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Culture method and application of isaria microsclerotia

A culture method and micro sclerotia technology, applied in the field of biological preparations, can solve the problems of lack of micro sclerotium of Corynebacterium, poor stress resistance, short shelf life, etc., and achieve good insecticidal activity, low production cost, and shelf life. long effect

Active Publication Date: 2016-06-22
CHONGQING UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The purpose of the present invention is to address the above problems, to provide a culture method and application thereof capable of large-scale production of I. The structure provides a biocontrol agent for pest control, provides high-quality mother drug or raw powder for the creation of insecticidal fungal agents, and solves the problem that there is currently no effective method for cultivating I. clavulas microsclerotia and traditional spore agents Common technical problems of poor stress resistance, short shelf life, and inconvenient storage and transportation

Method used

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  • Culture method and application of isaria microsclerotia
  • Culture method and application of isaria microsclerotia
  • Culture method and application of isaria microsclerotia

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0024] Example 1 Optimization of the formulation and induction process of the microsclerotium induction culture solution

[0025] 1. Preparation of Inoculum of Corynespora:

[0026] Inoculate the conidia of CQIF101 strain CQIF101 (from Chongqing Insecticidal Fungi Pesticide Engineering Technology Research Center) on a PDA plate, and cultivate under light at 25℃ (16 hours light: 8 hours dark) 12 Days, use 0.1~0.5% Tween-80 sterilized liquid to wash off the mature conidia on the plate, adjust the concentration of conidia, and prepare 1.0×10 8 The spore suspension of spores / ml is inoculated into 1 / 4SDA liquid medium, and cultured at a temperature of 25°C and a shaker speed of 250rpm for 24-36h to obtain an inoculum for microsclerotia induction culture.

[0027] 2. Screening of induction medium formula

[0028] 1) Concentration of inorganic nutrients:

[0029] To add KH 2 PO 4 :3.0~5.0g / L, CaCl 2 ·2H 2 O: 0.6~1.0g / L, MgSO 4 ·7H 2 O: 0.4~0.8g / L, CoCl 2 ·6H 2 O: 34~40mg / L, MnSO 4 ·H 2 O: 14~...

Embodiment 2

[0044] Example 2 Observation of the morphology and structure of microsclerotia

[0045] In the first implementation, when the Corynespora strain was cultured according to the liquid induction method, the microsclerotia production status in the liquid medium was observed microscopically: after 3 days of culture, a smaller microsclerotia structure began to be produced, with smooth edges and dense center. Reddish brown. When cultured for 3-4 days, the color of the fermentation broth became darker, the number of micro sclerotia continued to increase, and a large number of hyphae began to sprout around a small amount of micro sclerotia. After 5-6 days of cultivation, the number of micro sclerotia basically no longer increases, and a large number of slender hyphae sprouted around the micro sclerotia. After 7 days, the number of micro sclerotia does not increase significantly, and some sclerotia form elongated hyphae (such as figure 1 Shown).

Embodiment 3

[0046] Example Sporulation analysis of three corynespora microsclerotia

[0047] The Corynespora strains were cultured according to the liquid culture induction method in Example 1. The culture medium was the samples 2, 5, 8, and 11 in Example 1. Vitamin K 30.0010 g / L was added to the culture medium to obtain culture medium I , II, III, IV.

[0048] After 6 days, the fermented culture solution was added with 5% diatomaceous earth, dried at 35°C for 24-48 hours, and then pulverized. Afterwards, the mixture was treated with a 60-mesh standard sieve to separate the micro sclerotia from diatomaceous earth, and the micro sclerotia were sealed for storage. 20 mg of the harvested micro sclerotia were uniformly inoculated on a water agar plate and cultured at 25°C. After 14 days, the mature conidia on the plate were washed with 0.5% Tween-80 sterilized water and counted on a hemocytometer. It was found that per mg of medium I combined cultured microsclerotia can produce 8.3×10 5 Spores an...

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Abstract

The invention discloses a culture method and application of isaria microsclerotia. A fermentation method comprises the steps of (1) preparing an isaria spore inoculant; (2) carrying out induction culture on microsclerotia: adding the isaria spore inoculant prepared in the step 1 into an induction culture solution, and carrying out shake cultivation for 6-8 days; (3) filtering fermentation liquor, removing supernatant to obtain microsclerotia precipitate, drying and storing. The invention also discloses the application of the isaria microsclerotia in control of whitefly pests and a biocontrol agent used for controlling the whitefly pests. The isaria microsclerotia produced by the invention is capable of adapting external bad environments such as high temperature, drought, strong ultraviolet and the like, and has the characteristics of stress tolerance, storage stability, long shelf life and continuous pest control; the isaria microsclerotia is short in fermentation period, low in production cost, high in technology controllability and easy in raw material obtaining. The isaria microsclerotia can be used for preparing active ingredients of a novel insecticidal fungus preparation.

Description

Technical field [0001] The invention belongs to the field of biological preparations, and particularly relates to a method for culturing microsclerotia of Corynespora and its application in preventing and controlling pests. Background technique [0002] The pests of whitefly piercing and sucking mouthparts often cluster in the leaves, tender stems, flower buds, apical buds and other parts, piercing and sucking the juice, causing the leaves to shrink, curl, and deform. In severe cases, they can cause the branches and leaves to wither or even the whole plant death, which is harmful to A variety of crops, fruits, vegetables, etc. At present, the prevention and control of piercing and sucking mouthparts mainly rely on chemical prevention and control. The large-scale application of chemical pesticides leads to the development of insect resistance, which pollutes the ecological environment, affects the safety of vegetables and agricultural products, and threatens people's health. It al...

Claims

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Application Information

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IPC IPC(8): C12N1/14A01N63/04A01P7/04C12R1/645
CPCA01N63/30C12N1/14
Inventor 宋章永王中康殷幼平
Owner CHONGQING UNIV
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