US 4,735,801 A · Novel non-reverting salmonella live vaccines

Abstract

Live vaccines are provided and methods for preparing the live vaccines for protection of a host from a pathogenic microorganism. The vaccines are prepared by introducing at least one modification in a gene involved in at least one, normally at least two, biosynthetic pathways, involving the production of products which are unlikely to be found in the disease susceptible host. The modification results in a gene change which cannot be repaired by a single step e.g. polynucleotide deletions and inversions. Where the aro gene suffers such a change, the resultant auxotrophic mutants require aromatic amino acids, p-aminobenzoic acid and 2,3-dihydroxybenzoic acid or a highly concentrated source of absorbable iron. The auxotrophic mutations have substantially reduced or nonexistent virulence, while retaining the desired immunogenicity to initiate the immunogenic response. Various techniques can be employed for providing the desired change. Salmonella typhimurium strain SL1479 was deposited at the ATCC on Sept. 7, 1982 and given ATCC Accession No. 39183; Salmonella dublin strain SL1438 was deposited on Sept. 7, 1982 at the ATCC and given ATCC Accession No. 39184. Salmonella typhi strain 531Ty was deposited at the ATCC on Nov. 21, 1984, and granted ATCC Accession No. 39926.

Claims18

Dependent claims

A method for preparing a live attentuated vaccine from a virulent pathogenic bacterial microorganism, which vaccine has substantially zero probability of reverting to virulence in a vertebrate host, said method comprising:

introducing a transposon into a structural gene of a virulent strain of said microorganism, wherein said gene expresses a protein in the biosynthetic pathway of a metabolite essential for multiplication and normally unavailable in said vertebrate host, and wherein said transposon provides a phenotypic property allowing for selection of the presence of said transposon;
selecting for microorganisms having said phenotypic property provided by said transposon and isolating transposon-modified microorganisms;
selecting for, from said transposon-modified microorganisms, variant microorganisms lacking said phenotypic property due to excision of all or a part of said transposon and being auxotrophic due to a non-reverting mutation in said structural gene as a result of said excision, resulting in a culture having a non-reverting auxotrophic non-virulent mutant;
selecting for said non-reverting auxotrophic nonvirulent mutant;
and
isolating said auxotrophic non-reverting non-virulent mutant to provide a living vaccine.

3 more elements

A method for preparing a live non-virulent vaccine from a virulent pathogenic bacterium having substantially unaltered O antigenic character, which vaccine has a substantially zero probability of reverting to virulence in a vertebrate host susceptible to said bacterium, said method comprising:

transducing cells of a virulent strain of said bacterium with a first transducing phage comprising at least a portion of a first gene, which first gene expresses a protein utilized in a first biosynthetic pathway to a first essential metabolite normally unavailable in said vertebrate host, and which first gene includes a non-reverting mutation, resulting in a culture of a first auxotrophic non-virulent mutant;
selecting for said first auxotrophic mutant by means of said first auxotrophic mutation or other marker introduced by said first transducing phage;
transducing said first auxotrophic mutant with a second transducing phage comprising at least a portion of a second gene, which second gene expresses a protein utilized in a second biosynthetic pathway to a second essential metabolite normally unavailable in said vertebrate host, and which second gene includes a non-reverting mutation, resulting in a culture of a second auxotrophic non-virulent mutant;
selecting for said second auxotrophic mutant by means of said second auxotrophic mutation or other marker introduced by said second transducing phage;
and
isolating said second auxotrophic non-reverting mutant to provide a living vaccine.

3 more elements

Dependencies are inferred from the claim text.

Novel non-reverting salmonella live vaccines

Life of this patent

Novel non-reverting salmonella live vaccines

Life of this patent

Abstract

Claims18