93 results about "Fowl adenovirus" patented technology

A composition comprising an isolated fowl adenovirus (FAdV), wherein the FAdV is a strain selected from FAdV-2, FAdV-7, FAdv-8a, FAdV-8b, FAdV-8a / 8b or FAdV-11 serotype strains; and a suitable carrier and methods for inducing protective immunity in a subject and / or its progeny.

A fowl adenovirus group I serum type 4 genetic engineering subunit vaccine, and a preparing method and applications thereof are disclosed. The sequence of an antigen protein in the vaccine is shown as SEQ ID NO:1. The antigen protein has advantages of high safety, high immunity, no pathogenicity for chickens or other animals, and the like. The subunit vaccine can prevent chicken hydropericardium syndrome, inclusion body hepatitis and other diseases which are caused by infection of fowl adenovirus group I serum type 4.

The invention discloses a primer pair, a probe, a kit and a method for fluorescence quantitative PCR detection of FadV-4 type fowl adenoviruses. The detection method takes sample DNA as a template, SEQ ID No:1 and SEQ ID No:2 as primers and SEQ ID No:3 as the probe to perform fluorescence quantitative PCR amplification and acquire a fluorescence signal. The fluorescence quantitative PCR detection method disclosed by the invention can realize quick, accurate, convenient, rapid and specific detection of FadV-4 type fowl adenoviruse nucleic acids, is high in sensitivity, strong in specificity and good in repeatability, and has wide application prospects.

The invention provides a group I FAdV-4 (fowl adenovirus serotype 4) vaccine. An antigen of the group I FAdV-4 vaccine is an inactivated YBAV-4 virus. According to the prepared group I FAdV-4 inactivated vaccine, the immune effect of the vaccine is evaluated with a serology method and a vaccinated challenge method, a result shows that the prepared fowl adenovirus inactivated vaccine can provide effective immune protection for fowls, and the group I FAdV-4 inactivated vaccine has a good commercial development prospect.

The invention provides an I-group 4-type fowl adenovirus DNA vaccine and application thereof. According to the technical scheme, based on experimental means, research finds and shows that fiber protrusions have good immunity prototypes, in this way, with 4-type fowl adenovirus fibrous protein C-terminal genes being antigen substances, codon optimization is carried out on the basis of a natural sequence, an eukaryotic expression vector pCAGGS is cloned, and the DNA vaccine pCAGoptiFAV4C is constructed. According to the researched fowl adenovirus NDA vaccine, a method of gene engineering fermentation is adopted for preparing antigens, and therefore the I-group 4-type fowl adenovirus DNA vaccine is low in cost, pure in antigen and safe to use. By the utilization of a serology method and an immunity virus attack method, the immunity effect of the vaccine is evaluated. The result shows that the vaccine can provide effective immune protection for fowl, and has good commercial development prospects. Compared with a traditional vaccine, the DNA vaccine achieves the safety of subunit vaccines and inactivated vaccines, and also has the features of simultaneous induction of humoral immunity and cellular immune response, wherein the features only belong to attenuated vaccines or recombinant vaccines.

The invention discloses an RPA (recombinase polymerase amplification) primer for detecting a fowl adenovirus serotype 4. The nucleotide sequence of the RPA primer is as shown in SEQ ID No.1 and SEQ ID No.2 in the description. The invention further discloses a method for detecting the RPA of the fowl adenovirus serotype 4, and the method mainly comprises the steps of primer synthesis, extraction of DNA in a sample to be tested, RPA amplification reaction and analysis of an RPA amplification product. According to the RPA primer disclosed by the invention, the specificity is strong, the sensitivity is high, and the detection result is accurate. The detection method disclosed by the invention is simple in operation and good in stability, and a low-cost, rapid and specific on-site diagnosis method for effectively detecting and authenticating chicken cecum hepatitis-hydropericardium syndrome is provided.

The invention provides a combined inactivated vaccine for an avian influenza virus and a fowl adenovirus. An H9 subtype avian influenza virus QDY strain and an I-group 4-type aviadenovirus YBAV-4 new strain used by the vaccine are high in TCID50 / EID50 valence and good in immunogenicity and can well withstand attacks of the H9 avian virus and various local separate viruses. The vaccine prepared by the invention is good in safety, and free of local and whole-body adverse effects caused by the vaccine. Through analysis of characters, a safety test and efficacy test data in a storage-life test, the combined vaccine is free of an obvious difference from a single vaccine of a similar product in effect, and is stable and effective; the efficacy test result proves that the antibodies of the combined vaccine and two single vaccines are kept at a high level and are faster in antibody generation of similar products; and a control group antibody is negative.

