Bacillus preparation for preventing and controlling pathogenic bacteria in soil environments as well as preparation method and application thereof
A technology of bacillus and bacillus pumilus, which is applied in the field of preparation and application of bacillus for the prevention and treatment of pathogenic bacteria in the soil environment, and can solve problems such as the irrelevance of antibiotic abuse, the pollution of soil and water by disinfectants, and the threat to the survival of animals and plants.
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Embodiment 1
[0059] A bacillus preparation for preventing and controlling pathogenic bacteria in the soil environment: it is prepared by mixing equal volumes of bacterial liquids of three kinds of bacteria, and the three kinds of bacteria are all isolated by the inventor from soil contaminated by pathogenic bacteria in Hubei area. Three kinds of bacteria are respectively a kind of Bacillus pumilus D31 (abbreviated as D31 in the present invention), a kind of Bacillus amyloliquefaciens J3-1 (abbreviated as J3-1 in the present invention) and a kind of Bacillus amyloliquefaciens D66 (abbreviated as J3-1 in the present invention) referred to as D66), and their separation methods are the same. Its separation process is:
[0060] A. Prepare bilayer plates:
[0061] 1) inoculate Escherichia coli (the pathogenic bacteria used in the present invention is Escherichia coli, this bacterium is isolated from the pig farm soil, utilizes the mouse to carry out the animal regression test and confirms that...
Embodiment 2
[0081] A kind of preparation method of the bacillus preparation of preventing and treating soil pathogenic bacteria, its steps are:
[0082] The microbial preparation of the present invention contains three kinds of bacillus, D31, D66 and J3-1. The inventor mixed equal volumes of bacterial liquids of D31, D66, and J3-1, and then added peat and mixed thoroughly to obtain a soil microbial preparation for pollution prevention and control of pathogenic bacteria such as pathogenic Escherichia coli in livestock and poultry breeding areas. The preparation method of microbial preparation is specifically as follows:
[0083] A. Pick a single colony, inoculate D66, D31 and J3-1 into a PA bottle containing 5ml LB liquid medium, shake at 37°C, 180rpm, for 12h;
[0084] B. Transfer 1% of the inoculum to a new 500ml Erlenmeyer flask containing 250ml of LB culture medium, 37°C, 180rpm shaking culture until more than 90% of the spores are formed; the culture time of strains D66 and J3-1 is 7...
Embodiment 3
[0089] Plate inhibition zone test
[0090] 1 Experimental method
[0091] 1) Pick a single colony, inoculate antagonistic bacteria D31, D66, and J3-1 into PA bottles containing 5ml of LB, shake and culture for 48 hours at 28°C and 180rpm;
[0092] 2) Add 5ml of LB solid medium to the plate, and place the Oxford cup after solidification;
[0093] 3) Add 5ml of LB solid medium to the plate to fix the Oxford cup;
[0094] 4) After solidification, add 200 μl of antagonistic bacteria solution to the Oxford cup, and culture at 28°C for 48 hours;
[0095] 5) Inoculate Escherichia coli into a PA bottle containing 5ml of LB liquid medium, shake at 37°C, 180rpm, for 12h;
[0096] 6) Take 50 μl of Escherichia coli culture in 250ml of LB soft agar solid medium (agar content is 0.5% (agar mass as a percentage of medium volume) cooled to 50-60°C, and mix well;
[0097] 7) Add 4 ml of LB soft agar solid medium mixed with Escherichia coli to the plate in step 4, culture it statically at 2...
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