A Salmonella Phage with Wide Lysis Spectrum and Its Application

A technology of Salmonella and Salmonella enteritidis, applied in the direction of phage, virus/phage, medical raw materials derived from virus/phage, etc., can solve the problems affecting the treatment effect of infected patients, achieve good application development prospects, suitable for popularization and application, and high potency high effect

Active Publication Date: 2022-04-26
HUAZHONG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

According to statistics, Salmonella is highly resistant to streptomycin, sulfisoxazole, tetracycline, kanamycin, gentamicin, ceftriaxone, and ampicillin, and multidrug-resistant strains continue to appear in clinical cases. The increase will seriously affect the treatment effect of infected patients, which makes the prevention and control of salmonellosis face severe challenges

Method used

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  • A Salmonella Phage with Wide Lysis Spectrum and Its Application
  • A Salmonella Phage with Wide Lysis Spectrum and Its Application
  • A Salmonella Phage with Wide Lysis Spectrum and Its Application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0044] Example 1 Isolation and purification of Salmonella phage WXLSGP006

[0045] Centrifuge 30mL of fecal sewage samples from a chicken farm in Hubei at 8000r / min at 4°C for 10min, take 20mL of the supernatant and use filter paper for preliminary filtration to remove insoluble large particles of impurities and bacteria in the sample, and then use 0.22 The obtained filtrate was filtered and sterilized again by a microporous membrane of μm, and 5 mL of the filtrate was mixed with 20 mL of 2×TSB liquid medium and 2 mL of Salmonella HNSM2 bacterial liquid in the logarithmic phase (10 8 cfu / mL) were evenly mixed, and cultured at 37°C 180r / min for 14-18h to enrich the phage.

[0046] Centrifuge the sample enrichment solution at 8000r / min for 10min, take the supernatant and pass it through a 0.22μm microporous membrane, and observe whether there are phages targeting the host bacteria in the sample enrichment solution by dot spot method. The specific steps are: mix 200 μL of Salmon...

Embodiment 2

[0047] Example 2 The biological characteristics of Salmonella phage WXLSGP006

[0048] 1. Phage Morphological Characteristics

[0049] Phage particles were observed under a transmission electron microscope, as figure 1 As shown, the head of the phage has a regular polyhedral structure, the length of the head is 134.2nm, the transverse diameter is 126.6nm, and the length of the tail is about 232.6nm. The phage is preliminarily identified as a long-tailed phage family.

[0050] 2. Phage forms plaques on the double-layer plate

[0051] like figure 2 As shown, the Salmonella phage WXLSGP006 produced large circular plaques on the Salmonella lawn, with a bright center and a halo around the edge, with a diameter of about 5-6 mm.

Embodiment 3

[0052] Example 3 Salmonella bacteriophage WXLSGP006 to the determination of Salmonella optimum multiplicity of infection (MOI)

[0053] Salmonella pullorum HNSM2 was inoculated in 5 mL of TSB medium at a 1% inoculum amount, and cultured in a shaker at 37° C. with shaking at 200 r / min to the logarithmic phase to obtain a suspension of the host bacteria. The titer of the obtained bacterial suspension was detected by colony plate count after appropriate dilution. According to the set multiplicity of infection ratio (MOI=0.00001, 0.0001, 0.001, 0.01, 0.1, 1, 10, 100), add 100 μL of pure culture solution of phage WXLSGP006 (prepared by Example 1) and 100 μL of host bacterial suspension, and then An equal amount of TSB liquid medium was added to make the total volume of each experimental group the same. Cultivate in a shaker at 37°C at 180r / min for 3.5h, centrifuge the obtained culture solution at 8000r / min for 10min, collect the supernatant and pass it through a 0.22μm microporous...

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Abstract

The invention belongs to the field of biotechnology, and in particular relates to a Salmonella phage with a wide splitting spectrum and its application. The bacteriophage is a potent bacteriophage, has a strong lytic effect on Salmonella, and belongs to the family Long-tailed bacteriophage; the bacteriophage can form large, translucent plaques on solid medium with a halo around it; It can survive under the condition of ℃~70℃; cultured for 12h under the condition of MOI=0.001, its titer is 1.2×10 10 pfu / mL. The Salmonella phage provided by the invention has high titer and good safety, can be used alone or in combination, and has strong lytic activity against Salmonella with multiple serotypes or multiple drug-resistant phenotypes; Products and means of salmonella contamination. The bacteriophage can provide an excellent source of bacteriophage for the development of bacteriophage therapy, has good application and development prospects, and is suitable for popularization and application.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a Salmonella phage with a wide lysis spectrum and its application. Background technique [0002] Salmonella is an important zoonotic pathogen that can infect livestock and poultry and contaminate meat, eggs, vegetables and other foods through excretion, childbirth, egg production, etc., causing human foodborne diseases. According to statistics, 70%-80% of foodborne diseases in my country are caused by Salmonella. Due to the large number of Salmonella serotypes and their wide distribution in nature, it has brought great difficulties to the prevention and control of Salmonellosis. In recent years, the abuse of antibiotics in animal husbandry has led to the emergence of drug resistance in many bacteria, and the problem of Salmonella drug resistance has become increasingly serious. According to statistics, Salmonella is highly resistant to streptomycin, sulfisoxazole, tetra...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N7/00A61K35/76A61P31/04C12R1/92
CPCC12N7/00A61K35/76A61P31/04C12N2795/10321Y02A50/30
Inventor 王喜亮李越赵虹泽袁旭吕佩琳邰蓉金秀娥
Owner HUAZHONG AGRI UNIV
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