The invention discloses an F2 protein based indirect ELISA kit for detection of fowl adenovirus serotype 4 (FAdV-4) antibody. The kit includes an ELISA plate coated with a spike protein gene expression product, a positive and negative control, an HRP labeled rabbit anti-chicken secondary antibody, a sample diluent, a color development liquid and a washing liquid. The indirect ELISA kit for FAdV-4antibody established by the invention can specifically detect the FAdV-4 antibody, and does not react with anti-influenza virus (AIV) serum, anti-mardivirus (MDV) serum, anti-Newcastle disease virus (NDV) serum, anti-avian reticuloendotheliosis virus (REV) serum and SPF chicken serum (Figure 6). Therefore, the kit has good FAdV-4 specificity, and can be used for FAdV-4 infection and immune statusepidemiological investigation.

The invention relates to an LAMP primer pair used for I subgroup fowl adenovirus detection. The LAMP primer pair is composed of a pair of outer primers, a pair of inner primers and a pair of loop primers, wherein sequences of the outer primers are shown in SEQ ID NO.1 and SEQ ID NO.2; sequences of the inner primers are shown in SEQ ID NO.3 and SEQ ID NO.4; and sequences of the loop primers are shown in SEQ ID NO.5 and SEQ ID NO.6. Pathogens can be detected within about 60 minutes only by utilizing the LAMP primer pair provided by the invention, while the conventional PCR method needs 4-5 hours; meanwhile, an LAMP method has high sensitivity, minimum nucleic acid concentration is 8.4*10<-9> ng / muL, the sensitivity of the LAMP method is 10<4> times higher than that of the conventional PCR method, no expensive instrument is needed, detection result is easy to observe, and safety is good, so that the LAMP primer pair is applicable to being popularized to a primary veterinarian disease control and prevention organization or common farm.

The invention provides a flaggelline-fiber2 fusion protein as well as a preparation method and application thereof. The fusion protein is a fusion protein of a fowl adenovirus type 4 fiber2 protein with relatively high immunoprotection and a salmonella typhimurium flagellin. The preparation method comprises the following steps: cloning an artificially coded flaggin-fiber2 gene into a pFastBac-HA expression vector through fusion; carrying out gene transposition to form recombinant Bacmid, transfecting the recombinant Bacmid into Sf9 insect cells, expressing the fusion protein by utilizing a baculovirus system, and conducting identifying by virtue of indirect immunofluorescence IFA and Western blot. The period of obtaining the recombinant baculovirus through the system is short, and when SPFchicken are immunized by the flaggin-fibe2 fusion protein, results show that the flaggin-fibe2 fusion protein expressed by the baculovirus system has high immunoprotection capability.

The invention relates to a PCR detection kit for specific detection of fowl adenovirus group I and a detection method thereof. The detection kit comprises a fowl adenovirus group I PCR reaction liquid, 10*PCR buffer for UNG plus, hot start type DNA polymerase, dU plus dNTP mix, uracil DNA glycosylase, positive quality control serum and negative quality control serum, wherein the fowl adenovirus group I PCR reaction liquid comprises an upstream primer with a concentration of 5 micrometer and the sequence is shown as the specifications as well as a downstream primer with a concentration of 5 micrometer and the sequence is shown as the specifications. When detection is conducted through the PCR detection kit, conditions for a PCR amplified reaction are that UNG treatment is conducted at a temperature of 25 DEG C for 10 min, and predegeneration is conducted at a temperature of 95 DEG C for 2 min; degeneration is conducted at a temperature of 98 DEG C for 10 s, annealing is conducted at a temperature of 55 DEG C, extension is conducted at a temperature of 72 DEG C for 1 min, and all that are conducted for 30 work cycling; extension is conducted at a temperature of 72 DEG C for 10 min. The PCR detection kit for the specific detection of the fowl adenovirus group I has the advantages of being high in sensitivity (still positive when the reaction liquid is diluted to 1:105) and good in repeatability. The PCR detection method for the specific detection of the fowl adenovirus group I is easy, convenient and fast to operate, suitable for early diagnosis of fowl adenovirus group I virus, and can meet the demand for disease prevention and control in time.

The present invention relates to a novel fowl adenovirus and a vaccine thereof.

