Combination of high-throughput phenotyping with plant anatomical trait measurements to understand lettuce morpho-physiological acclimation under changing VPDs and watering regimes.

Nowadays, about 50% of the global yield loss is due to climate change. Increasing Vapor Pressure Deficit (VPD) and drought are among the principal environmental stressors, affecting stomatal regulation and reducing plant photosynthesis and biomass accumulation. Recent studies have revealed that the extent of plant acclimation is closely related to the anatomical traits of the leaves, which change with environmental conditions. It is not yet clear how the interaction between these environmental factors affects plant morpho-physiological development and plant capacity of acclimation under changing conditions. To fill this gap, in this study we used a high-throughput phenotyping facility (at the IPK-Gatersleben, Germany) to grow two lettuce cultivars (Lactuca sativa L. var. capitata) with green and red leaves under different VPDs (low and high) and watering regimes (well-watered, WW, and low watered, LW regimes). Two trials were performed: the first trial was conducted at a VPD of 0.7 kPa (low VPD) and the second at 1.4 kPa (high VPD), both with WW and WD conditions. After 12 days of cultivation in the phenotyping chamber, the environmental conditions were switched, and plants were kept for 5 days at the opposite VPD to evaluate their acclimation ability. RGB imaging was applied to track changes in morphological parameters, near-infrared camera (NIR) was used to estimate plant-water relationships, and FLUO made it possible to evaluate changes in photosystem II reflecting optimal/stressful conditions. At the end of the experimental trials, the leaf samples were characterized in terms of stomatal and mesophyll traits by light microscopy. A specific focus was d­edicated to exploring how stomata regulation and water use efficiency affect carbon gain and biomass allocation in pre-acclimated lettuces to different environmental conditions (VPDs) and hence undergoing sudden changes in the VPD. To test the influence of the different independent factors: i) VPD, ii) cultivar (C), iii) water (W) on the dependent variables, a three-way analysis of variance (ANOVA) was performed. Additionally, correlation plots and the principal component analysis were performed to explore correlations between morpho-anatomical and phenotypic data points. The results showed that WW plants at low VPD developed a morpho-anatomical structure in terms of mesophyll organization, stomatal and vein density which more efficiently guided acclimation to sudden changes in the environmental conditions and which was not detected by image-based phenotyping alone. Therefore, we emphasized the need to complement high-throughput phenotyping with the analysis of anatomical traits to unravel the mechanisms of crop acclimation under sudden fluctuation in environmental conditions due to climate change. Such an approach can help improving knowledge on how stomatal regulation and carbon allocation affect productivity in warmer areas and drier climates, with high impact also for the design of cultivation protocols for sustainable indoor farming.