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BACKGROUND: Unfavorable temperatures significantly constrain the quality formation of Dendrobium officinale, severely limiting its food demand. Salicylic acid (SA) enhances the resistance of D. officinale to stress and possesses various analogs. The impact and mechanism of the SA family on improving the quality of D. officinale under adverse temperature conditions remains unclear. RESULTS: Combined with molecular docking analysis, chlorophyll fluorescence and metabolic analysis after treatments with SA analogues or extreme temperatures are performed in this study. The results demonstrate that both heat and cold treatments impede several main parameters of chlorophyll fluorescence of D. officinale, including the ΦPSII parameter, a sensitive growth indicator. However, this inhibition is mitigated by SA or its chemically similar compounds. Comprehensive branch imaging of ΦPSII values revealed position-dependent improvement of tolerance. Molecular docking analysis using a crystal structure model of NPR4 protein reveals that the therapeutic effects of SA analogs are determined by their binding energy and the contact of certain residues. Metabolome analysis identifies 17 compounds are considered participating in the temperature-related SA signaling pathway. Moreover, several natural SA analogs such as 2-hydroxycinnamic acid, benzamide, 2-(formylamino) benzoic acid and 3-o-methylgallic acid, are further found to have high binding ability to NPR4 protein and probably enhance the tolerance of D. officinale against unfavorable temperatures through flavone and guanosine monophosphate degradation pathways. CONCLUSIONS: These results reveal that the SA family with a high binding capability of NPR4 could improve the tolerance of D. officinale upon extreme temperature challenges. This study also highlights the collaborative role of SA-related natural compounds present in D. officinale in the mechanism of temperature resistance and offers a potential way to develop protective agents for the cultivation of D. officinale.

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Soil salinization poses a critical challenge to global food security, impacting plant growth, development, and crop yield. This study investigates the efficacy of deep learning techniques alongside chlorophyll fluorescence (ChlF) imaging technology for discerning varying levels of salt stress in soybean seedlings. Traditional methods for stress identification in plants are often laborious and time-intensive, prompting the exploration of more efficient approaches. A total of six classic convolutional neural network (CNN) models-AlexNet, GoogLeNet, ResNet50, ShuffleNet, SqueezeNet, and MobileNetv2-are evaluated for salt stress recognition based on three types of ChlF images. Results indicate that ResNet50 outperforms other models in classifying salt stress levels across three types of ChlF images. Furthermore, feature fusion after extracting three types of ChlF image features in the average pooling layer of ResNet50 significantly enhanced classification accuracy, achieving the highest accuracy of 98.61% in particular when fusing features from three types of ChlF images. UMAP dimensionality reduction analysis confirms the discriminative power of fused features in distinguishing salt stress levels. These findings underscore the efficacy of deep learning and ChlF imaging technologies in elucidating plant responses to salt stress, offering insights for precision agriculture and crop management. Overall, this study demonstrates the potential of integrating deep learning with ChlF imaging for precise and efficient crop stress detection, offering a robust tool for advancing precision agriculture. The findings contribute to enhancing agricultural sustainability and addressing global food security challenges by enabling more effective crop stress management.

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In recent years, inorganic nanoparticles, including calcium hydroxide nanoparticles [Ca Ca(OH)2 NPs], have attracted significant interest for their ability to impact plant photosynthesis and boost agricultural productivity. In this study, the effects of 15 and 30 mg L-1 oleylamine-coated calcium hydroxide nanoparticles [Ca(OH)2@OAm NPs] on photosystem II (PSII) photochemistry were investigated on tomato plants at their growth irradiance (GI) (580 µmol photons m-2 s-1) and at high irradiance (HI) (1000 µmol photons m-2 s-1). Ca(OH)2@OAm NPs synthesized via a microwave-assisted method revealed a crystallite size of 25 nm with 34% w/w of oleylamine coater, a hydrodynamic size of 145 nm, and a ζ-potential of 4 mV. Compared with the control plants (sprayed with distilled water), PSII efficiency in tomato plants sprayed with Ca(OH)2@OAm NPs declined as soon as 90 min after the spray, accompanied by a higher excess excitation energy at PSII. Nevertheless, after 72 h, the effective quantum yield of PSII electron transport (ΦPSII) in tomato plants sprayed with Ca(OH)2@OAm NPs enhanced due to both an increase in the fraction of open PSII reaction centers (qp) and to the enhancement in the excitation capture efficiency (Fv'/Fm') of these centers. However, the decrease at the same time in non-photochemical quenching (NPQ) resulted in an increased generation of reactive oxygen species (ROS). It can be concluded that Ca(OH)2@OAm NPs, by effectively regulating the non-photochemical quenching (NPQ) mechanism, enhanced the electron transport rate (ETR) and decreased the excess excitation energy in tomato leaves. The delay in the enhancement of PSII photochemistry by the calcium hydroxide NPs was less at the GI than at the HI. The enhancement of PSII function by calcium hydroxide NPs is suggested to be triggered by the NPQ mechanism that intensifies ROS generation, which is considered to be beneficial. Calcium hydroxide nanoparticles, in less than 72 h, activated a ROS regulatory network of light energy partitioning signaling that enhanced PSII function. Therefore, synthesized Ca(OH)2@OAm NPs could potentially be used as photosynthetic biostimulants to enhance crop yields, pending further testing on other plant species.

