The comparative in vitro activity of the ketolide HMR 3647 (RU 66647) and those of structurally related macrolide-lincosamide-streptogramin compounds (erythromycin, roxithromycin, azithromycin, clarithromycin, josamycin, lincomycin, pristinamycin, and quinupristin-dalfopristin) as well as those of benzylpenicillin, doxycycline, vancomycin, teicoplanin, levofloxacin, and rifapentine against 247 aerobic and facultative non-spore-forming gram-positive bacilli were determined by an agar dilution method. The ketolide was active against most organisms tested exceptCorynebacterium striatum, coryneform CDC group I2, andOerskovia spp. The frequency of resistance to erythromycin and other macrolides as well as that to lincomycin was high. Pristinamycin and, to a lesser extent, quinupristin-dalfopristin were very active, but resistance to these agents was present in some strains of Rhodococcus equi, Listeria spp., C. striatum, Erysipelothrix rhusiopathiae, andOerskovia spp. HMR 3647 was very active against all erythromycin-sensitive and many erythromycin-nonsusceptible strains, especially Corynebacterium minutissimum,Corynebacterium pseudodiphtheriticum, Corynebacterium amycolatum, and Corynebacterium jeikeium. In vitro resistance to benzylpenicillin was common, but doxycycline, vancomycin, and teicoplanin were very active against most organisms tested exceptE. rhusiopathiae, against which glycopeptide antibiotics were not active. The in vitro activity of levofloxacin was remarkable, but resistance to this agent was common for C. amycolatum,Corynebacterium urealyticum, C. jeikeium, andOerskovia spp. strains. Rifapentine was also very active in vitro against many organisms, but resistance to this agent was always present in E. rhusiopathiae and was very common in C. striatum and C. urealyticum.

The efficacy of amoxicillin/clavulanate and cefuroxime was determined in a gerbil model of otitis media with a mixed Streptococcus pneumoniae plus Haemophilus influenzae middle ear (ME) infection. Results were compared with those obtained in a previous single H. influenzae model. All untreated animals inoculated with the mixed inoculum developed acute otitis media (AOM), whereas 86.7% of those inoculated with H. influenzae developed otitis media with effusion (OME). Antibiotics eradicated H. influenzae from the ME more efficiently in AOM than in OME, and this difference was highly significant (P80% of animals developed culture-negative OME.

Amoxicillin at doses of 0.2 to 5 mg/kg of body weight was administered for the treatment of pneumococcal otitis media in a gerbil model. Doses greater than or equal to 2.5 mg/kg, which resulted in concentrations in middle ear fluid of >1.4 g/ml and concentrations in serum higher than the MIC (1 g/ml) for >14% of the dosing interval, were both clinically and bacteriologically effective.The emergence of penicillin-insensitive Streptococcus pneumoniae strains is now a worldwide problem and causes great concern (2, 3, 12). To overcome this problem in cases of acute otitis media (AOM), a proposal has been made to increase the amoxicillin dose (5) in order to achieve concentrations in serum above the MIC for the pathogen during at least 40% of the dosing interval (4).The aim of this work was to determine, by means of an experimental model of AOM caused by a penicillin-resistant pneumococcus (penicillin MIC, 2 g/ml), the minimum dose of amoxicillin able to achieve both clinical and bacteriological success.One S. pneumoniae serotype 23F strain was used. Amoxicillin trihydrate (SmithKline Beecham Pharmaceuticals, Worthing, England) was used for in vitro studies. For in vivo studies, vials of commercially produced amoxicillin (Clamoxyl; SmithKline Beecham Pharmaceuticals, Toledo, Spain) were employed.MICs and minimum bactericidal concentrations were determined five times by a microdilution method (10, 11), and median values were considered. Eight-to 9-week-old adult female Mongolian gerbils were inoculated bilaterally with ϳ5 ϫ 10 6 S. pneumoniae CFU per 20 l, as previously reported (13). AOM and otitis media with effusion (OME) were defined as previously described (13).The antibiotic was tested at doses of 0.2, 0.4, 0.8, 1.25, 2.5, and 5 mg/kg of body weight and administered subcutaneously (s.c.) in 500-l volumes at 2, 10, and 18 h postinoculation (p.i.). Animals in the control group received apyrogen sterile distilled water. Treated and control animals were studied daily for otorrhea, weight, and behavior. On day 2 p.i., ears were examined with an otoscope and middle ear (ME) samples were obtained by washing the ME fossa with 20 l of saline solution (ME with wash fluid [MEWF] samples) for determining the volume and presence of bacteria and cells. Amoxicillin levels in serum at 15, 30, 60, and 120 min after drug administration were determined in groups of six healthy animals after single s.c. injections of the doses that had demonstrated the highest efficacies.Concentrations of the antibiotic in ME fluid without washing (MEF samples) were also determined for groups of 10 bilaterally inoculated animals. Single s.c. doses of the antibiotic or control were administered 46 h after bacterial inoculation. MEF samples were obtained 90 min later, and aliquots were pooled for determination of the antibiotic concentrations.Antibiotic concentrations were determined by using Micrococcus luteus ATCC 9341. Assay variability for individual samples was Ͻ10%.Differences in eradication (and presence of otorrhea at day 1) were analyzed by ...

