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A wide lytic spectrum Salmonella phage rdp-nsa-19050 and its application

A technology of Salmonella and phage, which is applied in the direction of phage, virus/phage, medical raw materials derived from virus/phage, etc., can solve the problems of ineffective medication, drug resistance of pathogenic bacteria, untimely treatment, etc., and achieve high lysis rate and high efficacy The effect of high price and broad cracking spectrum

Active Publication Date: 2022-04-22
RECOM QINGDAO BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The purpose of the present invention is to provide a strain of wide lysing Salmonella phage RDP-NSA-19050, which aims to solve the problems of acute onset of salmonella infection in farm poultry and untimely treatment, and ineffective medication due to drug resistance of pathogenic bacteria.

Method used

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  • A wide lytic spectrum Salmonella phage rdp-nsa-19050 and its application
  • A wide lytic spectrum Salmonella phage rdp-nsa-19050 and its application
  • A wide lytic spectrum Salmonella phage rdp-nsa-19050 and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] Example 1 Isolation and Identification of Pathogenic Bacteria and Phage

[0028] (1) Isolation and identification of pathogenic Salmonella NKS-50

[0029] Sampling was made from a white-feathered broiler chicken in Jimo Tianheng Broiler Farm, Qingdao. The liver of the diseased poultry was taken aseptically, and a line was drawn on the selective medium (SS agar). After culturing at 37°C for 18-24 hours, circles formed on the medium. Shaped, flat, with neat edges, smooth and moist surface, black center and transparent edges, pick typical colonies and continue to streak and purify for 3 times, then pick a single colony and inoculate in 5mL LB broth, shake and culture at 200rpm at 37°C for 6h to get A homogeneous turbid bacterial suspension. Through 16sRNA molecular identification and serotype identification, it was determined to be pathogenic Salmonella, named NKS-50, and stored in a -80°C refrigerator.

[0030] (2) Phage isolation and identification

[0031] Feces trea...

Embodiment 2

[0036] Example 2 Electron Microscopic Observation of Phage

[0037] Take 20 μL of the liquid containing crude phage particles and drop it on the copper grid, let it settle naturally for 15 min, absorb the excess liquid from the side with filter paper, add a drop of 2% phosphotungstic acid (PTA) on the copper grid to stain the phage for 10 min, and then use The staining solution was sucked from the side of the filter paper, and the morphology of the phage was observed with an electron microscope after the sample was dried.

[0038] Depend on figure 1It can be seen that the head of the phage is icosahedral, with a long diameter of about 60nm, a transverse diameter of about 70nm, a tail length of about 100nm, and a diameter of about 10nm; Myoviridae of the order Myoviridae.

Embodiment 3

[0039] Example 3 Determination of optimal multiplicity of infection

[0040] Add the phage proliferation solution and the host to the LB broth at the ratio of 10:1, 1:1, 1:10, 1:100, 1:1000, 1:10000 at the multiplicity of infection, and ensure that the total volume of the culture system is the same. After shaking and culturing at 200rpm at 37°C for 8h, centrifuge at 12000r / min for 5min at room temperature, take the supernatant and spread it on double plates to determine its titer. The results are shown in Table 1.

[0041] pipe number NKS50 Salmonella count (cfu / mL) RDP-NSA-19050 phage count (pfu / mL) multiplicity of infection 8h phage titer (pfu / mL) 1 1×10 8

[0042] It can be seen from the data in Table 1 that when the MOI is 1:10, the proliferation fluid is relatively clear and the titer is the highest after culturing for 8 hours, indicating that the optimal MOI of RDP-NSA-19050 is 1:10.

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Abstract

The invention discloses a Salmonella phage RDP‑NSA‑19050 with a wide lysis spectrum, the preservation number is CGMCCNo.21415, and the gene length is 87591bp. After sequencing the whole gene, it is found that it does not have virulence genes and lysogenic genes. The invention provides The Salmonella phage RDP‑NSA‑19050 can lyse a variety of species including Stanley, Salmonella gallinarum typhi, Salmonella typhimurium, Salmonella paratyphi A, Salmonella paratyphi B, Salmonella typhi, Salmonella enteritidis, and Salmonella Kentucky. Pathogenic bacteria have the characteristics of wide lysis spectrum and high lysis rate. The phage can be used alone or in combination with other Escherichia coli, Salmonella phages and probiotics, which can effectively reduce the mortality of chickens infected with Salmonella.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a broadly lytic Salmonella phage RDP-NSA-19050 and its application in pathogenic Salmonella infection of poultry animals. Background technique [0002] Salmonellosis is one of the important zoonotic diseases in public health, and it is a common foodborne pathogen. Chicken is the most important host of Salmonella. Salmonella can cause acute or chronic diseases in chickens. Chickens infected with Salmonella can also lead to vertical transmission of Salmonella by producing eggs with bacteria, thus forming a vicious circle. When salmonellosis breaks out in chicken flocks, the mortality rate can reach 80%, causing huge economic losses to the poultry industry. Once a person ingests animal food containing a large amount of Salmonella, it can lead to food-borne poisoning. According to statistics, among the types of bacterial food poisoning in various countries in the world, food poisoning c...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N7/00A61K35/76A61K45/06A61P31/04A23K10/18A23K20/195A23K50/75C11D3/38C11D3/48A01N63/40A01P1/00C12R1/92
CPCC12N7/00A61K35/76A61K45/06A61P31/04A23K10/18A23K20/195A23K50/75C11D3/381C11D3/48A01N63/40C12N2795/10121C12N2795/10132C12N2795/10131A61K2300/00
Inventor 杜新永李先胜罗成盛赵丹丹崔耐张庆刘玉庆盖春云马如霞王立坤刘爽
Owner RECOM QINGDAO BIOTECH CO LTD
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