Method for rapid screening of Bacillus spp with antibacterial activity

A technology for bacillus and antibacterial activity, applied in the field of rapid screening of bacillus with antibacterial activity, can solve the problems of complex screening methods, high screening labor intensity, long screening time, etc., and achieve low screening cost, simplified screening procedures, and high efficiency Screening effect

Inactive Publication Date: 2016-03-16
HUNAN AGRICULTURAL UNIV
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Problems solved by technology

Although there are some methods for screening bacteria with antibacterial activity without separation and purification, and preparing cell-free culture fluid, such as "A Method for Rapidly Screening Antibacterial Active Bacteria and Actinomycetes" (Application No.: 201410212912.5), the screening method It is too complicated, and the pipette tip needs to be intercepted, and the screening is labor-intensive
The Chinese patent application with the publication number CN105002114A discloses a method for isolating Bacillus oleolyticum, which uses high temperature combined with the method of indicator color development to screen Bacillus oleolyticum, but in this technical scheme, the samples need to pass two to three days. For the intermittent aerobic enrichment culture, the screened medium must be prepared on the basis of the prepared LB medium, add the BTB oil medium mixed with oil, and then cultivate for two days, and then judge the ability to dissolve oil by the size of the transparent circle. However, this method is cumbersome and takes a long time to screen

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  • Method for rapid screening of Bacillus spp with antibacterial activity

Examples

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Embodiment 1

[0029] The screening of the bacillus of embodiment 1 anti-escherichia coli

[0030] (1) Weigh 25.0g of the intestinal contents of the pig, add it into a conical flask containing 225mL sterile normal saline, shake it well at 37°C for 20min; In a water bath, treat in a water bath for 30 minutes, shake the Erlenmeyer flask 5 times during the water bath, then dilute the high-temperature-treated sample with sterile saline, and spread the 104-106-fold diluted bacterial suspension on the surface of the nutrient agar medium ;

[0031] (2) The composition of nutrient agar medium is: peptone 10.0g, beef extract 3.0g, sodium chloride 5.0g, agar 15.0g, distilled water 1000mL, pH7.4±0.2;

[0032] (3) The culture condition of bacillus is: culture temperature is 37 ℃, culture time is 12h;

[0033] (4) Inoculate the indicator bacterium Escherichia coli O157:H7ATCC43889 preserved in the slope into the LB liquid medium, the culture temperature is 37°C, and the culture time is 12h;

[0034] (...

Embodiment 2

[0039] Example 2 Screening of Bacillus against Listeria monocytogenes

[0040] (1) Weigh 25.0g of soil sample, add it to a conical flask containing 225mL of sterile saline, shake it well at 37°C for 20min; then place the conical flask containing the sample in a water bath at 80-90°C, Treat in a water bath for 30 minutes, shake the Erlenmeyer flask 5 times during the water bath, then gradiently dilute the high-temperature-treated sample with sterile saline, and spread the 104-106-fold diluted bacterial suspension on the surface of the nutrient agar medium.

[0041] (2) The composition of nutrient agar medium is: peptone 10.0g, beef extract 3.0g, sodium chloride 5.0g, agar 15.0g, distilled water 1000mL, pH7.4±0.2;

[0042] (3) The culture condition of bacillus is: culture temperature is 37 ℃, culture time is 12h;

[0043] (4) Inoculate the indicator bacterium Listeria monocytogenes CMCC54002 preserved in the slope into the BHI liquid medium, the culture temperature is 37°C, and...

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Abstract

The invention relates to the field of microbe screening and specifically discloses a method for rapid screening of Bacillus spp with antibacterial activity. According to the invention, high temperature treatment is carried out to rapidly eliminate competitors, agar overlaying is realized via an indication bacterium, and Bacillus spp with antibacterial activity is screened by observing an inhibition zone; and under a culture condition of 35 to 42 DEG C, the inhibition zone can be observed within 6 to 12 h of culture after agar overlaying, so high-efficiency screening of Bacillus spp with antibacterial activity is realized, the whole screening process can be completed within 48 h, screening procedures are substantially simplified, and labor intensity is mitigated. The method only utilizes a plurality of simple medium and does not need test materials like filter paper and an Oxford cup, so screening cost is low.

Description

technical field [0001] The invention relates to the field of microbial screening, in particular to a method for rapidly screening bacillus with antibacterial activity. Background technique [0002] Bacillus is a spore-forming Gram-positive bacterium that is widely distributed in the environment. During its growth, it can produce antimicrobial peptides such as subtilisin, polymyxin, and gramicidin, which are harmful to the intestinal tract or plants. Pathogenic bacteria have obvious inhibitory effects, and play a probiotic role by reducing free oxygen in the intestinal environment, promoting the growth of probiotic microorganisms such as lactic acid bacteria, and regulating intestinal immunity. Based on the broad antibacterial spectrum and high stability of the antibacterial peptides produced by Bacillus, the antimicrobial peptides produced by Bacillus are widely used in the production of feed additives and microecological preparations, and good application results have been ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12Q1/04
Inventor 周辉刘成国方俊田云葛龙
Owner HUNAN AGRICULTURAL UNIV
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