Prediction and Characterization of RXLR Effectors in Pythium Species.

RXLR effectors, a class of secreted proteins that are transferred into host cells to manipulate host immunity, have been reported to widely exist in oomycetes, including Phytophthora, Hyaloperonospora, Albugo and Saprolegnia. However, in Pythium species, no RXLR effector has been characterized yet, and the origin and evolution of such virulent effectors are still unknown. Here, we developed a modified regex method for de novo identification of RXLRs and characterized 359 putative RXLR effectors in nine Pythium species. Phylogenetic analysis revealed that all oomycetous RXLRs formed a single superfamily, suggesting that they might have a common ancestor. RXLR effectors from Pythium and Phytophthora species exhibited similar sequence features, protein structures and genome locations. In particular, there were significantly more RXLR proteins in the mosquito biological control agent P. guiyangense than in the other eight Pythium species, and P. guiyangense RXLRs might be the result of gene duplication and genome rearrangement events, as indicated by synteny analysis. Expression pattern analysis of RXLR-encoding genes in the plant pathogen P. ultimum detected transcripts of the majority of the predicted RXLRs, with some RXLR effectors induced in infection stages and one RXLR showing necrosis-inducing activity. Furthermore, all predicted RXLRs were cloned from two biocontrol agents, P. oligandrum and P. periplocum, and three of the RXLRs were found to induce a defence response in Nicotiana benthamiana. Taken together, our findings represent the first evidence of RXLR effectors in Pythium species, providing valuable information on their evolutionary patterns and the mechanisms of their interactions with diverse hosts.