Novel field-based approaches reveal wheat genotypic differences in nitrogen use efficiency and grain protein dynamics.

Achieving high yield and grain quality in wheat typically requires substantial nitrogen (N) fertiliser application. However, given economic and environmental constraints, it is critical to understand whether growers can reduce N inputs without compromising performance, and whether existing varieties differ in their ability to cope with lower N availability. Using a novel field-based experimental platform, we assessed the performance of fifteen registered wheat varieties under six N regimes and over two seasons with contrasting weather patterns. As expected, yields and grain protein contents both increased with N application, although protein content plateaued at a higher N threshold than yield. We noted higher genotypic differences in N use efficiency (NUE; defined as yield per unit of available N) under zero- N fertiliser applications, revealing intrinsic variation in low-N resilience. N-driven yield increase was more strongly associated with spike number rather than spike weight. Two varieties selected in Denmark where tight fertiliser regulations are in place were included for comparison and could achieve high yield with contrasting strategies; one with low and the other with high spike weight. In addition, using a novel stable isotope field-based method, we could show that under higher N levels, the post-anthesis N uptake was decreased and this trait is critical to achieving positive grain protein deviation (higher increase in grain protein content than expected given its yield). Our findings highlight the necessity of evaluating commercial and pre-breeding wheat germplasm under reduced N conditions to identify genotypes suited to sustainable, lower-input agricultural systems in a changing climate.