88 results about "Emoia loyaltiensis" patented technology

The invention discloses an E.tenella protein disulfide linkage isomerase gene EtPDI (Clone ID is BW1-E06,and the Genbank accession number of is EF552214). The gene is connected with a procaryon expression vector pGEX-4T-2; a procaryon expression recombination plasmid pGEX-4T-EtPDI is constructed and is expressed in a colibacillus system; and most of the expressed recombining protein exists in a soluble form. The recombining protein 4T-EtPDI is purified to carry out SPS-PAGE and is transferred to a PVDF film; and antiserum of E.tenella oocyst oral immunized chicken is used as first resistance and goat anti-chicken IgG is used as second resistance to carry out Western-blot analysis, thereby indicating that the gene has certain antigen. The gene is used for preparing an anti-chicken coccidiosis drug and an anti-chicken coccidiosis vaccine.

The invention discloses an application of a chloramine B solution in chicken coccidiosis oocyst culture and preservation. According to the invention, study finds out that the sporulation rate of the chicken coccidiosis oocysts can be effectively improved with the chloramine B aqueous solution as a chicken coccidiosis oocyst culture solution. The oocysts of the chicken eimeria tenella, eimeria acevulina, eimeria maxima and eimeria necatrix are not affected in activity if being preserved within 6 months; if being preserved within 8 months, the yield of the oocysts is reduced within 25%; the pathogenicity is not obviously affected. Therefore, the chloramine B aqueous solution can be used as the culture and preservation solution for the chicken coccidiosis oocysts in stead of potassium dichromate.

The invention discloses a method for batch preparation of chicken Eimeria coccidium oocysts, and particularly relates to a separation and purification method for producing oocysts (antigens) of chicken Eimeria coccidium vaccines. The method for preparing the chicken Eimeria coccidium oocysts comprises the following process steps: fecal sample processing, continuous screening, saturated brine float centrifuging, oocyst centrifugal depositing and oocyst purifying. According to the method, the industrial preparation of the chicken Eimeria coccidium oocysts can be realized, the production efficiency can be improved, and the cost can be reduced.

The present invention provides an isolated nucleic acid comprising a nucleotide sequence encoding a 250 kDa polypeptide from Sporozoites / Merozoites of Eimeria maxima, or encoding a homolog of the polypeptide, or a complement of the nucleic acid and a method of producing a recombinant 250 kDa polypeptide of the same. The present invention also provides an isolated nucleic acid comprising a nucleotide sequence encoding an immunodominant portion of a 250 kDa polypeptide from Sporozoites / Merozoites of Eimeria maxima having the amino acid sequence described herein, or encoding a homolog of the polypeptide, or a complement of the nucleic acid. The subject invention also provides a vaccine against Eimeria tenella, Eimeria maxima, Eimeria acervulina, Eimeria necatrix, Eimeriapraecax, Eimeria mitis or Eimeria brunetti, or a microorganism expressing an immunologically cross-reactive antigen, comprising the isolated nucleic acid comprising a nucleotide sequence encoding a 250 kDa polypeptide from Sporozoites / Merozoites of Eimeria maxima or the immunodominant portion of a 250 kDa polypeptide from Sporozoites / Merozoites of Eimeria maxima, or encoding a homolog of the polypeptide, or a complement of the nucleic acid which may also contain a 56 kDa, 82 kDa or 230 kDa protein isolated from the gametocytes of Eimeria maxima and a method of immunizing a subject against infection by Eimeria tenella, Eimeria maxima, Eimeria acervulina, Eimeria necatrix, Eimeria praecox, Eimeria mitis or Eimeria brunetti, or a microorganism expressing an immunologically cross-reactive antigen, comprising the step of administering to the subject any of the aforementioned vaccines.

The invention provides an immunoregulation type DNA vaccine with actions of preventing and treating chicken coccidiosis, which comprises calcium regulation bonding domain protein kinase genes pE+K2 of Eimeria tenella and chicken interleukin-2 genes, and a section of proteinase hydrolytic sequence is introduced to the 5' end of the chIL-2 gene.

