84 results about "Toxoplasma gondii Infections" patented technology

The invention provides a kit for detecting cysticercus cellulosae, swine trichinella and swine toxoplasmosis. The kit comprises sterile deionized water, polymerase chain reaction (PCR) liquid, thermus aquaticus deoxyribonucleic acid (Taq DNA) polymerase, a GoldView DNA dye, bromophenol blue sample loading buffer solution, a standard substance and a reference substance. In the detection kit provided by the invention, DNA of the cysticercus cellulosae, the swine trichinella and the swine toxoplasmosis in a specimen to be detected can be detected quickly and accurately only by one reaction, and the kit can also be used for the molecular epidemiological survey and curative effect monitoring of cysticercus cellulosae, swine trichinella and swine toxoplasmosis infection. The method has simple preparation steps and is low in cost, easy to operate, time-saving and labor-saving, a large number of reagents and consumables are saved, and the work efficiency is improved. The method is high in detection sensibility, specificity and accuracy, and the false positive rate is reduced.

The present invention discloses a diagnosis antigen for Toxoplasma gondii infection and a preparation method and applications thereof, belonging to the technical field of biological veterinary drugs. An antigen GRA8 suitable for diagnosis for Toxoplasma gondii infection is screened out by using methods combining Western blot, two-dimensional electrophoresis, and MALDI-TOF-TOF tandem mass spectrometry. The antigen can be used for a variety of molecular biology diagnosis and serological diagnosis for the Toxoplasma gondii infection.

The invention provides a vaccine for preventing toxoplasma infection. The vaccine contains nucleotide sequences selected from the followings: 1) 736-2385th nucleotide sequences as shown in SEQ ID No.1, and/or 759-2411th nucleotide sequences as shown in SEQ ID No.3; 2) nucleotide sequences complementary with the nucleotide sequences as shown in 1); and 3) nucleotide sequences which perform substituting, deletion and addition modification on one or a plurality of basic groups of the nucleotide sequences as shown in 1) or 2). The vaccine has the capability of effectively preventing and controlling the infection of a toxoplasma animal model (Kunming mice), and the combination of recombinant plasmids can effectively improve the immune effect of a single-gene vaccine, therefore, plasmids pVAX-ROP5 and pVAX-GRA15 can be used as DNA (Deoxyribose Nucleic Acid) vaccine candidate antigens for preventing the toxoplasma infection; and a combined immune vaccine of a recombinant plasmid based on such two genes can be used as a cocktail composite vaccine for preventing the toxoplasma infection.

The invention provides a nucleotide sequence for prevention of toxoplasma infection. The nucleotide sequence for prevention of toxoplasma infection contains nucleotide sequences selected from (1) a nucleotide sequence as shown in SEQ ID No.1 of a sequence table, (2) a nucleotide sequence complementary with the nucleotide sequence as shown in the (1), and (3) a nucleotide sequence obtained by replacing, deleting, adding and modifying one or multiple of basic groups of the nucleotide sequence as shown in the (1) or (2) and having a function of preventing toxoplasma infection. The invention also provides a vaccine which comprises the nucleotide sequence and a medically acceptable carrier. The plasmid DNA vaccine pVAX-ROP38 can be used for effectively preventing and controlling infection of toxoplasma animal models (Kunming strain mouse), namely can be used for effectively prolonging the survival time of the mouse and reducing the worm burden of brain capsules. Therefore, the plasmid pVAX-ROP38 can serve as a DNA vaccine candidate antigen for resisting toxoplasma infection.

