+

WO1992007582A1 - Corynebacteries et autres organismes apparentes utilises comme vecteurs de vaccin - Google Patents

Corynebacteries et autres organismes apparentes utilises comme vecteurs de vaccin Download PDF

Info

Publication number
WO1992007582A1
WO1992007582A1 PCT/AU1991/000471 AU9100471W WO9207582A1 WO 1992007582 A1 WO1992007582 A1 WO 1992007582A1 AU 9100471 W AU9100471 W AU 9100471W WO 9207582 A1 WO9207582 A1 WO 9207582A1
Authority
WO
WIPO (PCT)
Prior art keywords
pseudotuberculosis
corynebacterium
gene
vaccine vector
organism
Prior art date
Application number
PCT/AU1991/000471
Other languages
English (en)
Inventor
Anthony John Radford
Adrian Leslie Mark Hodgson
Original Assignee
Commonwealth Scientific And Industrial Research Organisation
Priority date (The priority date 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 date listed.)
Filing date
Publication date
Application filed by Commonwealth Scientific And Industrial Research Organisation filed Critical Commonwealth Scientific And Industrial Research Organisation
Priority to JP3516314A priority Critical patent/JPH06501847A/ja
Publication of WO1992007582A1 publication Critical patent/WO1992007582A1/fr

Links

Classifications

    • 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
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
    • C12N15/74—Vectors or expression systems specially adapted for prokaryotic hosts other than E. coli, e.g. Lactobacillus, Micromonospora
    • C12N15/77—Vectors or expression systems specially adapted for prokaryotic hosts other than E. coli, e.g. Lactobacillus, Micromonospora for Corynebacterium; for Brevibacterium
    • 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
    • A61K39/02—Bacterial antigens
    • A61K39/05—Actinobacteria, e.g. Actinomyces, Streptomyces, Nocardia, Bifidobacterium, Gardnerella, Corynebacterium; Propionibacterium
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/04—Antibacterial agents
    • 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
    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/14—Hydrolases (3)
    • C12N9/16—Hydrolases (3) acting on ester bonds (3.1)
    • C12N9/18—Carboxylic ester hydrolases (3.1.1)
    • C12N9/20—Triglyceride splitting, e.g. by means of lipase
    • 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

