| Summary: | <i>Leptosphaeria biglobosa</i> is a less virulent <i>Leptosphaeria</i> spp. that causes blackleg disease in canola. Previous studies from our lab have shown that inoculation with the less virulent <i>L. biglobosa</i> can boost the resistance of canola plants against the highly virulent <i>L. maculans</i>. The objective of this study was to confirm the effectiveness of <i>L. biglobosa</i> as a biocontrol agent against <i>L. maculans</i> utilizing morphology, fluorescence microscopy, gene quantification, and transcriptomic analysis. The in planta development of two <i>Leptosphaeria</i> species inoculated at different time points was assessed using fluorescent protein-tagged isolates which are GFP-tagged <i>L. maculans</i> and DsRed-tagged <i>L. biglobosa</i>. The growth inhibition of <i>L. maculans</i> by pre-and co-inoculated <i>L. biglobosa</i> was supported by no lesion development on cotyledons and no or weak fluorescence protein-tagged mycelia under the confocal microscope. The host defense-related genes, <i>WRKY33</i>, <i>PR1</i>, <i>APX6</i>, and <i>CHI,</i> were upregulated in <i>L. biglobosa</i> inoculated Westar cotyledons compared to <i>L. maculans</i> inoculated cotyledons. The quantification of each pathogen through qPCR assay and gene expressions analysis on host defense-related genes by RT-qPCR confirmed the potential of <i>L. biglobosa</i> “brassicae’ in the management of the blackleg disease pathogen, <i>L. maculans</i> ‘brassicae’, in canola.
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