The invention relates to a chicken toxic Eimeria acervulina immunoregulation type DNA vaccine, which belongs to the technical field of biological veterinary medicines. The molecular biology technology is used for connecting toxic Eimeria sporozoite surface antigen NA4 genes and chIL-2 genes in series to construct a fusion expressing carrier pVAX-LDH-IL-2. The vaccine comprises the inserted toxic Eimeria sporozoite surface antigen NA4 genes and the chIL-2 genes. The vaccine comprises a plurality of T cell immune response elements, and has the advantages of high safety, long keeping time in bodies, simple preparation, good heat stability, convenient storage and transportation, time saving and labor saving when in use and the like, thus preventing and curing chicken coccidiosis effectively. Tests prove that the immunoregulation type DNA vaccine pVAX-NA4-IL-2 has good immune protection effect in coccidian resistant infections.

Prevention and treatment of coccidiosis in chicken is accomplished by administering an anti-chicken coccidiosis composition adapted for oral administration. This composition contains an antibody obtained from an egg of a chicken immunized with an antigenic outermembrane protein or an immunogenic fragment thereof as an antigen having an immunogenicity common for sporozoit and merozoite of Eimeria acervulina, Eimeria tenella, and Eimeria maxima which are associated with chicken coccidiosis. The composition may optionally contain a lactic acid bacterium and / or an antibody obtained from an egg of a chicken immunized with Clostridium perfringens.

The invention discloses a preparation method of a live vector vaccine for controlling chicken coccidiosis, comprising the following steps of obtaining a recombinant expression vector by connecting a mature peptide of an eimeria tenella sporozoite antigen gene 3-1E gene and an expression vector of bacillus subtilis, obtaining a recombinant strain by transferring the recombinant expression vector into a bacillus subtilis host, and culturing the recombinant strain to finally obtain the live vector vaccine for controlling the chicken coccidiosis by. The live vector vaccine can be used for controlling and / or treating the chicken coccidiosis. The invention also discloses an oral biological preparation for controlling the chicken coccidiosis, prepared by utilizing the live vector vaccine. The oral biological preparation comprises the live vector vaccine and a vector. The oral biological preparation per gram contains the live vector vaccine with a living bacteria number of 108-109cfu.

The invention provides a new medicinal application of nitazoxanide and particularly discloses an application of nitazoxanide in preparing a drug for resisting eimeria coccidium. The application can serve for the prevention and treatment of coccidiosis of animals such as chickens and rabbits, and the like.

The invention discloses a method for extracting and purifying eimeria tenella merozoite. The method comprises the steps of: releasing the merozoite and purifying the merozoite, wherein the merozoite release comprises mechanical grinding release and enzymic digestion release, 0.0625% pancreatin and 0.175% sodium taurodeoxylate are adopted as digestive juice in the enzymic digestion release process; digestion is moderate, which is beneficial to the activation of the merozoite, can eliminate erythrocytes to obtain pure merozoite. The method is convenient and simple to operate, is low in cost, and is ideal in separation and purification effects; the separated merozoites are large in quantity, are pure with few impurities and keep vigorous energy; and the method is suitable for research on drug susceptibility detection, vaccine research and other biological researches.

The invention discloses an eimeria tenella Et CHP39 gene. The Et CHP39 gene includes an open reading frame nucleotide sequence represented as the SEQ ID No.1. The eimeria tenella conserved protein Et CHP39 gene is a new gene of the eimeria tenella and is relative to sporozoite invaded host cells. The eimeria tenella conserved protein Et CHP39 gene has quite high application value in development of a new vaccine or a new medicine capable of inhibiting the sporozoite invaded host cells and blocking a life history of the eimeria tenella.

The invention discloses a new Eimeria tenella heat shock protein 60 (Et HSP60) gene. The cloning number is ZB2-D07, the total length is 1802 bp, and the Et HSP60 gene contains a 1455 bp complete open reading frame (ORF) and can code 484 amino acids (Genbank accession number: FJ911605). The gene is connected with a prokaryotic expression vector pET28a(+) to construct a prokaryotic expression recombinant plasmid pET28a(+)-Et HSP60 and is expressed in an escherichia coli system. SDS-PAGE (Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis) is carried out by purifying the recombinant protein pET28a(+)-Et HSP60, and then the recombinant protein pET28a(+)-Et HSP60 is transferred to a PVDF (Polyvinylidene Fluoride) membrane. The antiserum of immuned chicken orally taking E. tenella oocyst is taken as a primary antibody, and goat-anti-chicken IgG (immunoglobulin G) is taken as a secondary antibody to carry out Western-blot analysis. The analysis result shows that the recombinant protein pET28a(+)-Et HSP60 has certain antigenicity and can be used for preparing a medicine for resisting chicken coccidiosis and a vaccine resisting chicken coccidiosis.

