Natural variation in stomatal dynamics drives divergence in heat stress tolerance and contributes to the seasonal intrinsic water-use efficiency in Vitis vinifera (subsp. sativa and sylvestris)

Stomata control CO2 uptake for photosynthesis and water loss through transpiration, thus playing a key role in leaf thermoregulation, water-use efficiency (iWUE) and plant productivity. In this work, we investigated the relationship between several leaf traits and hypothesized that stomatal behavior to fast (i.e. minutes) environmental changes co-determines along with steady-state traits the physiological response of grapevine to the surrounding fluctuating environment over the growing season. No relationship between iWUE, heat stress (HS) tolerance and stomatal traits was observed in field grown grapevine, suggesting that other physiological mechanisms are involved in determining leaf evaporative cooling capacity and the seasonal ratio of CO2 uptake (A) to stomatal conductance (gs). Indeed, cultivars that in the field had an unexpected combination of high iWUE but low sensitivity to thermal stress, displayed a quick stomatal closure to light, but a sluggish closure to increased vapor pressure deficit (VPD) levels. This strategy aiming both at conserving water under a high-to-low light transition and in prioritizing evaporative cooling under a low-to-high VPD transition, was mainly observed in Regina and Syrah. Moreover, cultivars with different known responses to soil moisture deficit or high air VPD (isohydric vs anisohydric) had opposite behavior under fluctuating environments, with the isohydric cultivar showing slow stomatal closure to reduced light intensity but quick temporal responses to VPD manipulation. We propose that stomatal behavior to fast environmental fluctuations can play a critical role on leaf thermoregulation and water conservation under natural field conditions in grapevine.

Melatonin Enhances Drought Tolerance by Regulating Leaf Stomatal Behavior, Carbon and Nitrogen Metabolism, and Related Gene Expression in Maize Plants

It is commonly known that exogenously applied melatonin can alleviate the impact of drought stress, but the mechanism used by melatonin to regulate stomatal behavior and carbon (C) and nitrogen (N) metabolism to increase drought resistance remains elusive. Herein, our aim was to investigate the influence of exogenous melatonin on the regulation of C and N metabolism in maize plants under water deficit. In this study, we analyzed stomatal behavior, the key components of C and N metabolism, and the gene expression and activity of enzymes involved in the C and N metabolism in maize plants. The results showed that the application of melatonin (100 μM) significantly increased maize growth and sustained the opening of stomata, and secondarily increased the photosynthetic capacity in maize. Under drought stress, foliar application of melatonin induced the gene transcription and activities of sucrose phosphate synthetase, ADP-glucose pyrophosphorylase, phosphoenolpyruvate carboxylase, and citrate synthase, resulting in the enhancement of sucrose and starch synthesis and the tricarboxylic acid (TCA) cycle. This enhancement in sugar biosynthesis and the TCA cycle might lead to stronger N assimilation. As anticipated, NO3– reduction and NH4+ assimilation were also strengthened after melatonin treatment under drought stress. An increase was observed in some key enzymatic activities and transcription involved in nitrogen metabolism, such as that of nitrate reductase, nitrite reductase, glutamate synthase, and glutamine synthetase, in melatonin-treated, drought-stressed maize. Moreover, melatonin attenuated the drought-induced damage by reducing protein degradation and increasing the level of proline. Conclusively, our results indicate that exogenous melatonin enhances drought tolerance in maize via promoting stomatal opening and regulating C and N metabolism and related gene expression.

Salt stress slows down dynamic photosynthesis mainly through osmotic effects on dynamic stomatal behavior

Salt stress affects stomatal behavior and photosynthesis, by a combination of osmotic and ionic components, but it is unknown how these components affect photosynthesis dynamics under fluctuating light. Tomato (Solanum lycopersicum) plants were grown using a reference nutrient solution (Control, EC: 2.3 dS m-1), the reference containing extra macronutrients (only osmotic effect; EC: 12.6 dS m-1), or the reference containing an additional 100 mM NaCl (osmotic and ionic effects; EC: 12.8 dS m-1). Steady-state and dynamic photosynthesis along with leaf biochemistry were characterized throughout leaf development. Osmotic effects resulted in increased leaf chlorophyll content per unit leaf area, induced stomatal closure along with rapid stomatal responses to changes in light intensity, and limited dynamic but not steady-state photosynthesis. Ionic effects were barely observed in plant growth and dynamic photosynthesis, but led to a reduction in leaf chlorophyll content and photosynthetic capacity in old leaves. Steady-state and dynamic photosynthesis traits decreased with leaf age, due to increases in stomatal and non-stomatal limitations. With increasing leaf age, rates of light-triggered stomatal movement decreased across treatments, which is more strongly for stomatal opening rather than closure. We conclude that osmotic effect strongly impacts dynamic stomatal and photosynthetic behavior under salt stress.

