Climate -related combined abiotic stress enhances dhurrin accumulation and drives metabolic reprogramming in Sorghum bicolor compromising forage safety

Sorghum (Sorghum bicolor (L.) Moench) is a forage crop in agricultural systems resilient to environmental stresses typical of hot periods such as water deficit, increased salinity and elevated temperatures. However, these environmental stress can increase the content of dhurrin, a cyanogenic glycoside that is potentially toxic to livestock. While single stresses are studied, integrated responses to combinations that include physiology, metabolism, and gene expression remain underexplored. Sorghum plants underwent single (water deficit at 40% field capacity, 200 mM NaCl salinity, 40°C heat for 4h) and combined stresses. We assessed photosynthetic pigments, stress biomarkers (proline, H2O2, antioxidant capacity), hormones (ABA, JA), lipid profiles/indices, phenolics/flavonoids, dhurrin (HPLC-MS/MS), and gene expression (qRT-PCR for oxidative, biosynthetic, and hormonal pathways), with multivariate analyses. Results showed that combined stresses reduced chlorophyll with carotenoid shifts; elevated proline, H2O2, ABA/JA, and dhurrin (doubling in combined stress). Lipid saturation lowered nutritional indices through phenolic/flavonoid reprofiling and gene modulation (e.g., UGT85B1 for dhurrin). Our results revealed non-additive stress synergies, trading resilience for quality/safety. Understanding sorghum stress responses might inform breeding and management strategies to mitigate dhurrin risks under climate change.