Primers used in this study.

<div><p><i>Zymoseptoria tritici</i> is the causal agent of Septoria tritici blotch (STB), one of the most economically destructive wheat foliar diseases. In this study, we explore the physiological and molecular changes elicited in two wheat cultivars with divergent responses (Taichung 29 = susceptible, and Shafir = resistant) upon infection by <i>Z</i>. <i>tritici</i>. Our aim is to uncover novel insights into the intricate mechanisms that govern wheat defense against <i>Z</i>. <i>tritici</i> infection. Our quantitative histopathological study showed that H<sub>2</sub>O<sub>2</sub> accumulated in the resistant cultivar to a higher degree compared to the susceptible cultivar at the biotrophic and switching phase. Additionally, we combined qPCR with a targeted quantitative HPLC technique to evaluate the expression profiles of 13 defense-related genes and profile the polyphenolic compounds induced differentially in the STB susceptible and resistant cultivar. Our finding indicated that five out of 13 genes were strongly up-regulated in the resistant cultivar compared with that of the susceptible one at eight days post-inoculation (dpi), corresponding to the transition phase present in the infection process of <i>Z</i>. <i>tritici</i>. Finally, our targeted HPLC analysis demonstrated that the traced phenolic compounds were highly elevated in the susceptible cultivar infected by <i>Z</i>. <i>tritici</i> compared with that of the resistant cultivar. In conclusion, our comprehensive analysis unveils a robust defense response in the resistant wheat cultivar Shafir, characterized by heightened H<sub>2</sub>O<sub>2</sub> accumulation, significant up-regulation of key defense-related genes during the transition phase, and a distinct profile of polyphenolic compounds, shedding light on the intricate mechanisms contributing to its resistance against <i>Z</i>. <i>tritici</i>, thereby providing valuable insights for the development of more resilient wheat varieties.</p></div>