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Water deficit is the major stress factor magnified by climate change that causes the most reductions in plant productivity. Knowledge of photosystem II (PSII) response mechanisms underlying crop vulnerability to drought is critical to better understanding the consequences of climate change on crop plants. Salicylic acid (SA) application under drought stress may stimulate PSII function, although the exact mechanism remains essentially unclear. To reveal the PSII response mechanism of celery plants sprayed with water (WA) or SA, we employed chlorophyll fluorescence imaging analysis at 48 h, 96 h, and 192 h after watering. The results showed that up to 96 h after watering, the stroma lamellae of SA-sprayed leaves appeared dilated, and the efficiency of PSII declined, compared to WA-sprayed plants, which displayed a better PSII function. However, 192 h after watering, the stroma lamellae of SA-sprayed leaves was restored, while SA boosted chlorophyll synthesis, and by ameliorating the osmotic potential of celery plants, it resulted in higher relative leaf water content compared to WA-sprayed plants. SA, by acting as an antioxidant under drought stress, suppressed phototoxicity, thereby offering PSII photoprotection, together with enhanced effective quantum yield of PSII photochemistry (ΦPSII) and decreased quantity of singlet oxygen (1O2) generation compared to WA-sprayed plants. The PSII photoprotection mechanism induced by SA under drought stress was triggered by non-photochemical quenching (NPQ), which is a strategy to protect the chloroplast from photo-oxidative damage by dissipating the excess light energy as heat. This photoprotective mechanism, triggered by NPQ under drought stress, was adequate in keeping, especially in high-light conditions, an equal fraction of open PSII reaction centers (qp) as of non-stress conditions. Thus, under water deficit stress, SA activates a regulatory network of stress and light energy partitioning signaling that can mitigate, to an extent, the water deficit stress on PSII functioning.

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Photosystem II (PSII) functions were investigated in basil (Ocimum basilicum L.) plants sprayed with 1 mM salicylic acid (SA) under non-stress (NS) or mild drought-stress (MiDS) conditions. Under MiDS, SA-sprayed leaves retained significantly higher (+36%) chlorophyll content compared to NS, SA-sprayed leaves. PSII efficiency in SA-sprayed leaves under NS conditions, evaluated at both low light (LL, 200 µmol photons m-2 s-1) and high light (HL, 900 µmol photons m-2 s-1), increased significantly with a parallel significant decrease in the excitation pressure at PSII (1-qL) and the excess excitation energy (EXC). This enhancement of PSII efficiency under NS conditions was induced by the mechanism of non-photochemical quenching (NPQ) that reduced singlet oxygen (1O2) production, as indicated by the reduced quantum yield of non-regulated energy loss in PSII (ΦNO). Under MiDS, the thylakoid structure of water-sprayed leaves appeared slightly dilated, and the efficiency of PSII declined, compared to NS conditions. In contrast, the thylakoid structure of SA-sprayed leaves did not change under MiDS, while PSII functionality was retained, similar to NS plants at HL. This was due to the photoprotective heat dissipation by NPQ, which was sufficient to retain the same percentage of open PSII reaction centers (qp), as in NS conditions and HL. We suggest that the redox status of the plastoquinone pool (qp) under MiDS and HL initiated the acclimation response to MiDS in SA-sprayed leaves, which retained the same electron transport rate (ETR) with control plants. Foliar spray of SA could be considered as a method to improve PSII efficiency in basil plants under NS conditions, at both LL and HL, while under MiDS and HL conditions, basil plants could retain PSII efficiency similar to control plants.

