In vitro susceptibilities of Bartonella henselae, B. quintana, B. elizabethae, Rickettsia rickettsii, R. conorii, R. akari, and R. prowazekii to macrolide antibiotics as determined by immunofluorescent-antibody analysis of infected Vero cell monolayers.

The in vitro susceptibilities of Bartonella (Rochalimaea) henselae, B. quintana, B. elizabethae, Rickettsia akari, R. conorii, R. prowazekii, and R. rickettsii to different concentrations of azithromycin, clarithromycin, dirithromycin, erythromycin, and roxithromycin in Vero cell cultures were evaluated. Bartonella and Rickettsia spp. were allowed to initiate infection of the antibiotic-free Vero cell monolayers, which were maintained in 16-chamber microscope slides in the absence of antibiotics at 32 degrees C in a CO2-enriched atmosphere. The monolayers were then incubated for 3 h to allow for initial host cell intracellular penetration by infecting species. After inoculation, inocula were replaced and tested with media containing 12 different concentrations of each antibiotic in replicate (10 wells of each antibiotic dilution) for each species, and the monolayers were reincubated. Tetracycline served as the control. Growth status of Bartonella spp. and Rickettsia spp. was determined by evaluation of immunofluorescent staining bacilli. Five days later, when antibiotic-free, control-infected cell monolayers demonstrated significant fluorescence, media were removed for all cell monolayers, the monolayers were fixed, and all specimens were stained with standard indirect immunofluorescent antibody reagents. Fluorescent foci were enumerated by counting such foci on random fields visualized with an epifluorescence microscope. The extent of antibiotic-induced focus inhibition was recorded for each dilution of antibiotic and compared with that of an antibiotic-negative control. Effective antibiotic dilution endpoints for inhibition of Bartonella and Rickettsia proliferation, as judged by absence of increase of significant fluorescence (as compared with no-growth controls), were enumerated by determining the number of cell culture chambers at various antibiotic dilutions that were negative or positive for significant Bartonella- or Rickettsia-specific fluorescence. All of the macrolide agents tested were readily active against all three Bartonella organisms, and azithromycin, clarithromycin, and roxithromycin may have potential in the treatment of Rickettsia infections. Animal model-based clinical trials are warranted to define the specific treatment role of the newer macrolide antibiotics.     

In vitro susceptibilities of Rickettsia rickettsii and Rickettsia conorii to roxithromycin and pristinamycin.

In vitro susceptibilities of Rickettsia rickettsii and Rickettsia conorii to roxithromycin, pristinamycin, and the pristinamycin compounds, P1 and P2, were determined by a dye uptake assay and a plaque assay. The MICs were 1 microgram/ml for roxithromycin, 2 micrograms/ml for pristinamycin, greater than 256 micrograms/ml for P1, and 2 micrograms/ml for P2. Compounds P1 and P2 did not share synergetic activity. The toxicity of each compound was determined by a dye uptake assay. Toxic concentrations were 128 micrograms/ml for roxithromycin, 32 micrograms/ml for pristinamycin, greater than 256 micrograms/ml for P1, and 32 micrograms/ml for P2. Roxithromycin and pristinamycin could be useful in the treatment of Rocky Mountain spotted fever and Mediterranean spotted fever.     

In vitro susceptibilities of suspected periodontopathic anaerobes as determined by membrane transfer assay.

Attempts to devise an antimicrobial approach to combating dentomicrobial infections such as periodontal diseases continue to be hampered by the lack of a relevant in vitro method for determining the susceptibility of suspected periodontopathogens to topically applied antimicrobial agents. Proposed here is a novel in vitro method called the membrane transfer technique, which acknowledges those aspects unique to localized pathogenic infections, particularly those associated with anaerobic bacteria. Bacterial lawns representing six suspected periodontopathic bacteria were prepared on membranes and then placed in contact with different concentrations of antimicrobial agents for 5 min. After incubation for 12 to 24 h, MBCs were determined with the aid of a tetrazolium chloride indicator. Four antimicrobial agents (chlorhexidine, iodine, stannous fluoride, and sodium fluoride) were used to test the applicability of the proposed in vitro method. MBCs were derived for each agent except sodium fluoride against all or most of the six bacterial strains tested. The proposed method may also be useful for examining the bactericidal action of topically applied antimicrobial agents against nonoral infections.     