The invention discloses a dog toxoplasma gondii antibody indirect ELISA detection kit, and belongs to the technical field of biological inspection and quarantine. According to the dog toxoplasma gondii antibody indirect ELISA detection kit, toxoplasma gondii ROP5 recombinant protein is taken as a detection antigen. According to the dog toxoplasma gondii antibody indirect ELISA detection kit, toxoplasma gondii recombinant protein ROP5 is obtained via gene cloning and prokaryotic expression; and a detection method comprises following steps: establishment of an indirect ELISA detection method, specificity testing, sensitivity testing, and comparison with other dog toxoplasma gondii detection kits. Large-scale standardized production of an envelope antigen of the dog toxoplasma gondii antibody indirect ELISA detection kit can be realized; detection specificity is high; sensitivity is high; repeatability is high; result determination is objective; operation process is simple; and large-scale detection can be realized. The dog toxoplasma gondii antibody indirect ELISA detection kit is suitable for dog toxoplasmosis epidemiological investigation, and detection and clinical diagnosis of dog toxoplasma gondii infection; and a rapid, simple, and specific detection method is provided for diagnosis and prevention, and epidemiological investigation of dog toxoplasmosis.

The invention discloses an immunologic adjuvant for preventing toxoplasma infection. The immunologic adjuvant is characterized in that the immunologic adjuvant is recombinant plasmids pVAX-IL21-IL15 formed by fusing interleukins IL21 and IL15 by using pVAX1 plasmids as carriers, and the base sequence of the immunologic adjuvant is represented by SEQ ID NO:1. The immunologic adjuvant for preventing the toxoplasma infection has a function of obviously improving the immune effect of MIC8 genes, so that the immune effect of an immune prevention vaccine against toxoplasma can be obviously enhanced.

The invention firstly provides a nucleotide sequence for preventing toxoplasma infection. The nucleotide sequence is selected from one of the following nucleotide sequences: (1) a nucleotide sequence as shown in SEQ ID No.1 in a sequence table, (2) a nucleotide sequence which is complementary with the nucleotide sequence in (1) and (3) a nucleotide sequence which is obtained by substituting, deleting and modifying one or more basic groups of the nucleotide sequence as shown in (1) or (2) and has a function of preventing toxoplasma infection. The invention also provides a vaccine comprising the nucleotide sequence and a medically acceptable carrier. The plasmid DNA vaccine pVAX-ROP7 provided by the invention can be used for effectively preventing and controlling toxoplasma animal model (Kunming mouse) infection, namely effectively prolonging the survival time of a rat and reducing the polypide quantity of a brain cyst. Therefore, the plasmid pVAX-ROP7 can be used as a candidate antigen of DNA vaccine to prevent toxoplasma infection.

The invention provides a nucleotide sequence for preventing sheep toxoplasmosis infection, a vector, a protein, a vaccine, a preparation method and application thereof. The nucleotide sequence comprises the following nucleotide sequences: (1) nucleotide sequence shown in SEQ ID NO.1; (2) nucleotide sequence which is complementary to the nucleotide sequence shown in (1); and (3) nucleotide sequence obtained by subjecting the nucleotide sequence in the (1) or (2) to replacing, deleting and adding and modifying of one or several base groups. The invention also provides the vaccine which is prepared by that toxoplasma antigen is subjected to double-enzyme digestion and then is connected with a pET28a expression vector, and then is transformed into a BL21 (DE3) engineering bacteria, high-efficiency expression is induced, the expression product is purified to obtain soluble proteins, and finally the soluble proteins are added with 206 adjuvant to obtain the vaccine. The prepared subunit inactivated vaccine has relatively strong cell immunity and humoral immunity effect, the congenital immune response of the body can be induced, various cell inflammatory factors are secreted, and therefore the toxoplasmosis is effectively prevented.

The invention discloses two CD8+T cell dominant epitopes of VPNSSLVEN and SQFLSLSLL based on toxoplasmagondii bradyzoite specific surface antigens, wherein six CD8+T cell epitopes with H-2 restriction and high affinity are screened according to the protein sequences of the bradyzoite specific surface antigens of SAG2C, -2C and -2X; BALB/c mice are actively immunized by the screened epitope polypeptides; determinations for a lymphopoiesis level and a cell factor content, and an immune protection evaluation for epitope vaccines are performed after the immunization. The result indicates that powerful cell immunization can be generated by inducing the mice via the epitope vaccinesm, wherein the two antigen peptides of VPNSSLVEN and SQFLSLSLL are provided to be great in the capacity of stimulating T cell proliferation, as well as capable of inducing the mice to secrete protective cell factors and immune protection, being used as the epitope vaccines effectively resisting toxoplasmagondii infection, and reducing an encystations rate in the brain tissue of a host.

