Postantibiotic effect of ceftaroline against gram-positive organisms.

The postantibiotic effects (PAEs), postantibiotic sub-MIC effects (PA-SMEs), and sub-MIC effects (SMEs) of ceftaroline, a novel injectable cephalosporin, were determined for 15 gram-positive organisms. The pneumococcal, staphylococcal, and enterococcal PAEs were 0.8 to 1.8 h, 0.7 to 2.2 h, and 0.2 to 1.1 h, respectively. The corresponding PA-SMEs (0.4 times the MIC) were 2.5 to 6.7 h, 2.9 to >0.0 h, and 7.9 to >10.3 h, respectively. The PA-SMEs were longer than the PAEs, suggesting that sub-MIC levels extend the PAE of ceftaroline against gram-positive cocci.

Postantibiotic effects of telavancin against 16 gram-positive organisms.

The in vitro postantibiotic effects (PAEs), postantibiotic sub-MIC effects (PA-SMEs), and sub-MIC effects of telavancin were determined for 16 gram-positive organisms. Telavancin staphylococcal, streptococcal, and enterococcal PAE ranges were 0.9 to 3.9 h, 0.4 to 6.7 h, and 0.3 to 2.2 h, respectively. The PA-SME ranges (0.4 times the MIC) for staphylococci, streptococci, and enterococci were 6.7 to >10.7 h, >10.7 to >11.0 h, and >10 to >10.8 h, respectively. The extended PAE of telavancin, together with its long elimination half-life in humans, supports once-daily dosing for this investigational drug.

Postantibiotic effect of tigecycline against 14 gram-positive organisms.

The in vitro postantibiotic effects (PAEs), postantibiotic sub-MIC effects (PA-SMEs), and sub-MIC effects of tigecycline were determined for 14 gram-positive and gram-negative organisms. The pneumococcal, staphylococcal, and enterococcal PAEs were 1.9 to 5.1, 2.9 to 5.7, and 3.9 to 6.1 h, respectively, and those for Haemophilus influenzae, Escherichia coli, Klebsiella pneumoniae, Enterobacter cloacae, and Acinetobacter baumannii were 1.1 to 5.0, 1.9 to 2.1, 1.7 to 1.8, 1.0 to 1.7, and 0.7 to 3 h, respectively. The PA-SMEs (four times the MIC) ranged from 6.7 to >11 h for gram-positive organisms and from 2.3 to >11.3 h for gram-negative organisms.

Postantibiotic effect of ceftobiprole against 12 Gram-positive organisms.

The in vitro postantibiotic effects (PAEs), postantibiotic sub-MIC effects (PA-SMEs), and sub-MIC effects of ceftobiprole were determined for 12 gram-positive organisms. Pneumococcal, staphylococcal, and enterococcal PAEs were 1.4 to 3.1 h, 0 to 1.8 h, and 0 to 0.9 h, respectively. The PA-SMEs (0.4 times the MIC) for pneumococci, staphylococci, and enterococci were 4.8 to >10.3 h, 1.5 to 9.6 h, and 3.8 to >10.7 h, respectively.

Activity of five quinolones, three macrolides and telithromycin against 12 Haemophilus influenzae strains with different resistance phenotypes.

Gemifloxacin MICs for 12 Haemophilus influenzae strains with different resistance phenotypes were 0.001-0.015 mg/L. Gemifloxacin was bactericidal against all 12 strains after 24 h at 2 x MIC. Ciprofloxacin, levofloxacin, gatifloxacin and moxifloxacin had MICs of 0.008-0.03 mg/L and similar kill kinetics. Macrolides and telithromycin had unimodal MICs (1.0-8.0 mg/L), except for two strains without efflux systems (0.0125-0.5 mg/L) and two with efflux systems and ribosomal protein mutations (> 64.0 mg/L), and were bactericidal against eight to ten strains tested at 2 x MIC after 24 h.

Postantibiotic effect of DX-619 against 16 gram-positive organisms.