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Climate change caused by global warming involves crucial plant growth factors such as atmospheric CO2 concentration, ambient temperature or water availability. These stressors usually co-occur, causing intricate alterations in plant physiology and development. This work focuses on how elevated atmospheric CO2 levels, together with the concomitant high temperature, would affect the physiology of a relevant crop, such as broccoli. Particular attention has been paid to those defence mechanisms that contribute to plant fitness under abiotic stress. Results show that both photosynthesis and leaf transpiration were reduced in plants grown under climate change environments compared to those grown under current climate conditions. Furthermore, an induction of carbohydrate catabolism pointed to a redistribution from primary to secondary metabolism. This result could be related to a reinforcement of cell walls, as well as to an increase in the pool of antioxidants in the leaves. Broccoli plants, a C3 crop, grown under an intermediate condition showed activation of those adaptive mechanisms, which would contribute to coping with abiotic stress, as confirmed by reduced levels of lipid peroxidation relative to current climate conditions. On the contrary, the most severe climate change scenario exceeded the adaptive capacity of broccoli plants, as shown by the inhibition of growth and reduced vigour of plants. In conclusion, only a moderate increase in atmospheric CO2 concentration and temperature would not have a negative impact on broccoli crop yields.

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Early detection of pathogens can significantly reduce yield losses and improve the quality of agricultural products. This study compares the efficiency of hyperspectral (HS) imaging and pulse amplitude modulation (PAM) fluorometry to detect pathogens in plants. Reflectance spectra, normalized indices, and fluorescence parameters were studied in healthy and infected areas of leaves. Potato virus X with GFP fluorescent protein was used to assess the spread of infection throughout the plant. The study found that infection increased the reflectance of leaves in certain wavelength ranges. Analysis of the normalized reflectance indices (NRIs) revealed indices that were sensitive and insensitive to infection. NRI700/850 was optimal for virus detection; significant differences were detected on the 4th day after the virus arrived in the leaf. Maximum (Fv/Fm) and effective quantum yields of photosystem II (ΦPSII) and non-photochemical fluorescence quenching (NPQ) were almost unchanged at the early stage of infection. ΦPSII and NPQ in the transition state (a short time after actinic light was switched on) showed high sensitivity to infection. The higher sensitivity of PAM compared to HS imaging may be due to the possibility of assessing the physiological changes earlier than changes in leaf structure.

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Taxus is a rare and endangered woody plant worldwide with important economic and ecological values. However, the weak environmental adaptability of Taxus species, in particular the unstable photosynthetic activity in different seasons, always affects its normal growth and development and limits its conservation and exploitation. To improve the survival of Taxus trees in cultivated areas, the seasonal dynamics of chlorophyll fluorescence (CF) and key physiological parameters were comprehensively investigated in T. media and T. mairei. The results demonstrated that the photosynthetic activity of both Taxus species was sensitive to local summer and winter environmental conditions, with the heterogeneity of fluorescence signatures intuitively presented on the needle surface by CF-Imaging detection, while images of maximum quantum efficiency of PSII photochemistry (Fv/Fm) demonstrated values below 0.7 in the blue-green sectors in winter. The distribution of light energy was regulated by the photosynthetic apparatus in both Taxus species to maintain a stable actual quantum yield of PSII photochemistry (φPSII), which was around 0.4-0.5. Based on a redundancy discriminant analysis, the interpretation rate of light intensity and air temperature ranked as the top two in both Taxus species, which were considered the main environmental factors affecting the photosynthetic performance of Taxus by disturbing the electron transport chain. In the winter, T. mairei exhibited weaker electron transport activity than T. media, thus caused lower photochemistry and more severe photosynthetic damages. Interestingly, both Taxus species demonstrated consistent response patterns, including diverse energy dissipation strategies and enhancement of osmoregulatory substances and antioxidative activities, thus maintaining stable photosynthetic functions in response to environmental changes.

