Recombinant Listeria monocytogenes expressing a cell wall-associated listeriolysin O is weakly virulent but immunogenic

Abstract

Listeriolysin O (LLO) is an essential virulence factor for the gram-positive bacterium Listeria monocytogenes. Our goal was to determine if altering the topology of LLO would alter the virulence and toxicity of L. monocytogenes in vivo. A recombinant strain was generated that expressed a surface-associated LLO (sLLO) variant secreted at 40-fold-lower levels than the wild type. In culture, the sLLO strain grew in macrophages, translocated to the cytosol, and induced cell death. However, the sLLO strain showed decreased infectivity, reduced lymphocyte apoptosis, and decreased virulence despite a normal in vitro phenotype. Thus, the topology of LLO in L. monocytogenes was a factor in the pathogenesis of the infection and points to a role of LLO secretion during in vivo infection. The sLLO strain was cleared by severe combined immunodeficient (SCID) mice. Despite the attenuation of virulence, the sLLO strain was immunogenic and capable of eliciting protec-tive T-cell responses. Listeria monocytogenes is a gram-positive facultative intra-cellular pathogen extensively used to understand host-patho-gen interactions (44, 51, 53). It expresses the highly conserved pore-forming toxin listeriolysin O (LLO), a member of a large family of cholesterol-dependent cytolysins found in many im