B-223. Accelerated Macrophage Apoptosis Induced by Group A Streptococcus

A. M. Timmer1, J. C. Timmer2, M. A. Pence1, L-C. Hsu1, M. Karin1, G. S. Salvesen2, V. Nizet1;
1Univ. of California, La Jolla, CA, 2Burnham Inst. for Med. Res., La Jolla, CA.

Background: Group A Streptococcus (GAS) is a Gram-positive bacterium responsible for millions of human infections each year worldwide, including invasive conditions such as necrotizing fasciitis and septicemia. As frontline components of the host innate immune system, macrophages play a key role in the control and clearance of GAS infections. Methods: Apoptosis of murine macrophages in vitro was assessed by TUNEL and a series of caspase inhibitor and activity assays. Isogenic single-gene deletion GAS mutants and heterologous gene expression were used to probe the role of individual GAS virulence determinants in the proapoptotic phenotype. Results: GAS induced rapid, dose-dependent apoptosis of primary murine macrophages and J774 macrophages (50% TUNEL positive cells by 4 h). GAS had to be viable and brought into the host cell by phagocytosis to activate the cell death pathway, which was shown to be caspase-dependent, and diminished in macrophages derived from caspase-1 knockout mice. From a panel of virulence factor mutants, streptolysin O (SLO) was identified to play a major role in GAS-induced macrophage apoptosis; heterologous expression of SLO in the nonpathogenic Lactococcus lactis and administration of purified SLO demonstrated this pore-forming cytolysin to be sufficient to confer the macrophage-inducing phenotype. SLO-deficient GAS mutants induced less macrophage apoptosis in vivo and were less virulent in a murine systemic infection model. Conclusions: Accelerated, caspase-dependent macrophage apoptosis induced by the pore-forming cytolysin SLO may contribute to GAS immune evasion and virulence.