75 results about "Brucella Vaccine" patented technology

The invention relates to a kit for recognizing a Brucella wild strain and vaccine strains A19 and S2. The kit comprises a special primer pair SEQ ID No.1 and SEQ ID No.2 for recognizing the Brucella vaccine strain A19 and a special primer pair SEQ ID No.3 and SEQ ID No.4 for recognizing the Brucella vaccine strain S2. The kit is used for conducting further sequencing analysis on a PCR amplification product after the Brucella and other conventional bacterial strains are distributed on a PCR amplification-electrophoresis detection area, and the Brucella A19 and S2 vaccine strains in a clinic sample can be rapidly and effectively recognized and diagnosed according to the sequencing result.

The invention relates to a kit for identifying a Brucella S2 vaccine strain and a wild strain. The kit comprises a specific primer pair SEQ ID No.1 and SEQ ID No.2 for identifying the Brucella vaccine strain S2. Sequencing analysis is further performed on a PCR amplification product after the kit is used for distributing Brucella and other conventional bacterial strains through a PCR amplification-electrophoresis detection area, and the Brucella S2 vaccine strain in a clinic sample can be rapidly and effectively identified and diagnosed according to the sequencing result.

Indirect ELISA of deer's brucellosis belongs to the veterinary field and is used for diagnosing deer's brucellosis to improve the diagnosis level of the prior art. Biomaterials used in the assay of the invention comprise pig brucella vaccine 2 (S2) as an antigen, an enzyme labeled antibody rabbit anti-deer IgG-HRP, a standard positive serum, a standard negative serum and a serum to be assayed; and the assay comprises the following steps: coating the antigen, washing, adding the serum, adding the rabbit anti-deer IgG-HRP, adding a substrate liquid, terminating a reaction, testing by an enzyme-linked detector, adjusting to zero at 490nm, testing the OD value of each hole, and determining the result is positive when the OD value is equal to or greater than 0.32. According to the invention, the S2 is used as the antigen, the rabbit anti-deer IgG-HRP is used as the enzyme labeled antibody, and the indirect ELISA is used to diagnose the deer's brucellosis, so compared with previous diagnosis methods, the indirect ELISA of the invention has the advantages of good specificity, sensitivity and repeatability, and accurate diagnosis.

The invention relates to a Brucella vaccine, in particular to the molecular marker and virulence gene deletion of Brucella vaccine strain. The study uses luciferase modified gene (Luc NF plus) to replace the partial fragment of Bp26 gene of Brucella attenuated vaccine S19 strain by constructing suicide plasmid and adopting the method of targeted homologous recombination (gene targeting), so as to damage the expression of the immunogenicity protein BP26 and construct the gene deletion mutant strain Delta S19-1 of the Brucella Bp26. The BMP18 protein is one of the main virulence factors of Brucella. The invention adopts the same method to exclude the Bmp 18 gene of Delta S19-1, so as to lead the Delta S19-1 not to express the Bmp 18 protein and the Brucella virulence gene deletion mutant strain Delta S19-2 is constructed. The invention solves the problems that the conventional Brucella vaccine can not distinguish between the artificial immunization and the wild bacteria infection of people and animals, the virus is strong and the vaccine is easy to cause the illness of inoculated people and animals. The invention has important significance and practical application value of the monitoring, diagnosis, purification and all the controls of Brucella.

The invention relates to recombinant brucella expressing a VP1 gene of O-type foot-and-mouth disease virus (FMDV) and a method for producing vaccines thereof. A recombinant strain is steadily integrated into the VPI gene of an O-type FMDV Myanmar strain from a brucella S2 strain genome, simultaneously, the condition of forming an O strand in a smooth type brucella cell wall LPS structure is destructed to ensure that the recombinant strain is changed from a smooth type into a rough type, the safety of the strain is further improved but a good immune effect on brucellosis is still kept, wherein the recombinant strain is named a recombinant brucella rS2-Mya strain. The recombinant strain can express the VP1 protein of the O-type FMDV Mya strain and induce to generate corresponding antibodies, and has good fundamental immunization effect on an O-type FMD. When the strain is used for producing the vaccines, the existing condition that brucella vaccine immune animals are difficult to distinguish from wild strain infected animals is changed, simultaneously the cellular immunity of FMD vaccines is realized, and good vaccines for preventing and controlling the brucellosis and the FMD are provided.