In vitro susceptibilities of Coxiella burnetii, Rickettsia rickettsii, and Rickettsia conorii to the fluoroquinolone sparfloxacin.

In vitro susceptibilities of Rickettsia rickettsii, Rickettsia conorii, and Coxiella burnetii to the new fluoroquinolone sparfloxacin (AT-4140; RP 64206) were determined. Plaque and dye uptake assays were used to measure the MICs against R. rickettsii and R. conorii. The susceptibilities of C. burnetii Nine Mile and Q 212 were determined in two acute-infection models and in two chronic-infection models. The MICs were 0.125 to 0.25 microgram/ml for R. rickettsii and 0.25 to 0.5 microgram/ml for R. conorii. Sparfloxacin (1 microgram/ml) cured cells recently infected with C. burnetii Nine Mile and Q 212 within 4 to 9 days and cured multiplying, persistently infected cells within 10 days. As previously described with other fluoroquinolones (D. Raoult, M. Drancourt, and G. Vestris, Antimicrob. Agents Chemother. 34:1512-1514, 1990), sparfloxacin failed to cure cells persistently infected with C. burnetii and blocked from dividing with cycloheximide. As determined by the dye uptake assay, no cellular toxicity was noted with sparfloxacin at up to 128 micrograms/ml. These results are consistent with those previously obtained with fluoroquinolones (D. Raoult, M. Yeaman, and O. Baca, Rev. Infect. Dis. 11[Suppl. 5]:S986, 1989), although sparfloxacin may be slightly more active.     

In vitro susceptibility of Rickettsia conorii to ciprofloxacin as determined by suppressing lethality in chicken embryos and by plaque assay.

We tested the susceptibility of Rickettsia conorii to ciprofloxacin, a new quinolone antibiotic. A final concentration of 1 microgram/g of egg was effective in suppressing chicken embryo lethality, and a concentration of 0.25 micrograms/ml inhibited plaque formation in a plaque assay; however, a concentration of 0.5 microgram/ml was necessary to obtain rickettsiacidal activity. These results support the idea that ciprofloxacin could be of clinical use in treating Mediterranean spotted fever.     

In vitro activities of telithromycin (HMR 3647) against Rickettsia rickettsii, Rickettsia conorii, Rickettsia africae, Rickettsia typhi, Rickettsia prowazekii, Coxiella burnetii, Bartonella henselae, Bartonella quintana, Bartonella bacilliformis, and Ehrlichia chaffeensis

In vitro activities of telithromycin compared to those of erythromycin against Rickettsia spp., Bartonella spp., Coxiella burnetii, and Ehrlichia chaffeensis were determined. Telithromycin was more active than erythromycin against Rickettsia, Bartonella, and Coxiella burnetii, with MICs of 0.5 microg/ml, 0.003 to 0.015 microg/ml, and 1 microg/ml, respectively, but was inactive against Ehrlichia chaffeensis.     

In vitro susceptibilities of the Bacteroides fragilis group to 14 antimicrobial agents in Korea.

The susceptibilities to 14 antimicrobial agents of 172 clinical isolates of the Bacteroides fragilis group from Korean patients during 1989 and 1990 were tested by an agar dilution method. All the isolates tested were susceptible to imipenem, chloramphenicol, and metronidazole. The rate of resistance to cefoxitin was 6%.     

In vitro susceptibilities of Mycoplasma genitalium to antibiotics.