Definitions

  • the present invention relates generally to the use of corynebacteria and related organisms and/or mutants or derivatives thereof as vaccine vectors.
  • Corynebacterium pseudotuberculosis which is the causative agent of caseous lymphadenitis (CLA), commonly known as “cheesy gland” . This disease affects mainly sheep and goats but can also affect horses.
  • the major virulence factor of C. pseudotuberculosis appears to be a.31 kDa exotoxin called phospholipase D (PLD). Sheep can be protected from CLA by vaccination with a formalin-treated precipitate of C. pseudotuberculosis culture supernatant or using a toxoid form of pure PLD.
  • PLD phospholipase D
  • one aspect of the present invention relates to a vaccine vector comprising a Corynebacterium or related organism which is incapable of expressing an active virulence factor or which synthesises an immunoprotection effective amount of an antigen from a pathogenic organism.
  • the term "vaccine vector" is used in its widest sense to include a biological means for antigen presentation.
  • the biological means is conveniently a microorganism and is generally viable although dead organisms could be employed.
  • the biological vector is non-pathogenic or rendered avirulent or is given in non-pathogenic or avirulent effective amounts.
  • antigen presentation means expression of naturally occurring antigens and/or expression of recombinant antigens such as by transforming the biological vector with a plasmid carrying a gene or genes encoding the antigen or antigenic parts thereof and which is then expressed; or where the plasmid and/or gene or genes and/or parts thereof are integrated into the host genome, which includes the chromosome and/or any naturally or non- naturally occurring extra-chromosomal element, wherein the gene or genes or parts thereof are expressed.
  • the biological vector is manipulated to prevent or reduce expression of an antigen, such as a virulence factor, or to produce an altered and non-toxic form of the antigen or parts thereof.
  • the biological vector presents other non-toxic antigens or toxic antigens in non-virulence or non-pathogenic effective amounts.
  • the cell or gene or both may need to be further manipulated to ensure the antigen is secreted in sufficient amounts.
  • the antigen expressed is recombinant and, if it is itself a virulence factor, the recombinant antigen will also be de-toxified.
  • the present invention is described using, as a vaccine vector, C. pseudotuberculosis carrying a deletion in the gene encoding PLD which, up to the presen_ time, provides the best model of the subject invention.
  • C. pseudotuberculosis carrying a deletion in the gene encoding PLD which, up to the presen_ time, provides the best model of the subject invention.
  • the present invention extends to any single or multiple nucleotide substitution, addition and/or deletion in the virulence gene which results in the inactivation of that gene or which results in an avirulent form of product, such as an inactive toxin.
  • a vaccine vector strain of C. pseudotuberculosis was obtained by genetically disrupting the PLD gene from a virulent strain using recombinant techniques as herein described in the Examples. It is to be appreciated, however, that other techniques such as chemical- or radiation- induced mutagenesis could also be employed without departing from the scope of the present invention.
  • the PLD gene was disrupted by deletion and an erythromycin resistance gene inserted within the gene. This was transferred into a virulent ______ pseudotuberculosis on a shuttle vector such as a pEP vector bearing a kanamycin resistance determinant (see Australian Patent Application No. PJ8815/90).
  • a variant pEP vector carrying a different antibiotic resistance marker was then also introduced into the bacteria and selection applied for the second plasmid. This encouraged the recombination of the deleted gene into the chromosomal PLD gene, thus conferring erythromycin resistance on the strain and eliminating PLD production. Selection for the second pEP plasmid acts against maintenance of the first plasmid carrying the mutant PLD.
  • the first plasmid will be lost, but in some cases, recombination of the mutant PLD and erythromycin resistance gene to the chromosome will have occurred.
  • PLD mediates clearing on blood plates, the loss of the clearing phenotype with the concomitant loss of kanamycin resistance and maintenance of erythromycin resistance indicates incorporation of the deleted gene into the chromosome of C. pseudotuberculosis.
  • deletion mutants are particularly useful since these are incapable of reversion to the wild type (virulent) phenotype.
  • the combined effect of the adjuvant properties associated with Corynebacterium and the rendering of a virulent strain avirulent make this a particularly potent vaccine vector, capable of stimulating an immunoprotective effect amount of antibody and/or other immune protective factors (including cells) against the virulent organism.
  • this aspect of the present invention relates to a vaccine vector comprising C. pseudotuberculosis carrying a single or multiple nucleotide substitution, addition and/or deletion in the gene encoding PLD.