The invention discloses a recombinant fowl adenovirus type 4 (FAdV-4) fiber2 protein as well as a preparation method and application thereof, and belongs to the technical field of gene engineering. The recombinant protein is obtained by removing 274 amino acid residues at the N end of the fowl adenovirus type 4 fiber2 protein and then recombining with the 21-55th amino acid sequence of the fowl adenovirus type 4 hexon protein; and can be used for preparing the avian adenovirus 4-type genetic engineering subunit vaccine. Aiming at the defects of poor solubility and immunogenicity of prokaryoticexpression natural fiber2 protein, the fiber2 protein coding gene is subjected to series modification, and compared with the unmodified fiber2 protein, the recombinant protein has the advantages thatthe gene coding sequence of 274 amino acid residues at the N end of the fiber2 protein is removed, and the gene coding sequences of two important antigen epitopes of the hexon protein are fused; so that the solubility and the immunogenicity are obviously improved, so that the subunit vaccine which is controllable in quality, safe and effective and can prevent highly pathogenic FAdV-4 virus infection can be prepared, and complete protection can be obtained by once immunization.

The invention relates to a recombinant serum type-4 fowl adenovirus for expressing serum type-8 fowl adenovirus spike protein and a preparation method thereof, and the principle and the core key technology of the invention are that sgRNA is used for replacing EGFP and then virus purification is carried out to obtain the recombinant fowl adenovirus capable of stably amplifying the fowl adenovirus type 8 spike protein. The recombinant FAdV-4 for expressing the fowl adenovirus type-8 spike protein is successfully obtained by inserting the fowl adenovirus type-8 spike protein gene F which is popular at present as a carrier, and an in-vitro experiment result shows that the replication capacity of the FAdV4-F1-F-F2 recombinant virus is almost the same as that of a FAdV-4 wild virus, and the recombinant virus can be used as an inactivated recombinant multivalent vaccine for preventing and controlling FAdV-4 and FAdV-8.

The invention discloses a RAA primer for detecting 12 serotypes of fowl adenovirus I group and a detection method of the RAA primer, and belongs to the technical field of fowl virus detection. The sequences of a primer and a probe in a RAA primer and probe combination for detecting 12 serotypes of the fowl adenovirus I group are respectively as shown in SEQ ID NO.1-SEQ ID NO.3. According to the invention, 12 different serotype FadV can be detected at the same time. In addition, only necessary reagents and isothermal amplification equipment are needed, operation is easy and convenient, and the method is particularly suitable for large-scale detection of chicken flocks in basic-level farms, so that purification of chicken flock FadV is facilitated.

The invention provides a method for large-scale suspension culture of fowl adenoviruses. The method provided by the invention can be used for obtaining high-titer fowl adenoviruses of I group 4 type, 8b type, D11 type, 8a type and the like, a domesticated LMH cell used in the method is a chicken liver cancer suspension LMH cell, and the collection number of the domesticated LMH cell is CCTCC NO: C2021202. According to the method disclosed by the invention, adherent LMH cells are domesticated into suspension cells capable of realizing stable and continuous passage by adopting a gradual serum reduction method. Then, full suspension culture is carried out on the LMH cells by using a bioreactor, then, step-by-step amplification suspension culture is carried out on the LMH cells, and then, aviadenovirus antigens are respectively inoculated and proliferated on the suspension cells. By use of the method disclosed by the invention, high-titer aviadenovirus suspension culture technology virus antigens can be obtained, and the titer of the antigens is about 5-10 times of that of existing vaccine regulation antigens.

The invention belongs to the technical field of biology, and mainly relates to a recombinant fowlpox virus transfer vector expressing a fowl adenovirus serotype-4 (FADV4) fiber2 gene, a constructing method thereof and applications of the transfer vector. A pMD19T-Simple vector is adopted as a base of the transfer vector. The FADV4 fiber2 gene, a lacz gene, and fowlpox virus genome replicated non-essential fragments which are LTYB and RTYB used for homologous recombination are inserted in a TA cloning site. The method includes constructing a plasmid pMD-TYB; constructing a plasmid pMD22; constructing a plasmid pMD22-lacz; constructing an intermediate vector pMD22-TYB-lacz; amplifying the FADV4 fiber2 gene; constructing a recombinant fowlpox virus transfer vector pMD22-TYB-lacz-F4; subjecting a chicken embryo Fibroblast to cotransfection with the transfer vector pMD22-TYB-lacz-F4 and a fowlpox virus; performing identification to select a positive product; performing subculture continuously; and identifying expression effects of the recombinant fowlpox virus. The recombinant fowlpox virus transfer vector constructed by the method lays a foundation for development of an efficient recombinant fowlpox virus genetic engineering living-vector vaccine expressing the fowl adenovirus serotype-4.