The invention discloses an attenuated live vaccine capable of preventing toxoplasma gondii infection. The vaccine contains a toxoplasma gondii vaccine strain with simultaneous deficiency of the orotidine-5-phosphate decarboxylase gene and the lactic dehydrogenase 1 gene, wherein the nucleotide sequence of the orotidine-5-phosphate decarboxylase gene is as shown in SEQ ID NO:1, and the nucleotide sequence of the lactic dehydrogenase 1 gene is as shown in SEQ ID NO:2. The attenuated live vaccine provided by the invention has the advantages that basically no virulence exists, the immunocompetencefor toxoplasma gondii of animals can be improved, the acute and chronic as well as congenital infection of toxoplasma gondii can be prevented, the vaccine strain cuts off the drug resistance screening label, and thus the vaccine strain has the potential of being prepared into the genetically engineered vaccine for resisting toxoplasma gondii.

The invention relates to the field of molecular biology, and provides specific primers for detecting Toxoplasma gondii. The sequence of an upstream primer in the specific primers is as shown in SEQ IDNo. 1; the sequence of a downstream primer in the specific primers is as shown in SEQ ID No. 2. The invention also provides probes for detecting Toxoplasma gondii. The sequences of the probes are asshown in SEQ ID No. 3-6. The invention further provides a kit for detecting Toxoplasma gondii. The kit comprises the primers and the probes for detecting Toxoplasma gondii. The invention further provides a chip for detecting Toxoplasma gondii. The chip comprises the probes for detecting Toxoplasma gondii. According to the invention, gene chip detection is carried out on Toxoplasma gondii by usingthe primers and the probes, and differential diagnosis can be specifically carried out on Toxoplasma gondii infection conditions.

The invention discloses an attenuated live vaccine capable of preventing toxoplasma gondii infection. The vaccine contains a toxoplasma gondii vaccine strain with deficiency of both the orotate phosphoribosyltransferase (OPRT) gene and the lactate dehydrogenase 1 (LDH1) gene, wherein the nucleotide sequence of the OPRT gene is as shown in SEQ ID NO:1, and the nucleotide sequence of the LDH1 gene is as shown in SEQ ID NO:2. According to the attenuated toxoplasma gondii live vaccine used for preventing the animal toxoplasmosis provided by the invention, for the vaccine strain, the OPRT and LDH1genes are knocked out, thus the vaccine strain has the advantages that the virulence is weak and basically no pathogenicity is caused, and the vaccine strain can be used for preventing toxoplasma gondii infection of animals, and has the potential of being prepared into a genetic engineering vaccine for resisting toxoplasma gondii.

The invention provides a nucleotide sequence for preventing infection of toxoplasmas. The nucleotide sequence contains any one group of the following nucleotide sequences: (1) a nucleotide sequence of profilin; (2) nucleotide sequences of profilin, rop16, rop18, MIC6 and CDPK3; (3) nucleotide sequences of profilin and IL15; (4) nucleotide sequences of rop16, rop18, MIC6 and CDPK3; (5) complementary sequences of the nucleotide sequences from (1) to (4); and (6) nucleotide sequences prepared through carrying out substitution, deletion, addition and modification on nucleotide sequences from (1) to (5). By virtue of a pVAX-profilin vaccine and a combined vaccine, the generation of brain cysts of the toxoplasmas can be effectively inhibited, and a relatively good immune effect is achieved.

The invention discloses an attenuated live vaccine used for preventing toxoplasma infection and application of the attenuated live vaccine. The attenuated live vaccine takes PBS as a solvent to prepare a tachyzoite suspension of a toxoplasmosis attenuative strain RHdetltaGRA17. An immune preparation for the attenuated live vaccine is established, the immunization procedure and the inoculum size of the vaccine are explored, and the animal experiment shows that the attenuated live vaccine has a stronger immunoprotection function for the acute infection, the chronic infection and the congenital infection of toxoplasmosis, the normal infection of the toxoplasmosis is prevented, meanwhile, the mother-to-fetus vertical propagation of the toxoplasmosis is effectively stopped, and the vaccine has a huge application value, and can be used for preventing the acute infection, the chronic infection and the congenital infection of toxoplasmosis.

