CN112494641B - Live vaccine for avian escherichia coli - Google Patents
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Abstract
The invention provides an avian Escherichia coli live vaccine, which is a low virulent strain prepared by removing aroA gene on the basis of obtaining a novel avian pathogenic Escherichia coli O78 by screening, and is used for preparing the vaccine; the low virulent strain keeps the natural immunogenicity of the epidemic strain Escherichia coli O78 strain, and simultaneously, the toxicity is obviously weakened. The avian escherichia coli live vaccine provided by the invention comprises an antigen and an adjuvant, wherein the antigen is a live YBO78-1 strain with a preservation number of CCTCC M2020383. The vaccine provided by the invention is also used for preparing products for controlling poultry; the poultry comprise chicken, duck, turkey, goose, bantam, quail, pheasant, pigeon, etc. The E.coli vaccine provided by the present invention can be used by conventional methods, such as large scale administration by economical and easy-to-handle spraying and drinking water, or by intramuscular injection or other means.
Description
Technical Field
The invention belongs to the technical field of veterinary biological products, and particularly relates to a live avian escherichia coli vaccine.
Background
Avian colibacillosis is an economically important infectious disease of poultry, the etiology being Avian Pathogenic Escherichia Coli (APEC). APECs are commonly found in the intestinal tract of poultry and wild birds and on the surface of other mucous membranes, and include serotypes O1, O2, O78, O8 and O35, with serotype O78 being the most common serotype in clinical isolation. The most severe manifestation of escherichia coli infection in poultry is septicemia, usually starting from primary mycoplasma or upper respiratory infection after viral infection, leading to infiltration of blood and internal organs, ultimately causing diseases such as pericarditis, hepatitis, air cystitis, and salpingitis.
The avian colibacillosis has short onset time and high mortality rate, and antibiotics are mainly used for treating bacterial infectious diseases in the breeding industry at present. However, the long-term use of antibiotics with a dose lower than the therapeutic dose in the animal body can greatly accelerate the appearance of drug-resistant bacteria, and if the drug-resistant bacteria are spread in a large range in the cultured animals, the cultured animals become a large drug-resistant gene storage bank. Therefore, prevention and control of APEC by a non-antibiotic method through vaccine is a necessary trend for future development, wherein vaccination is an important way for effectively preventing and controlling occurrence and prevalence of avian colibacillosis, reducing avian product pollution and ensuring food safety. However, most of the conventional escherichia coli vaccines are inactivated vaccines and are prepared into oil emulsion vaccines and aluminum hydroxide gel vaccines, the oil emulsion vaccines have large side reactions, the aluminum hydroxide gel vaccines have poor immune effect, and the use effect of the escherichia coli inactivated vaccines in the market is not ideal.
Disclosure of Invention
The invention aims to provide an avian Escherichia coli live vaccine, which is a low virulent strain prepared by removing aroA gene on the basis of screening to obtain a novel avian pathogenic Escherichia coli O78, and is used for preparing the vaccine; the low virulent strain keeps the natural immunogenicity of the epidemic strain Escherichia coli O78 strain, and simultaneously has obviously weakened toxicity, thereby making up the defects of the prior art.
The avian Escherichia coli live vaccine provided by the invention comprises an antigen and an adjuvant, wherein the antigen is a live YBO78-1 strain, and the preservation number of the avian Escherichia coli (Escherichia coli) is CCTCC M2020383;
the avian pathogenic Escherichia coli (Escherichia coli) YBO78-1 strain is prepared by deleting aroA gene of avian pathogenic Escherichia coli (Escherichia coli) YBO78 strain with the preservation number of CCTCC M2020382;
the YBO78-1 strain is prepared by deleting aroA genes of YBO78 strain through a Red/ET homologous recombination system.
The vaccine adjuvant is an aqueous vaccine adjuvant.
