Variation of anti-oomycete activity in Pseudomonas spp.: phenotypic characterization and comparative genomics

Abstract

Pathogenic aquatic oomycetes Aphanomyces astaci and Saprolegnia parasitica represent a major threat to biodiversity and aquaculture production, but their interactions with host-associated microbes remain poorly understood. From a collection of bacterial isolates (n = 328) obtained from fish and crayfish hosts, we focused on Pseudomonas spp. (n = 66) and confirmed their previously reported strong inhibitory potential against A. astaci and S. parasitica. However, our results also revealed substantial inter- and intra-species variation in antagonism. To capture this variation, we selected eight isolates belonging to different P. fluorescens, P. putida, and P. syringae species groups and displaying contrasting levels of anti-oomycete activity for further phenotypic assays and comparative genomic analysis. Across these isolates, mycelial inhibition was markedly stronger against A. astaci than against S. parasitica, indicating species-specific differences in susceptibility. Comparative genomic analysis revealed substantial variation in biosynthetic gene cluster (BGC) repertoires among the analysed strains. Strongly inhibitory isolates carried candidate BGCs with similarity to characterised bioactive pathways, including pyoluteorin, rhizoxin, pyrrolnitrin, DAPG, and orfamide, alongside with multiple uncharacterised clusters that were either shared among inhibitory isolates or restricted to individual strains. All analysed genomes also contained clusters related to siderophore and HCN biosynthesis. However, in vitro assays showed that siderophore production was not clearly associated with inhibitory activity and that inhibition was mediated mainly by diffusible rather than volatile compounds. Altogether, our results suggest that Pseudomonas anti-oomycete activity is species- and strain-dependent and likely reflects different combinations of multiple, predominantly diffusible metabolites rather than a single conserved mechanism. In conclusion, this study provides a foundation for future work aimed at resolving mechanisms underlying microbial antagonism toward aquatic oomycete pathogens.