Root assays to study pattern-triggered immunity in plant-nematode interactions

Abstract

Plants employ extracellular immune receptors to perceive conserved pathogen-associated molecular patterns (PAMPs), triggering the first layer of defense known as pattern-triggered immunity (PTI). The understanding of PTI is mainly based on studies focusing on leaves. Plants are vulnerable to attack by various root pathogens including plant-parasitic nematodes. Evidence is accumulating that phytonematodes utilize their secreted effectors to suppress PTI to enable infection. PTI assays used for characterizing nematode effectors are often conducted in a non-host plant or tissue, such as leaves, because of lacking of root assays. Thus, there is a need for PTI assays in roots of host plants. Here, we tested two bacterial PAMPs (flg22 and flgII-28) and two nonpathogenic bacteria (Pseudomonas fluorescens and P. syringae strain DC3000 ΔhrcQ-U) for their ability to induce PTI responses, including the induction of defense gene expression and callose deposition, in roots of tomato and potato. We found that flg22 and the two nonpathogenic bacteria are potent in inducing defense gene expression and callose deposition in tested roots, demonstrating for the first time induction of PTI in roots of solanaceous plants. Effectors GrCEP12 and Hs10A06 were previously indicated to be involved in PTI suppression. Consistently, upon elicitor treatment, roots of transgenic plants overexpressing GrCEP12 and Hs10A06, respectively, showed a reduced level of defense gene expression or no induction of callose deposition compared to control roots. Taken together, our established root PTI assays represent a valuable tool that will facilitate the study of phytonematodes and potentially other root pathogens in their manipulation of plant immunity.