Antifungal Activity Of Pseudomonas Choloeaphtis HL5-4 Against Tomato Gray Mold And Its Preliminary Study

A promising biocontrol agent HL5-4, a strain with strong effect on Tomato gray mold, which selected from the rhizosphere of healthy tea plant.In vitro and in vivo experiments were used to determine its antifungal effect and colonization ability. The morphological and physiological characteristics and 16S rDNA sequence analysis were used to identify the strain.Through orthogonal test, optimize the conditions of cultivate and nutrition to find the fermentation condition of HL5-4.Through colorimetric method to determine POD、PPO and PAL. For the biocontrol of Tomato gray mold disease laid a foundation.1. Antifungal Activity of HL5-4 Against Tomato Gray Mold:HL5-4 inhibited mycelial growth and conidia germination of Botrytis cinerea in vitro experiment, the inhibition zone of the mycelial growth was 15mm, the inhibition rate of conidiaspores germination was 70.74%; HL5-4 performed good control potential against tomato gray mold, the control efficiencies were 72.36% and 59.85% respectively on fruits and leaf in vitro experiment, and 62.88% on seedlings in pot; When the HL5-4 concentration was 109cfu·mL-1, the control efficiency was 62.36%;the treatment time was the more the better, HL5-4 performed good control efficiency was 75.23%; the control efficiency of prevention was better than treatment.2. Identification of HL5-4:Based on the morphological, physiological characteristics and 16S rDNA sequence analysis,the strain HL5-4 was identified Pseudomonas chlororaphis.3. The colonization density of HL5-4:HL5-4 successfully colonized on fruits and leaf of tomato, the colonized density HL5-4 were 6.26×104 cfu·g-1 and 4.35 X 103cfu·g-1 respectively on fruits and leafs at 1 days after treatment, and the colonization density were maintained 2.47×104 cfu·g-1 and 2.90×103cfu·g-1 at 7 days after treatment.4. Effect of HL5-4 against tomato defendant enymes:Through the treatment with HL5-4, PPO, POD and PAL activity was significantly increased in the tomato leaves, the peak of PPO, POD and PAL activity were increased by 22.57%.30.23% and 20.99%.5. The characteristics of HL5-4:HL5-4 can inhibit a variety of plant pathogens, such as Verticillium dahliae, Gaeumannomyces graminis var. tritici, Drechslera Sofokiniana and so on.On the plate culture, strain HL5-4 can excret proteinase, celluase and siderophore, but not chitinase and amylase.6. The optimization of fermentation conditions of HL5-4:Through orthogonal test, optimize the conditions of cultivate and nutrition of HL5-4. The fermentation condition of HL5-4 was in the temperature of 28℃ cultivate 52h, rotate speed of 160 rpm, ventilation of 60%, pH of 7, its carbon source was Glycerol 1%, nitrogen source was peptone 0.1% and inorganic salt was K2HPO4 0.7%、KH2PO4 0.2%. MgSO4·7H2O 0.01%.