Survival of Enterococcus faecalis in Mouse Peritoneal Macrophages

Abstract

ABSTRACT Enterococcus faecalis was tested for the ability to persist in mouse peritoneal macrophages in two separate studies. In the first study, the intracellular survival of serum-passaged E. faecalis 418 and two isogenic mutants [cytolytic strain FA2-2(pAM714) and non-cytolytic strain FA2-2(pAM771)] was compared with that of Escherichia coli DH5α by infecting BALB/c mice intraperitoneally and then monitoring the survival of the bacteria within lavaged peritoneal macrophages over a 72-h period. All E. faecalis isolates were serum passaged to enhance the production of cytolysin. E. faecalis 418, FA2-2(pAM714), and FA2-2(pAM771) survived at a significantly higher level ( P = 0.0001) than did E. coli DH5α at 24, 48, and 72 h. Internalized E. faecalis 418, FA2-2(pAM714), and FA2-2(pAM771) decreased 10-, 55-, and 31-fold, respectively, over the 72-h infection period, while internalized E. coli DH5α decreased 20,542-fold. The difference in the rate of survival of E. faecalis strains and E. coli DH5α was most prominent between 6 and 48 h postinfection ( P = 0.0001); however, no significant difference in killing was observed between 48 and 72 h postinfection. In the second study, additional E. faecalis strains from clinical sources, including DS16C2, MGH-2, OG1X, and the cytolytic strain FA2-2(pAM714), were compared with the nonpathogenic gram-positive bacterium, Lactococcus lactis K1, for the ability to survive in mouse peritoneal macrophages. In these experiments, the E. faecalis strains and L. lactis K1 were grown in brain heart infusion (BHI) broth to ensure that there were equal quantities of injected bacteria. E. faecalis FA2-2(pAM714), DS16C2, MGH-2, and OG1X survived significantly better ( P < 0.0001) than did L. lactis K1 at each time point. L. lactis K1 was rapidly destroyed by the macrophages, and by 24 h postinfection, viable L. lactis could not be recovered. E. faecalis FA2-2(pAM714), DS16C2, MGH-2, and OG1X declined at an equivalent rate over the 72-h infection period, and there was no significant difference in survival or rate of decline among the strains. E. faecalis FA2-2(pAM714), MGH-2, DS16C2, and OG1X exhibited an overall decrease of 25-, 55-, 186-, and 129-fold respectively, between 6 and 72 h postinfection. The overall reduction by 1.3 to 2.27 log units is slightly higher than that seen for serum-passaged E. faecalis strains and may be attributable to the higher level of uptake of serum-passaged E. faecalis than of E. faecalis grown in BHI broth. Electron microscopy of infected macrophages revealed that E. faecalis 418 was present within an intact phagocytic vacuole at 6 h postinfection but that by 24 h the infected macrophages were disorganized, the vacuolar membrane was degraded, and the bacterial cells had entered the cytoplasm. Macrophage destruction occurred by 48 h, and the bacteria were released. In conclusion, the results of these experiments indicate that E. faecalis can persist for an extended period in mouse peritoneal macrophages.