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A brucellosis protection strain and preparation method and application thereof

A technology for brucellosis and brucellosis, applied in the field of brucellosis protective strains and its preparation, can solve safety problems, weak immune protection S-type live vaccines, interference with field virus infection antibody diagnosis, etc. problems, achieve good safety and improve immune protection

Active Publication Date: 2022-07-22
CHINA INST OF VETERINARY DRUG CONTROL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] The S-type live vaccine has a better immune effect, but the immune antibody will interfere with the antibody diagnosis of wild virus infection, and there are certain safety problems; the R-type live vaccine can distinguish infection and immunity, which involves the diagnosis of LPS antibody and is safe. Improved, but less protective than S-type live vaccines

Method used

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  • A brucellosis protection strain and preparation method and application thereof
  • A brucellosis protection strain and preparation method and application thereof
  • A brucellosis protection strain and preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0051] Embodiment 1, the preparation of Brucella RS bivalent vaccine

[0052] 1. Preparation of Brucella RM227

[0053] 1. Inducer induction experiment

[0054] (1) R-type Brucella mutagenesis medium

[0055] Dissolve 0.1 g of the inducer acriflavine (purchased from sigma) in 100 mL of sterilized distilled water (1 mg / mL), which is the stock solution, which is stored at room temperature for future use. Dissolve 15g of TSB in 500mL of distilled water, sterilize by autoclaving at 121°C for 15min, wait for it to cool to room temperature, add 0.5μL of the above prepared stock solution, place it on a constant temperature shaker at 37°C, and take it out for use after 1 day, it is R-type Brucella Mutagenesis medium.

[0056] (2) Inducer-induced strain

[0057] Take 10 mL of R-type Brucella mutagenesis medium, inoculate a single colony of Brucella spp. Rev.1, place it on a constant temperature shaker at 37°C for 3 days, double-dilute it, and culture a single colony for colony crys...

Embodiment 2

[0107] Embodiment 2, RS bivalent vaccine mouse immunization

[0108] 1. Preparation of inducible strain RM227, deletion strain Rev.1-ΔwadC and Rev.1 bacterial solution

[0109] Culture the inducible strain RM227 and the deletion strain Rev.1-ΔwadC, and after counting, the deletion strain Rev.1-ΔwadC and Rev.1 strains were diluted with normal saline to 10 4 , 5×10 4 , 10 5 , 5×10 5 , 10 6 CFU / mL, the inducer strain RM227 was diluted to 10 11 CFU / mL, set aside.

[0110] 2. Grouping and immunization of mice

[0111] 120 BALB / C mice were divided into 12 groups, 10 mice in each group. The mice were immunized by subcutaneous injection in the groin, each 100 μL, and the specific immunization doses were matched according to Table 3.

[0112] Table 3 Mice grouping and immunization dose

[0113]

[0114]

[0115] Note: S monovalent vaccine represents the smooth type parental strain Rev.1 immune group; R monovalent vaccine represents the rough type induced strain M227 immu...

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Abstract

The invention discloses a brucellosis protection strain. The strain is obtained by random mutation deletion gene of S-type Brucella herba Rev.1 or deletion of wadC gene. The present invention induces S-type Brucella species Rev.1 random mutation deletion gene to obtain rough (R) type Brucella RM227 with good genetic stability; the homologous recombination deletion technology is adopted from S-type sheep species Brucella Bacteria Rev.1 obtained the smooth (S) type Brucella Rev.1‑ΔwadC lacking the wadC gene and having good genetic stability. The present invention also provides a brucellosis RS bivalent vaccine, comprising rough (R) type Brucella RM227 and smooth (S) type Brucella Rev.1-ΔwadC in a certain concentration ratio, with low immunity The mouse experiments prove that the RS bivalent vaccine of the present invention not only has good immune protection and safety, but also can realize the differential diagnosis between natural infection antibodies and lipopolysaccharide (LPS) antibodies involved in immunity.

Description

technical field [0001] The present invention relates to the technical field of brucellosis protection strains, in particular to a brucellosis protection strain and a preparation method and application thereof. Background technique [0002] Brucellosis, referred to as brucellosis, is a zoonotic infectious disease caused by Brucella bacteria, mainly infecting livestock. The World Organization for Animal Health (OIE) lists it as a variety of animal diseases, and my country lists it as a second-class animal disease. [0003] The source of infection related to human brucellosis in my country is mainly sick sheep and cattle. Infected animals can excrete pathogenic bacteria from milk, feces and urine, and carry bacteria for a long time or for life. When a sick female animal aborts, a large number of pathogens are excreted along with the aborted fetus, afterbirth and uterine secretions, becoming the most dangerous source of infection. [0004] Brucellosis vaccine is an important m...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/20C12N1/21C12N15/01C12N15/74C12N15/90C12N15/31A61K39/10A61K39/116A61P31/04C12R1/01
CPCC07K14/23C12N15/01C12N15/74C12N15/902A61K39/098A61P31/04A61K2039/52A61K2039/552A61K2039/70C12R2001/01C12N1/205
Inventor 王楠徐磊孙浩杰毛开荣辛凌翔丁家波蒋卉任小侠
Owner CHINA INST OF VETERINARY DRUG CONTROL
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