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The efficiency of photosynthesis in strawberry plants is measured to maintain the quality and quantity of strawberries produced. The latest method used to measure the photosynthetic status of plants is chlorophyll fluorescence imaging (CFI), which has the advantage of obtaining plant spatiotemporal data non-destructively. This study developed a CFI system to measure the maximum quantum efficiency of photochemistry (Fv/Fm). The main components of this system include a chamber for plants to adapt to dark environments, blue LED light sources to excite the chlorophyll in plants, and a monochrome camera with a lens filter attached to capture the emission spectra. In this study, 120 pots of strawberry plants were cultivated for 15 days and divided into four treatment groups: control, drought stress, heat stress, and a combination of drought and heat stress, resulting in Fv/Fm values of 0.802 ± 0.0036, 0.780 ± 0.0026, 0.768 ± 0.0023, and 0.749 ± 0.0099, respectively. A strong correlation was found between the developed system and a chlorophyll meter (r = 0.75). These results prove that the developed CFI system can accurately capture the spatial and temporal dynamics resulting from the response of strawberry plants to abiotic stresses.

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Chrysanthemum (Chrysanthemum morifolium) is among the most popular ornamental plants, propagated mainly through stem cuttings. There is a lack of information regarding the impact of the lighting environment on the successful production of cuttings and underlying mechanisms. The light spectrum affects plant morphology, growth, and photosynthesis. In the present study, chrysanthemum, cv. 'Katinka' cuttings, were exposed to five lighting spectra, including monochromatic red (R), blue (B) lights, and multichromatic lights, including a combination of R and B (R:B), a combination of R, B, and far red (R:B:FR) and white (W), for 30 days. B light enhanced areal growth, as indicated by a higher shoot mass ratio, while R light directed the biomass towards the underground parts of the cuttings. Monochromatic R and B lights promoted the emergence of new leaves. In contrast, individual leaf area was largest under multichromatic lights. Exposing the cuttings to R light led to the accumulation of carbohydrates in the leaves. Cuttings exposed to multichromatic lights showed higher chlorophyll content than monochromatic R- and B-exposed cuttings. Conversely, carotenoid and anthocyanin contents were the highest in monochromatic R- and B-exposed plants. B-exposed cuttings showed higher photosynthetic performance, exhibited by the highest performance index on the basis of light absorption, and maximal quantum yield of PSII efficiency. Although R light increased biomass toward roots, B light improved above-ground growth, photosynthetic functionality, and the visual performance of Chrysanthemum cuttings.

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Exposure of Salvia sclarea plants to excess Zn for 8 days resulted in increased Ca, Fe, Mn, and Zn concentrations, but decreased Mg, in the aboveground tissues. The significant increase in the aboveground tissues of Mn, which is vital in the oxygen-evolving complex (OEC) of photosystem II (PSII), contributed to the higher efficiency of the OEC, and together with the increased Fe, which has a fundamental role as a component of the enzymes involved in the electron transport process, resulted in an increased electron transport rate (ETR). The decreased Mg content in the aboveground tissues contributed to decreased chlorophyll content that reduced excess absorption of sunlight and operated to improve PSII photochemistry (ΦPSII), decreasing excess energy at PSII and lowering the degree of photoinhibition, as judged from the increased maximum efficiency of PSII photochemistry (Fv/Fm). The molecular mechanism by which Zn-treated leaves displayed an improved PSII photochemistry was the increased fraction of open PSII reaction centers (qp) and, mainly, the increased efficiency of the reaction centers (Fv'/Fm') that enhanced ETR. Elemental bioimaging of Zn and Ca by laser ablation-inductively coupled plasma-mass spectrometry (LA-ICP-MS) revealed their co-localization in the mid-leaf veins. The high Zn concentration was located in the mid-leaf-vein area, while mesophyll cells accumulated small amounts of Zn, thus resembling a spatiotemporal heterogenous response and suggesting an adaptive strategy. These findings contribute to our understanding of how exposure to excess Zn triggered a hormetic response of PSII photochemistry. Exposure of aromatic and medicinal plants to excess Zn in hydroponics can be regarded as an economical approach to ameliorate the deficiency of Fe and Zn, which are essential micronutrients for human health.

