Multi-Omics and Phenotypic Analysis Reveal Paenibacillus polymyxa JX-1 as a Broad-Spectrum Biocontrol Agent Against Clubroot Disease

<i>Paenibacillus polymyxa</i> is a promising biocontrol agent based on many applications in agriculture. In this study, characterization of a novel strain JX-1 was performed by various comparative phenotypic assays including cell wall-degrading enzyme activity assay, antagonism in vitro and in planta and multi-omics analysis consisting of whole-genome sequencing and metabolomic analysis. The results showed that JX-1 produced protease, cellulase, and pectinase, with protease activity exceeding other <i>P. polymyxa</i> strains. The supernatant and VOCs produced by JX-1 both contain components that effectively antagonize both bacteria and fungi. JX-1 inhibited <i>Fusarium verticillioides</i>, <i>Sclerotinia sclerotiorum</i>, <i>Botrytis cinerea</i> and <i>Sclerotium rolfsii</i>. Furthermore, it significantly reduced the severity of clubroot in <i>Arabidopsis thaliana</i> and <i>Brassica rapa</i> in planta, resulting in an improvement in the total shoot fresh weight. The closely related <i>P. polymyxa</i> strain JX-2 (ANI = 99.96% compared to strain JX-1) was used for comparative phenotyping and multi-omics analysis that implicated extracellular proteases and some small peptides could be essential for clubroot biocontrol. Genomic analysis of strain JX-1 confirmed that it is indeed <i>P. polymyxa</i> (ANI > 95% compared to reference strains). Moreover, JX-1 harbors predicted carbohydrate-active enzymes (CAZymes), 16 extracellular protease genes, and 17 gene clusters with biosynthetic potential (NRPS/PKS hybrids). Metabolomic profiling of the culture supernatant further identified differential accumulation of hydrophobic-amino-acid-containing small peptides, providing a metabolic basis for the observed antagonism and offering leads for future mechanistic studies. These findings establish JX-1 as a broad-spectrum biocontrol agent against clubroot disease.