The vaccine provided by the invention is also used for preparing products for controlling poultry;
the poultry include chicken, duck, turkey, goose, bantam, quail, pheasant, pigeon, etc., preferably commercially important poultry such as chicken, duck, goose, turkey, more preferably chicken and turkey, and particularly preferably chicken.
The E.coli vaccine provided by the present invention can be used by conventional methods, such as large scale administration by economical and easy-to-handle spraying and drinking water, or by intramuscular injection or other means.
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FIG. 1: PCR identification of Escherichia coli O78 and O78 aroA gene knock-out strains.
Detailed Description
The present invention uses conventional techniques and methods used in the fields of genetic engineering and molecular biology, such as the methods described in A new logic for DNA engineering recombination in Escherichia coli (zhang ET al, Nature genetics 1998), Rapid modification of bacterial microorganisms by ET-recombination (Muyres ET al, Nucleic acids research 1993).
The aroA gene of bacteria encodes 5-enolpyruvylshikimate-3-phosphate synthetase, the enzyme has important significance for anabolism of aromatic compounds such as aromatic amino acid of bacteria, and the deletion of the gene can block an aromatic biosynthesis pathway in bacteria.
The Red/ET homologous recombination technology is derived from lamda bacteriophage, can be used for gene knockout, insertion, point mutation and the like of escherichia coli genome, constructs a pathogenic escherichia coli aroA gene deletion strain by utilizing the Red/ET homologous recombination technology, and evaluates the potential of the gene deletion strain as a vaccine candidate strain on the basis of the biological characteristics and the pathogenicity of the pathogenic escherichia coli aroA gene deletion strain.
The avian Escherichia coli YBO78 provided by the invention is a virulent strain obtained by screening and culturing the organ tissues of Escherichia coli-infected chickens in a large chicken farm in Qingdao; YBO78-1 is an O78 strain aroA gene deletion strain obtained by a Red/ET homologous recombination system, which leads to attenuated virulence, and the strain is used for preparing live vaccines, thereby providing more complete immune protection for chickens.
The present invention will be described in detail with reference to examples.
Example 1: separation and purification of strain and analysis of physicochemical properties
1) A chicken farm in Shandong in 6 months in 2018 has a serious epidemic situation of escherichia coli, organs, such as hearts, livers, spleens and the like of dead chickens are collected and placed in a sampling bag for sealing, and the sampling bag is taken back to a laboratory and placed in an ice box at 4 ℃ for storage for later use.
2) Grinding collected chicken organs, spreading on chromogenic culture medium, inoculating 3 strains with different colors as a group to LB liquid culture medium for amplification culture, centrifuging at 10000rpm to collect thallus, resuspending the thallus with 0.5% sterile PBS, and turbidimetry to obtain bacterial concentration of 1 × 107CFU/ml。
3) Strong toxicity separation experiment:
culturing the separated strains in common broth medium for 37 hr respectively, and determining the number of the strains to be 1.0 × 10640 chickens aged 1 day were randomly divided into 4 groups of 10 chickens each, each group was intraperitoneally injected with 0.2ml of a serum type strain of bacteria per chicken, and a control group was injected with 0.2ml of a common broth, and after inoculation, observation was carried out for 72 hours in an isolator, the number of deaths of chickens was recorded and bacteria were isolated from their livers and spleens, and then the surviving chickens were dissected to observe the lesions (Table 1).
