CN105833263A - Bivalent vaccine of acian metapneumovirus and H9 subtype avian influenza virus - Google Patents
Bivalent vaccine of acian metapneumovirus and H9 subtype avian influenza virus Download PDFInfo
- Publication number
- CN105833263A CN105833263A CN201610236341.8A CN201610236341A CN105833263A CN 105833263 A CN105833263 A CN 105833263A CN 201610236341 A CN201610236341 A CN 201610236341A CN 105833263 A CN105833263 A CN 105833263A
- Authority
- CN
- China
- Prior art keywords
- vaccine
- protein
- metapneumovirus
- avian influenza
- influenza virus
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K39/12—Viral antigens
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K2039/51—Medicinal preparations containing antigens or antibodies comprising whole cells, viruses or DNA/RNA
- A61K2039/52—Bacterial cells; Fungal cells; Protozoal cells
- A61K2039/521—Bacterial cells; Fungal cells; Protozoal cells inactivated (killed)
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K2039/70—Multivalent vaccine
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2760/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA ssRNA viruses negative-sense
- C12N2760/00011—Details
- C12N2760/16011—Orthomyxoviridae
- C12N2760/16111—Influenzavirus A, i.e. influenza A virus
- C12N2760/16134—Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2760/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA ssRNA viruses negative-sense
- C12N2760/00011—Details
- C12N2760/18011—Paramyxoviridae
- C12N2760/18311—Metapneumovirus, e.g. avian pneumovirus
- C12N2760/18334—Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Microbiology (AREA)
- Chemical & Material Sciences (AREA)
- Immunology (AREA)
- Medicinal Chemistry (AREA)
- Virology (AREA)
- Mycology (AREA)
- Pharmacology & Pharmacy (AREA)
- Epidemiology (AREA)
- Animal Behavior & Ethology (AREA)
- General Health & Medical Sciences (AREA)
- Public Health (AREA)
- Veterinary Medicine (AREA)
- Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)
Abstract
The invention provides a bivalent vaccine of an acian metapneumovirus and an H9subtype avian influenza virus. The bivalent vaccine comprises an antigen and an vaccine adjuvant, wherein the antigen is an H9 subtype avian influenza virus strain and acian metapneumovirus F protein; thepreservation numberof the H9 subtype avian influenza virus strain is CCTCC NO.V201517; and the amino acid sequence of the acian metapneumovirus F protein is SEQ ID NO:1. The H9 subtype avian influenza virus strain QDY and the acian metapneumovirus F protein antigen solution are mixed at the ratio, and then an oil adjuvant is added to the mixture to be mixed and emulsified into the vaccine. According to the prepared vaccine, the immunized antibody level can be improved, the uniformity of the immunized antibody is improved, the immune effect of the vaccine is ensured, and the vaccine has the advantages of high efficiency and good safety.
Description
Technical field
The invention belongs to veterinary biologics technical field, be specifically related to a kind of fowl metapneumovirus and H9 hypotype fowl
Influenza virus bigeminy vaccine.
Background technology
Fowl metapneumovirus belongs to paramyxovirus Pneumovirus, and genome is the sub-thread strand RNA of non-segmented negative,
Turkey generation respiratory tract disease, referred to as Turkey Rhinotracheitis can be caused；Viral airsacculitis is caused after infected chicken,
Rhinotracheitis and Swollen head syndrome, egg drop reduction.Late 1970s, the disease that fowl metapneumovirus causes
Report in South Africa first, subsequently in North America, South America, the Middle East, the Far East have generation.The U.S. in 1989 is first
It is separated to this virus.While separating this virus, the most separable to bordetella bacilli, pasteurellosis bacillus, vacation
Pseudomonas bacillus, rhinotracheitis ornithosis bacillus, Bacillus foecalis alkaligenes etc..Fowl metapneumovirus can make the work of tracheal cilia
Moving and be suppressed, the dust in trachea discharges difficulty, and a large amount of antibacterials carried in dust are easy to occur
Secondary infection, encroaches on respiratory tract.Cause of disease fowl metapneumovirus can be divided into three types: A type infects turkey, Type B sense
Dye broiler, c-type only exists in the U.S., A and B amphitypy has cross-protection, makees without cross protection with c-type
With.Along with the continuous expansion of China's fowl industry cultivation scale, because fowl metapneumovirus infects the secondary infection caused also
Day by day serious, the harm to aviculture is very serious, and unique effective way is to prevent chicken by vaccination
The morbidity of group.Research to fowl metapneumovirus vaccine the most both at home and abroad has focused largely on the development of inactivated vaccine.
