Myxobacterial Outer Membrane β-1,6-Glucanase Induced the Cell Death of Fusarium oxysporum by Destroying the Cell Wall Integrity

The β-1,6-glucan is the key linker between mannoproteins in the outermost part of the cell wall and β-1,3-glucan/chitin polysaccharide to maintain the rigid structure of the cell wall. The β-1,6-glucanase GluM, which was purified from the fermentation supernatant of sp. EGB, was able to inhibit the germination of Fusarium oxysporum f. sp. conidia at a minimum concentration of 2.0 U/mL (0.08 μg/mL). The survival rates of GluM-treated conidia and monohyphae were 10.4% and 30.7%, respectively, which were significantly lower than that of β-1,3-glucanase treatment (Zymolyase, 20.0 U/mL; equate to 1.0 mg/mL) (72.9% and 73.9%). In contrast to β-1,3-glucanase treatment, the high-osmolarity glycerol (HOG) pathway of F. oxysporum f. sp. cells was activated after GluM treatment, and the intracellular glycerol content was increased by 2.6-fold. Moreover, the accumulation of reactive oxygen species (ROS) in F. oxysporum f. sp. cells after GluM treatment induced apoptosis, but it was not associated with the increased intracellular glycerol content. Together, the results indicate that β-1,6-glucan is a promising target for the development of novel broad-spectrum antifungal agents. Phytopathogenic fungi are the most devastating plant pathogens in agriculture, causing enormous economic losses to global crop production. Biocontrol agents have been promoted as replacements to synthetic chemical pesticides for sustainable agriculture development. Cell wall-degrading enzymes (CWDEs), including chitinases and β-1,3-glucanases, have been considered as important armaments to damage the cell wall. Here, we found that F. oxysporum f. sp. is more sensitive to β-1,6-glucanase GluM treatment (0.08 μg/mL) than β-1,3-glucanase Zymolyase (1.0 mg/mL). The HOG pathway was activated in F. oxysporum f. sp. cells after GluM treatment, and the intracellular glycerol content was significantly increased. Moreover, the decomposition of F. oxysporum f. sp. cell wall by GluM induced the burst of intracellular ROS and apoptosis, which eventually leads to cell death. Therefore, we suggest that the β-1,6-glucan of the fungal cell wall may be a better antifungal target compared to the β-1,3-glucan.