IP phase, which was shown experimentally to correlate with leaf PSI content in fully dark-adapted leaves, was always found to be enhanced during the noontime hours as well as in sun leaves Cascio et al.
However, the reason for this observation is different in these two cases. If leaves suffer from a significant amount of photoinhibition at noontime, the electron donation capacity of PSII will be lower. This is comparable to illumination at a lower light intensity cf.
Srivastava et al. An explanation for this observation may be that leaves kept overnight in darkness may have lower sink activities and a reduced stomatal opening Felle et al.
Leaves from a tree form a population, and each single leaf may differ from the others in terms of size, age and position in the tree canopy. The position of the leaf in the canopy top vs.
Sun and shade leaves differ significantly in their photosynthetic apparatus and performance Lichtenthaler et al. Furthermore, sun leaves have a higher Calvin—Benson cycle capacity relative to the capacity of the electron transport chains, and more efficiently dissipate excess energy as heat, compared to shade leaves.
Shade leaves, on the other hand, are more efficient at exploiting low PAR levels for photosynthesis, having a larger PSII antenna and more extensively stacked thylakoid membranes Lichtenthaler et al. NPQ induction in response to solar radiation is stronger in sun leaves than in shade leaves. Some stress factors act preferentially on a specific side of the canopy.
These factors include, e. Other environmental factors, such as soil properties and air pollutants, affect the canopy in a different way. The effects of these factors depend on the physiology, ontogeny and the position of leaves in the crown. We can thus have damaged leaves at specific levels in the tree crown, e. Finally, when leaves are lost from the branches due to senescence, or damaged by biotic or abiotic factors, the photosynthetic activity of the remaining foliage may increase Eyles et al.
Such a response affects the value of several Chl a fluorescence parameters, especially the amplitude of the IP phase of the OJIP transient Desotgiu et al. The usefulness of individual Chl a fluorescence parameters and protocols for evaluation of drought depends on the severity and duration of drought stress Suresh et al.
Recently, it was observed that blue-light-induced chloroplast movements are very sensitive to drought stress. These phenomena are also reflected in Chl a fluorescence and calculated fluorescence parameters.
However, it is highly insensitive to stomatal changes and other effects occurring under moderate drought stress see Fig. Response of Chl a fluorescence parameters to drought stress. Both the slow and fast Chl a fluorescence kinetics provide parameters sensitive to drought stress Fracheboud and Leipner ; Oukarroum et al.
In summary, measurements of slow fluorescence kinetics and calculation of quantum yields and electron transport rate ETR are useful for determination of drought stress effects, reflecting both stomatal and non-stomatal effects. However, such measurements during drought stress cannot be directly related to CO 2 assimilation Baker When a plant is stressed, F P decreases due to processes such as photoinhibition and under stress conditions F S can also increase, leading to a decrease of R Fd Lichtenthaler et al.
The simplicity of the fast Chl a measurements and automated analysis have sometimes led to misapplication. Analysis should be supported by parallel measurements using other methods, and the restrictions of the JIP test analysis should be kept in mind is the F M level still reached, is the F O truly measured, is the electron transport chain fully reoxidized?
Photosynthesis is very sensitive to high temperatures and can be partially or even completely inhibited before other stress symptoms are observed. High temperatures affect photosynthesis by their effect on the rates of chemical reactions and on the structural organization of the photosynthetic apparatus Pastenes and Horton This decrease is reversible and leads to a decrease in the linear electron transport rate, which can be detected using the saturating pulse method.
This may be due to high-temperature-induced changes in the properties of thylakoid membranes Sharkey and Zhang ; Yamauchi and Sugimoto , the dissociation of the manganese-stabilizing protein from the PSII reaction center complex and the release of Mn atoms Yamane et al. Damage to PSII can be observed in slow Chl a fluorescence kinetics using the saturating pulse method e.
However, the basal and maximum fluorescence values are rather variable between different samples, even under non-stressed conditions, and their use can, therefore, become a source of uncertainty.
Under high-temperature conditions, F O can slightly increase as high temperatures enhance the process of chlororespiration, leading to a partial reduction of Q A in the dark due to a more reduced PQ pool Sazanov et al.
In high-temperature-stressed samples with a large population of PSII with an impaired oxygen-evolving complex OEC , electron transport rate is lower and it takes longer to reach the maximum fluorescence intensity. This is frequently accompanied by a slowdown of the J — I rise and by an increase in the amplitude of the I — P rise Fig. Comparison of the effect of high-temperature stress applied in the light and in darkness on the OJIP kinetics. The K step has only been described in response to high-temperature stress or manganese deficiency, and its occurrence is thus a very specific symptom of high-temperature-induced damage of PSII.
This is clearly illustrated in Fig. High-temperature stress effects. Demonstration of the critical temperature T C : gradually increasing the leaf temperature leads above a certain critical temperature to an increase of the F O value. The K step is much more apparent if the leaves are heated in the dark Fig.
