Metarhizium anisopliae MAYX130921 and application thereof
A technology of scarab metarhizium anisopliae and black beetle, applied in the field of microorganisms, to achieve the effects of rapid growth, strong pathogenicity, and high spore germination rate
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Embodiment 1
[0010] Embodiment one, the isolation of pathogenic bacteria, identification
[0011] 1.1 Materials and methods
[0012] 1.1.1 Materials
[0013] In Eshan County, Yuxi City, Yunnan Province, the diseased Diablo spp. infected by an entomogenic fungus was collected.
[0014] Gill beetle larvae.
[0015] Sabouraud medium (SDAY): 1% peptone + 1% yeast powder + 4% glucose + 1.5-2% agar powder + 1000ml water.
[0016] Sterile operating conditions: All utensils and utensils are sterilized in a high-temperature autoclave (121°C, 30min), and inoculation and other operations are performed in an ultra-clean workbench.
[0017] Culture conditions: Culture in a 28°C light (12L:12D) incubator. After the colonies are formed, transfer to the SDAY slant of the test tube, cultivate for 3 to 5 days, and transfer to a 4°C refrigerator for storage.
[0018] 1.1.2 Isolation and purification of pathogenic bacteria
[0019] Isolation: Isolate the pathogenic bacteria from the dead body of the sick...
Embodiment 2
[0028] Embodiment two, the biological characteristics of the purified bacterial strain of Metarhizium anisopliae
[0029] 2.1 Materials and methods
[0030] 2.1.1 Tested strains
[0031] Select a plate with vigorous growth and uniform growth after purification as the test strain. The hyphae were inoculated on SDAY again, and cultivated in a constant temperature light (12L:12D) incubator at 28°C.
[0032] 2.1.2 Determination of colony growth rate and sporulation
[0033] Take a plate of Metarhizium anisopliae cultured in advance and punch holes with an 8mm puncher, inoculate it on a new medium, do three repetitions, and place it in a constant temperature light (12D:12L) incubator at 28°C for cultivation , measure its diameter regularly every day and record it until the colony is overgrown with the culture medium. Use a hole puncher with a diameter of 8mm to take the bacterium cake at the same position of the culture medium, add 1% Tween-80 and 20ml sterile water, wash the s...
Embodiment 3
[0044] Example 3, Indoor Toxicity Determination of Metarhizium anisopliae MAYX130921 to Scaraba larvae
[0045] 3.1. Materials and methods
[0046] 3.1.1 Source of tested insects
[0047] The 2nd instar larvae of the black beetle were reared with potato cubes in an incubator with a temperature of (25±1)°C, a humidity of (70±5)%, and a photoperiod of 0L:24D in our laboratory. The second-instar larvae of the black beetle, which are disease-free and of the same size, were selected as test insects.
[0048] 3.1.2 Preparation of spore suspension
[0049] Get the well-growing Purified Scarab anisopliae Metarhizium anisopliae, wash the conidia with sterile water and filter to configure 10 8 Spores / ml conidia suspension, use a sterile capillary pipette to take a drop of the filtrate onto a hemocytometer, count the spores under a microscope and record the data, and dilute to 10 with sterile water containing 0.5% Tween 80 8 、10 7 、10 6 、10 5 、10 4 The spore suspensions with five...
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