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The development of automated, image-based, high-throughput plant phenotyping enabled the simultaneous measurement of many plant traits. Big and complex phenotypic datasets require advanced statistical methods which enable the extraction of the most valuable traits when combined with other measurements, interpretation, and understanding of their (eco)physiological background. Nutrient deficiency in plants causes specific symptoms that can be easily detected by multispectral imaging, 3D scanning, and chlorophyll fluorescence measurements. Screening of numerous image-based phenotypic traits of common bean plants grown in nutrient-deficient solutions was conducted to optimize phenotyping and select the most valuable phenotypic traits related to the specific nutrient deficit. Discriminant analysis was used to compare the efficiency of groups of traits obtained by high-throughput phenotyping techniques (chlorophyll fluorescence, multispectral traits, and morphological traits) in discrimination between nutrients [nitrogen (N), phosphorus (P), potassium (K), magnesium (Mg), and iron (Fe)] at early and prolonged deficiency. Furthermore, a recursive partitioning analysis was used to select variables within each group of traits that show the highest accuracy for assigning plants to the respective nutrient deficit treatment. Using the entire set of measured traits, the highest classification success by discriminant function was achieved using multispectral traits. In the subsequent measurements, chlorophyll fluorescence and multispectral traits achieved comparably high classification success. Recursive partitioning analysis was able to intrinsically identify variables within each group of traits and their threshold values that best separate the observations from different nutrient deficiency groups. Again, the highest success in assigning plants into their respective groups was achieved based on selected multispectral traits. Selected chlorophyll fluorescence traits also showed high accuracy for assigning plants into control, Fe, Mg, and P deficit but could not correctly assign K and N deficit plants. This study has shown the usefulness of combining high-throughput phenotyping techniques with advanced data analysis to determine and differentiate nutrient deficiency stress.

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Potato, Solanum tuberosum L., one of the most commonly cultivated horticultural crops throughout the world, is susceptible to a variety of herbivory insects. In the present study, we evaluated the consequence of feeding by the sap-sucking insect Halyomorpha halys on potato leaf photosynthetic efficiency. By using chlorophyll fluorescence imaging methodology, we examined photosystem II (PSII) photochemistry in terms of feeding and at the whole leaf area. The role of reactive oxygen species (ROS) in potato's defence response mechanism immediately after feeding was also assessed. Even 3 min after feeding, increased ROS generation was observed to diffuse through the leaf central vein, probably to act as a long-distance signalling molecule. The proportion of absorbed energy being used in photochemistry (ΦPSII) at the whole leaf level, after 20 min of feeding, was reduced by 8% compared to before feeding due to the decreased number of open PSII reaction centres (qp). After 90 min of feeding, ΦPSII decreased by 46% at the whole leaf level. Meanwhile, at the feeding zones, which were located mainly in the proximity of the leaf midrib, ΦPSII was lower than 85%, with a concurrent increase in singlet-excited oxygen (1O2) generation, which is considered to be harmful. However, the photoprotective mechanism (ΦNPQ), which was highly induced 90 min after feeding, was efficient to compensate for the decrease in the quantum yield of PSII photochemistry (ΦPSII). Therefore, the quantum yield of non-regulated energy loss in PSII (ΦNO), which represents 1O2 generation, remained unaffected at the whole leaf level. We suggest that the potato PSII response to sap-sucking insect feeding underlies the ROS-dependent signalling that occurs immediately and initiates a photoprotective PSII defence response to reduce herbivory damage. A controlled ROS burst can be considered the primary plant defence response mechanism to herbivores.

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Duckweeds (Lemnaceae species) are extensively used models in ecotoxicology, and chlorophyll fluorescence imaging offers a sensitive and high throughput platform for phytotoxicity assays with these tiny plants. However, the vast number of potentially applicable chlorophyll fluorescence-based test endpoints makes comparison and generalization of results hard among different studies. The present study aimed to jointly measure and compare the sensitivity of various chlorophyll fluorescence parameters in Spirodela polyrhiza (giant duckweed) plants exposed to nickel, chromate (hexavalent chromium) and sodium chloride for 72 h, respectively. The photochemistry of Photosystem II in both dark- and light-adapted states of plants was assessed via in vivo chlorophyll fluorescence imaging method. Our results indicated that the studied parameters responded with very divergent sensitivity, highlighting the importance of parallelly assessing several chlorophyll fluorescence parameters. Generally, the light-adapted parameters were more sensitive than the dark-adapted ones. Thus, the former ones might be the preferred endpoints in phytotoxicity assays. Fv/Fm, i.e., the most extensively reported parameter literature-wise, proved to be the least sensitive endpoint; therefore, future studies might also consider reporting Fv/Fo, as its more responsive analogue. The tested toxicants induced different trends in the basic chlorophyll fluorescence parameters and, at least partly, in relative proportions of different quenching processes, suggesting that a basic distinction of water pollutants with different modes of action might be achievable by this method. We found definite hormetic patterns in responses to several endpoints. Hormesis occurred in the concentration ranges where the applied toxicants resulted in strong growth inhibition in longer-term exposures of the same duckweed clone in previous studies. These findings indicate that changes in the photochemical efficiency of plants do not necessarily go hand in hand with growth responses, and care should be taken when one exclusively interprets chlorophyll fluorescence-based endpoints as general proxies for phytotoxic effects.

