Higher stomatal density and smaller stomatal size associate with higher grain yield in winter wheat

In wheat ( Triticum aestivum L.), flag leaf is crucial in determining grain-filling rate and final grain yield. Stomata, microscopic pores on plant surfaces, balance CO 2 uptake and evaporative water loss; their numbers and aperture sizes depend on environmental factors. Previous studies have reported associations between stomatal traits and grain yield; mostly under controlled conditions. Our study aimed to investigate flag leaf stomatal anatomical and physiological traits, and their relationships with winter wheat grain yield under field conditions. We measured stomatal density and size, and stomatal conductance on adaxial and abaxial flag leaf surfaces in a three-year field study with twenty-five winter wheat varieties. Stomatal density ratio remained constant despite different stomatal densities occurring in different years, indicating coordinated adjustments of stomatal numbers between leaf surfaces in response to environmental conditions. Stomatal conductance ratios varied widely, indicating leaf-surface specific stomatal responses to environmental conditions. Grain yield was positively associated with stomatal density and negatively with stomatal size. Flag leaf removal reduced grain weight per spike and did not affect grain numbers per spike, unlike in spring wheat. Our field study provides valuable insights into the relationships between winter wheat stomatal traits and grain yield that can be used in breeding.