The invention relates to brucella and a production method of a brucella vaccine. In the vaccine, a recombinant brucella rS2-deltaWboA strain is taken as a production strain. A smooth recombinant strain is transformed into a rough recombinant strain by deleting base sequences among WboA genes 1-897bp of a brucella S2 strain and eliminating the forming condition of an O chain in a smooth brucella cell wall LPS (Lipopolysaccharide) structure with a non-resistant gene screening technology. The safety of the rough recombinant strain is improved remarkably, and a good immune effect on brucelliasis is still kept. Due to the adoption of the strain to the production of the vaccine, the current situation that a brucella vaccine immune animal cannot be distinguished from a wild strain infected animal easily is changed, and the safety of the conventional vaccine is effectively improved.

The invention discloses a method to prepare antigen protein of abortion brucellosis subunit vaccine and usage, which comprises the following steps: abstracting bacteria total DNA from abortion brucellosis; using the total DNA as mold; adopting property primer; proceeding polyose chain reaction; cloning; getting OMP25 protein gene; transferring into pronucleus expressing carrier; getting restructuring express carrier; leading-in bacillus coli; getting reconstructing large intestine Escherichia; proceeding inducing expression; culturing reconstructing large intestine Escherichia; preparing antigen protein. This protein can be used to prepare abortion brucellosis subunit vaccine.

The invention relates to a colloidal gold test paper bar for antibody detection of sheep Brucella. In the invention, lipopolysaccharide (LPS) extracted from a Brucella vaccine strain (S2) is taken as an envelope antigen of the colloidal gold test paper bar and improves the sensitivity of detection. Monoclonal-antibody-labeled colloidal gold of a mouse anti-sheep IgGFc terminal is prepared into a gold colloid pad of the colloidal gold test paper bar for antibody detection, and nonspecific adsorption of antibody protein by SPA in the tradition is changed. The monoclonal-antibody-labeled colloidal gold is taken as a colloidal gold labeled protein, so that specificity of detection is further improved. A set of serum for controlling quality of the test paper bar is prepared, and specificity and sensitivity of the test paper bar is ensured. The test paper bar can be used for on-site sampling, is convenient and rapid to use, and is quite suitable for application to a basic farm or a basic veterinarian.

The invention relates to a monogene deletion rough-type Brucella and a production method of a Brucella vaccine. According to the recombinant strain, base sequences between 1-897bp of the WboA gene of a Brucella 2308 strain are deleted by virtue of a non-resistance gene screening technology so as to lose the conditions for the formation of O-chain in a smooth-type Brucella cell wall LPS (Lipopolysaccharide) structure, and thus the smooth-type Brucella is transformed into a rough -type Brucella. The safety of the rough-type recombinant strain is significantly improved, but a good immune effect on brucellosis is still maintained, the recombinant bovine Brucella is named Brucella r2308-delta WboA strain (CGMCC No.8885). The current situation that Brucella vaccine immune animals are hard to distinguish from wild-strain-infected animals is changed by a vaccine produced by the strain, and the safeties of existing vaccines are effectively improved.

The invention discloses a brucella vaccine applied to livestock and a method for preparing antigen protein used for same. The antigen protein used for the brucella vaccine is brucella bcsap31 protein, and in particular brucella bcsp31 restructured protein. The method for preparing the restructured protein has steps of: taking bacterium total DNA extracted from the brucella as a template, designing a pair of primers according to ribotide complete sequence of brucella bcsp31 gene to carry out polymerase chain reaction, obtaining bcsp31 protein gene by clone and then transferring the bcsp31 protein gene to a yeast expression vector to obtain a restructured expression vector, importing the obtained restructured expression vector to yeast expression bacterium to obtain restructured pichia pastoris expression bacterium, and carrying out proliferation and induced expression of the restructured pichia pastoris expression bacterium so as to obtain the required antigen protein by separation and purification.

