Yield losses to waterlogging are expected to become an increasingly costly and frequent issue in some regions of the world. Despite the extensive work that has been carried out examining the molecular and physiological responses to waterlogging, phenotyping for waterlogging tolerance has proven difficult. This difficulty is largely due to the high variability of waterlogging conditions such as duration, temperature, soil type, and growth stage of the crop. In this review, we highlight use of phenotyping to assess and improve waterlogging tolerance in temperate crop species. We start by outlining the experimental methods that have been utilized to impose waterlogging stress, ranging from highly controlled conditions of hydroponic systems to large-scale screenings in the field. We also describe the phenotyping traits used to assess tolerance ranging from survival rates and visual scoring to precise photosynthetic measurements. Finally, we present an overview of the challenges faced in attempting to improve waterlogging tolerance, the trade-offs associated with phenotyping in controlled conditions, limitations of classic phenotyping methods, and future trends using plant-imaging methods. If effectively utilized to increase crop resilience to changing climates, crop phenotyping has a major role to play in global food security.

Waterlogging is expected to become a more prominent yield restricting stress for barley as rainfall frequency is increasing in many regions due to climate change. The duration of waterlogging events in the field is highly variable throughout the season, and this variation is also observed in experimental waterlogging studies. Such variety of protocols make intricate physiological responses challenging to assess and quantify. To assess barley waterlogging tolerance in controlled conditions, we present an optimal duration and setup of simulated waterlogging stress using image-based phenotyping. Six protocols durations, 5, 10, and 14 days of stress with and without seven days of recovery, were tested. To quantify the physiological effects of waterlogging on growth and greenness, we used top down and side view RGB (Red-Green-Blue) images. These images were taken daily throughout each of the protocols using the PSI PlantScreen™ imaging platform. Two genotypes of two-row spring barley, grown in glasshouse conditions, were subjected to each of the six protocols, with stress being imposed at the three-leaf stage. Shoot biomass and root imaging data were analysed to determine the optimal stress protocol duration, as well as to quantify the growth and morphometric changes of barley in response to waterlogging stress. Our time-series results show a significant growth reduction and alteration of greenness, allowing us to determine an optimal protocol duration of 14 days of stress and seven days of recovery for controlled conditions. Moreover, to confirm the reproducibility of this protocol, we conducted the same experiment in a different facility equipped with RGB and chlorophyll fluorescence imaging sensors. Our results demonstrate that the selected protocol enables the assessment of genotypic differences, which allow us to further determine tolerance responses in a glasshouse environment. Altogether, this work presents a new and reproducible image-based protocol to assess early stage waterlogging tolerance, empowering a precise quantification of waterlogging stress relevant markers such as greenness, Fv/Fm and growth rates.