The invention provides an immunoregulation type DNA vaccine with actions of preventing and treating chicken coccidiosis, which comprises sporozoite surface antigen genes TA4 of Eimeria tenella and chicken interleukin-2 genes, and a section of proteinase hydrolytic sequence is introduced to the 5' end of the chIL-2 gene.

The invention relates to a chicken Eimeria acervulina immunoregulation type DNA vaccine, which belongs to the technical field of biological veterinary medicines. The molecular biology technology is used for connecting lactate dehydrogenase genes of schizont antigen of the first generation of chicken Eimeria acervulina and chIL-2 genes in series to construct a fusion expressing vaccine pVAX-LDH-IL-2. The vaccine comprises a plurality of T cell immune response elements, thus preventing and curing chicken coccidiosis effectively.

The invention relates to a fusion gene for expressing Fc fragment of chicken IgY molecules and avian influenza virus antigen and an application of the same. The fusion gene is constructed by means of an HA1 gene fragment of avian influenza virus H9N2 sub-type and the Fc gene fragment of the chicken IgY molecule and then is transfected to eimeria mitis to obtain transgenic coccidium. The transgenic coccidium is inoculated through an oral method for enhance immune reaction aiming to an avian influenza specific antigen in chickens. The application is simple to carry out, is low in cost and has an excellent application prospect.

The invention discloses an eimeria tenella Et CHP559 gene. The Et CHP559 gene includes an open reading frame nucleotide sequence represented as the SEQ ID No.1. The eimeria tenella conserved protein Et CHP559 gene is a new gene of the eimeria tenella and is relative to sporozoite invaded host cells. The eimeria tenella conserved protein Et CHP559 gene has quite high application value in development of a new vaccine or a new medicine capable of inhibiting the sporozoite invaded host cells and blocking a life history of the eimeria tenella.

The invention belongs to the field of genetic engineering vaccines, and in particular relates to a gametophyte antigen gene of chicken Eimeria necatrix, expression and application of an expression product of the gene. An open reading frame of the gene has the size of 561 bp. The gene is constructed into a prokaryotic expression vector pET28a-Engam22 to transform BL21 host bacteria to perform inducible expression. The expression product can be identified by chicken rehabilitation serum, and the fact shows that recombinant protein has immunogenicity. The recombinant eukaryotic expression plasmid pcDNA3.1-Engam22 constructed by the gene is used for immunizing chicks and can induce to generate humoral immunity and cellular immunity, and the fact shows that the gene can be applied to development of a chicken coccidiosis gene engineering vaccine.

The invention discloses a coccidiosis trivalent live vaccine which is prepared by performing first-stage attenuation on spore ball egg sacs which are obtained by performing separation, purification and cultivation on three types of eimeriida, namely eimeria tenella, eimeria necatrix and eimeria maxima, through alkylating agent chemical mutagen and mixing three second-stage attenuation plants obtained by performing second-stage attenuation by irradiation of physical mutagen ultraviolet on oocysts of chicks cultivated by first-stage attenuation plants. Furthermore, the invention also discloses a preparation method and application of the coccidiosis trivalent live vaccine. By adopting dual attenuation, the coccidiosis trivalent live vaccine disclosed by the invention is high in induction rate, low in toxicity and stable in immunity.

The invention relates to a method for expressing an eimeria acevulina 3-1E protein in lactococcus lactis, and relates to a method for expressing an eimeria acevulina 3-1E protein in order to solve the problem that the eimeria acevulina sporozoite / merozoite stage surface antigen 3-1E can not be expressed in the lactococcus lactis. The method comprises the following steps of optimizing an eimeria acevulina 3-1E gene sequence according to the preference of the condon of the lactococcus lactis, inserting a cloning vector and transforming escherichia coli cells to obtain positive transformants; carrying out enzyme digestion and connection of the positive transformants and a lactic acid bacteria expression vector, and constructing a recombinant expression vector; transforming competent lactic acid bacteria cells, and identifying; and culturing and inducing recombinant bacteria to complete the expression of the 3-1E protein in the lactococcus lactis NZ9000. The artificially synthesized codon-optimized 3-1E protein is successfully expressed in the lactococcus lactis. The method provided by the invention is used for the prevention field of coccidiosis.

