Preparation and application of composite microecological feed additive
A feed additive and compound micro-ecology technology, applied in the field of micro-ecological feed additives, can solve the problems of antibiotic residues, superinfection, etc., and achieve the effects of improving the feed conversion rate and reducing the production of toxic substances in the gastrointestinal tract.
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Embodiment 1
[0025] Isolation, Purification and Physiological and Biochemical Identification of Clostridium butyricum CPU-1
[0026] 1. Isolation and purification of Clostridium butyricum CPU-1
[0027] (1) Dissolve 1 g of fresh pig feces in 9 ml of normal saline, put it in an 80°C water bath for 10 minutes to kill non-spore bacteria, then put it into the proliferation medium, and incubate at 37°C for 36 hours.
[0028] (2) Place the above-mentioned culture solution in a water bath at 80°C for 10 minutes, and transfer it to the Clostridium selective culture medium, the composition of which is: tryptone 15g / L, yeast extract powder 10g / L, sodium sulfite 1g / L, ferric citrate 0.5g / L, novobiocin 0.02g / L, polymyxin 0.05g / L, pH7.2, sterilized at 121°C for 20min, anaerobic selective enrichment culture at 37°C for 48h.
[0029] (3) Separate the samples from 10 -1 to 10 -7 Doubling dilution, spread on the GAM medium plate, the medium composition is as follows: 15g / L leucone, 10g / L tryptone, 3g / L ...
Embodiment 2
[0039] Isolation, Purification and Physiological and Biochemical Identification of Lactobacillus Acidophilus CPU-2
[0040] For the isolation and purification method of Lactobacillus acidophilus CPU-2, refer to the isolation and purification of Clostridium butyricum, and the selection medium is added with 3% CaCO3 MRS medium; see the morphological characteristics of Lactobacillus acidophilus figure 2 , Lactobacillus acidophilus is a facultative anaerobic bacteria, Gram-positive, blunt-rounded ends, medium-sized, non-spore-free, non-motile bacilli, appearing in single, double or short chains.
[0041] Table 2 Physiological and biochemical characteristics of Lactobacillus acidophilus
[0042]
[0043]
[0044] Note: "+" indicates a positive reaction or can be used; "—" indicates a negative reaction or cannot be used.
[0045] According to the classification and identification of the ninth edition of Bergey's Bacteria Identification Manual, the strain is Lactobacillus aci...
Embodiment 3
[0047] Study on Optimum Proportion of Clostridium butyricum CPU-1 and Lactobacillus acidophilus CPU-2
[0048] Activate Clostridium butyricum CPU-1 and Lactobacillus acidophilus CPU-2 respectively, adjust the bacterial concentration to 10 with MRS medium 8 CFU / ml, respectively inoculate Clostridium butyricum CPU-1 and Lactobacillus acidophilus CPU-2 in 5ml MRS medium for 24 hours according to the inoculation pattern in the table below, and centrifuge the bacterial culture solution at 12000rpm for 5 minutes in a refrigerated centrifuge, and then use 0.22μm filter membrane and store in a -20°C refrigerator. Using Shigella as indicator bacteria, the inhibition zone of the supernatant was measured by the double-layer agar diffusion method.
[0049]
[0050] Depend on image 3 It can be seen that the different ratios of the two strains are different in the size of the inhibition zone produced by Shigella as the indicator bacterium, and the maximum inhibitory zone is reached wh...
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