CN101248799B - Verticillium lecanii pesticides and uses thereof - Google Patents

Abstract

The invention relates to a verticillium lecanii pesticide as well as the application thereof, and belongs to the plant protection and pesticide technology field. The bacterial strain provided by the invention is verticillium lecanii K1207 with CGMCC No.2326. The invention produces the bacterial strain via liquid and solid two-phase fermentation, and is characterized in that a wheat bran and bran coat culture medium is adopted as the solid culture medium. The cultured product can be added with and mixed with a wetting agent, a bulking agent and a stabilizing agent to obtain wettable powder after being parched and being crumbled. The inventive verticillium lecanii bacterial strain has high virulence, thereby having good control effects on pests such as green peach aphids, urolcueon formosanums, acyrthosiphon dirhodums, cabbage aphids, cabbage worms, diamondback moths, and greenhouse whiteflies. The materials in fermentation process are cheap and have wide sources, which is suitable for industrialized production without environmental pollution.

Description

A kind of verticillium wax scale insecticide and its application

Technical field:

The invention relates to the technical field of plant protection and pesticides, in particular to an insecticide for Verticillium lecanii and its application.

Background technique:

Verticillium lecanii (zimm) viegas is an entomopathogenic fungus of the class Hyphospora. The fungus was first discovered in the world by Neitner in 1861 on the Ceylon coffee wax scale (Lecanii coffeee). Regarding the applied basic research of this fungus, Dr. Hall RA (1976-1985) of the British Greenhouse Crop Research Institute carried out earlier, and presided over the research on the commercialization of this fungal preparation in the early 1980s. Since the commercialization of Vertalec ^R , the first V.lecanii aphidicide product developed by Hall in 1981, there have been more than 8 kinds of commodities in many countries with Vertalec, Mycotal, Thriptal, Vertisol, AgroBiocontrol Verticillium 50, Verticon, Verticillin and Mycolabs Verticillium lecanii are produced and sold under the names of Verticillin and Mycolabs Verticillium lecanii, and are used for the control of pests such as aphids, whiteflies and thrips in greenhouses and other places. Due to the slow effect of this fungal preparation product, it has certain requirements for the use environment; in the development process (especially in the application process of aphid control), it has been affected by the competitive pressure of special chemical pesticides such as pirimicarb, resulting in market share has declined. In recent years, due to the increasingly prominent problem of residual pollution and impact of chemical pesticides, under the influence and promotion of the global environmental protection wave, the promotion and application of the fungal preparation products have developed rapidly.

Invention content:

The object of the present invention is to provide a kind of Verticillium lecanii insecticide and application thereof.

Verticillium lecanii (Verticillium lecanii) K1207 provided by the present invention, preservation unit: General Microbiology Center of China Microbiological Culture Collection Management Committee; address: Datun Road, Chaoyang District, Beijing, Institute of Microbiology, Chinese Academy of Sciences; preservation date: 2008 January 07; the number CGMCCNo.2326 in the deposit register.

Verticillium lecanii K1207 of the present invention is isolated from the sick aphid body in Kunming, Yunnan, and the bacterial strain has the following characteristics: the 10d colony diameter on PDA and CzA medium is 24-27mm, the thickness is 2-3mm, the colony is round, unfolded, White from the front, aerial hyphae are erect and fluffy in the early stage, densely flocculent in the later stage, and the edges are sparse. The colony has inconspicuous radial folds from the middle to the surroundings. In the later stage of cultivation, there are obvious concave rings in the middle and edge of the colony, and the back of the colony is milky yellow. , no soluble pigments. Aerial hyphae are colorless, with septal branches, width 1.2-2.5 μm, conidiophores swollen at the base, tapered upwards and conical, solitary, opposite or whorled in aerial mycelia, aerial mycelia There can be up to 4 to 6 phialides at the top, and the conidiophores are rarely branched. Conidia grow from the top of the stalk and coalesce into a spherical shape. The diameter of the spore ball is 5-12 μm. When the ball meets water, it disintegrates and releases spores (10-25). And symmetrical, the spore size is 2.5～8.0×0.75～2.8μm. Compared with the existing known strains, the bacterial strain A1207 of the present invention has better temperature adaptability, the temperature range is 15-33 DEG C, and the conidia germination rate can reach 91.78-100%.

