Can Phosphite Substitute for Phosphate in ptxD Rice? Evidence From Uptake and Agronomic Performance

Engineering crops to metabolize phosphite (Phi) as a phosphorus (P) source has been reported as a promising strategy to enhance nutrient use efficiency and enable innovative crop management. Here, we evaluated rice ( Oryza sativa L.) plants engineered with a codon‐optimized ptxD gene from Pseudomonas stutzeri for their capacity to utilize Phi under soil and hydroponic conditions. Two independent transgenic lines (L1 and L2), constitutively expressing ptxD, were cultivated in a Ferralsol with low available P and supplied with phosphate (Pi) or phosphite (Phi) at graded fertilization doses. Parallel hydroponic trials tested a wide range of P concentrations. To monitor uptake dynamics with high precision, we applied a fluorometric assay based on thermostable phosphite dehydrogenase (17X‐PTDH), enabling real‐time quantification of Pi and Phi depletion. Under Phi fertilization, transgenic lines displayed enhanced growth and phosphorus accumulation compared with wild‐type (WT) plants, confirming the functional expression of ptxD. However, agronomic performance with Phi was consistently lower than with Pi, especially at reduced doses. Uptake kinetics revealed slower Phi absorption even following P starvation, and biomass accumulation was restricted at low Phi concentrations. This work provides the first direct evidence, obtained through depletion assays, that Phi acquisition is intrinsically less efficient than Pi uptake. Together, these findings establish that while ptxD rice can metabolize Phi as a P source, its acquisition efficiency remains substantially inferior to Pi. Phi should therefore be regarded as a complementary fertilizer or as a selective tool for weed management in ptxD‐based systems, rather than a complete substitute for phosphate.