Table 1: experimental table of 1 day old chicken attacked by Escherichia coli O1, O2 and O78
| Name of drug | O1 | O2 | O78 |
| Number of deaths | 7 | 8 | 10 |
| Number of survivors | 3 | 2 | 0 |
4) Extracting the DNA of the bacteria by using a bacterial genome DNA extraction kit, performing high-fidelity PCR amplification by using universal primers 1492F (5-CGG TTA CCT TGT TAC GAC TT-3) and 27F (5-AGA GTT TGA TCM TGG CTC AG-3) for 16S rRNA bacterial identification, cutting a target band after recovery by agarose gel electrophoresis, sending the product to Shanghai Biometrics Limited company for sequencing, and performing NCBI blast analysis on the sequencing result, wherein the 16S rRNA sequence is as follows (SEQ ID NO: 1):
TGCAGTCGAACGGTAACAGGAAGCAAGCTTGCTTCTTCGCTGACGAGTGGCGGACGGGTGAGTAATGTCTGGGAAACTGCCTGATGGAGGGGGATAACTACTGGAAACGGTAGCTAATACCGCATAACGTCGCAAGACCAAAGAGGGGGACCTTCGGGCCTCTTGCCATCGCATGTGCCCAGATGGGATTAGCTAGTAGGTGGGGTAACGGCTCACCTAGGCGACGATCCCTAGCTGGTCTGAGAGGATGACCAGCCACACTGGAACTGAGACACGGTCCAGACTCCTACGGGAGGCAGCAGTGGGGAATATTGCACAATGGGCGCAAGCCTGATGCAGCCATGCCGCGTGTATGAAGAAGGCCTTCGGGTTGTAAAGTACTTTCTGCGGGGAGGAAGGGAGTAAAGTTAATACCTTTGCTCATTGACGTTACCCGCAGAAGAAGCACCGGCTAACTCCGTGCCAGCAGCCGCGGTAATACGGAGGGTGCAAGCGTTAATCGGAATTACTGGGCGTAAAGCGCACGCAGGCGGTTTGTTAAGTCAGATGTGAAATCCCCGGGCTCAACCTGGGAACTGCGTCTGATACTGGCAAGCTTGAGTCTCGTAGAGGGGGGTAGAATTCCAGGTGTAGCGGTGAAATGCGTAGAGATCTGGAGGAATACCGGTGGCGAAGGCGGCCCCCTGGACGAAGACTGACGCTCAGGTGCGAAAGCGTGGGGAGCAAACAGGATTAGATACCCTGGTAGTCCACGCCGTAAACGATGTCGACTTGGAGGTTGTGCCCTTGAGGCGTGGCTTCCGGAGCTAACGCGTTAAGTCGACCGCCTGGGGAGTACGGCCGCAAGGTTAAAACTCAAATGAATTGACGGGGGCCCACACAAGCGGTGGAGCATGTGGTTTAATTCGATGCAACGCGAAGAACCTTACCTGGTCTTGACATCCACAGAACTTTCCAGAGATGGATTGGTGCCTTCGGGAACTGTGAGACAGGTGCTGCATGGCTGTCGTCAGCTCGTGTTGTGAAATGTTGGGTTAAGTCCCGCAACGAGCGCAACCCTTATCCTTTGTTGCCAGCGGTCCGGCCGGGAACTCAAAGGAGACTGCCAGTGATAAACTGGAGGAAGGTGGGGATGACGTCAAGTCATCATGGCCCTTACGACCAGGGCTACACACGTGCTACAATGGCGCATACAAAGAGAAGCGACCTCGCGAGAGCAAGCGGACCTCATAAAGTGCGTCGTAGTCCGGATTGGAGTCTGCAACTCGACTCCATGAAGTCGGAATCGCTCGTAATCGTGGATCAGAATGCCACGGTGAATACGTTCCCGGGCCTTGTACACACCGCCCGTCACACCATGGGAGTGGGTTGCAAAAGAAGTAGGTAGCTTAACCTTCGGGAGGGCGC。
the separated bacteria are determined to be escherichia coli, and serotype detection is utilized to confirm that the screened strains are avian pathogenic escherichia coli of three serotypes, namely escherichia coli 01, O2 and O78. Experimental results the isolated escherichia coli serotype O78 was a virulent strain, designated escherichia coli YBO 78. The strain YBO78 was preserved in China center for type culture Collection, located in Lophania japonica, Wuhan, Wuchang, 31/7/2020, with the preservation number of CCTCC M2020382.