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Chlorophyll fluorescence imaging was used to study potato virus X (PVX) infection of Nicotiana benthamiana. Infection-induced changes in chlorophyll fluorescence parameters (quantum yield of photosystem II photochemistry (ΦPSII) and non-photochemical fluorescence quenching (NPQ)) in the non-inoculated leaf were recorded and compared with the spatial distribution of the virus detected by the fluorescence of GFP associated with the virus. We determined infection-related changes at different points of the light-induced chlorophyll fluorescence kinetics and at different days after inoculation. A slight change in the light-adapted steady-state values of ΦPSII and NPQ was observed in the infected area of the non-inoculated leaf. In contrast to the steady-state parameters, the dynamics of ΦPSII and NPQ caused by the dark-light transition in healthy and infected areas differed significantly starting from the second day after the detection of the virus in a non-inoculated leaf. The coefficients of correlation between chlorophyll fluorescence parameters and virus localization were 0.67 for ΦPSII and 0.76 for NPQ. In general, the results demonstrate the possibility of reliable pre-symptomatic detection of the spread of a viral infection using chlorophyll fluorescence imaging.

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To investigate the importance of light on healing and acclimatization, in the present study, grafted watermelon seedlings were exposed to darkness (D) or light, provided by blue (B), red (R), a mixture of R (68%) and B (RB), or white (W; 35% B, 49% intermediate spectra, 16% R) LEDs for 12 days. Survival ratio, root and shoot growth, soluble carbohydrate content, photosynthetic pigments content, and photosynthetic performance were evaluated. Seedling survival was not only strongly limited in D but the survived seedlings had an inferior shoot and root development, reduced chlorophyll content, and attenuated photosynthetic efficiency. RB-exposed seedlings had a less-developed root system. R-exposed seedlings showed leaf epinasty, and had the smallest leaf area, reduced chlorophyll content, and suppressed photosynthetic apparatus performance. The R-exposed seedlings contained the highest amount of soluble carbohydrate and together with D-exposed seedlings the lowest amount of chlorophyll in their scions. B-exposed seedlings showed the highest chlorophyll content and improved overall PSII photosynthetic functioning. W-exposed seedling had the largest leaf area, and closely resembled the photosynthetic properties of RB-exposed seedlings. We assume that, during healing of grafted seedlings monochromatic R light should be avoided. Instead, W and monochromatic B light may be willingly adopted due to their promoting effect on shoot, pigments content, and photosynthetic efficiency.

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In addition to direct tissue consumption, herbivory may affect other important plant processes. Here, we evaluated the effects of short-time leaf feeding by Spodoptera exigua larvae on the photosynthetic efficiency of tomato plants, using chlorophyll a fluorescence imaging analysis. After 15 min of feeding, the light used for photochemistry at photosystem II (PSII) (ΦPSII), and the regulated heat loss at PSII (ΦNPQ) decreased locally at the feeding zones, accompanied by increased non-regulated energy losses (ΦNO) that indicated increased singlet oxygen (1O2) formation. In contrast, in zones neighboring the feeding zones and in the rest of the leaf, ΦPSII increased due to a decreased ΦNPQ. This suggests that leaf areas not directly affected by herbivory compensate for the photosynthetic losses by increasing the fraction of open PSII reaction centers (qp) and the efficiency of these centers (Fv'/Fm'), because of decreased non-photochemical quenching (NPQ). This compensatory reaction mechanism may be signaled by singlet oxygen formed at the feeding zone. PSII functionality at the feeding zones began to balance with the rest of the leaf 3 h after feeding, in parallel with decreased compensatory responses. Thus, 3 h after feeding, PSII efficiency at the whole-leaf level was the same as before feeding, indicating that the plant managed to overcome the feeding effects with no or minor photosynthetic costs.