The invention belongs to the field of biotechnical detection, and particularly relates to an indirect ELISA (Enzyme Linked Immunosorbent Assay) kit for detecting a fowl adenovirus antibody based on a hexon protein N-terminal conservative area. The kit comprises an ELISA enzyme standard plate coated with the expression product of the hexon protein N-terminal conservative area, an HRP (Horseradish Peroxidase) marked rabbit anti-chicken antibody, a sample diluting solution and a washing solution; the sequence of the hexon protein N-terminal conservative area is as shown by SEQ ID NO. 5. By using the indirect ELISA kit for the fowl adenovirus antibody, which is established by the invention, an antibody resisting a fowl adenovirus can be detected specifically, but an antibody resisting other pathogens cannot be detected. Therefore, the kit has favorable specificity on the fowl adenovirus, and can be used for the epidemiological investigation of the infection status of the fowl adenovirus.

The invention provides a genetically-engineered vaccine against fowl adenovirus type 4, and a preparation method and application thereof. According to the invention, protective antigens, namely pentonand hexon proteins of fowl adenovirus serotype 4 are connected by a linker, and are expressed by an insect cell-baculovirus expression system to form virus-like particles of fowl adenovirus type 4 onspatial conformation; an adjuvant is added; and emulsification is carried out so as to prepare the genetically-engineered vaccine against fowl adenovirus type 4. The preparation method for the vaccine provided by the invention is simple in process, capable of preparing a large amount of fowl adenovirus type 4 antigen protein, short in time consumption, high in expression quantity and beneficial to large-scale production; and the obtained genetically-engineered vaccine is good in immune effect and capable of effectively preventing infection of the fowl adenovirus type 4.

The invention relates to a selecting method for duck adenovirus 1 type virus replicating non-necessary section segment and the recombination fowl adenovirus gained through transporter gene and the universal transporter gene. It uses duck adenovirus 1 type virus as material to expand the sequence at right side of E4 area by PCR method. Inducing loxP sequence to the two sides of GFP gene expression box and inserting into the expanded segment, the universal transporter gene would be constructed. The transporter gene would gain the recombination adenovirus of stable expression GFP, thus, a replicating non-necessary area would be determined. The universal transporter gene loxP-GFP-loxP sequence side contains a unique enzyme Restriction Enzyme cutting site, and cold insert relative target gene to gain recombination virus without GFP gene. The gene engineer medicine developed from the universal transporter gene would not contain selecting marker gene.

The invention relates to a heat-resistant serum 4 type fowl adenovirus genetic engineering vaccine candidate strain and a construction method thereof. The candidate strain is a thermal-stability newcastle disease virus (Newcastle Disease Virus) rAHR09-4F2 expressing serum 4 type fowl adenovirus spike protein 2, and the preservation number is CCTCC NO: V201932. The candidate strain is obtained through using a heat-resistant newcastle disease attenuated virus strain rAHR09 as a carrier, inserting a serum 4 type fowl adenovirus (FAdV4) spike protein (fiber2) gene between a P intergenic region andan M intergenic region, and adopting a backward heredity technique. The result of evaluating the biological characteristics and the immunoprotection efficacy of the candidate strain displays that therecombinant virus has favorable heat stability and immunogenicity and provides favorable immunoprotection on FAdV4 infection.

The present disclosure relates to a vaccine composition, wherein the vaccine composition comprises an immune amount of fowl adenovirus Fiber-2 protein or an immune amount of a live vector recombined with gene of the fowl adenovirus Fiber-2 protein, and a pharmaceutically acceptable carrier. The vaccine composition can provide effective immune protection against different serotypes of adenoviruses and provide a protection rate of 100% at low levels of immunogenic components, showing good immunological efficacy.

The invention discloses a primer group and a kit for detecting a fowl adenovirus serotype 4 (FAdV-4) as well as a detection method and an application of the primer group. The primer group comprises outer primers and inner primers, wherein the outer primers include F3 and B3, the inner primers include FIP and BIP, the nucleotide sequence of the F3 is shown as SEQ ID NO.1, and the nucleotide sequence of the B3 is shown as SEQ ID NO.2; and the nucleotide sequence of the FIP is shown as SEQ ID NO.3, the nucleotide sequence of the BIP is shown as SEQ ID NO.4, the FIP is a 5-end biotin labeled sequence, and the BIP is a 5-end fluorescein isothiocyanate labeled sequence. By combination of a loop-mediated isothermal amplification (LAMP) technology with a lateral flow dipstick (LFD) technology, a set of primers are designed for an important factor 52 k gene of the FAdV-4, and an LAMP-LFD rapid detection technology of the FAdV-4 is established. The LAMP-LFD kit provided by the invention is convenient to operate, safe, reliable, sensitive, efficient, good in specificity and particularly suitable for field rapid visual detection.