5) Drug susceptibility testing
The drug sensitivity test adopts a paper method, and the cultured strain is diluted to 1 × 10 with normal saline10After CFU/ml, 0.1ml of bacterial liquid is sucked and evenly coated on a nutrient agar culture medium; the drug sensitive paper pieces were clipped with sterile forceps and attached to a flat plate (6 pieces/plate, not less than 3cm apart), incubated at 37 ℃ for 24h, and the results were observed and the zone size was measured with a vernier caliper (Table 2). Drug sensitivity experiments show that the YBO78 strain is sensitive to kanamycin and chloramphenicol and is sensitive to amoxicillin, streptomycin, cefazolin and the likeAntibiotics are resistant.
Table 2: separated YBO78 drug sensitivity experiment table
| Name of drug | Dosage mu g/tablet | Bacterial inhibition zone diameter/mm of isolate |
| Tetracycline derivatives | 20 | 7 |
| Amoxicillin | 20 | 0 |
| Kanamycin | 30 | 10 |
| Streptomycin | 20 | 0 |
| Cefmazole | 20 | 0 |
| Ciprofloxacin | 20 | 0 |
| Enrufloxacin hydrochloride | 20 | 0 |
| Gentamicin | 20 | 9 |
| Ofloxacin | 20 | 0 |
| Chloromycetin | 30 | 15 |
| Hygromycin | 20 | 0 |
| Ampicillin | 20 | 5 |
| Streptomycin | 20 | 21 |
Note: the judgment standard of the drug sensitivity test is as follows: the phi bacteriostatic ring has the diameter phi larger than 19mm, is highly sensitive, has the phi not less than 15mm and not more than 19mm, is moderately sensitive, and has the phi less than 15mm, and is resistant.
Example 2: knockout and detection of aroA gene of avian Escherichia coli YBO78
1) According to the aroA gene sequence of avian Escherichia coli O78 on NCBI GenBank, designing sequencing primers (table 3), carrying out PCR amplification on the aroA gene of a clinical isolate YBO78, and carrying out sequencing and identification.
Table 3: sequence information Table of sequencing primers
| Upstream sequencing primer | Downstream sequencing primer |
| O78-seq-F1 | O78-seq-R1 |
| O78-seq-F2 | O78-seq-R2 |
| O78-seq-F3 | O78-seq-R3 |
2) Primers for aroA gene knock-out were designed based on the sequencing results (Table 4).
Table 4: primer sequence table for aroA gene knockout
3) Constructing a recombinant plasmid pSC101-BAD-gbaA-cm, and electrically transferring the plasmid into an escherichia coli YBO78 strain after the sequencing is correct.
4) Arabinose induction, the kanamycin gene cassette loxp-kan-loxp was electrotransferred into E.coli YBO78 to replace aroA gene, and spread on kanamycin-resistant plates. Screening positive clones, extracting genome, PCR, sequencing and identifying. The correct single clone was picked and named O78-loxp-kan-loxp.
5) The recombinant plasmid pSC101-BAD-cre-cm was electrotransferred into O78-loxp-kan-loxp strain and plated on chloramphenicol plates.
6) Adding 1mM arabinose to induce the expression of cre enzyme, allowing the cre enzyme to recognize loxp sites at two sides of the kanamycin gene cassette so as to delete the middle kan gene, culturing overnight, coating on a plate without an antibody, and picking out a monoclonal to extract a genome for detection.
7) And verifying whether the Escherichia coli aroA deletion strain is successfully constructed or not through PCR specificity detection and DNA sequencing results.
8) And (3) carrying out multiple passages on the strains with correct sequencing verification, and taking the strains with stable properties as the vaccine strains to be screened, wherein the strains are named as YBO78-1 (figure 1).
The strain YBO78-1 is preserved in China center for type culture Collection, with the preservation number of CCTCC M2020383, in Wuhan, Wuchang Lojia mountain, 7 and 31 months in 2020.