The present invention is achieved in this way: test tube culture of Verticillium lecanii (Verticillium lecanii) K1207 bacterial strain, solid and liquid two-phase fermentation, is prepared into wettable powder, and it is used for preventing and treating various aphids (Peach aphid, red aphid, wheat aphid) Aphids without nets, cabbage aphids, etc.), cabbage caterpillars, diamondback moths, whitefly in greenhouses.

The culture method of verticillium lecanii (Verticillium lecanii) K1207 bacterial strain of the present invention:

1. Test tube culture

The formula of test tube slant medium is: 20% potato, 1.7-2% glucose, 1.4-1.5% agar, natural pH

Move Verticillium lecanii K1207 to the slant medium of the test tube, and culture it at a constant temperature of 28°C for 3-4 days to obtain the test tube species.

2. Liquid culture

The liquid fermentation base formula is improved SDAY medium: 1% peptone, 4% glucose, 0.5% yeast extract; 0.136% KH ₂ PO ₄ , 0.12%, MgSO ₄ 7H ₂ O; pH6.0～6.5

Inoculate the test tube seeds into 500ml Erlenmeyer flask liquid culture medium, fill each bottle with 200ml, shake and cultivate at 28°C for 4-5 days at a constant temperature of 100-120rpm, mix the seed solution in each bottle, and add 20ml/bottle to the secondary In the culture medium (improved SDAY), culture on a shaker for 4-5 days (100-120 rpm) to obtain a liquid preparation.

3. Solid culture

The formula of the solid medium is wheat bran-bran medium: 56% wheat bran, 24% fine bran, 20% chaff, 0.136% KH ₂ PO ₄ , 0.12% MgSO ₄ ·7H ₂ O.

After diluting the liquid preparation 3 times with sterilized water, add solid medium at a ratio of 6:10, mix well and place in a solid fermentation room fumigated and sterilized by peracetic acid for 6 days in the dark at 25±1°C, with continuous light 3d, Obtain a solid culture.

4. Preparation of wettable powder

The cultivated product is dried, pulverized, mixed with wetting agent, filler and stabilizer to make wettable powder. The preferred composition and ratio of each component of wettable powder are: spore powder (50～60%), wetting agent (sodium dodecylbenzenesulfonate, 2～6%), stabilizer (glycerin, 0.5%～1 %), filler (diatomaceous earth, 30-40%).

Determination of finished product quality indicators: The determination of effective spore concentration, sieving rate, fineness, wetting time, suspension rate, water content, and pH value physical and chemical indicators are all carried out with reference to the compilation of Chinese agricultural standards. The results are shown in Table 1

Table 1 Physical indicators of Verticillium lecanii K1207 insecticide

Physical indicators Effective spore concentration Sieving rate Fineness Wetting time Suspension rate Water content pH value 3.0×10 ⁹ pieces/g 31.2% More than 97% passed through a 300-mesh sieve 2 minutes and 57 seconds 75.6% 3.0% 6～7

The determination of viable spore rate is determined by the spore germination method: take 0.5g of the preparation and 30mL of sterilized water and filter it with 6 layers of gauze, after the filtrate settles for 1min, take the supernatant and centrifuge at 500rpm for 1min, then take the supernatant and centrifuge at 1500rpm for 10min, remove the supernatant , dilute the precipitate with 1% glucose solution to a concentration of 10 ⁴ spores/mL, spread the diluted solution evenly on the clear water agar medium, and observe the spore germination after 18-24 hours. After the preparation was made, samples were taken and stored in a refrigerator at 4°C and in a cool place at room temperature, and the viable spore rates were measured after 1 month and 8 months. The test results are as follows:

The viable spore rate when the preparation is made: 63.5%; the viable spore rate of the preparation stored at room temperature after 1 month: 60.8%; the viable spore rate of the prepared preparation stored in a refrigerator at 4°C after 8 months: 54%; The live spore rate of the preparation under preservation: 46.6%.

Drug efficacy test of verticillium lecanii insecticide of the present invention

1. Preparation of insecticide for Verticillium lecanii: preparation for test was prepared according to the culture method of Verticillium lecanii K1207 strain mentioned above.