H9 subtype avian influenza belongs to low pathogenicity epidemic disease, but mixed infection special with other cause of diseases can improve it
Pathogenicity, causes egg production degradation and mortality rate to significantly improve, and provisions fowl industrial belt carrys out serious harm.
Existing bird flu (H9 hypotype) vaccine is widely used in clinic, but has that antibody horizontal is relatively low and antibody
Problem in terms of regularity, thus have influence on the prevention effect of vaccine.But it is the most effective in the market
Prevention fowl metapneumovirus and the bigeminy vaccine of H9 subtype avian influenza.
Summary of the invention
It is an object of the invention to provide a kind of fowl metapneumovirus and H9 subtype avian influenza virus bigeminy vaccine, thus
Make up the deficiencies in the prior art.
The fowl metapneumovirus of the present invention and H9 subtype avian influenza virus bigeminy vaccine, include antigen and vaccine assistant
Agent, wherein antigen be deposit number be the H9 subtype avian influenza virus strain of CCTCC NO:V201517 and fowl inclined
Pneumovirinae F protein；
Described fowl metapneumovirus F protein, includes:
1) aminoacid sequence is the albumen of SEQ ID NO:1,
2) 1) albumen on replace, lack, add one or several aminoacid, and have 1) in egg
The albumen of contour painting energy；
Encoding the gene of above-mentioned F protein, its nucleotides sequence is classified as SEQ ID NO:2；
Described fowl metapneumovirus F protein is by pichia pastoris phaff X33-F (Pichia pastoris
X33-F), the China typical culture collection center being preserved in Wuhan University on March 9th, 2016, protect
Hide numbered CCTCC NO:M 2016097.
H9 subtype avian influenza virus and fowl metapneumovirus F protein in above-mentioned vaccine inactivate through formalin；
In above-mentioned vaccine, the content of H9 subtype avian influenza virus is not less than 106.0EID50/0.3ml；The inclined pneumonopathy of fowl
The content of poison F protein is not less than 50 μ g/0.3ml.
H9 subtype avian influenza virus QDY strain is mixed in proportion by the present invention with fowl metapneumovirus F protein antigen liquid
After, the adjuvant mixing and emulsifying that refuels makes vaccine.Vaccine prepared by the present invention can improve the antibody horizontal after immunity,
The regularity of antibody after raising immunity, it is ensured that the immune effect of vaccine, this vaccine has efficiently, safety
Good advantage.
Detailed description of the invention
Accompanying drawing explanation
Figure is identified in the F gene PCR amplification of Fig. 1: the embodiment of the present invention 1；
Fig. 2: the nucleotide sequence comparison chart of F gene of the present invention；
The amino acid alignment figure of Fig. 3: F protein of the present invention；
The enzyme action of Fig. 4: embodiment of the present invention efficient expression vector identifies figure；
The PCR of Fig. 5: embodiment of the present invention recombinant bacterial strain X33-F identifies figure；
Fig. 6: the SDS-PAGE electrophoresis rating diagram of embodiment of the present invention recombinant bacterium fermentation inducement expression product.
Detailed description of the invention
The present invention will be described in detail with specific embodiment below in conjunction with the accompanying drawings.
The amplification of embodiment 1:F albumen and sequence analysis
Within 2010, in Shandong Province, multiple kinds of chicken houses occur in that the symptom of swollen head sydrome, and before the chicken group that falls ill
Through having injected existing avian pneumovirus vaccine, thus it is speculated that the virus of infection there occurs variation；Therefore from Affected individuals
In carry out the screening of avian pneumovirus.Finishing screen have selected a strain avian pneumovirus SHS/A3.