Example 4: evaluation of biological safety of YBO78-1 Strain
Firstly, streaking and inoculating a strain YBO78-1 of escherichia coli to a non-resistant LB solid plate culture medium, culturing overnight at 37 ℃, then selecting a single bacterial colony to be inoculated to 50ml of a non-resistant LB liquid culture medium, placing the culture medium in a small oscillator, culturing overnight at 37 ℃ and 200rpm, centrifuging at 10000rpm for 1min, collecting a precipitate, carrying out heavy suspension on the precipitate by using sterile PBS, carrying out viable bacteria counting on a bacterial suspension, and preparing for subsequent animal injection infection experiments.
25 SPF chickens of 30 days old were selected and randomly divided into 5 groups of 5 chickens each. Diluting the YBO78-1 bacterial liquid to 1.0 multiplied by 108CFU/ml、2.0×108CFU/ml、4.0×108CFU/ml、8.0×108Four different CFU/ml gradients were used to inoculate 5 test SPFs 1.0 ml/one, and 1 additional group was injected with 1.0ml of isolated wild YBO78 strain as a control.
The isolator was observed for 14 days and clinical morbidity was recorded. The result shows that SPF chickens injected with the separated wild YBO78 strain of the control group die in one chicken within 24 hours, and the rest four chickens die in 72 hours; while the experimental group was 1.0X108CFU/ml、2.0×108CFU/ml、4.0×108CFU/ml、8.0×108None of the four different concentrations of CFU/ml resulted in the development of SPF-chickens (Table 5).
Table 5: virulence detection experiment table of avian Escherichia coli YBO78-1 strain
The results show that the bacterial count of the avian Escherichia coli YBO78-1 strain can reach 8.0 multiplied by 108CFU also did not cause the onset of the disease in chickens, and the injection amount of the control group was 1.0X108The chicken is attacked and all the chicken die when CFU is used, which shows that the strain YBO78-1 screened by the invention has good biological safety.
Example 5: preparation of vaccine by using avian Escherichia coli strain YBO78-1
1) Preparing a live vaccine:
streaking avian Escherichia coli YBO78-1 on an improved Martin plate, standing and culturing at 37 deg.C for 20-24h, and taking out for use. 2-3 colonies from the cultured plate strains are picked in a test tube liquid culture medium, fixed in a shaking table at an inclination angle of 45 degrees, subjected to shake culture at 37 ℃ and 200rpm overnight (12-16h), and taken out for later use. The cultured cells were inoculated into a medium (baffle flask, 300 ml/flask) in a triangular flask at a ratio of 2%, and each was cultured in 1 flask. Culturing at 37 deg.C and 200rpm for 5 hr with shaking, and taking out the bacterial liquid. Adding a freeze-drying protective agent according to the ratio of the bacterial liquid to the protective agent of 9:1, freeze-drying and preserving strains, counting live bacteria of the freeze-dried strains, diluting with improved Martin broth, and then diluting with 1 ml/feather. Wherein the adjuvant and the protective agent can be selected from conventional agents.
2) Immune effect of vaccine
Randomly dividing SPF chickens of 30 days old into four groups, wherein each group comprises 10 SPF chickens; the experimental group contained 30 animals, and the control group contained 10 animals. Respectively intramuscular injecting 30 SPF chickens with YBO78-1 vaccine strains, intramuscular injecting normal saline into a control group, feeding and observing for 14 days in an isolator, and then respectively carrying out challenge experiments on the experimental group and the control group by using wild type O78.
The chickens were kept in the isolator for 14 days and the morbidity and mortality of each group of test chickens were recorded. The results of the experiment are shown in table 6 below.
Table 6: data table of immune effect by using YBO78-1 vaccine strain
| Group of | Route of attacking toxin | Number of attacking toxin (only) | Number of survivors (only) | Survival rate (%) |
| Vaccine group 1 | Intramuscular injection | 10 | 9 | 90 |
| |
Intramuscular injection | 10 | 8 | 80 |
| Control group | Intramuscular injection | 10 | 0 | 0 |
The experimental results show that the vaccine prepared by the strain YBO78-1 can completely protect the infection of the wild strain YBO78, and the experimental animals do not show any clinical symptoms and have no death phenomenon.
