Balthazar Carole, St-Onge Renée, Léger Geneviève, Lamarre Simon G, Joly David L, Filion Martin
Department of Biology, Université de Moncton, Moncton, NB, Canada.
Agriculture and Agri-Food Canada, Saint-Jean-sur-Richelieu Research and Development Center, Saint-Jean-sur-Richelieu, QC, Canada.
Front Microbiol. 2022 Aug 9;13:945498. doi: 10.3389/fmicb.2022.945498. eCollection 2022.
Pf-5 is an effective biocontrol agent that protects many crops against pathogens, including the fungal pathogen causing gray mold disease in crops. Previous studies have demonstrated the important role of antibiotics pyoluteorin (PLT) and 2,4-diacetylphloroglucinol (DAPG) in Pf-5-mediated biocontrol. To assess the potential involvement of PLT and DAPG in the biocontrol exerted by Pf-5 against in the phyllosphere of , two knockout Pf-5 mutants were generated by in-frame deletion of genes or , required for the synthesis of PLT or DAPG respectively, using a two-step allelic exchange method. Additionally, two complemented mutants were constructed by introducing a multicopy plasmid carrying the deleted gene into each deletion mutant. confrontation assays revealed that deletion mutant ∆ inhibited growth significantly less than wild-type Pf-5, supporting antifungal activity of PLT. However, deletion mutant ∆ inhibited mycelial growth significantly more than the wild-type, hypothetically due to a co-regulation of PLT and DAPG biosynthesis pathways. Both complemented mutants recovered inhibition levels similar to that of the wild-type. In subsequent growth chamber inoculation trials, characterization of gray mold disease symptoms on infected cannabis plants revealed that both ∆ and ∆ significantly lost a part of their biocontrol capabilities, achieving only 10 and 19% disease reduction respectively, compared to 40% achieved by inoculation with the wild-type. Finally, both complemented mutants recovered biocontrol capabilities similar to that of the wild-type. These results indicate that intact biosynthesis pathways for production of PLT and DAPG are required for the optimal antagonistic activity of Pf-5 against in the cannabis phyllosphere.
Pf-5是一种有效的生物防治剂，可保护多种作物免受病原体侵害，包括导致作物灰霉病的真菌病原体。先前的研究已经证明了抗生素绿脓菌素（PLT）和2,4-二乙酰基间苯三酚（DAPG）在Pf-5介导的生物防治中的重要作用。为了评估PLT和DAPG在Pf-5对大麻叶际中[具体病原体未提及]的生物防治中可能发挥的作用，使用两步等位基因交换方法，通过分别缺失PLT或DAPG合成所需的基因[具体基因未提及]或[具体基因未提及]，构建了两个Pf-5基因敲除突变体。此外，通过将携带缺失基因的多拷贝质粒导入每个缺失突变体，构建了两个互补突变体。对峙试验表明，缺失突变体∆对[具体病原体未提及]生长的抑制作用明显小于野生型Pf-5，这支持了PLT的抗真菌活性。然而，缺失突变体∆对菌丝体生长的抑制作用明显大于野生型，推测这是由于PLT和DAPG生物合成途径的共同调节。两个互补突变体恢复了与野生型相似的抑制水平。在随后的生长室接种试验中，对感染大麻植株上灰霉病症状的观察表明，∆和∆都显著丧失了部分生物防治能力，与接种野生型所实现的40%的病害减轻相比，它们分别仅实现了10%和19%的病害减轻。最后，两个互补突变体恢复了与野生型相似的生物防治能力。这些结果表明，完整的PLT和DAPG生物合成途径是Pf-5在大麻叶际中对[具体病原体未提及]发挥最佳拮抗活性所必需的。
