Selective maintenance of a recombinant gene in a population of vaccine cells

A composition for stimulating an immune response in an individual comprising a live avirulent derivative of a pathogenic strain of bacteria characterized by:

• a) a lack of a functioning native chromosomal gene encoding a first enzyme which is a .beta.-aspartic semialdehyde dehydrogenase (Asd);
• b) the presence of a first recombinant gene encoding a second Asd enzyme wherein the first recombinant gene cannot recombine to replace the defective chromosomal gene;
• c) the presence of a second recombinant gene encoding a desired polypeptide;
• and physical linkage between the first recombinant gene and the second recombinant gene, wherein loss of the first recombinant gene causes the bacteria to lyse when in an environment which requires expression of said first recombinant gene for cell survival.

A method of preparing a bacterial carrier for a vaccine for immunization of an individual, said method comprising:

• a) providing an avirulent derivative of a pathogenic strain of bacteria characterized by:
• 1) a lack of a functioning native chromosomal gene encoding a first enzyme which is a .beta.-aspartic semialdehyde dehydrogenase (Asd)
• 2) the presence of a first recombinant gene encoding a second Asd enzyme wherein the first recombinant gene cannot recombine to replace the defective chromosomal gene;
• 3) the presence of a second recombinant gene encoding a desired polypeptide;
• and
• 4) physical linkage between the first recombinant gene and the second recombinant gene, wherein loss of the first recombinant gene causes the bacteria to lyse when in an environment which requires expression of said first recombinant gene for cell survival;
• b) providing a suitable excipient;
• and
• c) mixing the bacteria with the excipient in a suitable pharmacologic dose.

An immunogenic composition comprising an avirulent derivative of a pathogenic strain of bacteria characterized by:

• a) a lack of a functioning native chromosomal gene encoding a first enzyme which is a .beta.-aspartic semialdehyde dehydrogenase (Asd);
• b) the presence of a first recombinant gene encoding a second Asd enzyme wherein the first recombinant gene cannot recombine to replace the defective chromosomal gene;
• c) the presence of a second recombinant gene encoding a desired polypeptide;
• and
• d) physical linkage between the first recombinant gene and the second recombinant gent, wherein loss of the first recombinant gene causes the bacteria to lyse when in an environment which requires expression of said first recombinant gene for cell survival.