A gerbil model of otitis media induced by a beta-lactamase producing and non-serotypeable isolate of Haemophilus influenzae was used to assess the in-vivo efficacy of co-amoxiclav and cefuroxime at low (5 mg/kg) and high (20 mg/kg) doses. The MIC of the antibiotics tested against the pathogen was 1 mg/L (1/0.5 mg/L for co-amoxiclav). The organism was inoculated (+/-10(6) cfu) by transbullar challenge directly in the middle ear and antibiotic treatment was commenced 2 h post-inoculation and continued at 8 h intervals for three doses. Only high dose co-amoxiclav significantly reduced the number of culture-positive specimens as compared with untreated animals or with other treatment groups (91.7% as compared with 36.7% for high dose cefuroxime). The results obtained in any treatment group were related to middle ear antibiotic level/MIC. Antibiotic concentrations in the middle ear 90 min after administration were about 10% of serum levels at 15 min, probably related to a slight inflammatory response. Only after high dose co-amoxiclav did the concentration in the middle ear exceed the MIC by a factor of four. In otitis media with effusion, if indicated, antibiotics active in vitro should be administered in high doses and, to avoid side effects, probably in short courses.

The comparative efficacies of amoxicillin and cefuroxime against acute otitis media caused by a penicillin-resistant (MIC, 2 μg/ml) Streptococcus pneumoniae strain were assessed in a gerbil model by challenging each ear with 107 bacteria through transbullar instillation. Each antibiotic was tested at two doses (5 and 20 mg/kg of body weight) administered at 2, 10, and 18 h postinoculation. Samples were obtained from the middle ear (ME) on days 3 and 7 postinoculation for determination of bacterial counts. Only amoxicillin, at both doses, was able to significantly halt the weight loss in animals, reducing both the number of culture-positive animals and the bacterial concentration in ME samples versus the values for untreated animals. Comparison of the efficacies between the antibiotics, determined by their ability to achieve culture-negative ME specimens, showed that amoxicillin at 5 mg/kg was significantly more active than cefuroxime at the same dose. The use of higher doses of either amoxicillin or cefuroxime did not produce significantly better results than those obtained with the lower dose but caused a greater inflammatory response. The more favorable results obtained with amoxicillin compared with those obtained with cefuroxime could be related to the antimicrobial susceptibility of the pneumococcal strain (MICs and minimum bactericidal concentrations of 1 and 1 μg/ml and 4 and 4 μg/ml for amoxicillin and cefuroxime, respectively) as well as to the better pharmacokinetic parameters obtained with amoxicillin.

