A Salmonella phage resistant to high temperature and wide cracking spectrum and composition thereof

A Salmonella and phage technology, applied in the field of phage, can solve the problems of Salmonella phage lysis range and limited tolerance to high temperature, and achieve good high temperature resistance, strong lysis, and wide host range.

Active Publication Date: 2022-02-08
PHAGELUX (NANJING) BIO-TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The above-mentioned existing technical solutions have the following defects: the range of cracking of the above-mentioned Salmonella phage and the tolerance to high temperature are all limited, and it is urgent to provide a Salmonella phage with a wide cracking spectrum of high temperature resistance

Method used

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  • A Salmonella phage resistant to high temperature and wide cracking spectrum and composition thereof
  • A Salmonella phage resistant to high temperature and wide cracking spectrum and composition thereof
  • A Salmonella phage resistant to high temperature and wide cracking spectrum and composition thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0057] Example 1: Screening and purification of phage

[0058] (1) Sample collection and processing

[0059] The samples in the present invention are collected from farm sewage and nearby farmland soil in Shandong, Henan and Jiangsu.

[0060] The collected samples were centrifuged at 5000r / min for 10min and passed through a 0.22μm filter membrane.

[0061] (2) Enrichment of phages targeting Salmonella in samples

[0062] Mix the above filtrate with 2 times LB liquid medium at a ratio of 1:1, inoculate 100 μL of the target Salmonella strain, and enrich overnight.

[0063] (3) Phage screening

[0064] Centrifuge the above-mentioned enrichment solution to get the supernatant and pass it through a 0.22μm membrane, mix 1mL with 5mL LB semi-solid medium containing the target Salmonella, pour it on a petri dish containing LB solid medium, and wait for the semi-solid medium to solidify Afterwards, culture overnight at 37°C, observe whether there are plaques the next day, and recor...

Embodiment 2

[0068] Example 2 Salmonella phage SG8P3 (Salmonella pullorum phage SG8P3) to the determination of the optimal multiplicity of infection (MOI) of Salmonella

[0069]1. Phage counting method: the obtained Salmonella phage SG8P3 (Salmonella pullorum phageSG8P3) sample is diluted in a 10-fold ratio, and 100 μL of a sample of a certain dilution ratio is taken, and double layers are laid according to the method in step (four) described in Example 1 For the plate, take an appropriate proportion of the plate to count the number of plaques, and one plaque represents one phage monomer.

[0070] 2. Pick a single colony of Salmonella, inoculate it into a test tube filled with 3 mL of LB culture solution, culture it with shaking at 150 rpm at 37°C for 12 hours, and obtain a suspension of the host bacteria. The bacterial suspension was transferred to 10 mL LB culture medium at a ratio of 1:100, and cultured at 37°C with shaking at 150 rpm to the pre-logarithmic phase. Add the pure culture ...

Embodiment 3

[0074] Embodiment 3: The virulence gene of Salmonella phage SG8P3 or the detection test of bad gene deletion select 65 kinds of virulence genes (table 2) that are derived from lysogenic phage in pathogenic bacteria through identification, by measuring the whole genome of Salmonella phage SG8P3 and It undergoes bioinformatics analysis to determine whether it contains the above-mentioned virulence genes. The results showed that Salmonella phage SG8P3 did not contain the following virulence genes. The tested phages had no adverse genes.

[0075] Table 2 The main known virulence genes of lysogenic phages in pathogenic bacteria

[0076]

[0077]

[0078] The Salmonella phage SG8P3 of the present invention does not contain virulence genes or adverse genes, wherein the absence of virulence genes or adverse genes refers to the virulence genes or adverse genes recorded in Table 2.

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Abstract

The invention relates to the field of phage technology, in particular to a Salmonella phage with high temperature resistance and wide cracking spectrum and a composition thereof. Described Salmonella phage is Salmonella phage SG8P3 ( Salmonella pullorum Phage SG8P3). The Salmonella bacteriophage SG8P3 of the present invention has good high temperature resistance characteristics, 65 ℃ of water baths 2h still have 80% survival rate, 75 ℃ of lower water baths 2h have 10 6 PFU/mL titer. At a concentration of 10 2 ~10 3 cfu/mL Salmonella medium, 10 4 ~10 6 PFU/mL of Salmonella phage SG8P3 can sterilize more than 99% of the above-mentioned concentrations of Salmonella, and has a broad-spectrum bactericidal ability against Salmonella; the composition of Salmonella phage SG8P3 can crack more Salmonella, and the cleavage rate can reach more than 98%, with Stronger disintegration.

Description

technical field [0001] The invention relates to the field of phage technology, in particular to a high-temperature-resistant and wide-split-spectrum Salmonella phage and its composition, kit and application. Background technique [0002] Salmonella generally colonizes the intestines of animals. If it is not handled properly in food processing, it can easily contaminate food, cause food safety hazards, and endanger human health. In recent years, with the abuse of antibiotics by farmers and feed manufacturers, more and more drug-resistant strains of the pathogen have emerged, which has rendered numerous antibiotics for the treatment of Salmonella infection ineffective. According to reports, the resistance of Salmonella in China to sulfonamide antibiotics is close to 100%, and people have to seek other effective prevention and control methods against this bacterial infection. [0003] Phages are a class of viruses that obligately parasitize or infect bacteria, and they are wid...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N7/00A61K35/76A61P31/04A01N63/40A01P1/00A23K10/18A23K50/75C12R1/92
CPCC12N7/00A61K35/76A61P31/04A01N63/40A23K10/18A23K50/75C12N2795/10121C12N2795/10132C12N2795/10131A61K2300/00
Inventor 黄杰费文斌胡怿林丁良肖逍何四龙陈海刘墨乔欢丛郁谢晓莉徐旭凌
Owner PHAGELUX (NANJING) BIO-TECH CO LTD
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