Non-pyrogenic bacterial strains and use of the same

A method for producing a non-pyrogenic bacterial strain, said method comprising the steps of:

• (a) genetically modifying a gram negative bacterial strain, said step of genetically modifying comprising introducing into the genome of the gram negative bacterial strain a conditional mutation which affects the biosynthesis of lipid A under non-permissive culture conditions, and results in the synthesis of non-pyrogenic lipid A;
• (b) culturing the genetically modified gram negative bacterial strain of (a) in a medium under non-permissive culture conditions, wherein the medium comprises a modified gram-negative bacterial strain producing non-pyrogenic lipid A;
• and
• (c) isolating the modified gram negative bacterial strain producing non-pyrogenic lipid A from the medium.

A method for preparing a culture comprising a non-pyrogenic bacterial strain, said method comprising the steps of:

• (a) genetically modifying a gram-negative host bacterial strain that is suitable for use as a vaccine, said step of genetically modifying comprising introducing a conditional mutation in one or more genes selected from the group consisting of kdsA, kdsB, kdtA, lpxA, lpxB, lpxC, lpxD, ssc, pmr, htrB and msbB of the gram-negative host strain, wherein the conditional mutation affects the biosynthesis of lipid A under non-permissive culturing conditions, and results in the synthesis of non-pyrogenic lipid A;
• and
• (b) culturing the genetically modified gram-negative host strain in non-permissive culture conditions to produce non-pyrogenic lipid A thereby generating a non-pyrogenic vaccine,
• wherein the non-permissive culture conditions include temperature of 35° C. to 44° C. and quaternary cationic compound presence.