The invention discloses an eimeria tenella gene. The eimeria tenella gene is an eimeria tenella elF3d gene which can code an amino acid sequence shown in SEQ ID NO.1. The invention also discloses a protein coded by the eimeria tenella eIF3d gene. Recombinant protein coded by the eimeria tenella eIF3d gene has a good immune protection effect on avian coccidiosis and is applicable to being developed into a new vaccine or new drug which can be used for preventing and treating the avian coccidiosis.

The invention relates to an Eimeria maxima DNA vaccine of chicken, belonging to the field of biological veterinary drugs. A gene fragment Em8 of an antigen EmTFP250 at the spore stage of the Eimeria maxima of the chicken is cloned by utilizing the molecular biology technology, and the fragment has higher antigen index and the centralized T cell epitope. The gene fragment is connected with a chicken interleukin-2 (chIL-2) gene in series, thereby constructing a vaccine expression vector pVAX-Em8-IL-2. The vaccine contains a plurality of T cell immune response elements and has the advantages of high safety, long maintenance time in vivo, simple preparation, good thermal stability, convenient storage and transport, time-saving and effort-saving use, and the like. Experiments prove that the immune regulatory DNA vaccine pVAX-Em8-IL-2 has good immune protection effect on the anti-Eimeria maxima infection.

The invention provides an eimeria tenella AN1-like Zinc finger domain-containing protein and an application thereof in inhibiting invasion of coccidium. The eimeria tenella AN1-like Zinc finger domain-containing protein is cloned, expressed and purified. The transcription level of EtAN1-ZnFP in different developmental stages of polypides is detected by fluorescence quantitative PCR (qPCR). The immune protective effect of EtAN1-ZnFP after chickens are infected with coccidium is evaluated by immunoprotection experiments. Research results indicate that the EtAN1-ZnFP is possible to participate ininvasion and growing development of eimeria tenella.

The invention discloses an application of secoisolariciresinol diglucoside in preparation of a drug with resistance to eimeria tenella. Powder is prepared from the secoisolariciresinol diglucoside and acceptable auxiliary materials, is added to feed and is mixed uniformly, and chickens are fed according to the daily amount of feed; or water-soluble secoisolariciresinol diglucoside powder is prepared from the secoisolariciresinol diglucoside and dissolved in water, and the chickens are fed according to the daily drinking amount. Animal infection experiments prove that the ACI (anti-coccidial index) of the secoisolariciresinol diglucoside can be up to 165 or higher, and the secoisolariciresinol diglucoside has a better eimeria tenella resistant effect. Compared with a conventional anti-coccidial drug, namely, sulfachloropyrazin, the secoisolariciresinol diglucoside has a remarkable eimeria tenella resistant effect. According to the application of the secoisolariciresinol diglucoside, the problem of severe drug tolerance produced through adoption of the anti-coccidial drug in the current breeding industry can be effectively solved, the research and development speed of a new drug can be increased on the basis, and great benefits are provided for green and healthy development of the breeding industry.

The invention relates to a chicken coccidiosis prevention and treatment preparation, in particular to a preparation technology of a chicken Eimeria tenella egg yolk antibody targeted sustained release preparation so as to solve the prevention and treatment problems of chicken coccidiosis. The preparation technology of the chicken Eimeria tenella egg yolk antibody targeted sustained release preparation includes the steps of: preparation, separation and purification of a highly immunized egg yolk antibody (IgY), and preparation of a microcapsule. Preparation of the microcapsule has the reaction conditions that: the chitosan concentration is 0.05-0.8% (w / v), the concentration of CaCl2 is 0.5-1.5% (w / v), the concentration of sodium alginate is 1.0-3.0% (w / v), and the IgY and the sodium alginate are in a mass ratio of 1-4:8. The IgY microcapsule prepared by the technical scheme involved in the invention is a preventive medicine, and is also a therapeutic drug. The microcapsule prepared by the invention provides a very good example for antibiotic substitution therapy. And the determined technology can also be used to prepare IgY or IgG microcapsules of other pathogenic microorganisms, thus having promotional value, and being beneficial to sound development of the green and organic chicken raising industry.

