Abstract: A parasitic fungus with nematocidal activity, Clpse01, was found while pine wood nematodes (Bursaphelenchus xylophilus) were feeding on Botrytis cinerea media. Clpse01 morphologies and parasite evidences were seen under the microscope. Multiple genes, including ITS, large subunit ribosomal DNA, β-tublin and elongation factor, were used for further limiting by the phylogenic tree. Clpse01's nematocidal effectiveness against pine wood nematodes, as well as its effects on egg hatching and reproduction, were investigated. The Clpse01 fermentation fluid was analyzed for extracellular protein concentrations, β-glucosidase activity, and chitinase activity. Clpse01 was identified as Clonostachys pseudochloleuca using a combination of morphological and molecular phylogenetic approaches. C. pseudochroleuca Clpse01 was discovered entangling and infiltrating B. cinerea hyphae, acquiring nutrients from them while limiting B. cinerea normal growth. Inoculating 2000 pine wood nematodes into media with full growth of isolate Clpse01 resulted in only 6±4 live ones after 8 days, while nematodes isolated from the control medium of B. cinerea reached 33066±6882. The adjusted mortality rate of pine wood nematode treated with C. pseudochroleuca Clpse01 fermentation broth for 48 h was 88.0%, while that treated with 1╳10⁸ conidiospore suspensions per milliliter for 24 h was 88.33%. In double diluent fermentation broth for 24 hours, egg hatching was completely inhibited. After 6 hours of fermentation, the Clpse01 liquid contained 1782.917 μg/mL of total extracellular protein, 9.99 U/mL of β-glucosidase, and 0.8792 U/mL of chitinase. Protein concentrations increased with time. In general, C. pseudochroleuca Clpse01 functions by parasitizing B. cinerea, the feeding fungus, and influencing B. xylophilus reproduction, survival, and egg hatching.

Key words:

pine wood nematode, Botrytis cinerea, Clonostachys pseudochroleuca; parasitic fungus, biocontrol fungus