The susceptibilities of seven clinical isolates of Mycoplasma genitalium and three strains of Mycoplasma pneumoniae to a variety of antibiotics were examined by an agar dilution method. Macrolides, pristinamycin, and tetracyclines were very active against both species. Sparfloxacin was the most active quinolone tested. None of the 21 antibiotics tested had differential activity toward the two organisms.     

In vitro susceptibilities of Ehrlichia risticii to eight antibiotics.

Inhibition of the proliferation of Ehrlichia risticii cultured in murine macrophage P388D1 cells by eight antibiotics was evaluated by indirect fluorescent-antibody staining with an antiserum specific to E. risticii. There was a negative correlation between the percentage of infected cells and the log10 of the concentrations of all antibiotics examined. The ranks of the antibiotics in the order of 50% inhibitory concentrations (on a microgram-per-milliliter basis) after 48 h of exposure were as follows: demeclocycline, doxycycline, and oxytetracycline less than minocycline less than rifampin less than tetracycline less than erythromycin and nalidixic acid. When the antibiotics were removed after 48 h of incubation, continuous inhibition of proliferation was evident at 72 h. At 96 h regrowth of the organisms occurred in most of the cultures. The rate of regrowth was the highest with nalidixic acid, followed by erythromycin, at all concentrations of the antibiotic tested. Regrowth was observed with less than 0.1 microgram of minocycline per ml and less than 0.01 microgram of oxytetracycline, tetracycline, and doxycycline per ml. With more than 0.01 microgram of demeclocycline per ml, however, the inhibition persisted for up to 72 h after removal of the antibiotic. These results indicate that demeclocycline was slightly more effective than doxycycline, oxytetracycline, and minocycline in eliminating E. risticii in macrophages in vitro, whereas tetracycline and rifampin were less effective. Nalidixic acid and erythromycin were ineffective.     

In vitro susceptibilities of oral bacterial isolates to spiramycin.

Four hundred strains of oral bacteria were tested for their susceptibility to spiramycin. Actinobacillus actinomycetemcomitans and most species of Lactobacillus were resistant to the antibiotic. All strains of cariogenic Streptococcus mutans and most strains of bacterial species implicated in adult chronic periodontitis (Bacteroides gingivalis, B. intermedius, and Treponema denticola) were susceptible to spiramycin.     

In vitro susceptibilities of Aeromonas hydrophila to 32 antimicrobial agents.

Minimal inhibitory concentrations of 32 antimicrobial agents for 20 strains fo Aeromonas hydrophila were determined by a microdilution method. Moxalactam was the most active drug tested. All strains were also susceptible to clinically achievable concentrations of mecillinam, cefamandole, cefuroxime, cefotaxime, aminoglycosides (except streptomycin), tetracycline, chloramphenicol, and trimethoprim-sulfamethoxazole.     

In vitro susceptibilities of common pediatric pathogens to LY163892.

LY163892 is a carbacephem antibiotic for oral administration with an antibacterial spectrum similar to that of cefaclor and amoxicillin-clavulanic acid. It has greater stability than cefaclor and greater activity against beta-lactamase-producing Haemophilus influenzae and Escherichia coli. LY163892 is less active than amoxicillin against streptococci and less active than amoxicillin-clavulanic acid against Branhamella catarrhalis but comparable against other pathogens.     

In vitro susceptibilities of Pseudomonas pseudomallei to 27 antimicrobial agents.

Clinical isolates of Pseudomonas pseudomallei isolated in Thailand from 1981 to 1989 were tested for their in vitro susceptibilities to 27 antimicrobial agents, including older and newer quinolones, broad-spectrum cephems, carbapenems, monobactams, penicillins, tetracyclines, chloramphenicol, rifamycin, sulfamethoxazole, trimethoprim, and fosfomycin. Tosufloxacin, meropenem, CS-533, and minocycline were active against P. pseudomallei at levels comparable to or even greater than those of antimicrobial agents tested in previous studies, such as ciprofloxacin, ceftazidime, imipenem, carumonam, and piperacillin. Drug-resistant P. pseudomallei was found in only 1% of the isolates. The drug-resistant P. pseudomallei isolates displayed a unique pattern of susceptibility to the above-listed drugs.     

