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Virulence of Listeria monocytogenes, Listeria seeligeri, and Listeria innocua Assayed with In Vitro Murine Macrophagocytosis

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Abstract:

The survival of virulent and avirulent Listeria species internalized in cells of a murine macrophage-like cell line, RAW264.7, was monitored. Mouse macrophage cells (ca. 5 × 105/ml) suspended in fresh RPMI medium 1640 containing fetal bovine serum were mixed with 5 × 107 to 5 × 108 Listeria cells per ml and incubated 1 h at 37°C with CO2-enriched air. Gentamicin (10 μg/ml) was added to kill bacteria not internalized by the cells. At 2, 4, and 6 h postinfection, 10-μl amounts of the suspensions were lysed in microtiter plate wells during serial decimal dilution in water. Triplicate dilutions (10-μl each) were plated on trypticase soy agar, and colonies were counted after 48 h incubation at 35°C. About 0.1 to 1% of the added hemolytic pathogen L. monocytogenes Scott A and the avirulent nonhemolytic L. innocua were internalized at 2 h. The number of internal L. monocytogenes cells increased significantly by 6 h, but L. innocua cells showed no significant change. A strain of the hemolytic species L. seeligeri behaved like the nonhemolytic L. innocua. This distinction between the intracellular behavior of pathogenic and nonpathogenic species, if a general phenomenon, may be useful as an in vitro virulence assessment parameter.

Keywords: LISTERIA SPP; MACROPHAGOCYTOSIS ASSAY; MICROSCOPY; TISSUE CULTURE; VIRULENCE

Document Type: Research Article

Affiliations: 1: Division of Microbiological Studies, Center for Food Safety and Applied Nutrition, U.S. Food and Drug Administration, 200 C Street, S.W., Washington, D.C. 20204, USA 2: Center for Devices and Radiological Health, U.S. Food and Drug Administration, Rockville, Maryland 20857, USA

Publication date: January 1, 1996

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