Root system architecture profiling for aluminium tolerance in maize seedlings using an optimized high-throughput phenotyping

Aluminium (Al) toxicity is a potential constraint to maize productivity in acidic soils, primarily due to its inhibitory effect on root growth during its early establishment. In the present study, a hydroponic screening protocol was standardized using Modified Magnavaca-II solution at the seedling stage and applied to 250 tropical maize inbred lines. Five root traits-total root length (TRL), root surface area (RSA), root volume (RV), average root diameter (AD), and number of root tips (NRT)-were quantified using WinRHIZO. To assess differential tolerance, the Relative Root Tolerance Index (RRTI)-a ratio-based metric comparing root performance under stress versus control-was calculated along with percent reduction for all traits. Protocol optimization with seven elite inbreds exposed to graded AlCl₃ concentrations (0-1500 µM) identified 300 µM AlCl₃ at 11 days post-germination as optimal for differentiating genotypic responses. Under this optimized condition, the 250 inbreds showed highly significant genotypic variation and genotype × treatment interactions. Stress significantly reduced most root traits by 10-40%, while improving the average root diameter, indicating compensatory thickening. Substantial variability was observed for both RRTI and percent reduction indices, ranging from 3.83 to 533.88. Principal component analysis and composite indices identified IMR292, IMR592, IMR463, IMR621, IMR546, IMR534, IMR629 and IMR395 as tolerant due to high TRL, RSA and NRT under stress, while IMR388, IMR33, IMR58, IMR349 and IMR446 were highly susceptible. The tolerant inbreds offer promising genetic resources for breeding Al-tolerant maize, while the optimized hydroponic system provides a robust, scalable framework for future phenotyping and genetic dissection studies.