In order to detect the cross-immunity efficacy of the YBO78-1 vaccine strain on different serotypes of escherichia coli, vaccines prepared by the YBO78-1 vaccine strain for SPF chickens are respectively immunized, then virus challenge protection experiments are respectively carried out on the serotypes of escherichia coli such as O1, O2, O8 and O35, the vaccines are respectively fed in an isolator for 14 days, and the morbidity and mortality of each group of test chickens are recorded. The results of the experiment are shown in table 7 below.
Table 7: data table of cross-immunity effect by using YBO78-1 vaccine strain
| Group of | Route of attacking toxin | Number of attacking toxin (only) | Survival number (only) | Survival rate (%) |
| O1 | Intramuscular injection | 10 | 8 | 80 |
| O2 | Intramuscular injection | 10 | 9 | 90 |
| O8 | Intramuscular injection | 10 | 9 | 90 |
| O35 | Intramuscular injection | 10 | 9 | 90 |
The experimental results show that the vaccine prepared by the YBO78-1 strain can provide better cross protection for the infection of Escherichia coli strains O1, O2, O8 and O35 separated in the market.
In order to detect whether the vaccination mode of the YBO78-1 vaccine strain affects the immunization effect, three vaccination modes of intramuscular injection, spraying and drinking are respectively selected for vaccine immunization, a challenge experiment is carried out by using a wild strain O78 after 14 days, the wild strain is fed in an isolator for 14 days, and the morbidity and mortality of each group of test chickens are recorded. The results of the experiment are shown in table 8 below.
Table 8: immune effect data table of different inoculation modes by using YBO78-1 vaccine strain
The experimental results show that the vaccine strain prepared by the YBO78-1 strain can completely protect the infection of the wild YBO78 strain by three inoculation modes of intramuscular injection, spraying and drinking, and the experimental animals do not show any clinical symptoms and have no death phenomenon.
In order to evaluate the immune effect of the YBO78-1 vaccine strain and the Escherichia coli vaccine sold in the markets at home and abroad, the Escherichia coli vaccine of Shuiteng company is used
Coli as a control group. Are used separatelyYBO78-1 vaccine strain and
and E, immunizing SPF (specific pathogen free) chickens by using coli, performing a challenge experiment by using a wild strain O78 after 14 days, feeding and observing in an isolator for 14 days, and recording the morbidity and mortality of each group of test chickens. The results of the experiment are shown in table 9 below.
Table 9: use of the YBO78-1 vaccine strain and
coli immune effect data sheet
The result shows that the vaccine strain prepared by the YBO78-1 strain of the invention and the vaccine sold in the market
Coli can protect the infection of the wild YBO78 strain and is a vaccine
Coli cannot provide complete protection (9/10), but the vaccine strain prepared with YBO78-1 of the present invention can provide complete protection (10/10), and the group of experimental animals does not show any clinical symptoms and does not die.
In conclusion, the artificially modified avian escherichia coli YBO78-1 strain has good immunogenicity, and is found by being made into a live vaccine immune animal, the vaccine is safe for chickens, can induce and generate higher neutralizing antibodies, can protect the attack of serotype virulent strains such as escherichia coli O78 and O1 which are popular at present, generates an ideal protection effect, and has a wide application prospect.
