High-lysis klebsiella pneumoniae bacteriophage RDP-KP-20007 and application thereof

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: 2021-11-02
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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  • High-lysis klebsiella pneumoniae bacteriophage RDP-KP-20007 and application thereof
  • High-lysis klebsiella pneumoniae bacteriophage RDP-KP-20007 and application thereof
  • High-lysis klebsiella pneumoniae bacteriophage RDP-KP-20007 and application thereof

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 livers of chickens with Klebsiella pneumoniae disease, inoculated in DHL medium by aseptic operation, and cultured at 37°C for 18-24h. Klebsiella pneumoniae forms pale pink colonies on the DHL agar plate, with a large raised surface, smooth and moist, in the form of mucus. Adjacent colonies are easy to fuse into a thick juice, and long filaments can be pulled out when the inoculation needle is picked.

[0035] Select the pale pink colonies to streak and purify three times in a row for biochemical identification. Transplanted from DHL medium to Klebsiella medium and cultured at 37°C for 18-24h, the slant surface produced acid, the bottom layer produced acid and gas, and the hydrogen sulfide was negative. Biochemical identification showed positive V-P test, positive nitrate reduction test, negative M.R, positive Simon's citrate utilization test, positive ma...

Embodiment 2

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

[0038] The liver sample used in the experiment of the present invention was collected from a sick chicken in a broiler chicken farm in Jimo, Qingdao in 2020, and used as a sample for phage isolation.

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

[0040] Take 10㎎ of the liver and soak it in 10ml LB broth overnight at 4°C, centrifuge at 10,000rpm for 5min, and filter the supernatant with a 0.22μm microporous membrane to sterilize. Take 3ml of the filtrate and 3ml of the host bacterium culture solution in the logarithmic growth phase, add them together to 20ml of autoclaved LB broth, put them in a 37°C incubator, and culture them overnight to multiply the possible phages. Take 3ml of the sample culture solution, centrifuge at 10000rpm for 5min, and take the supernatant to filter and sterilize with a 0.22μm microporous membrane. Assume that the above filtrate contains phages. Th...

Embodiment 3

[0046] Morphological observation of embodiment 3 phage

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

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

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Abstract

The invention discloses a high-lysis klebsiella pneumoniae bacteriophage RKP-KP-20007 and application thereof, the host of the high-lysis klebsiella pneumoniae bacteriophage RKP-KP-20007 is klebsiella pneumoniae, observation through an electron microscope shows that the head of the bacteriophage is an eicosahedron and is in stereo symmetry, the length of the head is 72.640 nm, the length of the tail of the bacteriophage is 177.797 nm, the bacteriophage is a long-tail bacteriophage family with the tail, and the preservation number is CGMCC No. 22495. The Klebsiella pneumoniae bacteriophage RKP-KP-20007 has a strong cracking effect on Klebsiella pneumoniae in a poultry breeding environment, can be used for efficiently treating Klebsiella pneumoniae diseases caused by pathogenic bacteria in poultry breeding, and can be used as a disinfectant. A bacteriophage source is provided for industrial, large-scale and programmed production of bacteriophages and treatment of poultry breeding bacterial diseases caused by klebsiella pneumoniae.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a highly lytic Klebsiella pneumoniae phage and its application in poultry breeding Klebsiella pneumoniae infection. Background technique [0002] Klebsiella pneumoniae, belonging to the Enterobacteriaceae Klebsiella genus, is a common Gram-negative bacterium that widely exists in various environments in nature and is a very common foodborne pathogen. Klebsiella pneumoniae mainly infects humans and animals, can cause high mortality, and has caused significant economic losses to the aquaculture industry worldwide. Klebsiella pneumoniae-contaminated animal products (such as meat, eggs, milk, etc.) transmit pathogens to humans through the food chain, thereby endangering the health of humans and animals. It can produce septicemia, pneumonia and urinary tract infection, and cause death in humans and poultry in severe cases. It is one of the important conditional pathogenic bacteria and pa...

Claims

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Application Information

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Patent Type & Authority Applications(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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