  • the substitution, addition and/or deletion in the PLD gene must be effective to prevent expression of the gene altogether, to prevent synthesis of active PLD or to result in only a non-virulent effective amount of PLD being synthesized.
  • the vaccine will generally consist of a biological pure formulation of the organism and may additionally comprise one or more pharmaceutically acceptable carriers and/or diluents depending on the intended recipient and mode of administration.
  • Another aspect of the present invention contemplates a method for vaccinating a livestock animal against CLA and/or treating an animal infected with C___ pseudotuberculosis which method comprises administering to said animal an immunoprotective amount of a derivative of C. pseudotuberculosis carrying a single or multiple nucleotide substitution, addition and/or deletion in the gene encoding PLD for a time and under conditions sufficient to render the animal immunoresponsive to current or subsequent C. pseudotuberculosis infection.
  • Administration may be by any suitable route such as by oral or intravenous administration.
  • the preparation may also be in dry or liquid form.
  • the route of administration chosen may also necessitate additional components such as protease inhibitors and the like.
  • a method for the production of non-active or toxoid PLD comprises culturing C ⁇ pseudotuberculosis carrying a single or multiple nucleotide substitution, addition and/or deletion in the gene encoding PLD, and recovering non-active or toxoided PLD produced by expression of said gene.
  • the non-active or toxoided PLD produced by this method may, for example, be used as the active immunogen in a vaccine for stimulating a protective immune response against C. pseudotuberculosis.
  • a gene encoding a virulent factor such as PLD
  • a virulent factor can be mutated in vitro or in vivo by one or more insertion, deletion and/or substitution mutations so as to encode toxoid and then introduced into a suitable corynebacterium or related organism to produce a vaccine vector.
  • the virulent factor-encoding gene may be deleted or otherwise disrupted in a corynebacterium or related organism to obtain an avirulent vaccine vector. All such embodiments are included within the scope of the present invention.
  • Figure 1 is a pictorial representation of a site-specific recombination plasmid.
  • the PLD gene was sub-cloned as a 1.5 kb Sacl fragment into the Pstl site of pEP2.
  • the erythromycin-resistance gene was then cloned as a 1.7 kb HindiII fragment into the Pstl site of the PLD gene thereby creating a deletion.
  • FIG. 2 is a photographic representation depicting mutagenesis of the C. pseudotuberculosis PLD gene. Wild- type C. pseudotuberculosis were mutated using site- specific recombination. Cells were plated onto sheep blood plates. Greater than 90% of mutated cells failed to cause erythrocyte lysis suggesting that the PLD gene had been inactivated.
  • Figures 3 (A) and (B) is a photographic representation showing Southern blot analyses of C. pseudotuberculosis mutants.
  • Panel A Sacl digested genomic DNA hybridised with a PLD-specific probe.
  • G DNA from C. pseudotuberculosis retaining PLD activity after mutagenesis. Higher molecular weight and ghost bands are probably partially digested material.
  • Panel B As Panel A but filter hybridised with an erythromycin gene-specific probe. Note absence of the
  • Figure 3 (C) is a pictorial representation showing the nucleotide sequence of the 5' and 3' regions of C___ pseudotuberculosis phospholipase D gene. Boxed regions show overlaps with the published sequence; dots below STOP indicate putative translational terminator; inverted arrows define transcriptional terminator stem (S) and dotted line the loop (L) structure (-13.6 kcal).
  • Figure 4 is a graphical representation of the T cell responses of sheep injected with PLD-producing and Toxminus strains of C. pseudotuberculosis.
  • Figure 5 is a graphical representation of the serological response of vaccinating sheep with varying doses of C__ pseudotuberculosis Toxminus.
  • E. coli strain DH ⁇ alpha (BRL) was host for a pUC118 clone of a Corynebacterium erythromycin resistance gene (Hodgson et al 1990b) and the E. coli- vcobacterium-Corvnebacterium shuttle plasmids pEP2 and pEP3 (Radford and Hodgson, submitted for publication).
  • the C. pseudotuberculosis strain wild- type for PLD production was obtained from Dr Doug Burrell (CSIRO Division of Animal Health, Armidale, NSW).
  • Rhodococcus e ⁇ ui strain CC50 was a gift from Dr Keith Hughes (Melbourne University, Department of Veterinary Science, Werribee).
  • E. coli were grown in Luria broth (LB: 10 g tryptone, 5 g yeast extract, 10 g NaCl per litre) containing 50 ⁇ g ampicillin (Sigma), 50 ⁇ g kanamycin sulphate (Boehringer Mannheim), 100 ⁇ g erythromycin (Boehringer Mannheim) or 150 ⁇ g hygromycin B (Sigma) per ml as necessary.
  • Cells of C. pseudotuberculosis were cultured at 37°C in brain heart infusion media (Difco) containing 0.1% v/v Tween 80 (BHI) at 37°C.
