A strain of Clostridium butyricum with strong antibacterial ability and its application
A technology of Clostridium butyricum and bacterial suspension, applied in the field of microbiology, can solve the problems of different effects, different strain characteristics, Clostridium unbutyricum toxin gene and drug resistance spectrum safety evaluation, etc., and achieve strong bile salt resistance , safety assurance, good application promotion potential effect
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Embodiment 1
[0041] Example 1: Isolation and screening of strains
[0042] Take the feces of Babcock SPF laying hens for bacterial test: accurately weigh 5 g of fecal samples and add them to 50 mL of normal saline to prepare a suspension; place the suspension in a water bath at 85 °C for 10 min, and shake 1 mL of the mixed dilution solution was added to 5 mL of RCM medium, and anaerobic enrichment was carried out at 37 °C for 48 h; the above enrichment solution was taken in a water bath at 85 °C for 10 min, and the RCM enrichment was repeated once according to the above steps, and the three-line method was used for inoculation. On the RCM plate (the RCM plate is the addition of 15 g / L agar to the RCM medium), anaerobic culture for 24-48 h; after the colony grows, select the single colony with irregular colony shape, white or yellowish white on the RCM Streak purification was performed on the plate again and this was repeated twice. The morphology of the strain on the RCM plate is as follo...
Embodiment 2
[0043] Example 2: Identification of strains
[0044]The suspected Clostridium butyricum colonies with irregular edges were picked after 2 streak purification and cultured for 48 h, named as FW1, and the bacteria were identified according to the instructions for identification of Salmonella by microbial mass spectrometry. Pick a single colony with a toothpick and spread it on MSP 96target polished steel BC. First add 1 μL of standard solvent, let it dry, then add 1 μL of matrix solution, dry it, and use Bruker’s automatic rapid biological mass spectrometer to identify the results. shown as Clostridium butyricum ( figure 2 ).
[0045] The genome of strain FW1 was further extracted, and the 16S rDNA universal primers (27F: 5'-AGAGTTTGATCCTGGCTCAG-3'; 1492R: 5'-GGCTACCTTGTTACGACTT-3') were used for PCR amplification. The reaction system was 25 μL: MIX 12.5 μL, ddH 2 O 9.5 μL, upstream and downstream primers 1 μL each, DNA template 1 μL. PCR reaction conditions were: pre-denatu...
Embodiment 3
[0050] Example 3: Drug susceptibility test of Clostridium butyricum strain FW1 to evaluate drug resistance spectrum
[0051] Select 8 kinds of antibiotic susceptibility tablets that are effective against Gram-positive bacteria: ofloxacin, cefepime, neomycin, nitrofurantoin, florfenicol, erythromycin, vancomycin, tetracycline, and use sterile cotton swabs. The bacterial solution enriched for 24 h was evenly spread on the RCM plate, and each plate was equidistantly pasted with three kinds of drug-sensitive paper sheets. The recommended Kirby-Bauer method (K-B method) and the American Clinical Laboratory Standardization Institute (CLSI) handbook are used for adjudication of experimental results. The results are shown in Table 1. Clostridium butyricum strain FW1 was sensitive to the above eight antibiotics.
[0052] Table 1 Sensitivity results of Clostridium butyricum strain FW1 to 8 antibiotics
[0053]
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