Prediction and Characterization of RXLR Effectors in <i>Pythium</i> Species

Abstract

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 those from genera <i>Phytophthora</i>, <i>Hyaloperonospora</i>, <i>Albugo</i>, and <i>Saprolegnia</i>. However, in <i>Pythium</i> species, no RXLR effector has yet been characterized, and the origin and evolution of such virulent effectors are still unknown. Here, we developed a modified regular expression method for de novo identification of RXLRs and characterized 359 putative RXLR effectors in nine <i>Pythium</i> species. Phylogenetic analysis revealed that all oomycetous RXLRs formed a single superfamily, suggesting that they might have a common ancestor. RXLR effectors from <i>Pythium</i> and <i>Phytophthora</i> species exhibited similar sequence features, protein structures, and genome locations. In particular, there were significantly more RXLR proteins in the mosquito biological control agent <i>P. guiyangense</i> than in the other eight <i>Pythium</i> species, and <i>P. guiyangense</i> 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 <i>P. ultimum</i> detected transcripts of the majority of the predicted <i>RXLR</i> genes, with some RXLR effectors induced in infection stages and one RXLR showing necrosis-inducing activity. Furthermore, all predicted <i>RXLR</i> genes were cloned from two biocontrol agents, <i>P. oligandrum</i> and <i>P. periplocum</i>, and three of the <i>RXLR</i> genes were found to induce a defense response in <i>Nicotiana benthamiana</i>. Taken together, our findings represent the first evidence of RXLR effectors in <i>Pythium</i> species, providing valuable information on their evolutionary patterns and the mechanisms of their interactions with diverse hosts.