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Salmonella bacteriophage resistant to antiviral drugs and application of salmonella bacteriophage

An antiviral drug and Salmonella technology, applied to viruses/phages, medical raw materials derived from viruses/phages, phages, etc., can solve the problems of bacterial resistance, untimely treatment, and ineffective medication, etc. Good tolerance and titer stability

Active Publication Date: 2021-07-23
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 strain of Salmonella phage RDP-SA-20032 resistant to antiviral drugs, aiming to solve the problem of acute onset of salmonella infection in farm poultry and untimely treatment, and drug resistance caused by pathogenic bacteria. Invalid and other issues

Method used

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  • Salmonella bacteriophage resistant to antiviral drugs and application of salmonella bacteriophage
  • Salmonella bacteriophage resistant to antiviral drugs and application of salmonella bacteriophage
  • Salmonella bacteriophage resistant to antiviral drugs and application of salmonella bacteriophage

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0033] Embodiment 1 Isolation and identification of pathogenic bacteria and bacteriophage

[0034] 1. Isolation and identification of pathogenic Salmonella BS20133

[0035] Sampling from the diseased farm, aseptically take the liver of diseased poultry, streak it on the selective medium (SS agar), culture it at 37°C for 18-24 hours, and form a round, flat, neat edge and smooth surface on the medium Wet black colonies, pick typical colonies and continue to streak and purify 3 times, then pick a single colony and inoculate in 5mL LB broth, shake and culture at 200rpm at 37°C for 8h 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 the strains was named BS20133 and stored in a -80°C refrigerator.

[0036] 2. Isolation and identification of Salmonella phage RDP-SA-20032

[0037] (1) Manure treatment: Take manure from the farm, weigh 5g of chicken man...

Embodiment 2

[0043] Morphological observation of embodiment 2 bacteriophage

[0044] Using phosphotungstic acid negative staining method: drop phage liquid (titer 10 10 pfu / m L) 100μL, put the film side of the copper mesh on the phage droplet, take it off after 10min, let it dry naturally in the air for 2-3min, then drop a drop of 2% phosphotungstic acid (PTA) on the copper mesh The aqueous solution was stained, removed after 10 minutes, dried in the air for 10-15 minutes, observed with an electron microscope, and a clear phage image was selected for photographing. photo by electron microscope figure 1 It can be seen that the bacteriophage has a polyhedral stereosymmetric head, wrapped in nucleic acid, about 65-40nm in diameter, has a tail about 100nm long, has a tail sheath, and the neck is connected to the head and tail. Classified as Tauroviridae.

Embodiment 3

[0045] The mensuration of embodiment 3 phage thermostability

[0046] Dispense the phage stock solution into 50 ml sterilized Erlenmeyer flasks, incubate at 0°C, 30°C, 40°C, 50°C, 60°C, 70°C, 80°C and 90°C for 30min and 60min respectively, and then use After 10-fold serial dilution of normal saline, spread double plates to determine its titer; the results are as follows: figure 2 shown.

[0047] Depend on figure 2 It can be seen that the titer of bacteriophage RDP-SA-20032 is stable within 60°C and remains at 10 11 About pfu / mL; when treated at 60°C for 30min and 60min, the phage titer only decreased by 1 order of magnitude; when incubated at 80°C for 30min and 60min, it was still active, indicating that the phage has good high temperature resistance.

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Abstract

The invention discloses a salmonella bacteriophage resistant to antiviral drugs. The preservation number of the salmonella bacteriophage is CGMCC No.21994. After sequencing is carried out on whole genes, the salmonella bacteriophage does not have virulence genes and lyogenic genes. After the salmonella bacteriophage RDP-SA-20032 is treated for 60 minutes at 60 DEG C, the titer is still kept at 10<9> pfu / mL, and the salmonella bacteriophage RDP-SA-20032 has better tolerance to temperature; the titer is still kept at 10<11>pfu / mL when the bacteriophage is cultured for 1 hour at 37 DEG C in a pH range of 4-9, and the bacteriophage still survives when the pH is 12 or 2, so that the acid-base tolerance range of the bacteriophage is relatively wide; the salmonella bacteriophage has good titer stability under the coexistence condition of the salmonella bacteriophage and antiviral drugs; and after being redissolved, the phage spray-dried powder is still stable in titer after being kept at 4 DEG C for 72 hours, can tolerate higher temperature, and can be produced by adopting a conventional spray drying tower, the production efficiency is improved, and the production cost is reduced.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a Salmonella phage resistant to antiviral drugs and its application in poultry animal pathogenic Salmonella infection. Background technique [0002] Salmonella (Salmonella) is a kind of non-spore straight bacteria parasitic in the intestinal tract of humans and animals. It has more than 2500 serotypes, many of which can be cross-infected between animals and humans, and most of the serotypes have very Strong pathogenicity has caused great harm to the livestock and poultry industry and humans. Due to the abuse of antibiotics in the poultry industry, the drug resistance of Salmonella is getting stronger and stronger, and the multi-drug resistant strains are increasing. According to the different types of antibiotics used, the drug resistance spectrum of Salmonella is also constantly changing. In recent years, drug-resistant Salmonella has become the main pathogenic bacteria of chicken ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/00A61K35/76A61P31/04A23K10/18A23K50/75A01N63/40A01P1/00C12R1/92
CPCC12N7/00A61K35/76A61P31/04A23K10/18A23K50/75A01N63/40C12N2795/10321C12N2795/10332C12N2795/10331Y02A50/30
Inventor 杜新永张得彦高启兴李先胜马如霞刘玉庆张庆
Owner RECOM QINGDAO BIOTECH CO LTD
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