Transcriptional Profiling of Diffusible Lipopeptides and Fungal Virulence Genes During <i>Bacillus amyloliquefaciens</i> EZ1509-Mediated Suppression of <i>Sclerotinia sclerotiorum</i>

Abstract

<i>Sclerotinia sclerotiorum</i> is a devastating necrotrophic pathogen that infects multiple crops, and its control is an unremitting challenge. In this work, we attempted to gain insights into the pivotal role of lipopeptides (LPs) in the antifungal activity of <i>Bacillus amyloliquefaciens</i> EZ1509. In a comparative study involving five <i>Bacillus</i> strains, <i>B. amyloliquefaciens</i> EZ1509 harboring four LPs biosynthetic genes (viz. surfactin, iturin, fengycin, and bacilysin) exhibited promising antifungal activity against <i>S. sclerotiorum</i> in a dual-culture assay. Our data demonstrated a remarkable upsurge in LPs biosynthetic gene expression through quantitative reverse transcription PCR during in vitro interaction assay with <i>S. sclerotiorum</i>. Maximum upregulation in LPs biosynthetic genes was observed on the second and third days of in vitro interaction, with iturin and fengycin being the highly expressed genes. Subsequently, Matrix-assisted laser desorption/ionization-time of flight-mass spectrometry analysis confirmed the presence of LPs in the inhibition zone. Scanning electron microscope analysis showed disintegration, shrinkage, plasmolysis, and breakdown of fungal hyphae. During in planta evaluation, <i>S. sclerotiorum</i> previously challenged with EZ1509 showed significant suppression in pathogenicity on detached leaves of tobacco and rapeseed. The oxalic acid synthesis was also significantly reduced in <i>S. sclerotiorum</i> previously confronted with antagonistic bacterium. The expression of major virulence genes of <i>S. sclerotiorum</i>, including endopolygalacturonase-3, oxalic acid hydrolase, and endopolygalacturonase-6, was significantly downregulated during in vitro confrontation with EZ1509.