The invention discloses an abortus Brucella vaccine recombinant strain and an application thereof in preparing vaccine. The recombinant strain is a strain obtained by deactivating capsular polysaccharide synthesis protein gene and carboxyl uridine phosphate decarboxylase gene in the abortus Brucella strain S19. The obtained recombinant strain has small virulence and can be used as vaccines to immunize animals without needs of deactivation. The inoculated animal generates no lipopolysaccharide antibody and the virulent virus strain-infected animal can be distinguished from novel vaccine strain immunized animal by serological reaction. The recombinant strain is beneficial to improving the Brucella vaccine, researching diagnosis and identification reagents and developing the Brucella vaccine and matched diagnosis and identification reagents, and has extremely important significance in deracinating and purifying the global animal Brucella.

The invention discloses a method for identifying a brucella S2 vaccine strain by dual quantitative real-time PCR (Polymerase Chain Reaction) and a complete set of reagents used for the method. The complete set of the reagents consist of a primer Bru-F1, a primer Bru-R1, a probe Bru-Probe1, a primer S2-F2, a primer S2-R2 and a probe S2-Probe2, and the nucleotide sequences of the primer Bru-F1, theprimer Bru-R1, the probe Bru-Probe1, the primer S2-F2, the primer S2-R2 and the probe S2-Probe2 are sequentially shown as SEQ ID NO: 1-SEQ ID NO: 6. Compared with other brucellosis nucleic acid detection methods, the method established by the invention directly performs qualitative and quantitative analysis and identification on the brucellosis S2 vaccine strain through fluorescence information, has the advantages of convenience, accuracy, sensitivity, specificity and the like, greatly shortens the detection time, has a great application value for prevention and control of brucellosis, and hasa wide application prospect. Epidemic situations can be controlled from the source, large-scale outbreak of the brucellosis epidemic situations is effectively prevented, and development of animal husbandry and public health safety are guaranteed.

The invention provides a method for detecting a live attenuated Brucella vaccine strain and a wild strain and an application of the method, and belongs to the field of biotechnologies. The method fordetecting the live attenuated Brucella vaccine strain and the wild strain provided by the invention designs a primer according to a missing gene of the live attenuated Brucella vaccine strain, then adopts the primer to carry out PCR amplification on DNA of a sample to be detected, and judges the sample based on detection results. By the method, the primer is designed according to the missing geneof the live attenuated Brucella vaccine strain, namely the designed primer for amplification is a part or all of the missing gene, so that the primer effectively amplifies a gene of the wild strain without amplifying a gene of the live attenuated vaccine strain, thereby accurately and effectively distinguishing the live attenuated Brucella vaccine strain from the wild strain. The method has broaduniversality, high specificity and high accuracy.

The invention discloses protective antigen Omp25c protein of brucellosis and application thereof to preparation of a brucellosis vaccine. The test result shows that compared with a wild type vaccine strain, an omp25c mutant strain has reduced toxicity, so an omp25c gene plays a role in the toxicity of the vaccine strain. The immune level and the immune protection effect of body fluid and cells induced by an omp25c deletion strain are reduced, so the omp25c promotes the immune response and the immune protection effect of the vaccine strain. Accordingly, the omp25c protein is an important protective antigen of the brucellosis and can enhance the vaccine strain-induced immune response and the immune protection effect. The brucellosis vaccine is prepared from the omp25c protein serving as an active ingredient, so the immune protection effect of the vaccine can be improved.

The invention provides a serum-based rapid brucella vaccine strain infection detection method, and relates to the technical field of protein fingerprint spectrum detection. The invention provides characteristic protein combination for identifying serum infection of a brucella vaccine strain. The characteristic protein combination is used for constructing a standard detection model of vaccine strain infected brucellosis serum through a GA model algorithm of ClinProTools software, the recognition capability of the constructed model is 100%, and the cross validation value is 99.56%. MALDI-TOF MSmass spectrum data of to-be-detected serum are classified and analyzed by utilizing the standard detection model, and whether the to-be-detected serum is infected by a brucella vaccine strain or not can be accurately judged. The method is suitable for serum detection of high-risk groups infected by Brucella vaccine strains, is high in accuracy, good in repeatability and high in flux, detection of96 samples can be completed within 4 hours, the detection cost is low and the result is reliable, and the method has the broad application prospect.