2. The control test of Verticillium lecanii K1207 liquid preparation on various aphids (Peach aphid, red aphid, wheat aphid without net, cabbage aphid, etc.), cabbage caterpillar, diamondback moth, whitefly in greenhouse

The results of the indoor toxicity test carried out by the spore liquid dipping method are shown in Table 2-17

Table 2 LC ₅₀ and LC ₈₀ of conidia of Verticillium lecanii K1207 against Myzus persicae

Days after treatment (d) Virulence regression equation (y=a+bx) Correlation coefficient (r) LC ₅₀ (conidia/ml) LC ₈₀ (conidia/ml) 4 5 6 7 y＝3.1063+0.2178x y＝2.9137+0.3684x y＝2.8752+0.4235x y＝3.2307+0.4132x 0.9953 0.9718 0.9777 0.9582 9.27×10 ⁶ 4.60×10 ⁵ 1.04×10 ⁵ 1.91×10 ⁴ 10.07×10 ⁸ 8.98×10 ⁷ 1.02×10 ⁷ 2.11×10 ⁶

Table 3 LT ₅₀ and LT ₈₀ of conidia of Verticillium lecanii K1207 to Myzus persicae

Concentration (conidia/ml) Virulence regression equation (y=a+bx) Correlation coefficient (r) LT ₅₀ (conidia/ml) LT ₈₀ (conidia/ml) 1.8×10 ³ 1.8×10 ⁴ 1.8×10 ⁵ 1.8×10 ⁶ 1.8×10 ⁷ 1.8×10 ⁸ y＝2.4332+2.6959x y＝2.3993+2.3711x y＝2.8122+2.8219x y＝2.1884+4.1442x y＝1.7930+5.3626x y＝1.8340+6.0774x 0.9897 0.9924 0.9904 0.9995 0.9950 0.9961 9.0 8.5 6.0 4.8 4.0 3.3 18.4 19.2 11.9 7.6 5.7 4.6

According to the measurement results in Tables 2 and 3, when conidia of Verticillium lecanii K1207 are used to control peach aphids, the effective period is 5-7 days, and 80% is the control effect standard. The suitable concentration is 1.8×10 ⁶ Conidia/ml.

Table 4 LC ₅₀ and LC ₈₀ of conidia of Verticillium lecanii K1207 against Uroleucon formosanum

Days after treatment (d) Virulence regression equation (y=a+bx) Correlation coefficient (r) LC ₅₀ (conidia/ml) LC ₈₀ (conidia/ml) 4 5 6 7 y＝2.0066+0.2745x y＝1.8057+0.4394x y＝2.9095+0.3761x y＝3.3660+0.3779x 0.9083 0.9919 0.9973 0.9860 8.03×10 ¹⁰ 1.86×10 ⁷ 3.62×10 ⁵ 2.11×10 ⁴ 9.51×10 ¹ 1.55×10 ⁹ 6.33×10 ⁷ 3.60×10 ⁶

Table 5 LT ₅₀ and LT ₈₀ of conidia of Verticillium lecanii K1207 against Uroleucon formosanum

Concentration (conidia/ml) Virulence regression equation (y=a+bx) Correlation coefficient (r) LT ₅₀ (conidia/ml) LT ₈₀ (conidia/ml) 2.5×10 ³ 2.5×10 ⁴ 2.5×10 ⁵ 2.5×10 ⁶ 2.5×10 ⁷ 2.5×10 ⁸ y＝0.8153+4.2370x y＝1.3060+4.0023x y＝0.8391+4.8095x y＝0.8546+5.6775x y＝0.8210+8.3440x y＝0.7498+6.6632x 0.8953 0.9252 0.9072 0.9588 0.9935 0.9820 9.7 8.4 7.3 5.4 5.0 4.3 15.4 13.6 11.5 7.6 6.3 5.8

According to the measurement results in Tables 4 and 5, when using the conidia of the K1207 strain to control the white-tailed red aphid, the effective period is 5-7 days, the control effect standard is 80%, and the suitable concentration is 2.5× ¹⁰⁶ conidia /ml.

