Infection mechanisms and putative effector repertoire of the mosquito pathogenic oomycete Pythium guiyangense uncovered by genomic analysis

Abstract

Pythium guiyangense, an oomycete from a genus of mostly plant pathogens, is an effective biological control agent that has wide potential to manage diverse mosquitoes. However, its mosquito-killing mechanisms are almost unknown. In this study, we observed that P. guiyangense could utilize cuticle penetration and ingestion of mycelia into the digestive system to infect mosquito larvae. To explore pathogenic mechanisms, a high-quality genome sequence with 239 contigs and an N50 contig length of 1,009 kb was generated. The genome assembly is approximately 110 Mb, which is almost twice the size of other sequenced Pythium genomes. Further genome analysis suggests that P. guiyangense may arise from a hybridization of two related but distinct parental species. Phylogenetic analysis demonstrated that P. guiyangense likely evolved from common ancestors shared with plant pathogens. Comparative genome analysis coupled with transcriptome sequencing data suggested that P. guiyangense may employ multiple virulence mechanisms to infect mosquitoes, including secreted proteases and kazal-type protease inhibitors. It also shares intracellular Crinkler (CRN) effectors used by plant pathogenic oomycetes to facilitate the colonization of plant hosts. Our experimental evidence demonstrates that CRN effectors of P. guiyangense can be toxic to insect cells. The infection mechanisms and putative virulence effectors of P. guiyangense uncovered by this study provide the basis to develop improved mosquito control strategies. These data also provide useful knowledge on host adaptation and evolution of the entomopathogenic lifestyle within the oomycete lineage. A deeper understanding of the biology of P. guiyangense effectors might also be useful for management of other important agricultural pests.

Author summary

Utilization of biocontrol agents has emerged as a promising mosquito control strategy, and Pythium guiyangense has wide potential to manage diverse mosquitoes with high efficiency. However, the molecular mechanisms underlying pathological processes remain almost unknown. We observed that P. guiyangense invades mosquito larvae through cuticle penetration and through ingestion of mycelia via the digestive system, jointly accelerating mosquito larvae mortality. We also present a high-quality genome assembly of P. guiyangense that contains two distinct genome complements, which likely resulted from a hybridization of two parental species. Our analyses revealed expansions of kinases, proteases, kazal-type protease inhibitors, and elicitins that may be important for adaptation of P. guiyangense to a mosquito-pathogenic lifestyle. Moreover, our experimental evidence demonstrated that some Crinkler effectors of P. guiyangense can be toxic to insect cells. Our findings suggest new insights into oomycete evolution and host adaptation by animal pathogenic oomycetes. Our new genome resource will enable better understanding of infection mechanisms, with the potential to improve the biological control of mosquitoes and other agriculturally important pests.