The invention relates to a polypeptide capable of reacting specifically with an anti-Toxoplasma gondii P30 protein antibody, and comprising a peptide sequence, in which any succession of 6 contiguous amino acids exhibits at most 67 % homology with the peptide sequence of said P30 protein, identified by SEQ ID No.1, or comprising a sequence derived from said peptide sequence, and the applications of this polypeptide especially for detecting a Toxoplasma gondii infection, in a biological sample.

The invention provides a semi-nested PCR method for detecting toxoplasma gondii, which determines whether a sample is toxoplasma gondii infection positive or negative by arranging a PCR detection kit, extracting the DNA of a test sample and carrying out the semi-nested PCR amplification and the analysis of the amplified products. The method of the invention can rapidly and accurately detect the toxoplasma gondii in the test sample, and can also be used for the molecular epidemiological investigation and the efficacy monitoring of toxoplasmosis. The template DNA preparation steps of the method are simple, the cost is low, however, the conventional method needs to be processed by lysozyme, protease K, sodium dodecyl sulfate, cetyltrimethyl ammonium bromide and other reagents, the time is long and the cost is high. The method is established on the molecular biology, which can exclude the interferences of bacteria and impurity particles, thus greatly improving diagnostic accuracy and reducing false positive rate.

The invention discloses an attenuated live vaccine for preventing toxoplasma gondii infection and application of the attenuated live vaccine. The attenuated live vaccine is tachyzoite suspension, prepared by taking PBS as a solvent, of a toxoplasma gondii attenuated stain RH: delta gra17 delta npt 1. Toxoplasma gondii double gene knockout is conducted, the immune dosage form of the attenuated livevaccine is determined, the immune vaccination program and dosage of the attenuated live vaccine are verified, it is found that the attenuated live vaccine has a good immune protection effect on toxoplasma gondii acute infection, chronic infection and congenital infection, and toxoplasmosis can be effectively prevented.

The invention provides a nucleotide sequence for preventing toxoplasma infection. The nucleotide sequences selected from the followings are contained: 1) 724-1470th nucleotide sequences as shown in SEQ ID No.3, and/or 724-2472nd nucleotide sequences as shown in SEQ ID No.1; 2) nucleotide sequences complementary with the nucleotide sequences as shown in 1); and 3) nucleotide sequences which perform substituting, deletion and addition modification on one or a plurality of basic groups of the nucleotide sequences as shown in 1) or 2). The pVAX-CDPK (Calcium Dependent Protein Kinase)1 can effectively prevent and control the infection of a toxoplasma animal model (Kunming mice), and pVAX-IL21-IL15 not only can be independently used for effectively preventing and controlling the toxoplasma infection, but also can be used as a cytokines adjuvant for effectively improving the immune effect of a vaccine pVAX-CDPK1.

The invention provides application of lncRNA 11496 in diagnosis and treatment of toxoplasmosis and belongs to the technical field of biological medicine and molecular biology. It is found through research that NONMMUG011496 (lncRNA 11496) was significantly down-regulated in the brain tissue of mice infected with Toxoplasma gondii; meanwhile, the expression of lncRNA 11496 in brain tissues of micechronically infected with toxoplasma gondii affected the proliferation, differentiation and apoptosis of nerve cells, which was closely related to Toxoplasma gondii infection. The lncRNA 11496 is a noncoding RNA first found in brain tissues chronically infected with toxoplasma gondii. Therefore, the lncRNA 11496 can serve as an important biological target, provides a basis for diagnosis of toxoplasma encephalopathy and research and development of targeted anti-toxoplasma drugs based on the lncRNA 11496, and has a good practical application value.