The in vitro postantibiotic effects (PAEs), the postantibiotic sub-MIC effects (PA-SMEs), and the sub-MIC effects (SMEs) of DX-619 were determined for 16 gram-positive organisms. DX-619 pneumococcal, staphylococcal, and enterococcal PAE ranges were 1.7 to 5.0 h, 0.7 to 1.8 h, and 1.2 to 6.5 h, respectively. The PA-SME ranges (0.4x MIC) for pneumococci, staphylococci, and enterococci were 5.2 to >8.6 h, 2.1 to 8.3 h, and 4.9 to >10.0 h, respectively.

Activity of the new quinolone WCK 771 against pneumococci.

The activity of WCK 771, a new experimental quinolone being developed to overcome quinolone resistance in staphylococci, against quinolone-susceptible and -resistant pneumococci was determined. Comparative activities of ciprofloxacin, levofloxacin, gatifloxacin, moxifloxacin, clinafloxacin, vancomycin, linezolid, amoxycillin, cefuroxime, azithromycin and clarithromycin were determined with MIC and time-kill experiments. Animal experiments were also performed to test the in-vivo anti-pneumococcal activity of WCK 771 compared to levofloxacin. WCK 771 MIC50/90 values for 300 quinolone-susceptible Streptococcus pneumoniae isolates (108 penicillin-susceptible, 92 penicillin-intermediate and 100 penicillin-resistant) were 0.5/0.5 mg/L; the MICs of beta-lactams and macrolides rose with those of penicillin G, and all isolates were susceptible to vancomycin and linezolid. WCK 771 MIC50/90 values for 25 quinolone-resistant pneumococcal isolates were 4/8 mg/L, compared to 0.5/1 mg/L for clinafloxacin, 2/4 mg/L for gatifloxacin and moxifloxacin, 8/16 mg/L for levofloxacin, and 16/>32 mg/L for ciprofloxacin. Time-kill studies showed that WCK 771 was bactericidal against pneumococci after 24 h at 4 x MIC, as were the other quinolones tested. Animal model studies showed that WCK 771 had efficacy comparable to that of levofloxacin, by both the oral and subcutaneous routes, for systemic infection caused by three quinolone-susceptible isolates of pneumococci. Overall, WCK 771 was potent both in vivo and in vitro against quinolone-susceptible, but not quinolone-resistant, S. pneumoniae, regardless of penicillin susceptibility.

Postantibiotic effects of daptomycin against 14 staphylococcal and pneumococcal clinical isolates.

Daptomycin mean staphylococcal postantibiotic effects (PAEs) were 1.1 to 6.2 h, with a mean of 2.5 h. The mean pneumococcal PAEs were 1.7 h, ranging between 1.0 and 2.5 h. The staphylococcal and pneumococcal postantibiotic sub-MIC effects at 0.4 times the MIC ranged from 3.0 to >12.0 h and 1.9 to >12.0 h, respectively.

Postantibiotic effects of garenoxacin (BMS-284756) against 12 gram-positive or -negative organisms.

Conventional in vitro methods were used to determine the postantibiotic effects (PAEs), sub-MIC effects (SMEs), and postantibiotic sub-MIC effects (PA-SMEs) of garenoxacin for a range of organisms. The mean PAEs of garenoxacin for pneumococci, staphylococci, and enterococci were 0.3 to 2.2 h. For Escherichia coli and Pseudomonas aeruginosa, the PAEs were 0.9 to 1.6 h. The mean PA-SMEs (0.4 times the MIC) for pneumococci, staphylococci, and enterococci were 3.0 to >10 h, 1.8 to >10.7 h, and 5.8 h, respectively, while those for E. coli and P. aeruginosa were 7.6 and 4.4 h, respectively.

Susceptibilities of Haemophilus influenzae and Moraxella catarrhalis to ABT-773 compared to their susceptibilities to 11 other agents.

