Diagnostic system for tebuthiuron soil ecotoxicity using morphophysiological indicators of Mucuna pruriens validated by Lactuca sativa.

This study developed an integrated diagnostic system for tebuthiuron-induced soil ecotoxicity based on morphophysiological indicators of Mucuna pruriens, using the germination index (GI) of Lactuca sativa as a sensitive ecotoxicological validation endpoint. The experiment was conducted under greenhouse conditions using a completely randomized design with 12 treatments and 360 individual pots (independent samples evaluated via destructive sampling), which were distributed across five evaluation periods at 14, 28, 42, 56, and 70 days after sowing. Morphophysiological variables, including plant height, root length, shoot and root dry mass, chlorophyll content, nodule number, and visual phytotoxicity, were quantified and integrated with multivariate and probabilistic modeling approaches. Given the multifactorial nature of the germination index, Principal Component Analysis (PCA) was applied to identify ecological and physiological gradients associated with plant vigor, stress, and symbiotic functioning. The PCA outputs were subsequently used as inputs for Probabilistic Neural Networks (PNNs), enabling the classification and prediction of bioindicator-based ecotoxicological levels using mathematically defined low, medium, and high GI classes. Model performance was internally assessed using training and validation datasets, confusion matrices, overall accuracy, sensitivity, specificity, and ROC curves. Because no independent external dataset was available, the predictive performance should be interpreted as evidence of internal consistency rather than definitive generalizability across different soils, climates, herbicide doses, or field conditions. Multivariate analyses revealed that ecotoxicological attenuation trajectories in tebuthiuron-contaminated soils are inherently nonlinear, being structured by coordinated shifts in morphophysiological traits rather than isolated responses of individual variables. The integrated PCA-PNN framework demonstrated that aboveground traits. Particularly plant height, chlorophyll content, and shoot dry mass, were more sensitive indicators of tebuthiuron-induced stress than root traits alone. Higher GI values were associated with PCA regions characterized by increased shoot biomass, greater plant height, reduced phytotoxicity, and improved physiological performance, whereas lower GI classes corresponded to suppressed growth and multidimensional stress signatures. The progressive convergence between plant vigor and GI across evaluation periods suggests a gradual mitigation of ecotoxicological stress signals on the indicator plants, indicating transitions from acute injury to physiological adaptation states. These findings confirm that M. pruriens functions as an effective bioindicator for diagnosing soil ecotoxicological status and monitoring tebuthiuron-induced impacts. However, as tebuthiuron residues were not chemically quantified, these responses should not be interpreted as direct evidence of herbicide degradation, dissipation, or removal. These findings confirm that M. pruriens functions as an effective bioindicator for diagnosing soil ecotoxicological status and monitoring tebuthiuron-induced impacts. However, as tebuthiuron residues were not chemically quantified, the observed improvements should be interpreted as evidence of physiological adaptation and/or ecological attenuation rather than definitive proof of herbicide degradation or removal. Overall, this approach provides a robust framework for early detection of soil contamination and supports its application in monitoring and guiding soil rehabilitation processes, with potential for future validation under field conditions.