The invention discloses identified mmu-miR-217-5p capable of detecting toxoplasma gondii infection. In a toxoplasma gondii infected mouse model, cDNA (complementary deoxyribonucleic acid) is obtained by extracting general RNA (ribonucleic acid) of plasma and inversely transcribing; high-expression mmu-miR-217-5p is screened by adopting a Q-PCR (quantitative polymerase chain reaction) technology, is remarkably related to toxoplasma gondii infection, and can be used as identified miRNA for toxoplasmosis diagnosis. The mmu-miR-217-5p is used as a marker to establish a Q-PCR detection method of toxoplasma gondii infection, has relatively high sensitivity and specificity, provides a reliable detection basis and quick detection method to clinical diagnosis for toxoplasmosis, and has an important clinical application value.

The invention discloses a compound multivalent antigen gene, a recombinant plasmid and an adenovirus vaccine for preventing toxoplasmosis. The preparation process comprises the following steps: connecting a ubiquitin protein gene, a toxoplasmosis tachyzoite antigen gene, a bradyzoite antigen gene, a sporozoite antigen SPA gene and an AS15 gene, and synthetizing the compound multivalent antigen gene for preventing toxoplasmosis; carrying out double digestion on the synthesized target gene and then inserting the synthesized target gene between BamHI and SmaI of a vector to generate the recombinant plasmid; and carrying out digestion, PCR and sequencing identification on the recombinant plasmid, extracting plasmid and purifying; transferring the recombinant plasmid and skeleton plasmid into cells together, packing and removing toxins, so as to obtain the recombinant adenovirus vaccine. By adopting the adenovirus vaccine disclosed by the invention, toxoplasma infection of people or animals can be effectively prevented, formation of brain tissue cysts in the chronic infection process of toxoplasma is controlled, the immunologic reaction and the survival rate are improved in acute infection, and candidate components of an effective vaccine with comprehensive protection are provided.

The invention provides an immunogenicity composition which comprises erk7 recombinant protein and adjuvant. The invention further provides an immune vaccine for preventing toxoplasma gondii infection. The effective component of the immune vaccine is optional one of the immunogenicity compositions in claims 1-4. The immune vaccine has the advantages that the vaccine can effectively prolong the survival time of toxoplasma gondii GT1 acute infection BALB/c mice, the vaccine has no evident effect on PRU acute infection but can greatly reduce brain cyst number in toxoplasma gondii PRU acute infection, and a good immune effect is achieved.

The invention discloses a RH: deltaNPT1 live attenuated vaccine for preventing toxoplasma infection. The method comprises the steps that plasmid pSAG1::Cas9::U6sgNPT1 and DHFR resistant fragments areconstructed, a theoxoplasma gondii RH: delta NPT1 strain is constructed, and a tachyzoite suspension of toxoplasma gondii Attenuated strain RH: delta NPT1 is prepared in a cell culture medium using PBS as a solvent. The method establishes an immunization dosage form of the live attenuated vaccine, and proves the immunization procedure and dosage thereof. The animal experiment shows that the live attenuated vaccine prepared by the method has a strong immunoprotective effect on acute toxoplasma gondii infection or chronic toxoplasma gondii infection, can be applied to the immune protection of the acute toxoplasma infection or the chronic toxoplasma infection, and has a great application value in the prevention and control of toxoplasma gondii.

The invention relates to a toxoplasma gondii ribulose-5-phosphate isomerase TgRPI gene edited strain and an application thereof. The invention discloses a potential vaccine and drug design target and a gene knockout strain for preventing toxoplasma gondii infection. The design target of the vaccine and the drug is a toxoplasma gondii ribulose-5-phosphate isomerase TgRPI gene, and the nucleotide sequence of the toxoplasma gondii ribulose-5-phosphate isomerase TgRPI gene is as shown in SEQ ID NO: 2. The TgRPI gene is knocked out of the gene knockout strain, the growth speed of the gene knockout strain in vitro is slowed down, but the gene knockout strain can be subcultured, and the gene knockout strain has the characteristics of small toxicity, less reproduction and the like in animal bodies, and can improve the immunocompetence of animals to toxoplasma gondii. The TgRPI plays an important role in in-vivo and in-vitro growth of toxoplasma gondii, so that the TgRPI has the potential of becoming a vaccine and a drug target, and the gene knockout strain has the potential of becoming an anti-toxoplasma gondii genetic engineering vaccine.