For the antigenicity of the virus of checking screening, employ 5 differences comprising screening strain SHS/A3
The Strain in source prepares vaccine as antigen, carries out counteracting toxic substances with SHS/A3 strain virus liquid real after immunity SPF chicken
Testing, result shows compared to other avian pneumovirus vaccine；The vaccine itself prepared has preferably immunity effect
Really (p ＜ 0.05)；It is thus determined that it there occurs the variation in heredity.
1, amplification Type B fowl metapneumovirus SHS/A3 strain (APV) F gene
According to the F gene order delivered in NCBI, having designed and synthesized primer, the sequence information of primer is as follows:
Primer1:5 '-GGGATGTACCTCAAACTGCTACTAAT-3 '；
primer2:5′-TCAACTGATGTAGCCCATGTTGC-3′。
2, PCR amplification F gene clone and sequencing
The nucleic acid of extraction SHS/A3 strain, as template, carries out PCR amplification with primer primer1 and primer2
Purpose fragment, through sequencing the amplification qualification figure of Type B fowl metapneumovirus F protein gene PCR (Fig. 1 be),
Result nucleotides sequence is classified as SEQ ID NO:2；The aminoacid sequence of its coding is SEQ ID NO:1.With NCBI
In the F gene of Type B fowl metapneumovirus announced carry out nucleotide sequence comparison analysis, result homology exists
95.1%～the 99.2% nucleotide sequence comparison chart of SHS/A3 strain F gene (Fig. 2 be)；The aminoacid derived
Sequence homology be 95.0%～97.4% the amino acid alignment figure of SHS/A3 strain F protein (Fig. 3 be).
Result shows, the SHS/A3 strain of separation is new Type B fowl metapneumovirus, containing new F gene.
3, Type B fowl metapneumovirus (APV) F gene is designed and synthesized
According to the sequencing results of SHS/A3 strain F gene, design synthesis pair of primers, the sequence letter of primer
Cease as follows:
Primer3:5 '-GGGGGTACCATGTACTTGAAGTTGCAATTG-3 '；
primer4:5′-GCGGCCGCTCAACTGATGTAACCCATGT-3′。
Arrange the F gene into SEQ ID NO:2 as template with the nucleotides sequence of synthesis, with primer primer3 with
Primer4 carries out PCR amplification, and purpose fragment products reclaims and connects pMD18-T carrier, converts and screens sun
Sex clone pMD18-T-F.
Embodiment 2:F albumen recombinant expressed
1. the preparation method of Type B fowl metapneumovirus F protein of recombinating
Comprise the following steps: a. construction of expression vector；B. construction expression bacterial strain；The induction of F protein of c. recombinating
And extraction purification.
A. construction of expression vector:
Positive colony plasmid pMD18-T-F and expression vector pPICZ α carrier are used KpnI and NotI respectively
Double digestion product, after 1.2% agarose gel electrophoresis, reclaims test kit with DNA gel and reclaims, obtain respectively about
1.6kb and 3.3kb fragment, (Fig. 4 is the present invention to connect structure pPICZ α-F expression vector 16 DEG C of orientations
The enzyme action of embodiment efficient expression vector identifies figure)；Sequence verification sequence and reading frame errorless after, by plasmid line
After property, electricity is transformed into Pichia sp. competent cell.
B. construction expression bacterial strain:
After electricity converts, in F gene recombinaton to Pichia sp. genome, build Pichiapastoris expression strain X33-F
(being preserved in China typical culture collection center on March 6th, 2016, deposit number is CCTCC M
2016097；Fig. 5)；
The induction of F protein of c. recombinating and extraction purification:
Abduction delivering, the picking monoclonal colony inoculation containing X33-F is in BMGY fluid medium, 30 DEG C of vibrations
Overnight incubation, centrifugal collection thalline, after suspending with appropriate BMMY, addition final concentration 0.5% methanol, 30 DEG C
Induce 96 hours.4 DEG C, 9000rpm is centrifuged 5min, retains supernatant, after adding the ammonium sulfate precipitation of 30%,
12000rpm is centrifuged 5min and collects albumen precipitation, with the PBS molten albumen of weight.Add protein electrophoresis loading buffer
Liquid, boiling water boiling is after 8 minutes, and the separation gel with 12% carries out SDS-PAGE qualification.(in Fig. 61,2,3
Precipitating for F protein expression product, M is molecular weight marker proteins matter.)