The present invention discloses an eimeria acervulina nano-subunit vaccine, a preparation method and an application thereof. The eimeria acervulina nano-subunit vaccine is a nano-particle formed by PLGA encapsulating a recombinant protein EaMIC3, and the recombinant protein EaIMIC3 is an eimeria acervulina microneme protein 3 and has an amino acid sequence shown as SEQ ID NO.1. The recombinant protein EaMIC3 is coated by the nano-material PLGA to form a brand-new vaccine form and to obtain the eimeria acervulina PLGA nano-subunit vaccine with a relatively high immune protection effect.

The invention belongs to the field of gene engineering vaccines, and concretely relates to an eimeria tenella gametophyte antigen gene, expression and application of an expression product. The size of the open reading frame of the gene is 1617 bp, and the nucleotide sequence of the gene is shown as SEQ ID NO. 1. A prokaryotic expression vector pET28a-Etgam59 is constructed based on the gene, and is transformed into BL21 host bacterium for inducible expression. The expression product can be recognized by chicken convalescent serum, which shows that recombinant protein possesses immunogenicity. The obtained serum of an immunized mouse of a recombinant eukaryotic expression plasmid pcDNA3.1-Etgam59 constructed by amplifying the gene is capable of specifically combined with the expression product of pET28a-Etgam59, which shows that the gene is applicable to development of a chicken coccidiosis gene engineering vaccine.

The invention relates to an E.maxima recombinant protein subunit vaccine EmMIC3 and a nanometer subunit vaccine PLGA-EmMIC 3, and belongs to the technical field of biological veterinary medicines. Thenanometer subunit vaccine is prepared by wrapping recombinant protein subunit vaccine EmMIC3 with PLGA nanometer material. Animal immune protective tests prove that the recombinant protein subunit vaccine EmMIC3 and the nanometer subunit vaccine PLGA-EmMIC3 can effectively resist the infection of the E.maxima. The invention uses nanometer material PLGA to wrap the recombinant protein EmMIC3 to form a brand-new vaccine form, which is different from the single EmMIC3 recombinant protein, and the two components are different. After the recombinant protein EmMIC3 and the nanometer material PLGA are coated, the immune protection effect is obviously improved, and the protection effect is improved from general level to excellent level.

The invention relates to polypeptides for treating chicken coccidiosis, in particular to a polypeptide for preventing and treating eimeria tenella and a pharmaceutical composition containing the polypeptide. The polypeptide has no obvious toxic effect on a host and has no pollution to the environment. Tests prove that the MIC50 of the polypeptide to eimeria tenella is 63.15 [mu]g / ml, and the polypeptide has a good eimeria tenella resisting effect.

The invention discloses an Eimeria tenella gene which is Et STK gene and includes an amino acid sequence shown as SEQ ID NO.1. Moreover, the invention provides a preparation method of a vaccine for prophylaxis and treatment of coccidiosis in chicken. The method includes: connecting the mature peptide of Eimeria tenella conserved protein Et STK with an expression vector of escherichia coli or pichia pastoris cell to obtain a recombinant expression vector, transferring the recombinant expression vector into a host of escherichia coli or pichia pastoris cell to obtain a recombination strain, and cultivating the recombination strain to obtain the vaccine of coccidiosis in chicken. The method has quite high application value.

The invention relates to the technical field of detection and particularly discloses fluorescent quantitative PCR (polymerase chain reaction) detection primers and kit for Eimeria media-rabbit. The primers are Me-F and Me-R with the sequences shown as SEQ ID NO.1-2; a reaction system and reaction conditions are optimized on the basis of the primers, one fluorescent quantitative PCR kit for quantitative detection of Eimeria media-rabbit on the basis of a fluorescent dye method is developed, the kit operation is simple and programing, the method has high specificity and high sensitivity, and result determination is objective. The method can realize epidemiological investigation and clinical diagnosis of rabbit coccidiosis and provides technical support for diagnosis, prevention and treatment technologies of rabbit coccidiosis and rapid and accurate clinical detection of the rabbit coccidiosis infection condition.