Table 6 LC ₅₀ and LC ₈₀ of conidia of Verticillium lecanii K1207 on Acyrthosiphon dirhodum

Days after treatment (d) Virulence regression equation (y=a+bx) Correlation coefficient (r) LC ₅₀ (conidia/ml) LC ₈₀ (conidia/ml) 4 5 6 7 y＝3.5590+0.2151x y＝3.6237+0.2779x y＝3.5847+0.3298x y＝3.3453+0.4543x 0.9680 0.9603 0.9757 0.9818 5.00×10 ⁶ 8.96×10 ⁴ 1.96×10 ⁴ 4.39×10 ³ 4.18×10 ¹⁰ 9.73×10 ⁷ 7.07×10 ⁶ 3.16×10 ⁵

Table 7 LT ₅₀ and LT ₈₀ of conidia of Verticillium lecanii K1207 on Acyrthosiphon dirhodum

Concentration (conidia/ml) Virulence regression equation (y=a+bx) Correlation coefficient (r) LT ₅₀ (conidia/ml) LT ₈₀ (conidia/ml) 1.3×10 ³ 1.3×10 ⁴ 1.3×10 ⁵ 1.3×10 ⁶ 1.3×10 ⁷ 1.3×10 ⁸ y＝1.9516+3.4025x y＝2.2354+3.4840x y＝2.0157+4.5020x y＝1.8894+4.8953x y＝2.1657+5.0472x y＝2.3913+4.9647x 0.9806 0.9884 0.9969 0.9960 0.9837 0.9903 7.9 6.2 4.6 4.3 3.6 3.4 13.9 10.9 7.1 6.4 5.4 5.0

According to the measurement results in Tables 6 and 7, when using the conidia of the K1207 strain to prevent and control Aphid aphidica, 5-7 days is the effective period, 80% is the control effect standard, and the suitable concentration is 1.4×10 ⁶ Conidia/ml.

Table 8 LC ₅₀ and LC ₈₀ of the conidia of Verticillium lecanii K1207 to the cabbage aphid (Brevicoryne brassicae)

Days after treatment (d) Virulence regression equation (y=a+bx) Correlation coefficient (r) LC ₅₀ (conidia/ml) LC ₈₀ (conidia/ml) 4 5 6 7 y＝2.7488+0.2695x y＝3.4550+0.2294x y＝3.4649+0.2802x y＝3.6811+0.3147x 0.9847 0.9943 0.9946 0.9997 2.26×10 ⁸ 5.43×10 ⁶ 3.01×10 ⁵ 1.55×10 ⁴ 3.04×10 ¹¹ 2.58×10 ¹⁰ 3.09×10 ⁸ 7.44×10 ⁶

Table 9 LT ₅₀ and LT ₈₀ of Verticillium lecanii K1207 conidia to cabbage aphid (Brevicoryne brassicae)

Concentration (conidia/ml) Virulence regression equation (y=a+bx) Correlation coefficient (r) LT ₅₀ (conidia/ml) LT ₈₀ (conidia/ml) 1.3×10 ³ 1.3×10 ⁴ 1.3×10 ⁵ 1.3×10 ⁶ 1.3×10 ⁷ 1.3×10 ⁸ y＝1.3912+3.8370x y＝1.0350+4.7027x y＝2.0157+4.5020x y＝1.8894+4.8953x y＝2.1657+5.0472x y＝2.3913+4.9647x 0.9790 0.9976 0.9969 0.9960 0.9837 0.9903 8.7 7.0 4.6 4.3 3.6 3.4 14.5 10.5 7.1 6.4 5.4 5.0

According to the measurement results in Tables 8 and 9, when using conidia of the K1207 strain to control cabbage aphids, 5 to 7 days is the effective period, 80% is the control effect standard, and the suitable concentration is 1.3×10 ⁵ conidia/ ml.

Table 10 LC ₅₀ and LC ₈₀ of conidia of Verticillium lecanii K1207 on cabbage caterpillar (Artogeia rapae)

Days after treatment (d) Virulence regression equation (y=a+bx) Correlation coefficient (r) LC ₅₀ (conidia/ml) LC ₈₀ (conidia/ml) 4 5 6 7 8 y＝2.7275+0.2324x y＝2.8076+0.3021x y＝3.2593+0.2947x y＝3.2298+0.3452x y＝3.2590+0.3840x 0.9797 0.9844 0.9906 0.9958 0.9982 6.00×10 ⁹ 1.81×10 ⁷ 8.07×10 ⁵ 1.34×10 ⁵ 3.42×10 ⁴ 2.056×10 ¹³ 2.96×10 ¹² 5.88×10 ⁸ 3.73×10 ⁷ 5.38×10 ⁶

