In vitro and in vivo analyses of constitutive and in vivo-induced promoters in attenuated vaccine and vector strains of Vibrio cholerae

• Published in: Infection and immunity 2000-03, Vol.68 (3), p.1171-1175
• Main Authors: JOHN, M, CREAN, T. I, CALDERWOOD, S. B, RYAN, E. T
• Format: Article
• Language: English
• Subjects: Animals; Antibodies, Bacterial - blood; Bacterial Vaccines - genetics; Bacteriology; Biological and medical sciences; Cholera Toxin - genetics; Cholera Toxin - immunology; DNA vaccines; Female; Fundamental and applied biological sciences. Psychology; Genetic Vectors; htpC gene; irgA gene; Mice; Microbial Immunity and Vaccines; Microbiology; Promoter Regions, Genetic; tac gene; Vaccines, antisera, therapeutical immunoglobulins and monoclonal antibodies; Vaccines, Inactivated - genetics; Vibrio cholerae; Vibrio cholerae - genetics; Vibrio cholerae - immunology
• ISSN: 0019-9567; 1098-5522
• DOI: 10.1128/IAI.68.3.1171-1175.2000

The optimal promoter for in vivo expression of heterologous antigens by live, attenuated vaccine vector strains of Vibrio cholerae is unclear; in vitro analyses of promoter activity may not accurately predict expression of antigens in vivo. We therefore introduced plasmids expressing the B subunit of cholera toxin (CtxB) under the control of a number of promoters into V. cholerae vaccine strain Peru2. We evaluated the tac promoter, which is constitutively expressed in V. cholerae, as well as the in vivo-induced V. cholerae heat shock htpG promoter and the in vivo-induced V. cholerae iron-regulated irgA promoter. The functionality of all promoters was confirmed in vitro. In vitro antigenic expression was highest in vaccine strains expressing CtxB under the control of the tac promoter (2 to 5 microgram/ml/unit of optical density at 600 nm [OD(600)]) and, under low-iron conditions, in strains containing the irgA promoter (5 microgram/ml/OD(600)). We orally inoculated mice with the various vaccine strains and used anti-CtxB immune responses as a marker for in vivo expression of CtxB. The vaccine strain expressing CtxB under the control of the tac promoter elicited the most prominent specific anti-CtxB responses in vivo (serum immunoglobulin G [IgG], P