Salmonella phage preparation and preparation method thereof

A Salmonella, phage technology, applied in the field of bioengineering

Active Publication Date: 2018-03-09
OCEAN UNIV OF CHINA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there is no report on the optimal method of fermentation preparation for phage maturation at home and abroad.

Method used

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  • Salmonella phage preparation and preparation method thereof
  • Salmonella phage preparation and preparation method thereof
  • Salmonella phage preparation and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0055] Embodiment 1: Salmonella phage STP4-a optimum liquid propagation condition optimization

[0056] (1) Determination of optimal culture time for liquid proliferation of Salmonella phage STP4-a

[0057] Add 2 mL of overnight cultured ST14028 (bacteria count 10) to 200 mL NB 9 cfu / mL) and 2mL phage STP4-a (titer 10 9 pfu / mL), set up three parallel groups, shake culture at 37°C 150r / min, take a sample every 2h, centrifuge to remove bacterial sediment, measure its potency by double-layer plate method, the results are shown in figure 1 .

[0058] Depend on figure 1 It can be seen that when phage STP4-a was cultured for 4 hours, its titer had reached 10 9 pfu / mL, especially when the culture time reaches 8h, the titer has basically stabilized at 4×10 9 pfu / mL, by analysis of variance, the titer of phage STP4-a had no significant difference at 8h and 12h (P>0.05), and was significantly higher than that at 0, 2, 4, and 6h (P<0.05). Therefore, the optimal culture time of phag...

Embodiment 2

[0081] Embodiment 2 Salmonella phage STP4-a fermenter test

[0082] (1) Fermentation preparation: Taking the 5L fermentation system as an example, the prepared CaCl 2 After the NB is fully stirred and mixed, it is poured into the tank of the fermenter with a liquid volume of 3L (the maximum liquid volume of the fermenter), and then an appropriate amount of defoamer is added to eliminate the bubbles generated during the fermentation process. Place the body of the fermenter with the culture medium in a high-pressure steam sterilizer at 121°C for 15 minutes, and cool it down before use. The host bacterium ST14028 (bacteria solution concentration is about 10 9 cfu / mL) 30mL (the inoculation volume is 1% of the fermentation volume) and 30mL of bacteriophage STP4-a (the inoculation volume is 1% of the fermentation volume) were successively added into the fermenter. The flame inoculation method is used for inoculation. A certain amount of alcohol is added to the alcohol groove aroun...

Embodiment 3

[0089] The in vitro antibacterial application effect of embodiment 3 phage STP4-a

[0090]The standard strain of Salmonella typhimurium ATCC 14028 preserved in the laboratory and the single colony of Salmonella isolated from sick chickens in chicken farms were inoculated into the nutrient broth medium, cultured overnight at 37°C, and the bacterial solution was dipped with a sterile cotton swab After that, evenly spread it on the nutrient agar plate, drop 10 μL of phage STP4-a on each plate, and after the droplet is dry, place it upside down in a 37°C incubator and cultivate overnight, and observe whether a transparent circle is formed, thereby judging the lysis of the phage To determine the cleavage profile of phage STP4-a.

[0091] Table 4 shows the bacteriostatic effect of phage STP4-a on bacteria isolated from diseased chickens. It shows that phage STP4-a has lytic properties to 10 isolated strains of Salmonella, while it is lytic to Escherichia coli, Klebsiella pneumoniae,...

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Abstract

The present invention relates to a phage preparation, especially a Salmonella phage preparation and a preparation method thereof, and belongs to the field of bioengineering. According to the present invention, phage with a preservation number of CCTCCNO: M 2014145 adopts Salmonella as host bacteria to ferment so as to prepare the Salmonella phage preparation; the optimal Salmonella phage STP4-a liquid proliferation conditions such as the culture temperature, the time, the pH value, the ion concentration, the rotation speed, the inoculation amount and the like are explored so as to determine the optimal culture conditions, and the progressive enlargement culture is performed to determine the optimal phage preparation conditions; and the preparation can be separately used or used in a compounding manner, can specifically inactivate a variety of salmonella, and provides a safe and no toxic-side effect phage product source for current prevention and control of Salmonella contamination.

Description

technical field [0001] The invention relates to a phage preparation, in particular to a Salmonella phage preparation and a preparation method thereof, belonging to the field of bioengineering. Background technique [0002] Salmonella (Salmonella) is a Gram-negative straight bacillus, which is an important zoonotic pathogen. Studies at home and abroad have shown that foodborne diseases caused by Salmonella have repeatedly topped the list. With the extensive and long-term use of antibiotics, the increasingly serious drug-resistant Salmonella in food and the environment has affected the treatment of human Salmonella infection. Whether to develop new biological antibacterial agents to replace chemical antibiotics is currently a challenge for researchers. a challenge. Phage is a type of bacterial virus. After a virulent phage infects a bacterium, it can rapidly lyse the bacterium and lead to the death of the host bacterium. Salmonella phage can specifically infect and lyse Sal...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N7/00A61K35/76A61P31/04
CPCY02A50/30
Inventor 王静雪林洪李梦哲李萌
Owner OCEAN UNIV OF CHINA
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