575 results about "Pseudorabies" patented technology

The invention discloses a porcine pseudorabies virus virulent strain, and a gene deletion vaccine strain thereof and applications thereof. The porcine pseudorabies virus virulent strain is named as HeN1, the microbial preservation number of the porcine pseudorabies virus virulent strain is CGMCC NO.6656, the deleted gE gene obtaines the gene deletion vaccine strain rPRV-gE-EGFP+ on the basis of the virulent strain HeN1, and the microbial preservation number is CGMCC NO.6657. The virulent strain can be prepared into inactivated vaccine (single vaccine or combined vaccine), the gene deletion vaccine strain rPRV-gE-EGFP+ can be prepared into activated vaccine or inactivated vaccine (single vaccine or combined vaccine) and the like, so that porcine pseudorabies can be effectively prevented or cured, or the gene deletion vaccine strain rPRV-gE-EGFP+ can be prepared into a diagnosis reagent for diagnosing the porcine pseudorabies. The gene deletion vaccine strain rPRV-gE-EGFP+ has the advantages of being good in safety, high in protection efficiency, beneficial to differential diagnosis.

The invention provides a porcine pseudorabies virus gene deletion strain, a vaccine composition, and a preparation method and an application of the vaccine composition. The vaccine composition comprises an immunizing dose of an attenuated livetotivirus antigen and an inactivated totivirus antigen of the porcine pseudorabies virus gene deletion strain or its culture. The vaccine composition can effectively induce the antibody production, can effectively protect pigs, and can be used as a marking vaccine to effectively differentiate wild strains and vaccine strains.

The present invention discloses a construction of three-gene defected recombinant pseudorabies virus (PrV) strain, vaccine prepared by said strain, method for constructing said strain and method for preparing said vaccine. The described recombinant pseudorabies virus strain defects genes of TK.gE ang gI, and contains no exogenous gene. The described vacine belongs to the freeze-dried live vaccine made up by virus liquid containing said invention and gelatin. Said invented live vaccine can be inoculated on the piglet, slaughter pig and sow with farrow, and can obtain obious immune effect. Said vaccine has good biological safety, and can be used for preventing and curing pseudorabies.

The invention discloses a colloidal gold immunochromatographic test strip for detecting wild-type classical swine fever virus, which consists of water absorbent paper (1), a cellulose nitrate membrane (2), a colloidal gold pad (3), a sample pad (4) and a support (5), wherein the cellulose nitrate membrane contains a detection line which is formed by coating monoclonal antibody HQ06 of anti-classical swine fever virus E2 protein and a quality control line which is formed by coating rabbit anti-mouse IgG antibody; and the colloidal gold pad is combined with colloidal gold-labeled monoclonal antibody 6E10 of the anti-classical swine fever virus E2 protein. The test strip does not react with C-strain of classical swine fever virus, bovine viral diarrhea virus, porcine reproductive and respiratory syndrome virus, transmissible gastroenteritis virus, porcine epidemic diarrhea virus, porcine rotavirus, pseudorabies virus, porcine parvovirus and porcine circovirus type 2, and can accurately and sensitively identify the wild-type classical swine fever virus, thereby having good specificity, sensitivity and repeatability.

The invention discloses a colloidal gold immunochromatography test paper for antibodies of pseudorabies viruses of gE, gB and gD and a preparation method. The test paper comprises a sample absorption area, a gold mark probe area, an immobilization antigen and antibody area, a water absorption area and a supporting plate, and the sample absorption area, the gold mark probe area, the immobilization antigen and antibody area and the water absorption area are laid on the supporting plate and are partially overlapped in sequence. The gold mark probe area is covered with gold mark probes which are mouse anti-swine IgG antibodies marked by colloidal gold. The immobilization antigen and antibody area is provided with a detection line T1 covered with pseudorabies virus gE protein, a detection line T2 covered with pseudorabies virus gB protein, a detection line T3 covered with pseudorabies virus gD protein and a control line C covered with goat anti-mouse IgG antibodies. The test paper is fast in detection, high in accuracy, strong in specificity, easy and convenient to carry and operate and capable of being used for differential diagnosis on PRV wild poisonous infection and vaccine immunity and evaluation on the PRV vaccine immunity effect at the same time.