  • Transformed cells were selected on BHI supplemented with 50 ⁇ g kanamycin, 150 ⁇ g hygromycin B or 100 ⁇ g erythromycin per ml.
  • Putative C. pseudotuberculosis mutants were selected on BHI containing 100 ng erythromycin and 150 ⁇ g hygromycin B (Sigma) per ml.
  • C. pseudotuberculosis cells were cultured on sheep blood plates (LB containing 5% v/v defribinated whole sheep blood, 10% v/v filtered (0.2 ⁇ m) culture supernatant from Rhodococcus e ⁇ ui.
  • a recombinant plasmid was constructed by cloning a fragment carrying the PLD gene into the Pstl site of pEP2 (pBTB58).
  • a 1.7 kb HindiII fragment containing an erythromycin resistance gene was cloned into the PLD gene deleted with Pstl (pBTB58).
  • the orientation of the PLD and erythromycin genes was determined by Sail restriction analysis.
  • the recombination plasmid was electroporated into wild- type C. pseudotuberculosis and transformants were selected on BHI containing erythromycin and kanamycin. Cells of C. pseudotuberculosis harbouring the recombination plasmid were then electroporated with pEP3 and the transformants selected on BHI plates supplemented with erythromycin, kanamycin and hygromycin B. The presence of plasmid pEP3 was confirmed using Southern blot analysis. A transformant containing both plasmids was grown overnight in BHI containing all three drugs. The culture was sub-cultured 1:100 in BHI broth supplemented with only hygromycin B and shaken overnight.
  • the sub-culturing regime was repeated a total of three times.
  • the final broth culture was dilution plated onto sheep blood plates supplemented with erythromycin and hygromycin B. Single colonies were patched to BHI plates containing erythromycin and hygromycin B and only kanamycin.
  • Plasmid pBTB50 containing the wild-type PLD gene was electroporated into a Toxminus C. pseudotuberculosis. Transformants were serially diluted and plated onto sheep-blood plates containing kanamycin. Zones of erythrocyte lysis produced by wild-type and transformed Toxminus cells were measured.
  • the PLD gene was first sub-cloned into the pEP2 shuttle vector. The PLD gene was then deleted with Pstl and an erythromycin- resistance gene was introduced into the Pstl site to produce plasmid pBTB58 (Fig. 1). Wild-type C_ pseudotuberculosis were transformed with pBTB58 and subsequently with the hygromycin resistance shuttle plasmid pEP3. Cells transformed with both plasmids were selected on media containing kanamycin, erythromycin and hygromycin.
  • Plasmid pEP2 carrying the PLD gene was electroporated into Toxminus C. pseudotuberculosis. To detect PLD activity, transformants were plated onto blood plates. Transformed cells produced zones of lysis. This result suggests that the cloned PLD gene is expressed in the C. pseudotuberculosis mutant and that its product is secreted from the host cell.
  • a second PLD negative strain of C. pseudotuberculosis was generated in the same way as for Toxminus except: (1) a different cloned fragment of the PLD gene was used to create the recombinant cassette and (2) the erythromycin gene was cloned into the PLD Pstl site in the opposite direction with respect to the PLD gene shown in Figure 1.
  • the PLD gene was cloned as a 2.1 kb Sacl fragment into the Sacl site of pUCll ⁇ and the new sequence determined (Fig. 3C).
  • a major feature of the second PLD gene clone is the presence of a putative transcriptional terminator just downstream of the translational stop codon (Fig. 3C).
  • Toxminus II differs from Toxminus in that the erythromycin resistance gene is inserted into the chromosome in opposite orientations.
  • T cell response was measured by quantifying the release of gamma-interferon in a whole blood culture incubated with 5 ⁇ g per ml of the C. pseudotuberculosis sonicate. Levels of interferon were assayed using a capture-tag EIA.
  • Titre Serological titres of vaccinated sheep. Titre is the inverse of the last dilution giving an absorbance > 1.25 using strain 231 sonicate as coating antigen in an indirect EIA.
  • C. pseudotuberculosis Toxminus strain sheep were vaccinated with,varying doses of C. pseudotuberculosis Toxminus intradermally into the skin 3cm above the coronet of the left hind leg, and later challenged with virulent 4 x IO 6 C. pseudotuberculosis strain 231 inoculated into the same site on the right hind leg.
  • Five nine month old sheep were used for each vaccine doosage, and seven control sheep were challenged without vaccination (Table 2). Challenge was delivered 9 weeks following the vaccine doses, and protection was assessed a further 9 weeks later.
  • Post-mortem of the vaccinated animals inoculated with either 2 x IO 5 or 2 x IO 7 cfu of C. pseudotuberculosis Toxminus showed that there was no indication of infection or culturable vaccine strain present in the left (vaccinated) popliteal lymph node, or from any other tissue.
  • the virulent strain used for challenge was recovered from all unvaccinated control animals, and from two of the 10 vaccinated sheep. Pathology results are shown in Table 3; except where noted, the virulent strain was cultured from all macroscopic lesions.
  • Pop. Popliteal Lymph node
  • SOI Site of Inoculation ⁇ lesion size cm, (no. lesions) ⁇

