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Broad-spectrum phage for rapidly cracking livestock and poultry escherichia coli and application

A technology for Escherichia coli and colibacillosis, applied in the field of broad-spectrum phages, can solve the problems of lack of cross-immunity, restriction of antibiotics or antibacterial drug use effects, etc., and achieve high fermentation efficiency and wide cracking spectrum

Pending Publication Date: 2021-11-05
武汉观海生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] Due to the different dominant serotypes of Escherichia coli prevalent in different regions, and the lack of cross-immunity among the serotypes, it brings challenges to vaccine prevention and control
Factors such as changes in bacterial resistance and their potential threats to humans also restrict the use of antibiotics or antimicrobials

Method used

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  • Broad-spectrum phage for rapidly cracking livestock and poultry escherichia coli and application
  • Broad-spectrum phage for rapidly cracking livestock and poultry escherichia coli and application
  • Broad-spectrum phage for rapidly cracking livestock and poultry escherichia coli and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0022] Isolation and Purification of Escherichia coli Phage GRH-ECP052

[0023] Eight waste samples were collected from a large farmer’s market in Wuhan, Hubei. Each sample was resuspended and mixed with 20 mL of sterile PBS. After centrifugation at 5000 rpm for 10 min, the supernatant was filtered and sterilized. The filtrate was mixed with the same volume of 2×TSB liquid medium and 1mL of Escherichia coli in the logarithmic phase (10 8 cfu / mL) were evenly mixed, and cultured overnight at 180 rpm at 37°C to enrich the phage. The sample enrichment solution was centrifuged at 5000 rpm for 10 min, and the supernatant was sterilized through a 0.22 μm microporous membrane to obtain a filtrate containing phage. Take 100uL of the filtrate and mix it evenly with 300uL of the host Escherichia coli liquid, and let it stand for 15min to fully bind with the receptors on the surface of the bacteria. Add the above mixed solution to 10mL of TSB semi-solid agar medium cooled to 50°C, sprea...

Embodiment 2

[0026] Determination of Cleavage Profile of Escherichia coli Phage GRH-ECP052

[0027] Take the potency to be about 1.0x10 8 PFU / mL stock solution of Escherichia coli phage GRH-ECP052, using the drop method to determine the lysis profile of the phage.

[0028] Pick 571 strains of avian Escherichia coli and 385 strains of porcine Escherichia coli isolated from different regions of the country, inoculate them in centrifuge tubes filled with 3mL TSB, and incubate them at 37°C and 180rpm for 8h to prepare bacterial suspensions of each strain. Take 300uL of the bacterial suspension and mix it with the TSB semi-solid culture medium and spread it on the prepared TSA plate, and take 10uL of the phage GRH-ECP052 culture solution and drop it on the plate. After natural air-drying, culture at 37°C for 8-12 hours, and observe the results. The results are shown in Table 1. Coliphage GRH-ECP052 can lyse 557 strains of 571 avian E. coli strains, with a lysis rate of 97.5%; it can lyse 364 ...

Embodiment 3

[0033] Lysis Rate Determination of Escherichia coliphage GRH-ECP052

[0034] Take the potency to be about 5.0x10 8 PFU / mL stock solution of Escherichia coli phage GRH-ECP052, using the drop method to determine the lysis efficiency of the phage.

[0035] Pick single clones of Escherichia coli, inoculate them in centrifuge tubes filled with 3mL TSB, and incubate at 180rpm at 37°C for 8h to prepare bacterial suspensions of each strain. Take 300uL of the bacterial suspension and mix it with the TSB semi-solid culture medium and spread it on the prepared TSA plate, and take 10uL of the phage GRH-ECP052 culture solution and drop it on the plate. After natural air-drying, culture at 37°C, and take out every 0.5 hours to observe the lysis results. At the same time, phage GDCP17 (CCTCC NO: M2020664) was used as a control.

[0036] The results are shown in Table 2, coliphage GRH-ECP052 began to appear cracking circle at 2h, and the cracking circle was completely transparent after 3h ...

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Abstract

The invention belongs to the technical field of agricultural microorganism application, and particularly relates to a broad-spectrum phage for rapidly cracking livestock and poultry pathogenic escherichia coli and application. The escherichia coli phage GRH-ECP052 which is rapid in cracking and wide in cracking spectrum is obtained through screening, the phage is preserved in the China center for type culture Collection, and the preservation number is CCTCC NO: M20211009. The phage has a good prevention and treatment effect on poultry colibacillosis and piglet yellow-white dysentery.

Description

technical field [0001] The invention belongs to the field of biotechnology and is related to the field of biological prevention and control. The invention specifically relates to a broad-spectrum phage that rapidly lyses Escherichia coli of livestock and poultry and its application. Background technique [0002] Escherichia coli is an opportunistic pathogenic bacterium that is normally considered part of the normal intestinal flora. Once the body's resistance decreases, some highly pathogenic strains can cause gastrointestinal infections in humans or animals, as well as urinary tract infections, arthritis, meningitis, and sepsis. With the vigorous development and promotion of large-scale and intensive livestock and poultry breeding in my country in recent years, the economic impact of pathogenic Escherichia coli on livestock and poultry breeding has become increasingly obvious, and even threatens the healthy and sustainable development of animal husbandry. [0003] Avian co...

Claims

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Application Information

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IPC IPC(8): C12N7/00A61K35/76A61P31/04A61P1/12C12R1/92
CPCC12N7/00A61K35/76A61P31/04A61P1/12C12N2795/00021C12N2795/00032Y02A50/30
Inventor 王喜亮黄金梅李越薛素强田甲张秀玲付咏堂徐岳张晓东金秀娥
Owner 武汉观海生物科技有限公司
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