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Broad-spectrum high-temperature-resistant salmonella virulent bacteriophage and application thereof

A phage, high temperature resistant technology, applied in the direction of phage, virus/phage, medical raw materials derived from virus/phage, etc., can solve the inability to achieve the therapeutic effect of broad-spectrum antibiotics, restrictions on the wide application of antibacterial agents, easy inactivation, acid and alkali resistance and other problems, to achieve the effect of strong cracking ability, high potency and good thermal stability

Pending Publication Date: 2022-03-01
HUAZHONG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

(1) Phage is highly specific and can only lyse its susceptible host bacteria, that is, its treatment is specific and cannot achieve the therapeutic effect of broad-spectrum antibiotics, which limits the wide application of phage as an antibacterial agent
(2) It is difficult to deter

Method used

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  • Broad-spectrum high-temperature-resistant salmonella virulent bacteriophage and application thereof
  • Broad-spectrum high-temperature-resistant salmonella virulent bacteriophage and application thereof
  • Broad-spectrum high-temperature-resistant salmonella virulent bacteriophage and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] Example 1 Isolation and purification of Salmonella phage ph2-2

[0043] Sample processing: Pig anal swabs collected from slaughterhouses were soaked in normal saline overnight, centrifuged at 7000rpm for 10min, and the supernatant was sterilized through a 0.22μm microporous membrane to obtain a filtrate containing phage.

[0044] Preparation of host bacteria: Select a single colony of Salmonella paratyphi A 201007 and inoculate it in 1ml of LB broth, culture at 220r / min, 37°C for 16-18h to obtain a suspension of host bacteria, and set aside.

[0045]Phage proliferation: The host bacterium 201007 was transferred to LB medium at a ratio of 1:100, cultured at 220r / min, 37°C for 1.5-3h, until logarithmic phase. Add host bacteria, phage filtrate, and LB medium into a sterilized Erlenmeyer flask at a volume ratio of 1:1:2, mix well, and incubate at 220r / min at 37°C for 2.5-3.5h to enrich phage.

[0046] Phage isolation: After the above-mentioned phage value-proliferation sol...

Embodiment 2

[0050] The determination of embodiment 2 Salmonella phage ph2-2 potency

[0051] The phage ph2-2 stocks were diluted to 10 with PBS -1 , 10 -2 , 10 -3 , 10 -4 , 10 -5 , 10 -6 , 10 -7 , take 300uL of the host bacteria suspension in a sterile 10ml EP tube, then add the melted semi-solid agar medium to 8mL, quickly pour it into the prepared plate with the bottom layer of TSA medium, tilt and rotate the plate to make it evenly distributed After the agar solidifies, take 10uL of phage filtrate according to the dilution and spot on the plate, and after drying, incubate overnight at 37°C. Repeat the test 3 times, and take 10 when counting -7 Average of triplicate experiments for dilutions. Wherein, phage titer (PFU / ml) = average number of plaques × dilution factor × 100.

[0052] The titer of phage ph2-2 stock solution is 1.8×10 10 PFU / ml.

Embodiment 3

[0053] The determination of embodiment 3 Salmonella phage ph2-2 host spectrum

[0054] The test selects 106 strains of Salmonella isolates (see Table 1) and 8 kinds of other species strains (see Table 1) to do the host spectrum analysis of phage ph2-2, and the specific operations are as follows:

[0055] Cultivate the host bacterial strains to the logarithmic phase, and dilute the phage ph2-2 stock solution with PBS to 10 -1 , 10 -2 , 10 -3 , 10 -4 , 10 -5 , 10 -6 , 10 -7 . Take 300 μL of the above-mentioned bacterial solution in the logarithmic phase in a sterile 10ml EP tube, then add the melted semi-solid agar medium to 8mL, quickly pour it into the prepared plate with the bottom layer of TSA medium, tilt and rotate the plate to distribute Evenly, according to the dilution, take 10uL phage filtrate and spot on the plate, after drying, keep it upside down at 37°C overnight. Statistical results, calculate the spotting rate. Plaque forming rate (EOP) = measured strain...

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Abstract

The invention belongs to the technical field of animal infectious disease prevention and treatment, and particularly relates to a broad-spectrum high-temperature-resistant salmonella virulent bacteriophage and application thereof. The salmonella bacteriophage is preserved in the China Center for Type Culture Collection, and the preservation number is CCTCC NO: M20211353. The salmonella bacteriophage has a good killing effect on salmonella drug-resistant strains, also has a good treatment effect on salmonella infection, is suitable for being used as a biological agent for preventing and controlling salmonella, and can be applied to comprehensive prevention and control of salmonella.

Description

technical field [0001] The invention belongs to the technical field of animal infectious disease prevention and control, and relates to a strain of bacteriophage and its application in antibacterial aspect. Background technique [0002] Salmonella (Salmonella) belongs to Enterobacteriaceae Gram-negative bacteria, is a common food-borne pathogenic microorganisms, with many serotypes, more than 2,600 serotypes are currently known, and 26 bacterial groups and 161 serotypes have been found in my country type. Salmonella has a wide range of hosts and is widely distributed. Clinically, it is mostly manifested as sepsis and enteritis, and it can also cause abortion of pregnant animals. At present, the contamination of Salmonella in the pork supply chain is very common, accounting for about 70-80% of food-borne bacterial infections, which has caused significant economic losses to my country's aquaculture industry. In addition, Salmonella has a wide range of hosts in nature, and it ...

Claims

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Application Information

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IPC IPC(8): C12N7/00A61K35/76A61P31/04C12R1/92
CPCC12N7/00A61K35/76A61P31/04C12N2795/10121C12N2795/10132Y02A50/30
Inventor 吴斌赵梦菲彭忠谢锐华琳宋文博陈焕春
Owner HUAZHONG AGRI UNIV
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