Screening Of Nematicide Fungi And Identification, Fermentation Optimization And Analyzing Of Enzyme System Of Strain Df09002

A total of 29 soil samples were collected from Wuzhishan mountain, Alishan mountain, Leiming Dingan and all the 312 fungi were isolated from these samples by culture plate dilute method. A further study was needed for screening anti-nematode activity of all the isolates. Among them, strain DF09002 exhibited the strongest nematicidal activity at lethal rate. The results were as follows.we tested the nematicidal activity of the 312 isolates using 2nd instar larvae of Meloidogyne incognita as target nematode, and found that a total of 22 fungi laboured the anti-nematode activity (adjusted mortality rate above 70%), the percentage of active strains reached 5.6%. The strain DF09002 was isolated from soil sample collected at pepper rhizosphere, which showed the strongest anti-nematode activity, with adjusted mortality rate as high as 95.3%. The fermentation broth of the strain DF09002 also had a significant effect on nematode, all the tested nematodes were killed within 24h. When the fermentation broth was diluted 5-fold, the adjusted mortality rate still reached 68.5%.By studying the morphology, cultural characteristics, physiological and biochemical properties, rDNA-ITS and phylogenetic tree, DF09002 was identified as one of the genus of Aspergillus niger aggregate.The optimal liquid fermentation conditions and medium of DF09002 were found out. The optimum culture medium of DF09002 contained 20% potato juice, soy powder 1%, soluble starchl.5%, glucose 2%, yeast extract 0.75%.75mL medium (initial pH6.5), in 250mL Erlenmeyer flask with6% inoculum (36h) under orbital shaking at 200 r/min for 6d at 28℃.The types and characteristics of enzyme produced by the strain DF09002 were studied. The results showed that the strain DF09002 could produce chitinase, protease, but not cellulose. By using Folin-phenol reagent method, the protease activity of strain DF09002 was estimated as 41.63 U/mL, and by using DNS (3,5-dinitrosalicylic acid) method to test reducing sugar, the chitinase activity of strains DF09002 was measured as 0.67 U/mL. It is demonstrated that optimum temperature of the chitinase reaction was 40℃, and optimum temperature of the protease reaction was 55℃, Furthermore, their optimum pH of the reaction were ascertained, respectively 6.8 and 9.0.