Enhanced Phytopathogen Biofilm Control in the Soybean Phyllosphere by the Phoresy of Bacteriophages Hitchhiking on Biocontrol Bacteria

Phage-based biocontrol has shown notable advantages in protecting plants against pathogenic bacteria in agricultural settings compared to chemical-based bactericides. However, the efficiency and scope of phage biocontrol of pathogenic bacteria are limited by the intrinsic properties of phages. Here, we investigated pathogen biofilm eradication in the phyllosphere using the phoresy system of hitchhiking phages onto carrier biocontrol bacteria. The phoresy system efficiently removed the pathogen biofilm in the soybean phyllosphere, reducing the total biomass by 58% and phytopathogens by 82% compared to the untreated control. Biofilm eradication tests demonstrated a significant combined beneficial effect (Bliss independence model, CI < 1) as phages improved carrier bacteria colonization by 1.2-fold and carrier bacteria facilitated phage infection by 1.4-fold. Transcriptomic analysis showed that phoresy significantly enhanced motility (e.g., <i>fliC</i> and <i>pilD</i> genes) and energy metabolism (e.g., <i>pgm</i> and <i>pgk</i> genes) of carrier bacteria and suppressed the defense system (e.g., <i>MSH3</i> and <i>FLS2</i> genes) and energy metabolism (e.g., <i>petB</i> and <i>petC</i> genes) of pathogens. Metabolomics analysis revealed that the phoresy system stimulated the secretion of beneficial metabolites (e.g., flavonoid and tropane alkaloid) that could enhance stress response and phyllosphere protection in soybeans. Overall, the phoresy of phages hitchhiking on biocontrol bacteria offers a novel and effective strategy for phyllosphere microbiome manipulation and bacterial disease control.