In vitro susceptibilities of Mycobacterium tuberculosis to 10 antimicrobial agents.

After preliminary in vitro screening of 10 antimicrobial agents against Mycobacterium tuberculosis, the MICs of the 6 most promising agents against 27 clinical isolates were determined by agar dilution. The two quinolone compounds tested (difloxacin and A-56620) were the most active, each inhibiting 50% of the strains at concentrations of 4 micrograms/ml. M. tuberculosis strains previously shown to be resistant to isoniazid, streptomycin, rifampin, or ethambutol were as susceptible to these quinolone compounds as susceptible strains.     

In vitro susceptibilities of Nocardia species to newer antimicrobial agents.

The in vitro activities of various quinolones, two newer cephalosporins, and imipenem against 23 strains of Nocardia asteroides, 4 strains of Nocardia brasiliensis, and 4 strains of Nocardia caviae were determined by an agar dilution method. PD-117558, a newer carboxyquinolone, and imipenem were the most active agents tested, inhibiting 90% of N. asteroides isolates at an MIC of 2 micrograms/ml. Of the 23 strains of N. asteroides, 15 were susceptible to cefpirome and 10 were susceptible to ciprofloxacin. The N. brasiliensis and N. caviae strains were very susceptible to PD-117558 (MIC, less than or equal to 1 microgram/ml), moderately susceptible to ciprofloxacin, and resistant to most of the other tested drugs.     

In vitro susceptibilities of Campylobacter-like organisms to twenty antimicrobial agents.

We determined MICs of 20 antimicrobial agents for 50 representative strains of four subgroups of Campylobacter-like organisms (CLOs) by agar dilution. Ampicillin, gentamicin, doxycycline, tetracycline, ceftriaxone, rifampin, spectinomycin, nalidixic acid, and chloramphenicol were active against all strains of CLOs. Most CLO strains (83%) were inhibited by 4 micrograms of sulfamethoxazole per ml and by 8 micrograms of trimethoprim-sulfamethoxazole per ml. Of type 1 strains, 28% were resistant to 8 micrograms of erythromycin per ml. In addition, cross resistance between erythromycin and clindamycin was always present. Type 1 strains exhibited a broad distribution of MICs of metronidazole and streptomycin, whereas all type 2 strains were uniformly susceptible to metronidazole and resistant to streptomycin. Unlike type 1 and 3 strains, type 2 CLOs were susceptible to cephalothin and penicillin G and highly resistant to streptomycin. The type 3 strain was uniquely resistant to cefazolin. The majority of strains were not inhibited by cefoperazone; and all were resistant to trimethoprim. In contrast to Campylobacter jejuni and Campylobacter fetus subsp. fetus, all CLOs tested were susceptible to 0.5 microgram of rifampin per ml.     

In vitro susceptibilities of mycoplasmas and ureaplasmas to new macrolides and aryl-fluoroquinolones.

In vitro activities of the new macrolides clarithromycin, previously designated A-56268 (TE-031), and A-63075 and of the aryl-fluoroquinolones difloxacin (A-56619) and temafloxacin (A-62254) against 14 strains of Mycoplasma pneumoniae, 20 strains of Mycoplasma hominis, and 28 strains of Ureaplasma urealyticum were compared with that of erythromycin. All three macrolides inhibited growth of M. pneumoniae at less than 0.125 micrograms/ml. No macrolide was active against M. hominis. For five strains of U. urealyticum, MICs were greater than 256 micrograms/ml for all 3 macrolides. Excluding these, no other strain of U. urealyticum had an initial MIC of clarithromycin of greater than 1 microgram/ml, while five had initial MICs of erythromycin which were greater than 4 micrograms/ml. A-63075 was the least active of the three macrolides against ureaplasmas. Temafloxacin and difloxacin had similar activities against all three species, initially inhibiting 90% of M. pneumoniae strains at 2 and 8 micrograms/ml, 90% of M. hominis strains at 2 and 4 micrograms/ml, and 90% of U. urealyticum strains at 4 and 8 micrograms/ml, respectively. Additional pharmacokinetic and clinical trials with the new macrolides and quinolones with mycoplasmal or ureaplasmal infections are indicated.     