Embodiment 3: the preparation of antigen for vaccine
1. the preparation of seedling avian influenza venom is by sub-for the H9 that deposit number is CCTCC NO:V201517
Type bird flu virus QDY strain, through allantoic cavity inoculate 10～11 age in days SPF Embryo Gallus domesticus, every embryo 0.2ml, 36 DEG C～
Hatch for 37 DEG C, per sunshine embryo 2 times, Embryo Gallus domesticus dead after taking 24 hours, to 96 hours, no matter death was the most complete
Portion gathers in the crops, and puts 4～8 DEG C and cools down 12～24 hours, gathers in the crops Embryo Gallus domesticus liquid, be mixed in sterile chamber, put 2～8 DEG C
Preserve.Virus liquid after results measures viral level, and every 0.1ml viral level is not less than 108.0EID50；
2. inactivation is by checking qualified venom to be placed in sterilization container, adds 10% formalin solution to end
Concentration is 0.2%, and 37 DEG C inactivate 18 hours；
3. the seedling preparation of Type B fowl metapneumovirus F protein will produce Type B aviadenovirus fusion protein
Pichiapastoris expression strain X33-F strain is preserved in China typical culture collection center, and deposit number is CCTCC
M 2016097。
Strain is inoculated in the YPD fluid medium containing bleomycin by the preparation of 3.1 seedling bacterium solution,
30 DEG C of shaken cultivation 16～18 hours.Then streak inoculation is in the YPD solid medium added with bleomycin,
Choose colonies typical 2～3 be mixed in a small amount of YPD fluid medium, put shaken cultivation in 30 DEG C of shaking tables
18 hours, quantitative separating, after purely inspection, as first order seed.Take first order seed and be inoculated in BMGY
In fluid medium, 30 DEG C of shaken cultivation 16～18 hours, after microscopy, put 2～8 DEG C of preservations, as
Secondary seed.
The preparation of 3.2 seedling albumen is pressed fermenter volume 60% (V/V) and is added the training of BMGY incomplete liquid
Support base, add defoamer by culture medium 0.1% (V/V) simultaneously, be passed through high-temp steam sterilizing 30 minutes, treat
Culture medium temperature is down to 32 DEG C, adds YNB and biotin, the Type B fowl metapneumovirus of preparation in inoculation 3.1
F protein production secondary seed solution, fermentation tank parameter arranges and is respectively mixing speed 800r/min, temperature
30 DEG C, maintain DO value (dissolved oxygen amount) 20%.Bacterium solution after cultivating 24 hours adds methanol, methanol
Adding speed is that 2ml/h/L abduction delivering is cultivated, according to above ferment control parameter and technique abduction delivering 120
Hour；Fermentation culture bacterium solution is centrifuged 30 minutes through tube centrifuge 10000r/min, the supernatant of results, adds
After entering ammonium sulfate precipitated protein, it is centrifuged 30 minutes results precipitations by 12000r/min, adds appropriate physiology
Saline soluble protein precipitates.
4., in protein liquid is placed in inactivation bottle by inactivation, metering adds 10% formalin, with adding with shaking, makes
It is sufficiently mixed, and the ultimate density of formalin is 0.1%.Pour in another inactivation bottle after adding formalin,
Inactivator is contacted to avoid the virus adhered near bottleneck to fail.37 DEG C inactivation 16 hours after take out, put 2～
8 DEG C of preservations.
5. the inspection of semifinished product
(1) steriling test carries out steriling test by existing " Chinese veterinary pharmacopoeia " annex.
(2) viral level measures and virus liquid sterile saline is made 10 times of serial dilutions, takes 10-5、
10-6、10-7、10-8、10-95 dilution factors, respectively allantoic cavity inoculation 10～11 age in days SPF Embryo Gallus domesticus, every embryo
0.1ml, sets inoculation saline control 5 pieces, every embryo 0.2ml simultaneously.Put 36～37 DEG C to continue to hatch, often
Sunshine, embryo 2 times, observed 96 hours.Measuring the hemagglutinative titer of each chick embryo allantoic liquid, hemagglutinative titer is not less than
24, it is judged to infect and calculate EID50。
(3) determining the protein quantity presses Bradford method detection protein content.
