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Conditional Lethality Yields a New Vaccine Strain of Listeria monocytogenes for the Induction of Cell-Mediated Immunity

Li, Zhongxia, Zhao, Xinyan, Higgins, Darren E., Frankel, Fred R.
Infection and immunity 2005 v.73 no.8 pp. 5065-5073
Bacillus subtilis, Human immunodeficiency virus, Listeria monocytogenes, T-lymphocytes, bacteria, cell walls, cell-mediated immunity, genes, immune response, infectious diseases, mice, microbial growth, mutants, pathogens, plasmids, secondary immunization, splenocytes
Listeria monocytogenes is a gram-positive intracellular pathogen that can enter phagocytic and nonphagocytic cells and colonize their cytosols. Taking advantage of this property to generate an intracellular vaccine delivery vector, we previously described a mutant strain of L. monocytogenes, [Delta]dal [Delta]dat, which is unable to synthesize cell wall by virtue of deletions in two genes (dal and dat) required for D-alanine synthesis. This highly attenuated strain induced long-lived protective systemic and mucosal immune responses in mice when administered in the transient presence of D-alanine. We have now increased the usefulness of this organism as a vaccine vector by use of an inducible complementation system that obviates the need for exogenous D-alanine administration. The strain expresses a copy of the Bacillus subtilis racemase gene under the control of a tightly regulated isopropyl-{szligbeta}-D-thiogalactopyranoside (IPTG)-inducible promoter present on a multicopy plasmid. This bacterium demonstrates strict dose-dependent growth in the presence of IPTG. After removal of inducer, bacterial growth ceased within two replication cycles. Following infection of mice in the absence of IPTG or D-alanine, the bacterium survived in vivo for less than 3 days. Nevertheless, a single immunization elicited a state of long-lasting protective immunity against wild-type L. monocytogenes and induced a subset of effector listeriolysin O-specific CD11a⁺ CD8⁺ T cells in spleen and other tissues that was strongly enhanced after secondary immunization. This improved L. monocytogenes vector system may have potential use as a live vaccine against human immunodeficiency virus, other infectious diseases, and cancer.