Staphylococcus aureus phage and bacteriostatic application thereof

A staphylococcus, golden yellow technology, applied in the field of microorganisms, can solve the problems of narrow host spectrum, poor temperature and pH stability, etc., and achieve the effect of good pH stability, good temperature stability and high potency

Active Publication Date: 2021-11-09
SHANDONG AGRICULTURAL UNIVERSITY +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Some Staphylococcus aureus phages have been disclosed in the prior art, but many of these phages have limitations such as narrow host spectrum, poor temperature and pH stability, etc.

Method used

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  • Staphylococcus aureus phage and bacteriostatic application thereof
  • Staphylococcus aureus phage and bacteriostatic application thereof
  • Staphylococcus aureus phage and bacteriostatic application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0045] Example 1 Isolation, preparation and purification of phage

[0046] 1.1. Source of the strain

[0047] The host bacteria came from Staphylococcus aureus (107) isolated from milk samples submitted for mastitis in dairy cows in Shandong Province;

[0048] 1.2. Sewage source:

[0049] Staphylococcus aureus was detected in a mixed sample of fecal sewage and milking pond sewage around a dairy farm in Qufu;

[0050] 1.3. Main reagents:

[0051] LB broth, LB semi-solid medium, LB solid medium, SM buffer solution;

[0052] Main instruments: biological safety cabinet, electric heating incubator, constant temperature shaker, low temperature high-speed centrifuge, JEM-2100Plus transmission electron microscope, dynamic microplate reader.

[0053] 2. Method

[0054] 2.1. Enrichment of phage

[0055] The collected mixed sewage was centrifuged at 12000rpm for 15min to remove impurities, and the supernatant was drawn with a syringe. If the collected sewage contains a large amoun...

Embodiment 2

[0070] 2.7. Determination of phage titer

[0071] The purified phages were serially diluted 10 times, and 100 μl of each gradient concentration was taken, and the double-layer plate was coated according to the method in 2.2, and the appropriate concentration gradient plate was selected to calculate the number of phage plaques. Each concentration gradient was repeated three times, and the average value was finally calculated to calculate the phage titer.

[0072] Phage titer (PFU / mL) = average number of plaques × dilution factor × 10

[0073] The purified phages were serially diluted 10 times, and the number of plaques in each concentration gradient was calculated by the double-layer plate method. After multiple parallel experiments, the titer of s107 phage was 2.31×10 10 PFU / mL.

Embodiment 3

[0075] 2.8. Lysis ability of bacteriophage

[0076] The microplate phage virulence test was used to evaluate the lytic activity of phage on the host, that is, the lytic ability. Cultivate the host bacteria to the logarithmic growth phase (approximately 1×10 8 CFU / mL), dilute with broth, so that the final concentration of inoculated bacteria is 1×10 6 CFU / mL. The phage liquid was serially diluted 10 times, and each dilution concentration was co-inoculated with 100 μl of the host bacteria into a 96-microwell plate, and the multiplicity of infection was 0.001, 0.01, 0.1, 1, 10, 100, and 1000. Culture in air at 37°C 6-10h. Visually inspect the turbidity of the microwells and measure the absorbance at OD600, and record the highest dilution that completely dissolves the bacteria.

[0077] The MOI calculation method of phage-host experiment: the number of phage per well / the number of bacteria per well. Phage sensitivity is divided into: very sensitive (10 -6 ≤MOI-2 ); highly su...

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Abstract

The invention relates to the technical field of microorganisms, and particularly relates to a Staphylococcus aureus phage and bacteriostatic application thereof. The Staphylococcus aureus phages107 is sent to China Center for Type Culture Collection for Preservation, the classification name is Staphylococcus aureus phages107, the preservation number is CCTCC NO: M2021125, the preservation date is January 21, 2021, and the address is Wuhan University, Wuhan City, China. The phages107 has high titer up to 2.31 * 10<10>PFU / mL, good pH stability, good temperature stability and wide lysis spectrum, and has lysis ability on 9 strains of Staphylococcus aureus, wherein the 9 strains of Staphylococcus aureus have certain drug resistance and no lysis ability on escherichia coli. The Staphylococcus aureus phage has specific lysis spectrum for Staphylococcus aureus, and has strong lysis ability. The Staphylococcus aureus phages107 can be used for preparation of drugs for treating Staphylococcus aureus induced dairy cow mastitis and other diseases.

Description

technical field [0001] The invention relates to the technical field of microorganisms, in particular to a Staphylococcus aureus phage and its antibacterial application. Background technique [0002] The information disclosed in this background section is only intended to increase the understanding of the general background of the present invention, and is not necessarily taken as an acknowledgment or any form of suggestion that the information constitutes the prior art already known to those skilled in the art. [0003] Staphylococcus aureus is ubiquitous in nature and can be found in air, water, dust, and human and animal excrement, so food is often contaminated by Staphylococcus aureus, especially dairy products, frozen foods and fresh meat Classes, etc. are more susceptible to contamination. [0004] In the prior art, the prevention and treatment of Staphylococcus aureus is mainly treated with antibiotics. Due to some bad effects caused by the abuse of antibiotics, peopl...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/00A01N63/40A01P1/00C11D3/38A23K10/18A61K35/76A61P31/04C12R1/92
CPCC12N7/00A01N63/40C11D3/381A23K10/18A61K35/76A61P31/04C12N2795/00021C12N2795/00031Y02A50/30
Inventor 闫振贵张雪丽王亚东于浩淼张亮陈九张云飞杨宏军
Owner SHANDONG AGRICULTURAL UNIVERSITY
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