(4) inactivation inspection is by virus liquid allantoic cavity the inoculation 10～11 age in days SPF Embryo Gallus domesticus 10 after inactivation
Piece, every embryo 0.2ml, put 36～37 DEG C and continue to hatch, Continuous Observation 96 hours.Measure each chick embryo allantois
The hemagglutinative titer of liquid, hemagglutinative titer is not less than 24, it is judged to infect and calculate EID50。
Embodiment 4: vaccine preparation and inspection
(1) prepared by vaccine
Semi-finished product antigen through after the assay was approved carries out vaccine and prepares that (in following preparation, each liquid component is by body
Long-pending than meter).
1. oil phase preparation takes white oil for animals 95 parts, aluminium stearate 1 part, is placed in oil phase preparation tank heating
After 80 DEG C, then Jia Siben-80 5 parts, to temperature reach 115 DEG C time, maintain 30min, cool down standby
With.
2. the H9 subtype avian influenza virus liquid of inactivation is used normal saline dilution to being not less than 2 by aqueous phase preparation
×106.0EID50/0.1ml.Use normal saline dilution to being not less than 100 the fowl metapneumovirus F protein of inactivation
μg/0.1ml.Take H9 subtype avian influenza virus liquid and each 1 part of the fowl metapneumovirus F protein liquid of inactivation respectively,
Mixing is made into antigen for vaccine liquid.Take 5 parts of tween 80s after sterilizing, add in Agitation Tank, be simultaneously introduced system
Seedling antigen liquid 95 parts, stirs 20～30min, makes tween 80 be completely dissolved.
3. emulsifying takes oil phase 2 parts and is put in high-speed shearing machine, starts the stirring of motor slow rotation, simultaneously Xu
Xu adds aqueous phase 1 part, with 10000r/min, emulsifying 5 minutes.After emulsifying, take 10ml, with 3000r/min
Centrifugal 15 minutes, the aqueous phase separated out at the bottom of pipe should be less than 0.5ml.
(2) vaccine product inspection
1. character
Outward appearance vaccine should be milky Emulsion, free from admixture and outer package should be qualified.
Dosage form is water-in-oil type.Take a cleaning suction pipe, draw a small amount of vaccine and instill in cold water, except the 1st
Outward, all should indiffusion.
Stability is drawn vaccine 10ml and is added in centrifuge tube, is centrifuged 15 minutes, at the bottom of pipe with 3000r/min
The aqueous phase separated out should be less than 0.5ml.
Viscosity is carried out by existing " Chinese veterinary pharmacopoeia " annex, should meet regulation.
2. loading quantity inspection is carried out by existing " Chinese veterinary pharmacopoeia " annex, should meet regulation.
3. steriling test is carried out by existing " Chinese veterinary pharmacopoeia " annex, should meet regulation.
4. safety verification is with 7 age in days SPF chicken 10, every cervical region subcutaneous injection vaccine 1.0ml, simultaneously
If compareing 5, raise at identical conditions, Continuous Observation 14 days, record test chicken search for food, drink water and
Clinical setting.Any locally and systemically untoward reaction caused by vaccine should be occurred without.
5. efficacy test
With 21 age in days SPF chicken 20, every cervical region subcutaneous injection bigeminy vaccine, 0.3ml/ only, separately takes 20
Only compare with age in days chicken is the most immune, exempt from latter 28 days, counteracting toxic substances.
The counteracting toxic substances of 5.1 pairs of H9 subtype avian influenzas is protected in latter 28 days of immunity, and immune group and matched group respectively take
10, the separation strain of counteracting toxic substances 6 strain each place, every intravenous injection 0.1ml after 10 times of dilutions.Result shows,
Bivalent inactivated vaccine can be resisted each place and separate the attack (seeing table 1) of poison.
The table 1 counteracting toxic substances protection to local epidemic isolates
5.2B type fowl metapneumovirus protest test is in latter 28 days of immunity, the eye dripping counteracting toxic substances inclined lung of Type B fowl
Virus liquid, only, viral level is 10 to 0.1ml/6.5TCID50/0.1ml.Continuous Observation 7 days after counteracting toxic substances, in attacking
Within after poison the 5th day, gather nose swab, isolated viral.Result shows, single with Type B fowl metapneumovirus F protein Asia
Position vaccine immune chicken group energy enough resists the attack of virus, occurs without clinical symptoms, and virus purification is feminine gender.