The invention discloses a heat resisting protective agent of a bivalent live vaccine against highly pathogenic porcine reproductive and respiratory syndrome-pseudorabies, and a preparation method thereof. The heat resisting protective agent of the bivalent live vaccine against highly pathogenic porcine reproductive and respiratory syndrome-pseudorabies is prepared by the following materials by weight: 0.8 to 1.2% of gelatin, 4 to 6% of soybean peptone, 0.5 to 1.5% of glycine, 3 to 5% of sucrose, 2 to 3% of trehalose, 1 to 2% of polyvinyl pyrrolidone, 0.5 to 1% of mannitol, 0.1 to 0.2% of vitamin E and the rest being injection water. The preparation method comprises steps of high-temperature sterilization, filtration sterilization through a microporous membrane and mixing. The heat resisting protective agent of the bivalent live vaccine against highly pathogenic porcine reproductive and respiratory syndrome-pseudorabies of the invention reduces damage on virus activity caused by various physical and chemical factors during the freeze drying process, provides good protection and effectively solves the problems that a conventional protective agent of live vaccine needs refrigerated transport and storage is inconvenient.

The invention provides a detection reagent and a method for identifying a porcine pseudorabies virus vaccine strains and a wild strain, and belongs to the field of animal pathogen detection. The detection reagent comprises two primer pairs as shown in SEQ ID NO.1-4 and probes as shown in SEQ ID NO.5-6. The invention further provides a method for identifying a porcine pseudorabies virus vaccine strain gD gene and a wild strain gE gene by using the detection reagent. The porcine pseudorabies virus vaccine strains and the wild strain can be simultaneously identified through dual fluorogenic quantitative PCR, high-flux detection on large-scale samples can be achieved, the detection reagent has the characteristics of rapidness, specificity, sensitivity, accuracy, simplicity and convenience, provides an effective tool for surveying porcine pseudorabies infection sources and tracing infection environments and disease sources, and has relatively good application value in prevention and control on porcine pseudorabies.

The invention provides a heatproof lyoprotectant for a live vaccine against pseudorabies. The heatproof lyoprotectant comprises, by weight, 3 to 10 parts of gelatin, 1 to 5 parts of trehalose, 5 to 15 parts of sucrose, 0.1 to 2 parts bovine serum albumin, 1 to 8 parts of tryptone, 2 to 10 parts of enzyme-hydrolyzed casein, 1 to 5 parts of thiourea, 0.8 to 2 parts of L-monosodium glutamate, 0.1 to 3 parts of arginine, 0.5 to 5 parts of polyvinylpyrrolidone (PVP-K30) and 0.1 to 2 parts of mannitol. The invention also discloses a preparation method of the heatproof lyoprotectant and a lyophilized vaccine prepared from the heatproof lyoprotectant. When the heatproof lyoprotectant is used for protecting the vaccine and a specific lyophilization process is employed, lyophilization loss of viruses can be effectively reduced, the temperature tolerance of the viruses can be improved, and the vaccine can still maintain good physical properties and titer after long-term storage; i.e., the vaccine has stable characters and has the characteristics of heat resistance and long storage time.

The invention discloses a preparation method of a recombinant pseudorabies virus for expressing reverse neural circuit tracing of green fluorescin with high sensitivity, and application, including (1) the preparation of the recombinant pseudorabies virus for expressing green fluorescin with high sensitivity; (2) the application to marking of a neural circuit. The recombinant pseudorabies virus for expressing the green fluorescin with high sensitivity is successfully prepared by using a platform. The recombinant pseudorabies virus for expressing reverse neural circuit tracing of green fluorescin with high sensitivity is successfully obtained. Wide application value is realized in aspects of neural circuit marking, medicine screening platform building, medicine inhibition virus action mechanism, viral vaccine and diagnostic reagent invention and development, animal model building, virus replication, pathogenic mechanism analysis and the like.

