Activities of daptomycin and comparative antimicrobials, singly and in combination, against extracellular and intracellular Staphylococcus aureus and its stable small-colony variant in human monocyte-derived macrophages and in broth

Abstract

We investigated the antistaphylococcal activities of daptomycin, gentamicin, and rifampin against two Staphylococcus aureus strains and their stable small-colony variants, singly and in combination, in human monocyte-derived macrophages and in broth. Intracellularly, the three-drug combination and two-drug combinations with rifampin were most effective. Extracellularly, daptomycin, daptomycin plus gentamicin, gentamicin plus rifampin, and the three-drug combination had similar activities.

FIG. 1.

PFGE DNA fingerprinting of S. aureus parent strains 2546 and 2548 and the SVC 2547 and SCV 2549 strains (22).

FIG. 2.

Immunofluorescent staining of intracellular and extracellular S. aureus strains. Intracellular bacteria are red, extracellular bacteria are yellow-green, and nuclei are blue. Arrows indicate the intracellular parent strain 2546 (A), the intracellular SCV 2547 (B), and small clusters of extracellular strain 2546 (C). (Magnification, ×400).

FIG. 3.

Intracellular effects of single drugs and two- and three-drug combinations against S. aureus parent strain 2546 (A) and SCV 2547 (B), the parent strain 2548 (C), and SCV 2549 (D) in human MDM. Dotted lines represent results with single drugs. Numbers of viable bacteria at 0 h were 1 × 10⁵ to 2 × 10⁵ CFU/well. MDM were lysed at 0, 4, 24, and 48 h, and viable bacteria in the lysates were enumerated using the standard plate count method. The limit of detection was 20 CFU/ml. Results are from three independent experiments, each done in duplicate. Error bars represent standard errors of the means (some error bars are obscured by the symbols). Dap, daptomycin; Gent, gentamicin; Rif, rifampin.

FIG. 4.

Extracellular effects of single drugs and two- and three-drug combinations against the S. aureus parent strain 2546 (A) and SCV 2547 (B), the parent strain 2548 (C), and SCV 2549 (D) in MH-II broth. Dotted lines represent results for single drugs. Numbers of viable bacteria at 0 h were 1 × 10⁵ to 2 × 10⁵ CFU/ml. Prior to sampling at 0, 4, 24, and 48 h, broth cultures were thoroughly mixed by vortexing. Samples were then taken, and viable bacteria in the samples were enumerated using the standard plate count method. The limit of detection was 10 CFU/ml. Results are from three independent experiments, each done in duplicate. Error bars represent standard errors of the means (some error bars are obscured by the symbols). Dap, daptomycin; Gent, gentamicin; Rif, rifampin.