We have read with interest the article by Johnson et al. (7) about the variability of the results of Gram staining encountered with some anaerobes. Gram stain is one of the cornerstones for bacterial identification, but it also serves as a useful technique for rapid detection of organisms in clinical samples (6). Propionibacterium acnes is a non-spore-forming, pleomorphic, anaerobic gram-positive rod (3) that is frequently considered to be a contaminant in clinical samples but may also be implicated as the agent of human infection (2, 4, 8). In our experience, Gram stain directly performed on clinical samples from patients with documented infection by P. acnes frequently failed to detect this organism. In order to evaluate the usefulness of Gram stain for direct detection of P. acnes, the laboratory records of the Medical Microbiology Department of the Fundación Jiménez Díaz (a 600-bed university hospital in Madrid, Spain) between January 1988 and August 1995 were reviewed for culture evidence of the presence of P. acnes. Those samples with heavy (3-streak) or moderate (2-streak) growth of P. acnes together with evidence of an inflammatory reaction (as detected on Gram smear by the presence of polymorphonuclear leukocytes) and yielding pure or predominant growth of P. acnes without any other more probable pathogen were selected. Critical review of patient charts was done to evaluate the clinical significance of the isolate. In 39 cases, meeting the above mentioned selection criteria, P. acnes was judged to have clinical significance. For controlling these results we later selected randomly other charts and assessed the Gram stain utility for other infections caused by various grampositive and gram-negative organisms. Table 1 shows the results of Gram stain for P. acnes infections and controls, indicating where the ability of the Gram stain to detect P. acnes was poor, especially when growth was only moderate, as compared with the ability of the Gram stain to reveal other grampositive and gram-negative organisms. Murray et al. (5) attribute misleading Gram stain results in clinical samples to cell wall deterioration due to the inflammatory response. Alterations in the P. acnes cell wall because of interaction with leukocytes or monocytes have been reported previously (9), and this fact, together with the pleomorphic nature of this bacteria (occasionally misinterpreted as a mixed infection [1, 8]) plus the presence of fibrinous debris in the samples, may explain the observations made in our study. In conclusion, Gram stain may not be a reliable technique for the rapid diagnosis of P. acnes infections, and when evidence of an abundant inflammatory response is noted in the Gram-stained smear, a more careful evaluation of cultures must be performed, particularly when nosocomial infection is of concern.

Urine bactericidal titers (UBTs) against Escherichia coli ATCC 25922 and Staphylococcus saprophyticus ATCC 1970 were determined after the administration of single oral doses of gemifloxacin at 320 mg and trovafloxacin at 200 mg to healthy volunteers. Gemifloxacin presented significantly lower experimental versus mathematically predicted UBTs over 72 h, due to the effect of urine on the susceptibility of the E. coli strain. Experimental UBTs were significantly higher for gemifloxacin than trovafloxacin against both strains over 72 h."Ex vivo" activity in urine is important since the measurement of urine antibacterial activity correlates directly with the outcome of infection (4) more than the determination of MICs and the levels of antimicrobials in urine (2), probably because MICs are higher when tested in human urine than in conventional media (6).Urine bactericidal titers (UBTs) and the area under the urine bactericidal curve (AUBC) of gemifloxacin versus trovafloxacin were evaluated against Escherichia coli and Staphylococcus saprophyticus strains.Twelve healthy male volunteers (mean age, 26.2 Ϯ 3.5 years; height, 175.2 Ϯ 4.7 cm; weight, 73.2 Ϯ 4.2 kg) received single oral doses of 320 mg of gemifloxacin (SmithKline Beecham Pharmaceuticals, Harlow, United Kingdom) and 200 mg of trovafloxacin (Trovan; Pfizer Inc., New York, N.Y.), separated by a 14-day washout period. The protocol was approved by the Research Ethics Committee of Hospital La Paz, Madrid, Spain. Written informed consent was obtained from all subjects.Urine samples were collected prior to drug administration (0 h) and at intervals of 0 to 2, 2 to 4, 4 to 8, 8 to 12, 12 to 16, 16 to 24, 24 to 36, 36 to 48, 48 to 60, and 60 to 72 h after dosing for determination of urine concentrations of gemifloxacin and trovafloxacin by high-pressure liquid chromatography (HPLC) and bioassay and for determination of urine bactericidal titers (UBTs).In vitro susceptibility testing was performed using Laboratory reference standards of gemifloxacin and trovafloxacin (SmithKline Beecham Pharmaceuticals, Tonbridge, United Kingdom), five times each for E. coli ATCC 25922 and Staphylococcus saprophyticus ATCC 1970 by standardized methods (5). In addition, the same methodology was followed but using a pool of the volunteer's urine as test media to study the effect of urine on the antibacterial activity of the study drugs.Gemifloxacin and trovafloxacin concentrations were determined by bioassay in plates with nutrient agar (BBL; Becton Dickinson, Cockeysville, Md.) seeded with Bacillus subtilis NCTC 6633 spore suspension (Difco Laboratories, Detroit, Mich.). A total of 90 l of each sample was deposited into 7-mm-diameter wells in the inoculated plates and incubated at 30°C for 18 h. Gemifloxacin and trovafloxacin standards of from 32 to 0.125 g/ml were prepared in pooled urine obtained from volunteers before administration of the drugs. The lower limit of detection was 0.125 g/ml.For HPLC determination, frozen samples were thawed at ambient temperature, agitated at 37°C u...

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