The invention discloses a porcine pseudorabies virus strain and a porcine pseudorabies inactivated vaccine prepared by using the same. The porcine pseudorabies virus strain is separated from the brain, tonsil and other tissues of a still birth of a sow on a pig farm by virtue of subculture adaptation and has a microbial preservation number of GCMCC (China General Microbiological Culture Collection Center) No.5013. The virus strain separated in the invention has quick proliferation and high titer on ST (scheduled tribe) cells, can induce an animal to generate a high-titer neutralizing antibody, and has excellent immunogenicity. The invention also discloses a preparation method of the porcine pseudorabies inactivated vaccine, and the method comprises the following steps of: culturing the porcine pseudorabies virus strain to obtain a virus solution; adding an inactivator to inactivate the virus solution; and preparing an oil phase and a water phase, and emulsifying, thus obtaining the porcine pseudorabies inactivated vaccine. In the invention, each process parameter of the inactivated vaccine is optimized. Immune protective efficacy and safety tests prove that the inactivated vaccine has excellent immunogenicity and safety.

The invention provides a porcine pseudorabies virus vaccine composition. The porcine pseudorabies virus vaccine composition contains a porcine pseudorabies virus subunit antigen, or a recombinant Newcastle disease virus, namely a porcine pseudorabies virus vector. The invention further provides the preparation method and application of the vaccine composition. The vaccine composition can effectively prevent related diseases of a porcine pseudorabies virus and related diseases infected by the porcine pseudorabies virus. The combination of immunogenicity antigens in the porcine pseudorabies virus vaccine composition can be induced to generate a synergetic immune effect, thereby having good immune effect, and further lowering the immune usage amount to lower the immunization cost.

The invention relates to a method for industrially producing a pseudorabies vaccine by using a bioreactor. The method comprises the following steps of: (1) sterilizing a microcarrier and the bioreactor, inoculating cells for vaccine production, culturing the cells, and after the cells on the microcarrier form a compact monolayer, inoculating pseudorabies virus, and continuously culturing to allow the virus to reproduce; (2) after more than 80 percent of cells are subjected to a cytopathic effect, stopping culturing, and obtaining virus liquid to prepare the pseudorabies vaccine. In the method, the pseudorabies vaccine is produced by culturing the cells in high density by the bioreactor microcarrier culture technology; and compared with the conventional roller bottle production process, the invention has the advantages that: the method has high automation control degree, saves manpower, reduces cost and is small in production land, the production can be monitored in real time, the scale is easy to expand, the produced virus has high titer and small difference among batches, and the product has stable quality and small side effect.

The present disclosure provides vaccine compositions comprising a PRRSV vaccine and a second porcine vaccine, which are substantially free from immuno-inhibition against each other. The second porcine virus vaccine can be CSFV and/or PRV. The preparation methods for the vaccines and the formulations are also provided. The vaccine compositions provided herein confer protective immunity to pigs against porcine reproductive and respiratory syndrome, classical swine fever, and/or pseudorabies.

The invention discloses a method for generating a porcine pseudorabies virus by culturing an ST cell in a microcarrier of a bioreactor. The method comprises the following steps: (1) selecting the ST cell as a cell line for making vaccine; (2), transferring and culturing the ST cell; (3) reproducing a porcine pseudorabies virus cell virus seed; (4) culturing the ST cell in the microcarrier of the bioreactor in a suspension way; (5) reproducing porcine pseudorabies virus liquid; and (6) treating the obtained virus liquid. The method has the advantages that the cell vitality is more than and equal to 90%, the living cell density is more than and equal to 1*10<7> cells/mL, and the virus liquid tissue culture infectious dose 50 (TCID50) is more than and equal to 10<8.0>. The production process is intensive and smooth, the dimension is easy to amplify, the production period is short, a small space is occupied, little pollution is caused to the environment and is easy to treat, the virus liquid is convenient to obtain, the quality is easy to balance and stabilize, the production cost can be reduced obviously, and the yield and the quality of the vaccines are improved.

The invention discloses a primer, probe and assay kit used for detecting porcine circovirus, porcine pseudorabies virus and porcine parvovirus. The primer and probe used for detecting are shown as sequences from SEQIDNO:1 to SEQIDNO:9 in a sequence table. The invention also provides an assay kit used for detecting the porcine circovirus, the porcine pseudorabies virus and porcine parvovirus. The primer and the probe which are adopted by the invention have strong specificity, and three viruses can be detected in one step, thus cost is saved, steps are reduced, and efficiency is improved. The method provided by the invention has the characteristics of high specificity, high sensitivity, high efficiency and low cost and is applicable to analysis of a larger number of samples.

