Antimycobacterial spectrum of sparfloxacin and its activities alone and in association with other drugs against Mycobacterium avium complex growing extracellularly and intracellularly in murine and human macrophages

Abstract

The MICs and MBCs of the new difluorinated quinolone drug sparfloxacin against type strains belonging to 21 species of mycobacteria were screened. The MICs and MBCs were within the range of 0.1 to 2.0 and 0.1 to 4.0 ,ug/ml, respectively (with an MBC/MIC ratio of 1 to 2), and against 18 of the 21 species tested, the drug showed significant bactericidal activity (at least 99% killing or more of the initial inoculum added) at concentrations well within the reported peak concentrations in serum (Cm.) in humans. MICs of sparfloxacin for 7 of 10 Mycobacterium avium complex strains were below the Cm,,, with MBC/IMIC ratios within the range of 2 to 4. Enhancement of its activity by ethambutol, rifampin, amikacin, and clarithromycin (which were used at sublethal concentrations) assessed by using BACTEC radiometry revealed that its activity was further enhanced in 2 of 10 strains by rifampin and in 7 of 10 strains by ethambutol. The bactericidal effects of various drugs used alone as well as two-drug combinations used at Cm.. levels were also screened against four strains of M. avium complex growing intracellularly in two different macrophage systems, namely, mouse bone marrow-derived macrophages and peripheral blood monocyte-derived human macrophages. Our results showed a satisfactory correlation between the extracellular and intracellular drug activity data.Recently, newer fluoroquinolones have been reported to have broad antimicrobial spectra, including mycobacteria (3,14,25). Both the N-cyclopropyl analog ciprofloxacin and the tricyclic compound ofloxacin were found to penetrate actively and concentrate inside mammalian cells (6,13,35), and were found to be capable of killing intracellularly growing tubercle bacilli (19). A newly synthesized difluorinated quinolone, sparfloxacin [5 amino-1-cyclopropyl-6,8-difluoro-1,4-dihydro-(7 cis-3,5-dimethyl-1-piperazinyl)-4-oxoquinolone-3-carboxylic acid], was retained for this study because it is a broad-spectrum drug with in vitro and in vivo efficacies equal to or better than those of ofloxacin and ciprofloxacin (17), and compared with other fluoroquinolones, it has lower MICs against both tubercle bacilli (24) and the multiple-drug-resistant opportunistic pathogens belonging to the Mycobacterium avium complex (MAC) (37). Considering the intracellular parasitism of MAC organisms as well as that of other difficult-to-treat atypical mycobacteria, it was interesting that sparfloxacin attained levels in mouse and rat tissues which were 1 to 11 times higher than its concentrations in plasma (maximum concentration in serum [Cmax], 1.4 ,ug/ml) (16).After we made the observations described above, we decided to perform the present four-point study on sparfloxacin activity (i) to assess its antimycobacterial spectrum against 21 species of mycobacteria by MIC and MBC determinations with 7H9 broth; (ii) by radiometric determination of its MICs and MBCs against 10 MAC strains; (iii) to investigate the enhancement of its action (used at a sublethal concentration of 0.25 ,ug/ml) against 10 M...

A bone marrow-derived murine macrophage model for evaluating efficacy of antimycobacterial drugs under relevant physiological conditions

“…In support of the use of such models, a good correlation has previously been reported between the use of murine bone marrow-derived macrophages and the use of human monocytes for the evaluation of the bactericidal effects of therapeutic compounds (34). Moreover, studies that used different sources of macrophages (32,44) observed that compounds which are very active in broth also tended to be active in the macrophage cultures.…”

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

“…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).…”

Extra- and intracellular antimycobacterial activity of Arbutus unedo L.

“…The colony-forming units (CFU) were enumerated after about 4 days of the Petri dishes incubation at 37°C. As described by Rastogi et al (1991), this treatment with 0.25% (w/v) SDS did not affect the viability of mycobacteria. The results were compared with the growth of the bacilli in the control culture (untreated macrophages).…”

“…After phagocytosis, the medium from each well was retired and the cells were then treated during 1 h by a medium containing 10 µg/ml of amikacin. This treatment was followed by several changes of hanks balanced salt solution to remove all extracellular M. smegmatis MC2 (Rastogi et al, 1987(Rastogi et al, , 1991Sharbati-Tehrani et al, 2005). (iii) Treatment by plant ethanol extract: About 5 h after infection, necessary time of phagocytosis and elimination of extracellular M. smegmatis MC2, the ethanol leaves plant extract (6.02 ± 0.76 mg/ml) which was solubilized in RPMI-1640 as demonstrated above, was added to the wells containing infected macrophages.…”

“…The control used corresponded to discs impregnated with a liquid medium. (iii) Broth dilution assay: The MIC determination of ethanol extract part I for the mycobacterial species was realized according to the broth dilution method as described by Rastogi et al (1991). Tubes containing 6 ml of sauton liquid medium alone (control) or supplemented with increasing concentrations of ethanol extract part I, from 0.24 to 6.38 mg/ml, were inoculated with a freshly grown culture to give an OD595 about of 0.3 for M. bovis PPI, and 0.2 for M. smegmatis MC2 and M. aurum A +.…”