As a consequence, the K peak is much lower under such conditions Fig. RCs that lack a Mn cluster are very sensitive to light e. High-temperature susceptibility of PSII differs depending on species, age, physiological status, acclimation level, etc. The F O -T curve method is based on a continuous increase of sample temperature during which the F O value is continuously recorded. It is known that increasing the saturation level of membrane lipids increases the tolerance to high temperatures Murakami et al.
Critical temperatures represent the point at which a severe disorganization of structure and loss of main functions occur. On the other hand, the use of graduated temperatures with fast Chl a fluorescence not only gives a real-life T C measurement, but also enables the calculation of other parameters.
Moreover, the capacity to increase thermostability of the K step was higher than for the F O increase. High-temperature acclimation can occur quite quickly. PSII thermostability is only a small part of overall high-temperature tolerance.
The reversible inactivation of Rubisco activase occurs already at considerably lower temperatures than PSII inactivation. However, inactivation of the donor side of PSII takes considerably longer to repair than re-activation of Rubisco activase. As a consequence, PSII donor side inactivation may not so much play a role during a high-temperature episode; instead, it may have an effect on plant photosynthetic productivity in its aftermath.
By using Chl a fluorescence it is possible to detect the effects of environmental stressors e. Chl a fluorescence-based methods have been applied in ecotoxicological studies to examine the effects of pollutants on algae and plants Kumar et al.
These methods have many advantages over existing bioassays, especially with regard to sensitivity, rapidity, and the non-destructiveness and non-invasiveness of the methodology. Depending on the stressor, changes can be detected before visible symptoms appear Guidi et al.
While it is commonly assumed that parameters linked to PSII electron transport are indicators for toxicity in plants e. The effective quantum yield has been found to be a sensitive indicator for certain heavy metals, herbicides and petrochemicals Ralph and Burchet ; Haynes et al.
The tolerance to pollutants differs between plant species. Uptake reduction, compartmentalization or differential detoxification are strategies that have been demonstrated in plants Hall For every bioassay it is, therefore, important to choose plant or algal species that are sensitive to the pollutant or mixture of pollutants of interest.
In the case of crops, the comparison of sensitive and tolerant cultivars can also be interesting e. In summary, for a successful bioassay, the choice of a species sensitive to the pollutant and of a Chl a fluorescence parameter that is affected by the pollutant in the concentration range of interest is critical Choi et al.
Screening for herbicide efficacy and plant sensitivity is usually a lengthy process. The experiments take up a lot of space, and the evaluation of results, either quantitatively or qualitatively, is normally completed more than a week after the treatment Christensen et al. However, as shown by Krieger-Liszkay and Rutherford , the effect of these herbicides on the midpoint potential of Q A affects their working mechanism.
DCMU increases the midpoint potential of Q A , stabilizing the charge separation, whereas phenolic herbicides decrease the midpoint potential of Q A and destabilize the charge recombination see also Question The first type of herbicides reduces the probability that a charge recombination induces singlet oxygen, whereas the second type increases this probability.
In other words, the extent of inhibition is only half the story. The activities of a surprisingly large number of herbicides are directly or indirectly influenced by light Hess Herbicides that catalyze the formation of reactive oxygen species e. Glyphosate application leads to a depletion of free phosphate, leading to an inhibition of ATP synthesis, and isoxaflutole inhibits PQ synthesis. However, looking at the OJIP transients shown in that paper, these parameters seem to reflect mainly the destruction of either the photosynthetic system or alternatively PSII, since the main effect was observed on the F V amplitude.
The authors note that the fluorescence data obtained 48 h after the treatment gave the same information as the biomass measurements carried out 3 weeks after the treatment. The equipment works with low-intensity-modulated light that does not lead to the induction of fluorescence in the absence of herbicide, leads to the induction of fluorescence as a function of the fraction of PSII RCs inhibited by herbicide. Herbicides that inhibit the synthesis of amino acids or lipids have indirect effects on photosynthesis through their effect on the carbon metabolism or the stability of thylakoid membranes and may consequently alter PSII and PSI photochemical activity.
Their effects on the fluorescence kinetics will only be apparent after a much longer exposure time than in the case of PSII-type inhibitor herbicides Popovic et al. Olesen and Cedergreen and Yanniccari et al. This is true for OJIP transients, but it can be pointed out that a full Kautsky curve measured till the steady state is reached would also give information on the inhibition of CO 2 assimilation. As noted by Yanniccari et al. Chl a fluorescence imaging does not only allow the measurement of a whole plant or several small plants simultaneously, and it also makes it possible to follow the spread of herbicides in leaves or whole plants see Lichtenthaler et al.