An Integrated Method for Tracking and Monitoring Stomata Dynamics from Microscope Videos

Patchy stomata are a common and characteristic phenomenon in plants. Understanding and studying the regulation mechanism of patchy stomata are of great significance to further supplement and improve the stomatal theory. Currently, the common methods for stomatal behavior observation are based on static images, which makes it difficult to reflect dynamic changes of stomata. With the rapid development of portable microscopes and computer vision algorithms, it brings new chances for stomatal movement observation. In this study, a stomatal behavior observation system (SBOS) was proposed for real-time observation and automatic analysis of each single stoma in wheat leaf using object tracking and semantic segmentation methods. The SBOS includes two modules: the real-time observation module and the automatic analysis module. The real-time observation module can shoot videos of stomatal dynamic changes. In the automatic analysis module, object tracking locates every single stoma accurately to obtain stomatal pictures arranged in time-series; semantic segmentation can precisely quantify the stomatal opening area (SOA), with a mean pixel accuracy (MPA) of 0.8305 and a mean intersection over union (MIoU) of 0.5590 in the testing set. Moreover, we designed a graphical user interface (GUI) so that researchers could use this automatic analysis module smoothly. To verify the performance of the SBOS, the dynamic changes of stomata were observed and analyzed under chilling. Finally, we analyzed the correlation between gas exchange and SOA under drought stress, and the correlation coefficients between mean SOA and net photosynthetic rate (Pn), intercellular CO2 concentration (Ci), stomatal conductance (Gs), and transpiration rate (Tr) are 0.93, 0.96, 0.96, and 0.97.

Influence of Polyethylene Glycol on Leaf Anatomy, Stomatal Behavior, Water Loss, and Some Physiological Traits of Date Palm Plantlets Grown In Vitro and Ex Vitro

Few reports explain the mechanism of PEG action on stomatal behavior and anatomical structure and analyze the photosynthetic pigments of in vitro date palm plantlets for better tolerance to ex vitro exposure. The main challenge for in vitro micropropagation of date palm techniques remains restricted to high survival rates and vigorous growth after ex vitro transplantation. In vitro hardening is induced by Polyethylene glycol PEG (0.0, 10, 20, 30 g L−1) for 4 weeks. Leaf anatomy, stomatal behavior, water loss %, photosynthetic pigments, and reducing sugars were examined in date palm plantlets (Phoenix dactylifera L.) cv. (Sewi) after 4 weeks from in vitro PEG treatment and after 4 weeks from ex vitro transplanting to the greenhouse. Leaf anatomy and the surface ultrastructure of in vitro untreated leaves showed a thin cuticle layer, wide opened malfunctioning stomata, and abnormal leaf anatomy. Furthermore, addition of PEG resulted in increasing cuticle thickness, epicuticular wax depositions, and plastids density, improving the stomatal ability to close and decreasing the stomatal aperture length while reducing the substomatal chambers and intercellular spaces in the mesophyll. As a result, a significant reduction in water loss % was observed in both in vitro and ex vitro PEG treated leaves as compared to untreated ones, which exhibited rapid wilting when exposed to low humidity for 4 h. PEG application significantly increased Chlorophylls a, b and carotenoids concentrations, especially 10, 20 g L−1 treatments, which were sequentially reflected in increasing the reducing sugar concentration. However, leaves of plantlets treated with PEG at 30 g L−1 became yellow and had necrosis ends with death. In vitro hardening by 20 g L−1 PEG increased the survival rate of plantlets to 90% after ex vitro transfer compared to 63% recorded for the untreated plantlets. Therefore, this application provides normal date palm plantlets developed faster and enhances survival after ex vitro transfer.