, there is slight clinical symptoms in matched group, and 9/10 virus purification is positive.
And, use SHS/A3 strain virus to carry out challenge test, result shows exempting from of vaccine prepared by the present invention
Epidemic disease effect is better than other type of fowl metapneumovirus F protein vaccine, subunit vaccine counteracting toxic substances prepared by the present invention
After have 90% protection, and the protective rate of like product is 60%.Result above shows that use aminoacid sequence is
The vaccine that the fowl metapneumovirus F protein of SEQ ID NO:1 is made is same to being better than with the immune effect of SHS/A3 strain
Series products, sickness rate is far below its commercial available vaccines (p ＜ 0.05).Possible reason is the something lost due to F protein
The change of disease is different to be caused.
Claims (6)
1. a fowl metapneumovirus and H9 subtype avian influenza virus bigeminy vaccine, it is characterised in that described
Bigeminy vaccine includes antigen and vaccine adjuvant, wherein antigen be deposit number be CCTCC NO:V201517
H9 subtype avian influenza virus strain and fowl metapneumovirus F protein.
2. bigeminy vaccine as claimed in claim 1, it is characterised in that described fowl metapneumovirus F protein
Include:
1) aminoacid sequence is the albumen of SEQ ID NO:1,
2) 1) albumen on replace, lack, add one or several aminoacid, and have 1) in egg
The albumen of contour painting energy.
3. bigeminy vaccine as claimed in claim 2, it is characterised in that described F protein, it encodes base
The nucleotides sequence of cause is classified as SEQ ID NO:2.
4. bigeminy vaccine as claimed in claim 1 or 2, it is characterised in that described fowl metapneumovirus F
Albumen is by the X33-F recombinant expressed production of bacterial strain that deposit number is CCTCC M 2016097.
5. bigeminy vaccine as claimed in claim 1, it is characterised in that described H9 subtype avian influenza is sick
Poison and fowl metapneumovirus F protein inactivate through formalin.
6. bigeminy vaccine as claimed in claim 1, it is characterised in that described H9 subtype avian influenza is sick
The content of poison is not less than 106.0EID50/0.3ml；The content of fowl metapneumovirus F protein is not less than 50 μ g/0.3ml.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610236341.8A CN105833263B (en) | 2016-04-14 | 2016-04-14 | Bivalent vaccine of avian metapneumovirus and H9 subtype avian influenza virus |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610236341.8A CN105833263B (en) | 2016-04-14 | 2016-04-14 | Bivalent vaccine of avian metapneumovirus and H9 subtype avian influenza virus |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN105833263A true CN105833263A (en) | 2016-08-10 |
| CN105833263B CN105833263B (en) | 2020-11-27 |
Family
ID=56589578
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201610236341.8A Active CN105833263B (en) | 2016-04-14 | 2016-04-14 | Bivalent vaccine of avian metapneumovirus and H9 subtype avian influenza virus |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN105833263B (en) |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106754752A (en) * | 2017-01-06 | 2017-05-31 | 广东华南联合疫苗开发院有限公司 | One plant of new RSV virus and its application |
| CN107412762A (en) * | 2017-08-09 | 2017-12-01 | 青岛易邦生物工程有限公司 | A kind of ewcastle disease, bird flu, the bursa of farbricius and aviadenovirus quadruple vaccine |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104922663A (en) * | 2015-07-07 | 2015-09-23 | 青岛易邦生物工程有限公司 | New castle disease and H9 subtype bird flu bivalent vaccine |
| CN104946600A (en) * | 2015-07-07 | 2015-09-30 | 青岛易邦生物工程有限公司 | H9 subtype avian influenza virus strain |
-
2016
- 2016-04-14 CN CN201610236341.8A patent/CN105833263B/en active Active