The invention discloses application of protein fiber2 and recombinant protein thereof in detection of a 4-type fowl adenovirus antibody of serum. According to the application, firstly, recombinant FAdV-4 fiber2 protein capable of being specifically combined with a 4-type fowl adenovirus specific antibody and a horse radish peroxidase labeled FAdV-4 enzyme-labeled antibody are obtained based on theprotein fiber2, and a method for specifically blocking the 4-type fowl adenovirus antibody in the serum and a detection kit are established by further using the recombinant protein as coating protein. The detection kit is high in sensitivity, high in specificity and good in stability, can be used for reflecting a neutralizing antibody level, is used for rapidly detecting the 4-type fowl adenovirus antibody in the serum, has the characteristics of simplicity in operation, high specificity, high sensitivity and low cost, can be used for simultaneously detecting a large number of samples and canbe used for monitoring an overall immunizing effect and neutralizing antibody level, so that powerful foundations and supports are provided for the diagnosis, vaccine prevention and control and the like of diseases, and the application has a very good application prospect.

The invention discloses a fowl adenovirus group I immunity related fusion protein as well as an encoding gene and application of the fowl adenovirus group I immunity related fusion protein. The fusion protein provided by the invention comprises Hexon protein and chicken interleukin-2. The Hexon protein is the protein composed of an amino acid sequence from the 1st position to the 914th position of N' tail end of the sequence 6 in a sequence table, and the chicken interleukin-2 is the protein composed of the amino acid sequence from the 933rd position to the 1075th position of the N' tail end of the sequence 6 in the sequence table. Experiments show that a DNA (deoxyribonucleic acid) molecule composed of a Hexon gene and a chicken interleukin-2 (chIL-2) gene, which contain fowl adenovirus group I and are connected in series, is provided, a Hexon specific antibody can be generated when the DNA molecule is injected to chicken muscle by an expression vector, and the DNA molecule has good immunity protection effect and can be used for effectively preventing the fowl adenovirus group I after fowl adenovirus group I challenge experiments.

The invention belongs to the field of animal genetic engineering vaccines, and particularly relates to a fowl adenovirus serum type 4 (FAdV-4) recombinant virus for expressing fowl adenovirus serum type 8b (FAdV-8b) spike protein (Fiber) as well as a construction method and application of the fowl adenovirus serum type 4 (FAdV-4) recombinant virus. The recombinant virus is obtained by replacing a FAdV-4 Fiber1 gene with a Fiber gene of FAdV-8b, the recombinant virus can be used for preparing a bivalent vaccine for preventing and controlling chicken hepatitis-hydropericardium syndrome and / or chicken inclusion body hepatitis, and the bivalent vaccine prepared by utilizing the recombinant virus can achieve the effect of preventing two epidemic diseases at the same time by injecting a needle of vaccine. Therefore, the purpose of preventing and controlling two or more diseases by one needle can be achieved by inserting the FAdV-4 serving as a carrier into an exogenous gene.

The invention belongs to the technical field of traditional Chinese medicines, and particularly relates to a traditional Chinese medicine composition for treating adenoviral infection of poultry. The composition is prepared from the following medicinal raw materials in parts by weight: 15-25 parts of radix isatidis, 10-20 parts of radix paeoniae alba, 20-30 parts of herba artemisiae scopariae, 10-15 parts of felwort, 7.5-15 parts of codonopsis pilosula, 7.5-15 parts of poria cocos, 10-20 parts of scutellaria baicalensis, 10-20 parts of sophora flavescens and 10-30 parts of licorice root. The traditional Chinese medicine composition is reasonable in combination, is safe and effective, can be used for treating both symptom and root cause of the adenoviral infection of poultry, and basically prevents relapse.

The invention relates to the technical field of veterinary biological products, in particular to an avian adenovirus 4, 8 and 11 type trivalent vaccine as well as a preparation method and application thereof. The fowl adenovirus trivalent vaccine comprises a fowl adenovirus type 4 vaccine strain QYH2019-YN, a fowl adenovirus type 8 vaccine strain QYH2020-HB and a fowl adenovirus type 11 vaccine strain QYH2020-SY. The invention provides a fowl adenovirus trivalent vaccine which is prepared from a fowl adenovirus type 4 vaccine strain, a fowl adenovirus type 8 vaccine strain and a fowl adenovirus type 11 vaccine strain which are obtained by newest separation, and has an effective prevention effect on fowl adenovirus type 4, fowl adenovirus type 8 and fowl adenovirus type 11 as well as chicken inclusion body hepatitis and hydropericardium-hepatitis syndrome caused by the fowl adenovirus type 4, fowl adenovirus type 8 and fowl adenovirus type 11.