Table 11 LT ₅₀ and LT ₈₀ of Verticillium lecanii K1207 conidia to cabbage caterpillar (Artogeia rapae)

Concentration (conidia/ml) Virulence regression equation (y=a+bx) Correlation coefficient (r) LT ₅₀ (conidia/ml) LT ₈₀ (conidia/ml) 2.0×10 ³ 2.0×10 ⁴ 2.0×10 ⁵ 2.0×10 ⁶ 2.0×10 ⁷ 2.0×10 ⁸ y＝1.3936+3.5128x y＝1.3544+3.9444x y＝1.4358+4.3712x y＝1.2506+4.8540x＝0.9420+5.7688x y＝1.6288+5.2165x 0.9596 0.9852 0.9971 0.9996 0.9974 0.9919 10.6 8.4 6.5 5.9 5.0 4.4 18.5 13.7 10.2 8.8 7.1 6.4

According to the measurement results in Tables 10 and 11, when using the conidia of the K1207 strain to control Pieris rapae, 5 to 7 days is the effective period, 80% is the control effect standard, and the suitable concentration is 2.0×10 ⁷ conidia/ ml.

Table 12 LC ₅₀ and LC ₈₀ of conidia of Verticillium lecanii K1207 to 2nd instar larvae of diamondback moth (Plutella xylostella)

Days after treatment (d) Virulence regression equation (y=a+bx) Correlation coefficient (r) LC ₅₀ (conidia/ml) LC ₈₀ (conidia/ml) 4 5 6 7 8 y＝2.0045+0.3253x y＝2.8853+0.2958x y＝3.1601+0.3070x y＝2.9892+0.3839x y＝3.1835+0.4204x 0.9810 0.9599 0.9908 0.9909 0.9947 1.62×10 ⁹ 1.41×10 ⁷ 9.84×10 ⁵ 1.73×10 ⁵ 2.09×10 ⁴ 6.33×10 ¹¹ 1.00×10 ¹⁰ 5.51×10 ⁸ 2.72×10 ⁷ 2.13×10 ⁶

Table 13 LT ₅₀ and LT ₈₀ of conidia of Verticillium lecanii K1207 to 2nd instar larvae of diamondback moth (Plutella xylostella)

Concentration (conidia/ml) Virulence regression equation (y=a+bx) Correlation coefficient (r) LT ₅₀ (conidia/ml) LT ₈₀ (conidia/ml) 2.2×10 ³ 2.2×10 ⁴ 2.2×10 ⁵ 2.2×10 ⁶ 2.2×10 ⁷ 2.2×10 ⁸ y＝1.2292+3.7285x y＝0.7531+4.5672x y＝0.6399+5.3313x y＝0.3879+4.1519x y＝0.0478+6.9544x y＝0.3047+7.0017x 0.9683 0.9679 0.9841 0.9941 0.9928 0.9926 10.3 8.5 6.6 5.6 5.2 4.7 17.3 13.0 9.5 7.7 6.8 6.2

According to the measurement results in Tables 12 and 13, when the conidia of the K1207 strain are used to control Plutella xylostella, 5 to 7 days is the effective period, 80% is the control effect standard, and the suitable concentration is 2.2×10 ⁷ conidia/ ml.

Table 14 LC ₅₀ and LC ₈₀ of conidia of Verticillium lecanii K1207 against whitefly (Trialeurodes vaporariorum) nymphs

Days after treatment (d) Virulence regression equation (y=a+bx) Correlation coefficient (r) LC ₅₀ (conidia/ml) LC ₈₀ (conidia/ml) 4 5 6 7 8 y＝3.0068+0.2160x y＝3.1839+0.2636x y＝3.1709+0.3173x y＝3.1888+0.3403x y＝3.3458+0.3429x 0.9851 0.9941 0.9990 0.9605 0.9140 1.69×10 ⁹ 7.76×10 ⁶ 5.82×10 ⁵ 2.10×10 ⁵ 6.67×10 ⁴ 1.36×10 ¹³ 1.23×10 ¹⁰ 2.56×10 ⁸ 6.33×10 ⁷ 1.92×10 ⁷

