Salmonella bacteriophage with wide lysis spectrum and high temperature resistance and application thereof

A Salmonella and phage technology, 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 strong The effect of cracking performance and good high temperature resistance

Active Publication Date: 2021-09-24
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 wide-lysed Salmonella phage RDP-SA-20049, which aims to solve the problems of ac...

Method used

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  • Salmonella bacteriophage with wide lysis spectrum and high temperature resistance and application thereof
  • Salmonella bacteriophage with wide lysis spectrum and high temperature resistance and application thereof
  • Salmonella bacteriophage with wide lysis spectrum and high temperature resistance and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] Embodiment 1 Isolation and identification of pathogenic bacteria and bacteriophage

[0035] (1) Isolation and identification of pathogenic Salmonella BS-20036

[0036] Sampling was taken from a broiler chicken farm in Hebei, the liver of the diseased poultry was taken aseptically, and a line was drawn on the selective medium (SS agar). For colonies with black spots in the center with a smooth and wet surface, pick typical colonies and continue to streak and purify them for 3 times, so that the size and character of the colonies are basically the same, then pick a single colony and inoculate it in 5mL LB broth, and culture it with shaking at 37°C and 200rpm for 6h , to obtain a uniform turbid bacterial suspension. Then through 16sRNA molecular identification and serotype identification, it was determined to be pathogenic Salmonella, and one of them was named BS-20036 and stored in a -80°C refrigerator.

[0037] (2) Phage isolation and identification

[0038] Sample pr...

Embodiment 2

[0043] Electron microscope observation of embodiment 2 phage

[0044] Take 20 μL of the liquid containing crude phage particles and drop them on the copper grid, let it settle naturally for 15 minutes, and 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 minutes, and then use the filter paper to remove the The staining solution was sucked off from the side, and the morphology of the phage was observed with an electron microscope after the sample was dried.

[0045] Depend on figure 1 It can be seen that the head of the phage RDP-SA-20049 is icosahedral, the long diameter is about 61nm, the transverse diameter is about 67nm, the tail is about 112nm long, and the diameter is about 10nm; The phage is classified in the family Myoviridae of the order Cauviridae.

Embodiment 3

[0046] The drawing of embodiment 3 phage growth curves

[0047] Take the Salmonella suspension in the logarithmic growth phase, inoculate Salmonella BS-20036 and bacteriophage RDP-SA-20049 according to the ratio of the optimal multiplicity of infection, put it in a water bath at 37°C for 10 minutes, then centrifuge at 12000r / min for 5 minutes at room temperature, discard The supernatant was used to remove the free phage not adsorbed on the host, and thus washed twice with LB broth. The pellet was then resuspended in LB broth medium at 37°C, quickly placed in a shaker at 37°C at 200 r / min for shaking culture and timed. Sampling and counting every 5 minutes for the first 30 minutes, counting every 20 minutes for 30-60 minutes, counting every 30 minutes for 60-300 minutes, taking time as the abscissa and phage titer logarithm as the ordinate, draw the growth curve to obtain the incubation period and lysis of the phage period, and calculate the average lysis amount. Average lysi...

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Abstract

The invention discloses a salmonella bacteriophage with a wide lysis spectrum and high temperature resistance. The preservation number is CGMCC No.21995. After whole genes are sequenced, the salmonella bacteriophage does not have virulence genes and lyogenic genes. The salmonella bacteriophage RDP-SA-20049 acts for 60 min at the high temperature of 90 DEG C, the titer is kept at 1.0 * 10 < 7 > pfu/mL or above, and the salmonella bacteriophage RDP-SA-20049 has high biological activity and good high temperature resistance. The salmonella bacteriophage can be used for lysing pathogenic bacteria including 5 strains of salmonella typhimurium, 4 strains of salmonella pullorum, 1 strain of salmonella paratyphi B, 1 strain of salmonella typhimurium, 7 strains of salmonella enteritidis and salmonella BS-20036, and has the characteristic of high cracking rate, the cracking rate is up to 99%, the cracking performance is stronger, and the salmonella bacteriophage has good advantages in treatment of salmonella infection diseases, and is high in titer and good in prevention and treatment effect.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a strain of Salmonella phage with wide lysis spectrum and high temperature resistance 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 c...

Claims

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Application Information

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IPC IPC(8): C12N7/00A23K10/18A61K35/76A61P31/04A01N63/40A01P1/00A01P3/00C11D3/38C12R1/92
CPCC12N7/00A23K10/18A61K35/76A61P31/04A01N63/40C11D3/381C12N2795/10121C12N2795/10131C12N2795/10132Y02A50/30
Inventor 杜新永崔耐朱建君李先胜马如霞张得彦刘爽
Owner RECOM QINGDAO BIOTECH CO LTD
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