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Genetics & Genomics (AREA)
  • Organic Chemistry (AREA)
  • Engineering & Computer Science (AREA)
  • Wood Science & Technology (AREA)
  • General Health & Medical Sciences (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Zoology (AREA)
  • Biotechnology (AREA)
  • Medicinal Chemistry (AREA)
  • Biomedical Technology (AREA)
  • Microbiology (AREA)
  • General Engineering & Computer Science (AREA)
  • Animal Behavior & Ethology (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Mycology (AREA)
  • Veterinary Medicine (AREA)
  • Public Health (AREA)
  • Molecular Biology (AREA)
  • Biochemistry (AREA)
  • Communicable Diseases (AREA)
  • General Chemical & Material Sciences (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Oncology (AREA)
  • Plant Pathology (AREA)
  • Biophysics (AREA)
  • Physics & Mathematics (AREA)
  • Immunology (AREA)
  • Epidemiology (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)
  • Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)

Abstract

La présente invention concerne un vecteur de vaccin, comprenant une corynébactérie virulente ou un organisme virulent apparenté incapable d'exprimer un facteur virulent actif ou une corynébactérie non virulente ou un organisme non virulent apparenté qui synthétise une certaine quantité efficace de protection immune d'un antigène d'organisme pathogène; et son utilisation pour stimuler la réponse de protection immunitaire contre les membres virulents de ce groupe.
PCT/AU1991/000471 1990-10-25 1991-10-14 Corynebacteries et autres organismes apparentes utilises comme vecteurs de vaccin WO1992007582A1 (fr)

Priority Applications (1)

Application Number Priority Date Filing Date Title
JP3516314A JPH06501847A (ja) 1990-10-25 1991-10-14 ワクチンベクターとしてのコリネバクテリウム及び関連する生物

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
AUPK3005 1990-10-25
AUPK300590 1990-10-25

Publications (1)

Publication Number Publication Date
WO1992007582A1 true WO1992007582A1 (fr) 1992-05-14

Family

ID=3775033

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/AU1991/000471 WO1992007582A1 (fr) 1990-10-25 1991-10-14 Corynebacteries et autres organismes apparentes utilises comme vecteurs de vaccin

Country Status (5)

Country Link
EP (1) EP0554335A4 (fr)
JP (1) JPH06501847A (fr)
CA (1) CA2094718A1 (fr)
NZ (1) NZ240326A (fr)
WO (1) WO1992007582A1 (fr)

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2012080696A1 (fr) * 2010-12-13 2012-06-21 Moredun Research Institute Vaccin
CN112011496A (zh) * 2020-08-25 2020-12-01 西南大学 一种伪结核棒状杆菌pld基因无痕缺失株的构建方法

Citations (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
AU7459987A (en) * 1986-06-23 1988-01-07 Schweiz. Serum - & Impfinstitut Und Institut Zur Erforschung Der Infektionskrankheiten ( Swiss Serum and Vaccine Institute and Institute for the Research of Infectious Diseases) DNA fragments encoding the chromosomal nucleotide sugar synthetases and glycosyltransferases involved in the biosynthesis of the o-antigen of shigella dysenteriae, recombinant DNA molecules containing said fragments, host organisms containing said recombinant DNA molecules vaccines for preventing bacillary dysentery and method for preventing bacillary dysentery
AU1955088A (en) * 1987-06-04 1989-01-04 Washington University Avirulent microbes, incapable of producing functional adenylate cyclase and cyclic AMP
AU3867789A (en) * 1988-07-07 1990-02-05 Albert Einstein College Of Medicine Of Yeshiva University Recombinant mycobacterial expression vehicles and uses therefor
US4925792A (en) * 1983-02-08 1990-05-15 Sclavo, S.P.A. Process for producing proteins correlated with the diphtheric toxin
AU5239290A (en) * 1989-03-29 1990-10-04 Csl Limited Purification of c.pseudotuberculosis toxin, and cloning and expression of toxin gene
AU5262690A (en) * 1989-03-08 1990-10-09 Commonwealth Scientific And Industrial Research Organisation Expression system for actinomycetes and related organisms