The invention provides a B.melitensis Delta UGPase and a construction method thereof and belongs to the field of novel Brucella molecular marker vaccine research. The preservation number of the B.melitensis Delta UGPase is CGMCC No.11908, and a Brucella virulent strain M16 and UDP-glucose pyrophosphorylase (UGPase) gene related to virulence are lost. Upstream and downstream homologous arm sequences of the UGPase gene are amplified by PCR to obtain two gene segments, the gene segments and suicide carrier vector pBK-CMV-SacB are connected to pBK-CMV-SacB UGPase gene-NC losing mutation carriers after fusion and double enzyme digestion, the mixture is electro-transformed to Brucella for screening culture, and the B.melitensis Delta UGPase is finally obtained. The vaccine strain can be taken as a novel brucellosis vaccine candidate bacterial strain for controlling brucellosis spreading, and a basis is laid for novel Brucella vaccine development.

The invention provides a serum-based rapid identification method for brucella vaccine strain infection and wild strain infection. The method comprises the following steps: acquiring mass spectrometricdata of human serum infected by a brucella vaccine strain and human serum infected by a wild strain; the method comprises the following steps: constructing a standard detection model of serum of a vaccine strain and a wild strain infected with brucella through a supervised neural network (SNN) model algorithm of ClinProTools software; a characteristic protein combination for identifying serum infection of a brucella vaccine strain and a wild strain is obtained, the recognition capability of the constructed model is 100%, and the cross validation value is 95.13%. MALDI-TOF MS mass spectrum data of to-be-detected serum is classified and analyzed by utilizing the standard detection model, so that whether the to-be-detected serum is Brucella vaccine strain infection or wild strain infection or not can be accurately judged. The method is high in accuracy, good in repeatability, high in flux, low in detection cost and reliable in result, and has a good application prospect.

Provided herein is a multivalent Brucella vaccine expressing at least one heterologous M. tuberculosis antigen. The vaccines described herein serve as an environmentally safe bivalent vaccine for protection against Brucella and Mycobacterium infections simultaneously. In particular, a multivalent vaccine comprising a Brucella strain transformed with a vector that expresses at least one M. tuberculosis antigen, where the M. tuberculosis antigen(s) is codon optimized for the Brucella strain is provided. In some aspects, the Brucella strain is B. abortus strain RB51 leuB and the M. tuberculosis antigen is one or more of Ag85B, Rv2660c, and ESAT6.

The invention discloses an application of lipopolysaccharide extracted from a brucella ovis vaccine strain M5 as a human brucellosis diagnosis antigen. According to the invention, the reactivity of anLPS antigen extracted from the brucella ovis vaccine strain M5 as a diagnosis antigen is obviously superior to that of an LPS antigen extracted from a brucella cattle strain S19 and a brucella pig strain S2 commonly used in a brucella antibody rapid detection kit on the market at present, therefore, the brucella ovis vaccine strain M5 can be used as a preferred source strain of a new brucella gold-labeled diagnosis LPS antigen, the extracted LPS has a good diagnosis value on human brucellosis, and a new way is provided for selection of a human brucellosis infection diagnosis antigen. The invention also finds that the antigenicity of the LPS antigen extracted from the brucella ovis vaccine strain M5 subjected to innocent treatment is not influenced.

The invention discloses an abortus Brucella vaccine strain S19 marked recombinant strain and an application thereof. The abortus Brucella recombinant strain is a strain obtained by deactivating the glucosyltransferase coding gene. The obtained recombinant strain has small virulence and can be used as vaccines to immunize animals without needs of deactivation. When the recombinant strain is used for immunizing rats, the inoculated animal generates no lipopolysaccharide antibody and the virulent virus strain-infected animal can be distinguished from novel vaccine strain immunized animal by serological reaction. The recombinant strain is beneficial to improving the Brucella vaccine, researching diagnosis and identification reagents and developing the Brucella vaccine and matched diagnosis and identification reagents, and has extremely important significance in deracinating and purifying the global animal Brucella.