The activity of the ketolide ABT-773 against Haemophilus and Moraxella was compared to those of 11 other agents. Against 210 Haemophilus influenzae strains (39.0% beta-lactamase positive), microbroth dilution tests showed that azithromycin and ABT-773 had the lowest MICs (0.5 to 4.0 and 1.0 to 8.0 microg/ml, respectively), followed by clarithromycin and roxithromycin (4.0 to >32.0 microg/ml). Of the beta-lactams, ceftriaxone had the lowest MICs (32.0 microg/ml). Against 50 Moraxella catarrhalis strains, all of the compounds except amoxicillin and cefprozil were active. Time-kill studies against 10 H. influenzae strains showed that ABT-773, at two times the MIC, was bactericidal against 9 of 10 strains, with 99% killing of all strains at the MIC after 24 h; at 12 h, ABT-773 gave 90% killing of all strains at two times the MIC. At 3 and 6 h, killing by ABT-773 was slower, with 99.9% killing of four strains at two times the MIC after 6 h. Similar results were found for azithromycin, with slightly slower killing by erythromycin, clarithromycin, and roxithromycin, especially at earlier times. beta-Lactams were bactericidal against 8 to 10 strains at two times the MIC after 24 h, with slower killing at earlier time periods. Most compounds gave good killing of five M. catarrhalis strains, with beta-lactams killing more rapidly than other drugs. ABT-773 and azithromycin gave the longest postantibiotic effects (PAEs) of the ketolide-macrolide-azalide group tested (4.4 to >8.0 h), followed by clarithromycin, erythromycin, and roxithromycin. beta-Lactam PAEs were similar and shorter than those of the ketolide-macrolide-azalide group for all strains tested.

In vitro development of resistance to ceftriaxone, cefprozil and azithromycin in Streptococcus pneumoniae.

Approval of ceftriaxone for the treatment of otitis media has led to fear of selection of resistant mutants owing to widespread use. To test this, we examined the ability of sequential subcultures in sub-MICs of ceftriaxone, cefprozil and azithromycin to select resistant mutants in 12 pneumococci. Daily subculturing was performed 50 times or until mutants with raised ceftriaxone, cefprozil or azithromycin MICs were selected. Of eight ceftriaxone-susceptible parents, ceftriaxone did not select for any resistant mutants, while cefprozil selected for four mutants (MICs 2-4 mg/L after 21-50 subcultures). Among four ceftriaxone-resistant parents, subculturing in ceftriaxone selected for one stable mutant with raised ceftriaxone MIC (>16 mg/L after 21 subcultures) and subculturing in cefprozil selected for one mutant with raised cefprozil MIC (64 mg/L after 44 subcultures). Mutations were observed in pbp2x and pbp1a. Among six azithromycin-susceptible parents, subculturing in azithromycin selected for five resistant mutants (MIC 0.5-32 mg/L after 10-42 passages) and among six azithromycin-resistant strains, subculturing selected for mutants with raised azithromycin MICs in all six strains (MIC 16-32 mg/L after 4-18 passages). All azithromycin-resistant mutants derived from azithromycinsusceptible parents had mutations in domain V of 23S rRNA while all azithromycin-resistant parents and derived mutants had mefE. Single-step mutation rates among the 12 strains at the MIC ranged from 1.5 x 10(-6) to <6.2 x 10(-10) for ceftriaxone, >1.3 x 10(-5) to 8.9 x 10(-8) for cefprozil and >1.1 x 10(-6) to 6.7 x 10(-10) for azithromycin. Multi-step and single-step testing showed that ceftriaxone selected for resistant mutants less often than cefprozil and azithromycin.

In vitro selection of resistance to clinafloxacin, ciprofloxacin, and trovafloxacin in Streptococcus pneumoniae.