Patent Citations (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US4925792A (en) * 1983-02-08 1990-05-15 Sclavo, S.P.A. Process for producing proteins correlated with the diphtheric toxin
AU7459987A (en) * 1986-06-23 1988-01-07 Schweiz. Serum - & Impfinstitut Und Institut Zur Erforschung Der Infektionskrankheiten ( Swiss Serum and Vaccine Institute and Institute for the Research of Infectious Diseases) DNA fragments encoding the chromosomal nucleotide sugar synthetases and glycosyltransferases involved in the biosynthesis of the o-antigen of shigella dysenteriae, recombinant DNA molecules containing said fragments, host organisms containing said recombinant DNA molecules vaccines for preventing bacillary dysentery and method for preventing bacillary dysentery
AU1955088A (en) * 1987-06-04 1989-01-04 Washington University Avirulent microbes, incapable of producing functional adenylate cyclase and cyclic AMP
AU3867789A (en) * 1988-07-07 1990-02-05 Albert Einstein College Of Medicine Of Yeshiva University Recombinant mycobacterial expression vehicles and uses therefor
AU5262690A (en) * 1989-03-08 1990-10-09 Commonwealth Scientific And Industrial Research Organisation Expression system for actinomycetes and related organisms
AU5239290A (en) * 1989-03-29 1990-10-04 Csl Limited Purification of c.pseudotuberculosis toxin, and cloning and expression of toxin gene

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
See also references of EP0554335A4 *

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2012080696A1 (fr) * 2010-12-13 2012-06-21 Moredun Research Institute Vaccin
CN112011496A (zh) * 2020-08-25 2020-12-01 西南大学 一种伪结核棒状杆菌pld基因无痕缺失株的构建方法

Also Published As

Publication number Publication date
CA2094718A1 (fr) 1992-04-26
EP0554335A4 (en) 1994-09-14
JPH06501847A (ja) 1994-03-03
NZ240326A (en) 1993-04-28
EP0554335A1 (fr) 1993-08-11

Similar Documents

Publication Publication Date Title
Hodgson et al. Rational attenuation of Corynebacterium pseudotuberculosis: potential cheesy gland vaccine and live delivery vehicle
Curtiss III et al. Nonrecombinant and recombinant avirulent Salmonella live vaccines for poultry
Burnette et al. Direct expression of Bordetelia pertussis toxin subunits to high levels in Escherichia coli
US5747309A (en) Bacterial vaccines using vaccine strains of pathogenic bacteria
US5840312A (en) Recombinant Bacillus anthracis strains unable to produce the lethal factor protein or edema factor protein
JP2004536106A (ja) マイコプラズマ・ボビスワクチンおよび動物の肺炎を減少させる方法
EP2320943B1 (fr) Vaccins pour le bétail
TWI221847B (en) Clostridium perfringens vaccine
US6924118B2 (en) Recombinant intracellular pathogen immunogenic compositions and methods for use
US5637303A (en) Use of a phospholipase D mutant of Corynebacterium pseudotuberculosis for vaccination
JP2006501304A5 (fr)
Tabatabaei et al. Vaccine efficacy in cattle against hemorrhagic septicemia with live attenuated aroA mutant of Pasteurella multocida B: 2 strain
WO1992007582A1 (fr) Corynebacteries et autres organismes apparentes utilises comme vecteurs de vaccin
AU643842B2 (en) Corynebacteria and related organisms as vaccine vectors
US10406219B2 (en) Multivalent Brucella vaccine for protection against mycobacterial infections and methods of using the same
US6149920A (en) Over-expressing homologous antigen vaccine and a method of making the same
Galan et al. Expression and localization of the Streptococcus equi M protein in Escherichia coli and Salmonella typhimurium
US20100284963A1 (en) Growth regulatable recombinant bcg immunogenic compositions
US20050249755A1 (en) Mutant strains of Brucella melitensis and immunogenic compositions
Tafesse et al. Novel approaches toward the development of an oral post-exposure DNA vaccine for latent tuberculosis using Salmonella typhimurium ΔaroA vector
PL188925B1 (pl) Szczepionka przeciwko brucelozie, osłabiona lub awirulentna odmiana szczepu RB51 B. abortus, zastosowanie szczepionki, sposób wytwarzania szczepionki

Legal Events

Date Code Title Description
AK Designated states

Kind code of ref document: A1

Designated state(s): AU CA JP US

AL Designated countries for regional patents

Kind code of ref document: A1

Designated state(s): AT BE CH DE DK ES FR GB GR IT LU NL SE

WWE Wipo information: entry into national phase

Ref document number: 1991919246

Country of ref document: EP

WWE Wipo information: entry into national phase

Ref document number: 2094718

Country of ref document: CA

WWP Wipo information: published in national office

Ref document number: 1991919246

Country of ref document: EP

WWW Wipo information: withdrawn in national office

Ref document number: 1991919246

Country of ref document: EP

点击 这是indexloc提供的php浏览器服务，不要输入任何密码和下载