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104922663A (en) * | 2015-07-07 | 2015-09-23 | 青岛易邦生物工程有限公司 | New castle disease and H9 subtype bird flu bivalent vaccine |
| CN104946600A (en) * | 2015-07-07 | 2015-09-30 | 青岛易邦生物工程有限公司 | H9 subtype avian influenza virus strain |
Non-Patent Citations (3)
| Title |
|---|
| KC542808: "Avian metapneumovirus strain aMPV/B/IT/GuineaFowl/1818/12 fusion protein (F) gene, complete cds", 《GENBANK》 * |
| 胡海霞等: "禽偏肺病毒分子生物学及基因工程疫苗研究进展", 《中国家禽》 * |
| 薛聪等: "1株B亚型禽偏肺病毒的分离与鉴定", 《中国兽医学报》 * |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106754752A (en) * | 2017-01-06 | 2017-05-31 | 广东华南联合疫苗开发院有限公司 | One plant of new RSV virus and its application |
| CN106754752B (en) * | 2017-01-06 | 2020-03-24 | 广东华南疫苗股份有限公司 | RSV (respiratory syncytial virus) and application thereof |
| CN107412762A (en) * | 2017-08-09 | 2017-12-01 | 青岛易邦生物工程有限公司 | A kind of ewcastle disease, bird flu, the bursa of farbricius and aviadenovirus quadruple vaccine |
| CN107412762B (en) * | 2017-08-09 | 2020-08-14 | 青岛易邦生物工程有限公司 | Newcastle disease, avian influenza, bursa of fabricius and avian adenovirus quadruple vaccine |
Also Published As
| Publication number | Publication date |
|---|---|
| CN105833263B (en) | 2020-11-27 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN107899008B (en) | Sick three subunit vaccines of a kind of pig epidemic diarrhea, transmissible gastroenteritis of swine, pig fourth type coronavirus | |
| CN107412762B (en) | Newcastle disease, avian influenza, bursa of fabricius and avian adenovirus quadruple vaccine | |
| CN104946600A (en) | H9 subtype avian influenza virus strain | |
| CN105733987A (en) | Mycoplasma synoviae | |
| CN105833263A (en) | Bivalent vaccine of acian metapneumovirus and H9 subtype avian influenza virus | |
| CN103937817B (en) | Newcastle disease virus YT strain, its whole genome sequence and application thereof | |
| CN105770881A (en) | Mycoplasma gallisepticum (MG) and mycoplasma synoviae (MS) combined inactivate vaccine | |
| CN103468647B (en) | Swine flu H1N1 and H3N2 subtype bivalent inactivated vaccine | |
| CN106146626A (en) | A kind of erysipelothrix ruhsiopathiae subunit vaccine and preparation method and application | |
| CN104530230A (en) | Duck hepatitis A virus VP1 protein gene and application thereof | |
| CN104448005B (en) | The fusion rotein antigen of duck hepatitis A virus (HAV) 3 type VP1 albumen and LTB and application thereof | |
| CN104274829B (en) | A kind of vaccine combination and its preparation method and application | |
| CN103789272B (en) | H9 subtype avian influenza virus separation strain and the vaccine combination prepared by it | |
| CN106854647A (en) | Duck virus hepatitis divalence yolk antibody and its preparation method and application | |
| CN104940921A (en) | H9 subtype avian influenza virus inactivated vaccine including chicken a-interferon protein | |
| CN104195114A (en) | Avian pneumovirus and application thereof | |
| CN105861349B (en) | A kind of recombinant bacterial strain producing Type B fowl metapneumovirus F protein | |
| CN106039304A (en) | Porcine parvovirus, porcine epizootic diarrhea and Escherichia coli triple-antigen vaccine | |
| CN105753949A (en) | Erysipelothrix rhusiopathiae antigen protein and application thereof | |
| CN105770884A (en) | Type B acian metapneumovirus subunit vaccine | |
| CN110013549A (en) | A kind of Hepatitis E subunit vaccine | |
| CN105693829A (en) | B-type avian metapneumovirus F protein | |
| CN105879023B (en) | Bivalent inactivated vaccine for porcine parvovirus and porcine epidemic diarrhea | |
| CN105950492A (en) | Recombination strain for producing porcine parvovirus VP2 protein | |
| CN103937752B (en) | A kind of H3N2 hypotype canine influenza virus inactivated vaccine and its preparation method and application |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |