Screening and application of enterococcus faecium G12
A technology of Enterococcus faecium and G12, applied in the field of microbial additives for aquatic products, can solve the problems of increased overall drug resistance and unfavorable treatment of animal diseases, and achieve the effect of sustainable development, significant probiotics, and high resistance to stress.
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[0027] Example 1: Screening, isolation and identification of strains
[0028] Screening and isolation: A strain of Enterococcus faecium was selected from the hepatopancreas of shellfish, especially the hepatopancreas of Philippine clam. The specific operation of the screening is as follows: the body surface of the clam clams is sterilized with 75% medical alcohol and then dissected, the liver and pancreas are taken, and then 1 mL of pre-chilled sterile normal saline is added, homogenized thoroughly, centrifuged, and the supernatant is taken 10 times series dilution. Take stock solution, 10 -1 , 10 -2 The samples of the three dilutions were each 100uL, and they were coated on the MRS plate. Each dilution gradient was set to 3 parallel, and the samples were incubated at 37°C for 24h. Pick a single colony and purify the bacteria by streaking multiple times. The purified strain was made into a bacterial suspension with 40% glycerol and MRS liquid medium 1:1, and stored at -80°C for...
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[0035] Example 2: Bacteriostatic test
[0036] Aeromonas victorini, Pseudomonas aeruginosa and Vibrio vulnificus (the above-mentioned strains are provided by the Aquatic Animal Disease and Immunology Laboratory of the Fisheries College of Tianjin Agricultural College) as indicator bacteria. Methods Refer to the literature (Evidence for the competitive exclusion of Aeromonas salmonicida from fish with a tressinducible furunculosis by a fluorescent pseudomonad. Smith P and Dabey S, "Journal of Fish Diseases", 1993 Issue 16), and the activated indicator bacteria and the test Strains were cultured in LB liquid medium for 24h, diluted with saline to 1×10 7 cfu / mL, take 0.2mL of indicator strain and spread it on fresh LB solid medium, and then take 2μL of test strain to spot on the above plate, and observe whether there is obvious inhibition zone around the spot seeding area for 24h.
[0037] The Oxford cup method (using the Oxford cup method to determine the antibacterial activity of pr...
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[0040] Example 3: Safety test
[0041] Hemolysis test: Streak the tested strain on LB solid medium for 24h, scrape a single colony with an inoculation loop and spot it on the blood plate, incubate at 37℃ for 24h, observe whether a transparent circle is formed around the colony, and judge the production of hemolysin . The experimental results showed that: no hemolysis was observed on the blood plate, indicating that the strain did not produce hemolysin and had no potential pathogenicity.
[0042] Animal test: refer to the literature, select the immersion test of Penaeus monodon juveniles, select 1×10 7 , 1×10 5 The concentration of cfu / mL bacterial solution was soaked, the positive control was selected as Vibrio vulnificus, and the negative control group was selected as normal saline. After being soaked in the bacterial suspension for four hours, the juvenile shrimp were transferred to violent water. Each treatment group made three parallel observations for 7 days, and recorded th...
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