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A highly lytic Klebsiella pneumoniae phage rdp-kp-20007 and its application

A technology of RDP-KP-20007, Klebsiella pneumoniae bacteriophage, applied in bacteriophage, virus/phage, medical raw materials derived from virus/phage, etc. properties, etc., to achieve good application prospects and strong cleavage activity.

Active Publication Date: 2022-08-02
RECOM QINGDAO BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The purpose of the present invention is to provide a highly lytic Klebsiella pneumoniae phage RDP-KP-20007 to replace the application of antibiotics in the prevention and treatment of poultry breeding, and to solve the problem of acute onset of poultry animals and untimely treatment, and the Drug resistance leading to drug ineffectiveness and other problems

Method used

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  • A highly lytic Klebsiella pneumoniae phage rdp-kp-20007 and its application
  • A highly lytic Klebsiella pneumoniae phage rdp-kp-20007 and its application
  • A highly lytic Klebsiella pneumoniae phage rdp-kp-20007 and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0033] Example 1 Isolation and identification of pathogenic Klebsiella pneumoniae BK-20007

[0034]Samples were taken from the livers of chickens with Klebsiella pneumoniae bacterial disease, inoculated in DHL medium by aseptic operation, and cultured at 37°C for 18-24h. Klebsiella pneumoniae forms pale pink colonies on DHL agar plates, the large surface is raised, smooth and moist, and it is mucus-like.

[0035] Select pale pink colonies for continuous streak purification three times for biochemical identification. Transfer from DHL medium to Klebsiella disaccharide medium and culture at 37℃ for 18-24h, the slant surface produces acid, the bottom layer produces acid and gas, and it is negative for hydrogen sulfide. Biochemical identification showed that V-P test was positive, nitrate reduction test was positive, M.R was negative, Simon's citrate utilization test was positive, malonate test was positive, urease was positive, lysine decarboxylase was positive, ornithine decarb...

Embodiment 2

[0037] Example 2 Isolation and identification of Klebsiella pneumoniae phage RKP-KP-20007

[0038] The liver samples used in the experiments of the present invention were collected from sick chickens in a broiler farm in Jimo, Qingdao in 2020, as samples for phage isolation.

[0039] (1) Isolation of phage RKP-KP-20007:

[0040] Take 10㎎ of liver and soak it in 10ml LB broth at 4°C overnight, centrifuge at 10,000rpm for 5min, and filter the supernatant to sterilize it with a 0.22μm microporous membrane. Take 3ml of the filtrate and 3ml of the host bacteria culture solution in logarithmic growth phase, add them to 20ml of autoclaved LB broth, and place them in a 37°C incubator overnight to culture to proliferate possible phages. Take 3 ml of the sample culture solution, centrifuge at 10,000 rpm for 5 min, and take the supernatant to filter and sterilize it with a 0.22 μm microporous membrane. Assume that the above filtrate contains bacteriophages. Use LB broth for gradient d...

Embodiment 3

[0046] Example 3 Morphological observation of bacteriophage

[0047] Using phosphotungstic acid negative staining method: 20 μL of phage proliferation solution (titer 10 11 pfu / mL) was dropped on the paraffin sheet, and the copper mesh was placed on it for 10 min and then removed to adsorb phage. Dry at room temperature for 2-3 min, stain with 2% phosphotungstic acid aqueous solution, absorb water after 2 min, dry naturally for 5 min, and observe the phage morphology by electron microscope.

[0048] Photo by electron microscope ( figure 2 ), it can be seen that the head of phage RKP-KP-20007 is icosahedral and is three-dimensionally symmetrical. The length of the head is 72.640 nm, and the length of the phage tail is 177.797 nm. .

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Abstract

The invention discloses a highly lytic Klebsiella pneumoniae bacteriophage RKP-KP-20007 and an application thereof. Its host is Klebsiella pneumoniae. Observed through an electron microscope, the head of the bacteriophage is an icosahedron and is three-dimensionally symmetrical. The length of the head is 72.640nm, and the length of the tail of the phage is 177.797nm. Klebsiella pneumoniae bacteriophage RKP‑KP‑20007 has a strong lytic effect on Klebsiella pneumoniae in poultry breeding environment, can effectively treat Klebsiella pneumoniae disease caused by this pathogen in poultry breeding, and can be used as a disinfectant It provides a phage source for industrialized, large-scale and programmed production of phage and the treatment of bacterial diseases in poultry breeding caused by Klebsiella pneumoniae.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a highly lytic Klebsiella pneumoniae bacteriophage and its application in the infection of Klebsiella pneumoniae in poultry breeding. Background technique [0002] Klebsiella pneumoniae, belonging to the Enterobacteriaceae Klebsiella genus, is a common Gram-negative bacterium that exists widely in various environments in nature, and is a very common food-borne pathogen. Klebsiella pneumoniae mainly infects humans and animals, can cause high mortality, and cause significant economic losses to aquaculture around the world. Klebsiella pneumoniae-contaminated animal products (such as meat, eggs, milk, etc.) transmit the pathogen to humans through the food chain, thereby causing harm to human and animal health. It can produce sepsis, pneumonia and urinary tract infection, and in severe cases, it can cause human and poultry morbidity and death. It is one of the important opportunistic pat...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N7/00A61K35/76A61P31/04A61P11/00A23K10/18A23K50/75A01N63/40A01P1/00C11D3/48C11D3/38C12R1/92
CPCC12N7/00A61K35/76A61P31/04A61P11/00A23K10/18A23K50/75A01N63/40C11D3/48C11D3/381C12N2795/10321C12N2795/10332C12N2795/10331
Inventor 杜新永王晓铃马如霞李先胜崔耐
Owner RECOM QINGDAO BIOTECH CO LTD
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