Reduction of <i>Plasmodiophora brassicae</i> Infection on <i>Brassica rapa</i> Through Host-Induced Gene Silencing of Two Secreted Genes

Clubroot disease, caused by the biotrophic pathogen <i>Plasmodiophora brassicae</i>, is one of the most serious threats to cruciferous crops production worldwide. <i>P. brassicae</i> is known for rapid adaptive evolution to overcome plant resistance. The current prevention and control strategies are not effective against <i>P. brassicae.</i> Additionally, lack of genetic transformation has impeded the functional characteristic disclosure of virulence genes. In this study, we have identified two effectors, Pb48 and Pb52, that impact plant defense and are upregulated during the infection stage. To characterize the function of these virulence genes, we employed a transient method, host-induced gene silencing (HIGS). By instantaneously expressing a hairpin RNA interference construct with sequence homology to <i>P. brassicae Pb48</i> or <i>Pb52</i> in susceptible <i>Brassica rapa</i>, we successfully silenced the corresponding gene, resulting in reduced root gall size or enhanced host resistance to <i>P. brassicae.</i> Silencing <i>Pb48</i> led to a decrease in the numbers of zoosporangia within root hair and epidermal cells, and silencing either <i>Pb48</i> or <i>Pb52</i> led to downregulated expressions of cytokinin biosynthesis gene <i>IPT1</i> and auxin homeostasis <i>GH3.5</i>, which are associated with hormone regulation pathways involved in clubroot development. These findings validate HIGS as a practical tool for studying <i>P. brassicae</i> virulence genes. HIGS, by transiently expressing short interfering RNAs of <i>P. brassicae</i>, demonstrates its potential as an effective strategy against this pathogen. In the future, we can obtain durable disease resistance in susceptible host crops by developing a stable transformant.