In vitro susceptibilities of zygomycetes to conventional and new antifungals

In vitro susceptibilities of 36 zygomycete isolates, belonging to six genera, to itraconazole, posaconazole, voriconazole, terbinafine, amphotericin B and 5-fluorocytosine were determined by using a broth microdilution adaptation of the National Committee for Clinical Laboratory Standards M-38P reference method. The influence of incubation time on MIC values, and the performance of a spectrophotometric method for MIC determination in comparison with the visual reference method, were also evaluated. Amphotericin B was active against most of the isolates. All the isolates were highly resistant to 5-fluorocytosine (MICs > 256 mg/L). Voriconazole was significantly less active than the other drugs with an overall MIC(90) (MIC at which 90% of the isolates were inhibited) of 32 mg/L. In contrast, posaconazole showed good activity (MIC(90) 1 mg/L). A wide range of MICs, from 0.03 to > or =32 mg/L, was obtained for itraconazole and terbinafine. Differences in susceptibility between and within genera were noted. Rhizopus spp. were significantly less susceptible to itraconazole, posaconazole, terbinafine and amphotericin B than Absidia spp., and less susceptible than Mucor spp. to amphotericin B. Terbinafine appeared to be more active against Rhizopus microsporus than against Rhizopus oryzae (geometric mean MIC of 0.15 and 64 mg/L, respectively). The activity of the drugs was dependent on the incubation period. A significant increase in MICs was noted between 24 and 48 h of incubation. On the other hand, the two methods used for MIC determination (visual and spectrophotometric readings) showed good agreement. These results suggest that the zygomycetes are a heterogeneous group for antifungal susceptibility. Some of the conventional and new antifungals are effective in vitro; their efficacies in vivo remain to be determined. The spectrophotometric method appears to be a valuable alternative to the visual method for MIC determination for zygomycetes.     

In vitro susceptibilities of Bordetella pertussis and Bordetella parapertussis to seven fluoroquinolones.

The in vitro susceptibilities of Bordetella pertussis and Bordetella parapertussis to seven fluoroquinolones were assessed by the agar dilution method. Ciprofloxacin and temafloxacin were the most active compounds (MIC for 90% of isolates tested [MIC90], 0.06 microgram/ml), while enoxacin and pefloxacin were the least active (MIC90, 0.5 microgram/ml). Fleroxacin, lomefloxacin, and ofloxacin showed intermediate activities (MIC90s, 0.125 to 0.25 microgram/ml). These results suggest a possible role for the fluoroquinolones in the treatment of pertussis, at least in adult patients.     

In vitro susceptibilities of Borrelia burgdorferi to five oral cephalosporins and ceftriaxone.

We determined the in vitro susceptibilities of eight Borrelia burgdorferi isolates to five oral cephalosporins. MICs for B. burgdorferi 297 were 23 micrograms/ml (cephalexin), 45 micrograms/ml (cefadroxil), 91 micrograms/ml (cefaclor), 0.13 microgram/ml (cefuroxime), 0.8 microgram/ml (cefixime), and 0.02 microgram/ml (ceftriaxone). When B. burgdorferi isolates were exposed to concentrations twice the MIC of cefuroxime, cefixime, or ceftriaxone, at least 72 h of incubation was required to kill 99% of the organisms.