Table 15 LT ₅₀ and LT ₈₀ of conidia of Verticillium lecanii K1207 to whitefly (Trialeurodes vaporariorum) nymphs in greenhouse

Concentration (conidia/ml) Virulence regression equation (y=a+bx) Correlation coefficient (r) LT ₅₀ (conidia/ml) LT ₈₀ (conidia/ml) 1.8×10 ³ 1.8×10 ⁴ 1.8×10 ⁵ 1.8×10 ⁶ 1.8×10 ⁷ 1.8×10 ⁸ y＝1.8601+3.0707x y＝1.9303+3.3668x y＝1.2291+4.6573x y＝0.9629+5.4528x y＝1.0085+5.7903x y＝1.4678+5.5597x 0.9964 0.9547 0.9967 0.9860 0.9988 0.9996 10.5 8.2 6.5 5.5 4.9 4.3 14.6 14.5 9.8 7.9 6.8 6.1

According to the measurement results in Tables 14 and 15, when using conidia of the K1207 strain to control whitefly in greenhouses, the period of effective effect is 5-7 days, and the control effect standard is 80%. The suitable concentration is 1.8× ¹⁰⁷ conidia /ml.

Table 16 LC ₅₀ and LC ₈₀ of Verticillium wax scale K1207 to (on cycad) red wax scale (Ceroplastes rubens)

Days after treatment (d) Virulence regression equation (y=a+bx) Correlation coefficient (r) LC ₅₀ (conidia/ml) LC ₈₀ (conidia/ml) 7 8 9 10 11 12 y＝2.8761+0.2258x y＝3.0904+0.2432x y＝3.2195+0.2532x y＝3.6102+0.2456x y＝3.6551+0.2801x y＝3.6323+0.3351x 0.9794 0.9531 0.9884 0.9956 0.9983 0.9980 2.55×10 ⁹ 7.11×10 ⁷ 1.08×10 ⁷ 4.55×10 ⁵ 6.33×10 ⁴ 7.11×10 ³ 1.39×10 ¹³ 2.09×10 ¹¹ 2.31×10 ¹⁰ 1.24×10 ⁹ 6.51×10 ⁷ 1.69×10 ⁶

Table 17 LT ₅₀ and LT ₈₀ of Verticillium wax scale K1207 conidia to (on cycad) red wax scale (Ceroplastes rubens)

Concentration (conidia/ml) Virulence regression equation (y=a+bx) Correlation coefficient (r) LT ₅₀ (conidia/ml) LT ₈₀ (conidia/ml) 1.76×10 ³ 1.76×10 ⁴ 1.76×10 ⁵ 1.76×10 ⁶ 1.76×10 ⁷ 1.76×10 ⁸ y＝-0.5250+4.9318x y＝-0.4559+3.1312x y＝-0.4481+5.3611x y＝-1.7369+6.9387x y＝-1.7115+7.1850x y＝-1.1757+6.9330x 0.9833 0.9771 0.9774 0.9864 0.9844 0.9924 13.2 11.6 10.4 9.4 8.6 7.8 19.6 16.9 14.9 12.4 11.3 10.3

According to the measurement results in Tables 16 and 17, when conidia of the K1207 strain are used to control the red wax scale on cycads, 10-12 days is the effective period, 80% is the control effect standard, and the suitable concentration is 1.76×10 ⁶ ~1.76×10 ⁸ conidia/ml.

Overall results show that the verticillium lecanii (Verticillium lecanii) insecticide of the present invention is effective in preventing and controlling green peach aphid, red aphid white-tailed, long-tube aphid without net, cabbage aphid, etc.), cabbage caterpillar, diamondback moth, and greenhouse whitefly . Its effect of controlling plant pests is related to the frequency and method of use, and at the same time is related to the types of pests to be controlled.

Detailed ways:

The biocontrol agent can be mixed with other insecticidal pesticides and plant growth regulators, but it must be ensured that the mixed agents have no side effects on the Verticillium lecanii K1207 used in the present invention.

The verticillium lecanii of the present invention has obvious control effects on green peach aphid, red aphid white-tailed, long tube aphid, cabbage aphid, cabbage caterpillar, diamondback moth, and greenhouse whitefly, and has the advantages of low cost and easy use. Features such as simplicity and long validity period.