Herbicide-induced perturbations of plant metabolism have been detected using changes in the derived images of fluorescence parameters before any visual effects on growth were observed Barbagallo et al. Konishi et al. This allowed the authors to show in detail how the herbicide, arriving in the leaf via the xylem, spread inside the leaf. Saura and Quiles , using Chl a fluorescence imaging, to compare the uptake of DCMU and paraquat in Chrysanthemum morifolium, Rosa meillandina and Spathiphyllum wallisii , showed 1 that paraquat, an herbicide acting on the acceptor side of PSI, can also be monitored by Chl a fluorescence imaging and 2 that the more water-soluble paraquat affected the leaves more homogeneously than DCMU.
Muller et al. Chl a fluorescence imaging has not only been used to detect the effects of herbicides on the photosynthetic performance of plants but also of algae as recently reviewed by Kumar et al. QTLs or quantitative trait loci refer to a location on a chromosome coding for one or more genes that affect a certain characteristic or process, e. It is likely that, e. The word quantitative refers to the fact that the genes linked to a QTL only have partial control over a characteristic.
With respect to photosynthesis there is one more peculiarity. QTLs are located in the nuclear genome, whereas many important photosynthetic genes are found in the chloroplast genome. Several studies have been carried out that had as goal to identify QTLs related to fluorescence parameters see Question 30 for a discussion of a rational choice of parameters for such studies. There are two main approaches to the identification of QTLs, but for the study of QTLs related to fluorescence parameters only one of them, association mapping, has been used see Flood et al.
Separately, a genetic map was made based on the DNA of 90 lines. In addition, genes related to the photosynthetic light reactions, pigment metabolism. The authors then tested, which of the studied parameters were linked, in the sense that they were inherited together in the different daughter lines. By studying the extent of linkage between the parameters and the genetic markers, the genes controlling the variability in the studied parameters could be assigned to areas on particular chromosomes.
Similar studies were carried out by several other groups. Stamp and coworkers published several QTL studies related to chilling tolerance in maize. Fracheboud et al. In a second study, Fracheboud et al. Apart from an assignment of several QTLs, these studies also yielded a lot of biological variability allowing the study of the relationship between different parameters. Yang et al. The authors noted that for each of the two conditions studied different QTLs were found. An observation that is made in several other studies as well.
Yin et al. The authors planted at different times in order to be able to measure all the plants at the same age. The authors identified 10 significant QTLs, but also observed that different QTLs were found for different environmental conditions. With respect to the JIP test parameters chosen in the above-mentioned studies, it can be noted that most of them have not been characterized physiologically.
See the next question for a discussion about the choice of parameters for this type of study. In most of the other cited studies the link with the physiology of the studied plants is less evident.
Fluorescence parameters reflect an underlying process or processes. If we would assume, for example, that a decrease in PSI content during stress is a marker for oxidative damage, then the resistance of the IP amplitude to stress could be a good candidate for a QTL study. Although the meaning of changes in the ratio between the amplitude of the photochemical phase and the amplitude of the thermal phase has not been established, it probably reflects a fundamental property of the chloroplast and may, therefore, be another candidate.
For the q E several genes have already been identified Jung and Niyogi and it may, therefore, be a less interesting parameter for a QTL study. The parameter q P , as a rough indicator of the balance between excitation pressure and electron flow, in response to a certain stress may be another candidate parameter to screen for stress resistance genes. Parameters that have not been characterized physiologically are less rational choices for QTL studies. Many JIP test parameters are only used conceptionally and have so far not been characterized physiologically in a proper way.
Quite a few of the JIP test parameters mentioned in the previous question belong to this category of conceptional parameters. Other parameters are so fundamental that they are unlikely to change much. QTL studies are, in principle, not so different from mutant screening. If you do not have a well-thought-out strategy, it is unlikely that you will obtain interesting information. In summary, in our opinion, QTL studies can yield more interesting information if a more rational approach, making better use of our knowledge of the meaning of fluorescence parameters, is applied.
As discussed in response to several questions in this paper e. Therefore, fluorescence parameters can, in principle, be used as selection tools in plant breeding programs and for analyzing genotype—environment interactions Araus et al. In quite a few studies this approach has been advocated Greaves and Wilson ; Baker and Rosenqvist ; Kalaji and Pietkiewicz There are a few points that should be considered: 1 It is important to obtain Chl a fluorescence-related traits showing a high correlation with yield or plant performance in addition to Chl a fluorescence-related traits that are specific for resistance to the stress of interest; 2 the measurements should cause only minimal perturbations in growth conditions: The shorter the measurements the better; 3 short measurements that are easy to carry out, to allow the accumulation of many measurements in a short time—plant characteristics change during growth in response to both age and environmental factors.
To allow comparisons between different cultivars, varieties or crosses of all plants of interest, measurements have to be made within a short time interval. With respect to point 1 , it may be noted that our knowledge of plant stress responses is often too limited to decide with certainty which trait will improve both stress resistance and plant yield. For example, in the case of photoinhibition it has been argued widely that this represents damage.
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