Clostridium butyricum with strong antibacterial ability and application thereof

A technology of Clostridium butyricum and bacterial suspension, applied in the field of microorganisms, can solve the problems of different effects, different strain characteristics, safety evaluation of Clostridium butyricum toxin gene and drug resistance spectrum, etc., and achieves strong bile salt resistance. , safety assurance, the effect of good application promotion potential

Active Publication Date: 2021-07-23
SHANDONG AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The current domestic Clostridium butyricum products have different strain characteristics and different effects, and there is no safety assessment of the toxin gene and drug resistance spectrum of Clostridium butyricum

Method used

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  • Clostridium butyricum with strong antibacterial ability and application thereof
  • Clostridium butyricum with strong antibacterial ability and application thereof
  • Clostridium butyricum with strong antibacterial ability and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] Embodiment 1: Isolation and screening of bacterial strains

[0042] Take Babcourt SPF laying hens feces for bacterial separation test: Accurately weigh 5g of feces samples and add them to 50mL of normal saline to prepare a suspension; put the suspension in a water bath at 85°C for 10min, take the shaken and mixed Add 1mL of the diluent to 5mL RCM medium, and enrich the bacteria anaerobically at 37°C for 48h; take the above enrichment solution and bathe in water at 85°C for 10min, repeat the above steps for RCM enrichment once, and inoculate on the RCM plate by the three-line method (RCM plate is Add 15g / L agar to the RCM medium), and culture anaerobically for 24-48 hours; after the colony grows, select a single colony with irregular colony shape, white or yellowish white, and streak and purify it again on the RCM plate. Repeat twice. The morphology of the strains on the RCM plate is as follows: figure 1shown.

Embodiment 2

[0043] Embodiment 2: the identification of bacterial strain

[0044] The suspected Clostridium butyricum colony with irregular borders after being cultured for 48 hours after two times of streak purification was picked and named as FW1, and the species identification was carried out according to the instruction manual for the identification of Salmonella by microbial mass spectrometry. Pick a single colony with a toothpick and spread it on MSP96target polished steel BC, add 1 μL of standard solvent first, and then add 1 μL of matrix solution after drying, and use Bruker’s automatic rapid biological mass spectrometer for identification. Clostridium acid ( figure 2 ).

[0045] The genome of strain FW1 was further extracted, and 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, 1 μL of upstream and downstream primers, 1 μL of DNA template. ...

Embodiment 3

[0050] Example 3: Drug susceptibility test of Clostridium butyricum strain FW1 to evaluate drug resistance spectrum

[0051] Select 8 kinds of antibiotic-sensitive tablets that are effective against Gram-positive bacteria: ofloxacin, cefepime, neomycin, nitrofurantoin, florfenicol, erythromycin, vancomycin, tetracycline, and use sterile cotton swabs to The bacterial solution enriched for 24 hours was evenly spread on the RCM plate, and three kinds of drug-sensitive paper sheets were pasted equidistantly on each plate. The medium was cultured anaerobically at 37°C for 18 hours, and then the diameter of the inhibition zone was measured. According to the World Health Organization recommended The Kirby-Bauer method (K-B method) and the American Clinical Laboratory Standards Institute (CLSI) manual were used to determine the experimental results. The results are shown in Table 1. Clostridium butyricum strain FW1 was sensitive to the above eight antibiotics.

[0052] Table 1 Suscep...

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Abstract

The invention discloses clostridium butyricum with strong bacteriostatic ability and application of the clostridium butyricum. The clostridium butyricum is preserved in China General Microbiological Culture Collection Center on April 29, 2021, and the biological preservation number of the clostridium butyricum is CGMCC NO.22258. The clostridium butyricum screened in the invention does not have clostridial toxin and botulinum toxin genes, is narrow in drug resistance spectrum, high in safety and high in tolerance to gastrointestinal fluid, cholate and high temperature, has the characteristic of high bacteriostatic ability to pathogenic bacteria such as escherichia coli, salmonella, morganella morganii and klebsiella pneumonia, and has good application potential for animal feed probiotics.

Description

technical field [0001] The invention relates to the technical field of microorganisms, in particular to a strain of Clostridium butyricum with strong antibacterial ability and its application. Background technique [0002] Clostridium butyricum belongs to the genus Clostridium and is a strictly anaerobic Gram-positive bacillus with perinatal flagella, straight or slightly curved cells, and endospores oval, which produce acid and gas during fermentation. The bacterium is a normal flora of the intestinal tract of humans and animals and is also commonly found in the environment. Clostridium butyricum can produce a large amount of short-chain fatty acids, such as butyrate and acetate. Studies have shown that butyric acid can exert anti-inflammatory activity by inhibiting the activation of NF-κB or up-regulating the expression of peroxisome proliferator-activated receptor γ, thereby controlling the growth of pathogenic bacteria and improving intestinal flora disturbance. Clostr...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12P7/52A23L33/135A23K10/18C12R1/145
CPCC12N1/20C12P7/52A23L33/135A23K10/18A23V2002/00A23V2200/30
Inventor 孙淑红郝桂娟
Owner SHANDONG AGRICULTURAL UNIVERSITY
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