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Pickling cucumbers are susceptible to chilling injury (CI) during postharvest refrigerated storage, which would result in quality degradation and economic loss. It is, thus, desirable to remove the defective fruit before they are marketed as fresh products or processed into pickled products. Chlorophyll fluorescence is sensitive to CI in green fruits, because exposure to chilling temperatures can induce detectable alterations in chlorophylls of tissues. This study evaluated the feasibility of using a dual-band chlorophyll fluorescence imaging (CFI) technique for detecting CI-affected pickling cucumbers. Chlorophyll fluorescence images at 675 nm and 750 nm were acquired from pickling cucumbers under the excitation of ultraviolet-blue light. The raw images were processed for vignetting corrections through bi-dimensional empirical mode decomposition and subsequent image reconstruction. The fluorescence images were effective for ascertaining CI-affected tissues, which appeared as dark areas in the images. Support vector machine models were developed for classifying pickling cucumbers into two or three classes using the features extracted from the fluorescence images. Fusing the features of fluorescence images at 675 nm and 750 nm resulted in overall accuracies of 96.9% and 91.2% for two-class (normal and injured) and three-class (normal, mildly and severely injured) classification, respectively, which are statistically significantly better than those obtained using the features at a single wavelength, especially for the three-class classification. Furthermore, a subset of features, selected based on the neighborhood component feature selection technique, achieved the highest accuracies of 97.4% and 91.3% for the two-class and three-class classification, respectively. This study demonstrated that dual-band CFI is an effective modality for CI detection in pickling cucumbers.

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We evaluated photosystem II (PSII) functionality in potato plants (Solanum tuberosum L.) before and after a 15 min feeding by the leaf miner Tuta absoluta using chlorophyll a fluorescence imaging analysis combined with reactive oxygen species (ROS) detection. Fifteen minutes after feeding, we observed at the feeding zone and at the whole leaf a decrease in the effective quantum yield of photosystem II (PSII) photochemistry (ΦPSII). While at the feeding zone the quantum yield of regulated non-photochemical energy loss in PSII (ΦNPQ) did not change, at the whole leaf level there was a significant increase. As a result, at the feeding zone a significant increase in the quantum yield of non-regulated energy loss in PSII (ΦNO) occurred, but there was no change at the whole leaf level compared to that before feeding, indicating no change in singlet oxygen (1O2) formation. The decreased ΦPSII after feeding was due to a decreased fraction of open reaction centers (qp), since the efficiency of open PSII reaction centers to utilize the light energy (Fv'/Fm') did not differ before and after feeding. The decreased fraction of open reaction centers resulted in increased excess excitation energy (EXC) at the feeding zone and at the whole leaf level, while hydrogen peroxide (H2O2) production was detected only at the feeding zone. Although the whole leaf PSII efficiency decreased compared to that before feeding, the maximum efficiency of PSII photochemistry (Fv/Fm), and the efficiency of the water-splitting complex on the donor side of PSII (Fv/Fo), did not differ to that before feeding, thus they cannot be considered as sensitive parameters to monitor biotic stress effects. Chlorophyll fluorescence imaging analysis proved to be a good indicator to monitor even short-term impacts of insect herbivory on photosynthetic function, and among the studied parameters, the reduction status of the plastoquinone pool (qp) was the most sensitive and suitable indicator to probe photosynthetic function under biotic stress.

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Singlet oxygen (1O2) is an important damaging agent, which is produced during illumination by the interaction of the triplet excited state pigment molecules with molecular oxygen. In cells of photosynthetic organisms 1O2 is formed primarily in chlorophyll containing complexes, and damages pigments, lipids, proteins and other cellular constituents in their environment. A useful approach to study the physiological role of 1O2 is the utilization of external photosensitizers. In the present study, we employed a multiwell plate-based screening method in combination with chlorophyll fluorescence imaging to characterize the effect of externally produced 1O2 on the photosynthetic activity of isolated thylakoid membranes and intact Chlorella sorokiniana cells. The results show that the external 1O2 produced by the photosensitization reactions of Rose Bengal damages Photosystem II both in isolated thylakoid membranes and in intact cells in a concentration dependent manner indicating that 1O2 plays a significant role in photodamage of Photosystem II.

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