In vitro and ex vivo activities of antimicrobial agents used in combination with clarithromycin, with or without amikacin, against Mycobacterium avium

Abstract

MICs of clarithromycin, amikacin, isoniazid, rifabutin, ciprofloxacin, sparfloxacin, ethambutol, and clofazimine were determined for six isolates of Mycobacterium avium complex (MAC) from AIDS patients both by the radiometric method and by an ex vivo model of infection in human macrophages. The median MICs in macrophages were similar or slightly lower than values found in broth, except for amikacin, which had slightly higher MICs inside the cells. Combinations of clarithromycin with other antimicrobial agents showed that clarithromycin-clofazimine and clarithromycin-rifabutin were synergistic on five of six strains while clarithromycin-amikacin and clarithromycin-isoniazid were antagonistic on one and two strains, respectively. The addition of amikacin made the combinations of clarithromycin-clofazimine and clarithromycin-ethambutol synergistic against all the MAC strains. In the macrophage model, the combination of clarithromycinclofazimine (mean survival, 21%) and clarithromycin-rifabutin (mean survival, 29%) showed a strong reduction in viable counts compared with single drugs, while clarithromycin-amikacin was less active than single drugs alone. In general, the addition of amikacin did not improve the activity of the combinations, except for clarithromycin-isoniazid-amikacin (mean survival, 19%), which was significantly more active than either clarithromycin-isoniazid or clarithromycin-amikacin. The use of the macrophage model can suggest new combinations of antimicrobial agents with anti-MAC activity which, on the basis of their in vitro effectiveness, would probably be disregarded for assay in animal models.Mycobacterium avium complex (MAC) infections are very common in AIDS patients, with an estimated prevalence of 43% in the 2 years following the diagnosis of AIDS (25). Therapy for MAC diseases usually involves the combination of several drugs, not only to obtain a greater antimycobacterial effect but also to decrease the probability of selecting resistant mutants (13,15). In some studies, the efficacy of different antimicrobial combinations has been evaluated in vitro (12,34), while in others the activity has been investigated with human macrophages (MP) (33,35,37) or in the murine MP cell line J774 (33,35,36). The Public Health Service Task Force on Prophylaxis and Therapy for MAC recently recommended clarithromycin (CLA)-containing regimens for therapy for disseminated infections by these organisms in AIDS patients (20). At the present time, only for a few reports has the activity of CLA-containing combinations been studied, by in vitro and ex vivo methods, on the same strains (27,28).In this study, we assessed the susceptibility of MAC strains isolated in Italy from AIDS patients (7) by determining the MICs of eight antimicrobial agents both by the radiometric method (11) and by a model of human monocyte-derived MP infection (22,23). This was adopted to evaluate the relationship between the extracellular and intracellular activities of drugs with different capacities for accumulating within MP (31)....

Antimycobacterial Agents Differ with Respect to Their Bacteriostatic versus Bactericidal Activities in Relation to Time of Exposure, Mycobacterial Growth Phase, and Their Use in Combination

“…Only AMK had a substantial effect by improving the activity of the combination CLR-EMB tenfold within only 3 days of exposure. This is in agreement with the findings of Fattorini et al, demonstrating that the addition of AK made the combination of CLR-EMB synergistic against a number of MAC strains in vitro (17).…”

“…Aminoglycosides have been used in clinical studies in successful treatment regimens, whereas other investigators failed to show a beneficial effect of aminoglycosides (36). However, Fattorini et al demonstrated in in vitro assays that AMK significantly improved the activity of the CLR-EMB combination (17). The potential risks and benefits of treatment with aminoglycosides should be weighed carefully.…”

Clofazimine Prevents the Regrowth of Mycobacterium abscessus and Mycobacterium avium Type Strains Exposed to Amikacin and Clarithromycin

“…The combination clarithromycin-clofazimine also showed synergy against MAC strains in checkerboard evaluation (7). These checkerboard titrations offer no information on the mechanism of synergistic activity, the exact killing activity of these combinations, or its concentration dependence.…”

Virulence and drug susceptibility of Mycobacterium celatum

“…Leukocyte buffy coats obtained from healthy donors were separated over Ficoll-Hypaque (Histopaque 1077 ; Sigma), as previously described (Fattorini et al, 1995). Briefly, suspensions of isolated peripheral blood mononuclear cells prepared in complete RPMI 1640 medium (Life Technologies) containing 10 % heat-inactivated (56 mC, 30 min) fetal calf serum, 25 mM HEPES and 2 mM -glutamine were adjusted to 1i10( cells ml − " and distributed in 0n5 ml volumes in 24-well tissue culture plates.…”

“…Cell viability was determined by the trypan blue exclusion method. At various times, infected macrophages were lysed to determine the number of c.f.u., as previously described (Fattorini et al, 1995).…”