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Agave ( Agave tequilana var. azul) is considered a crop with low genetic diversity because it has been propagated vegetatively for centuries for commercial purposes, and consequently, it could be equally susceptible to pests and diseases. However, the present study employs plant material derived from field-grown plants exhibiting phenotypic variability in susceptibility to agave wilt. The offshoots from rhizomes of these plants were reproduced in vitro and classified as potentially resistant or susceptible. Amplified fragment length polymorphism analysis confirmed wide genetic differences among individuals, but these differences were not correlated with the observed phenotypic variability in resistance. Propagated plantlets were inoculated with Fusarium solani in two time-lapse confrontations for 72 h and 30 days. The early biochemical response showed statistically superior levels in the accumulation of shikimic acid, phenolic compounds, and chitinase activity in potentially resistant plantlets. There was an inverse correlation of these early biochemical responses and salicylic acid and the incidence of diseased root cells in isogenic plantlets in the 30-day confrontation with F. solani , suggesting that these activities and accumulation of molecules were primordial in the defence against this pathogen., (© The Author(s) 2022. Published by Oxford University Press on behalf of the Annals of Botany Company.)