The invention discloses a kit for detecting pig pseudorabies. The kit comprises an assay plate coated with pig pseudorabies virus gE antigen, a rabbit anti-pig IgG antibody marked by HRP (horseradish peroxidase), fast coating buffer, fast blocking buffer, sample diluents, TMB substrate, stop buffer, 20 times wash solution, negative control, and a positive control. Through adopting pig pseudorabies virus specificity gE antigen with high purity and high activity expressed through gene engineering to coat the assay plate, and through adopting HRP-conjugate of rabbit anti-pig IgG monoclonal antibody containing HRP, the kit for detecting pig pseudorabies, provided by the invention, has the advantages of strong specificity, high sensibility, less susceptibility to misjudge of false positive or false negative and the like; moreover, the kit is simple in structure, low in detection cost, convenient and fast in operation and strong in timeliness, has a wide market prospect, and can create larger social benefits.

The invention discloses a heat-resisting protective agent for freeze-dried vaccine and a preparing method and application thereof. The heat-resisting protective agent is prepared from, by mass, 0.3-0.6% of L-sodium glutamate, 8-14% of saccharose, 2-5% of D-sorbitol, 1-3% of bovine serum albumin, 0.1-0.3% of polyvinylpyrrolidone, 0.2-1% of thiourea, 3-5% of NZ-Amine AS, 0.8-1.5% of hydrolyzed gelatin and the balance PBS with PH of 7.2. The heat-resisting protective agent for freeze-dried vaccine can be applied to freeze-drying of porcine pseudorabies live vaccine and porcine reproductive and respiratory syndrome live vaccine after being subjected to asepsis treatment. By adopting the same heat-resisting protective agent and freeze-drying technology for the two viruses, freeze-drying loss is lower than 0.2 titer, virus content titer is lower than one titer after the viruses are stored at 37 DEG C for seven days, and the viruses can be stored for 24 months at 2-8 DEG C; convenience is brought for production of freeze-dried vaccine, and product quality and competitiveness are improved.

The invention relates to a pig pseudorabies virus natural low-virulent C strain and a heatproof preservation method thereof. The microbe collection registry number of the strain is CCTCC NO: V201114. According to the invention, various cryoprotectant substrates are mixed and blended according to a certain ratio; the cryoprotectants are respectively subject to aseptic processing; cryoprotectant components that can be subject to high-pressure sterilization are dissolved in bi-distilled water, and are sterilized for 15-30min under a temperature of 108-121 DEG C; cryoprotectant components that can not be subject to high-pressure sterilization are dissolved in bi-distilled water according to a certain formula, and are sterilized by using a filter membrane with a size of 0.22mum; all the cryoprotectant components are then mixed into a heatproof cryoprotectant; the heatproof cryoprotectant is mixed with a pseudorabies virus liquid according to a ratio of 1:1-1.2; and the mixture is lyophilized with a corresponding lyophilization curve. Compared to prior arts, the C strain provided by the invention has good stability. In a fifth generation animal of continuous back-procreation, the genetic sequence is stable, and no mutation is found. Therefore, the strain provided by the invention provides a good resource for the researches of pig pseudorabies low-virulent vaccines. The strain can bepreserved for 24 months under a temperature of 2-8 DEG C, and the virus titer is reduced by no more than 1 titer.

The invention relates to a fluorescence quantitative PCR rapid detection kit used for specifically detecting pig pseudorabies virus (PRV) infection, and an application thereof. The kit comprises a specific primer pair used for amplifying PRV gE gene conserved region, and a specific TaqMan fluorescent-labeled probe. An upstream primer sequence of the specific primer pair is 5'-GAGGACGACGGGCTGTAC-3', and a downstream primer sequence of the specific primer pair is 5'-GGACATCAACAGGCGGTTG-3'. The sequence of the TaqMan probe is 5'-TGGGTCCATTCGTCACTTCCG-3'. The 5' end of the TaqMan probe is labeled with a fluorescent reporter group FAM. The 3' end of the TaqMan probe is labeled with a fluorescence quenching group BHQ1 which is not intrinsically fluorescent. The kit can be used for rapid qualitative and quantitative detections of PRV infection, and can be used in identification detections of PRV gE gene-deleted vaccine strains and/or PRV wild strains.