Sequence listing
<110> Qingdao Yibang bioengineering Co., Ltd
<120> avian Escherichia coli live vaccine
<160> 1
<170> SIPOSequenceListing 1.0
<210> 1
<211> 1407
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 1
tgcagtcgaa cggtaacagg aagcaagctt gcttcttcgc tgacgagtgg cggacgggtg 60
agtaatgtct gggaaactgc ctgatggagg gggataacta ctggaaacgg tagctaatac 120
cgcataacgt cgcaagacca aagaggggga ccttcgggcc tcttgccatc gcatgtgccc 180
agatgggatt agctagtagg tggggtaacg gctcacctag gcgacgatcc ctagctggtc 240
tgagaggatg accagccaca ctggaactga gacacggtcc agactcctac gggaggcagc 300
agtggggaat attgcacaat gggcgcaagc ctgatgcagc catgccgcgt gtatgaagaa 360
ggccttcggg ttgtaaagta ctttctgcgg ggaggaaggg agtaaagtta atacctttgc 420
tcattgacgt tacccgcaga agaagcaccg gctaactccg tgccagcagc cgcggtaata 480
cggagggtgc aagcgttaat cggaattact gggcgtaaag cgcacgcagg cggtttgtta 540
agtcagatgt gaaatccccg ggctcaacct gggaactgcg tctgatactg gcaagcttga 600
gtctcgtaga ggggggtaga attccaggtg tagcggtgaa atgcgtagag atctggagga 660
ataccggtgg cgaaggcggc cccctggacg aagactgacg ctcaggtgcg aaagcgtggg 720
gagcaaacag gattagatac cctggtagtc cacgccgtaa acgatgtcga cttggaggtt 780
gtgcccttga ggcgtggctt ccggagctaa cgcgttaagt cgaccgcctg gggagtacgg 840
ccgcaaggtt aaaactcaaa tgaattgacg ggggcccaca caagcggtgg agcatgtggt 900
ttaattcgat gcaacgcgaa gaaccttacc tggtcttgac atccacagaa ctttccagag 960
atggattggt gccttcggga actgtgagac aggtgctgca tggctgtcgt cagctcgtgt 1020
tgtgaaatgt tgggttaagt cccgcaacga gcgcaaccct tatcctttgt tgccagcggt 1080
ccggccggga actcaaagga gactgccagt gataaactgg aggaaggtgg ggatgacgtc 1140
aagtcatcat ggcccttacg accagggcta cacacgtgct acaatggcgc atacaaagag 1200
aagcgacctc gcgagagcaa gcggacctca taaagtgcgt cgtagtccgg attggagtct 1260
gcaactcgac tccatgaagt cggaatcgct cgtaatcgtg gatcagaatg ccacggtgaa 1320
tacgttcccg ggccttgtac acaccgcccg tcacaccatg ggagtgggtt gcaaaagaag 1380
taggtagctt aaccttcggg agggcgc 1407
Claims (2)
1. The live avian Escherichia coli vaccine is characterized by comprising an antigen and an adjuvant, wherein the antigen is a live YBO78-1 strain with the preservation number of CCTCC M2020383.
2. The avian escherichia coli live vaccine according to claim 1, wherein the vaccine adjuvant is an aqueous vaccine adjuvant.
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Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1822855A (en) * | 2003-05-14 | 2006-08-23 | 惠氏公司 | Avian e. coli vaccine for protection against colibacillosis |
| CN104774796A (en) * | 2015-04-29 | 2015-07-15 | 中国农业科学院上海兽医研究所 | Inactivated vaccine for poultry pathogenic escherichia coli and preparation method thereof |
| CN105861404A (en) * | 2016-04-22 | 2016-08-17 | 中国农业科学院上海兽医研究所 | Avian pathogenic escherichia coli strain and application thereof in vaccine preparation |
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2020
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Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1822855A (en) * | 2003-05-14 | 2006-08-23 | 惠氏公司 | Avian e. coli vaccine for protection against colibacillosis |
| CN104774796A (en) * | 2015-04-29 | 2015-07-15 | 中国农业科学院上海兽医研究所 | Inactivated vaccine for poultry pathogenic escherichia coli and preparation method thereof |
| CN105861404A (en) * | 2016-04-22 | 2016-08-17 | 中国农业科学院上海兽医研究所 | Avian pathogenic escherichia coli strain and application thereof in vaccine preparation |
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