The present invention is further described in detail below with examples, but the content of the present invention is not limited thereto.

Embodiment one:

Verticillium lecanii (Verticillium lecanii) K1207 was moved to the test tube slant medium, the formula of the test tube slant medium was: 20% potato, 1.7-2% glucose, 1.4-1.5% agar, natural pH.

Cultivate at a constant temperature of 28°C for 3-4 days to obtain test tube species.

Inoculate the test tube seed into a 500ml Erlenmeyer flask (each bottle contains 200ml) liquid medium, the medium formula is: 1% peptone, 4% glucose, 0.5% yeast extract; 0.136% KH ₂ PO ₄ , 0.12% MgSO ₄ 7H ₂ O; pH 6.0-6.5, cultivated at 28°C for 4-5d with constant temperature shaking, with a rotation speed of 100-120rpm, added 20ml/bottle into the secondary culture medium (improved SDAY), and cultured on a shaker for 4-5d (100 ~120rpm) to obtain the liquid formulation of the present invention.

Embodiment two:

First obtain the liquid preparation (same as Example 1), after the liquid preparation is diluted 3 times with sterilized water, add solid culture medium in the ratio of 6:10, the formula of solid culture medium is wheat bran--grain bran culture medium: 56 % wheat bran, 24% fine bran, 20% chaff, 0.136% KH ₂ PO ₄ , 0.12% MgSO ₄ 7H ₂ O, mix well and place in a solid fermentation room after peracetic acid fumigation and sterilization at 25±1 Cultivate in the dark at ℃ for 6 days, with continuous light for 3 days, then take out the glass plate and spread it in the fermentation room, and air-dry it at 35±1℃ and 100 gear wind speed for 5 days. Obtain a solid formulation.

Embodiment three:

First carry out liquid-solid two-phase fermentation (same as Example 1 and Example 2), and then dry, pulverize, add a wetting agent, a filler and a stabilizer to mix the cultured product to prepare a wettable powder.

The preferred composition and ratio of each component of wettable powder are: spore powder (50～60%), wetting agent (sodium dodecylbenzenesulfonate, 2～6%), stabilizer (glycerin, 0.5%～1 %), filler (diatomaceous earth, 30-40%).

Determination of finished product quality indicators: The determination of effective spore concentration, sieving rate, fineness, wetting time, suspension rate, water content, and pH value physical and chemical indicators are all carried out with reference to the compilation of Chinese agricultural standards. The results are shown in Table 1

Table 1 Physical indicators of Verticillium lecanii K1207 insecticide

Physical indicators Effective spore concentration Sieving rate Fineness Wetting time Suspension rate Water content pH value 3.0×10 ⁹ pieces/g 31.2% More than 97% passed through a 300-mesh sieve 2 minutes and 57 seconds 75.6% 3.0% 6～7

The determination of the viable spore rate is determined by the spore germination method: take 0.5g of the preparation and 30mL of sterilized water and filter it with 6 layers of gauze. After the filtrate settles for 1min, take the supernatant and centrifuge at 500rpm for 1min, then take the supernatant and centrifuge at 1500rpm for 10min, and remove the supernatant , dilute the precipitate with 1% glucose solution to a concentration of 10 ⁴ spores/mL, spread the diluted solution evenly on the clear water agar medium, and observe the spore germination after 18-24 hours. After the preparation was made, samples were taken and stored in a refrigerator at 4°C and in a cool place at room temperature, and the viable spore rates were measured after 1 month and 8 months. The test results are as follows:

The viable spore rate when the preparation is made: 63.5%; the viable spore rate of the preparation stored at room temperature after 1 month: 60.8%; the viable spore rate of the prepared preparation stored in a refrigerator at 4°C after 8 months: 54%; The live spore rate of the preparation under preservation: 46.6%.

The biological preparation of the present invention is obtained.

Claims (2)

1. a Verticillium lecanii insecticide, characterized in that: the production strain of the preparation is Verticillium lecanii (Verticillium lecanii) K1207, preservation number CGMCC No.2326. 2. the application of a kind of Verticillium lecanii insecticide described in claim 1 in the prevention and control of green peach aphid, red aphid, wheat aphid without net, cabbage aphid, cabbage caterpillar, diamondback moth, greenhouse whitefly pest .