Ability of daily sequential subcultures in subinhibitory concentrations of clinafloxacin, ciprofloxacin, and trovafloxacin to select resistant mutants was studied in 10 pneumococci (ciprofloxacin MICs, 1 to 4 microg/ml, and clinafloxacin and trovafloxacin MICs, 0.06 to 0.125 microg/ml [n = 9]; ciprofloxacin, clinafloxacin, and trovafloxacin MICs, 32, 0.5, and 2 microg/ml, respectively [n = 1]). Subculturing was done 50 times, or until MICs increased fourfold or more. Mutants for which MICs were fourfold (or more) higher than those for parent strains were selected in five strains by clinafloxacin, in six strains by trovafloxacin, and nine strains by ciprofloxacin. Sequence analysis of type II topoisomerase showed that most mutants had mutations in ParC at Ser79 or Asp83 and in GyrA at Ser81, while a few mutants had mutations in ParE or GyrB. In the presence of reserpine, the MICs of ciprofloxacin and clinafloxacin for most mutants were lower (four to eight times lower), but for none of the mutants were trovafloxacin MICs lower, suggesting an efflux mechanism affecting the first two agents but not trovafloxacin. Single-step mutation rates were also determined for eight strains for which the MICs were as follows: 0.06 microg/ml (clinafloxacin), 0.06 to 0.125 microg/ml (trovafloxacin), and 1 microg/ml (ciprofloxacin). Single-step mutation rates with drugs at the MIC were 2.0x10(-9) to <1.1x10(-11), 5.0x10(-4) to 3.6x10(-9), and 4.8x10(-4) to 6.7x10(-9), respectively. For two strains with clinafloxacin MICs of 0.125 to 0.5 microg/ml trovafloxacin MICs of 0. 125 to 2 microg/ml, ciprofloxacin MICs of 4 to 32 microg/ml mutation rates with drugs at the MIC were 1.1x10(-8)-9.6x10(-8), 3.3x10(-6)-6. 7x10(-8), and 2.3x10(-5)-2.4x10(-7), respectively. Clinafloxacin was bactericidal at four times the MIC after 24 h against three parent and nine mutant strains by time-kill study. This study showed that single and multistep clinafloxacin exposure selected for resistant mutants less frequently than similar exposures to other drugs studied.

Antipneumococcal activity of ABT-773 compared to those of 10 other agents.

MICs, time-kills, and postantibiotic effects (PAEs) of ABT-773 (a new ketolide) and 10 other agents were determined against 226 pneumococci. Against 78 ermB- and 44 mefE-containing strains, ABT-773 MICs at which 50% of the isolates tested were inhibited (MIC(50)s) and MIC(90)s were 0.016 to 0.03 and 0.125 microgram/ml, respectively. Clindamycin was active only against macrolide-resistant strains containing mefE (MIC(50), 0.06 microgram/ml; MIC(90), 0.125 microgram/ml). Activities of pristinamycin (MIC(90), 0.5 microgram/ml) and vancomycin (MIC(90), 0.25 microgram/ml) were unaffected by macrolide or penicillin resistance, while beta-lactam MICs rose with those of penicillin G. Against 19 strains with L4 ribosomal protein mutations and two strains with mutations in domain V of 23S rRNA, ABT-773 MICs were 0.03 to 0.25 microgram/ml, while macrolide and azalide MICs were all >/=16.0 microgram/ml. ABT-773 was bactericidal at twice the MIC after 24 h for 8 of 12 strains (including three strains with erythromycin MICs greater than or equal to 64.0 microgram/ml). Kill kinetics of erythromycin, azithromycin, clarithromycin, and roxithromycin against macrolide-susceptible strains were slower than those of ABT-773. ABT-773 had longer PAEs than macrolides, azithromycin, clindamycin, or beta-lactams, including against ermB-containing strains. ABT-773, therefore, shows promising in vitro activity against macrolide-susceptible as well as -resistant pneumococci.

Antipneumococcal activities of GAR-936 (a new glycylcycline) compared to those of nine other agents against penicillin-susceptible and -resistant pneumococci.

GAR-936, a new glycylcycline, had lower MICs (< or =0.016 to 0.125 microg/ml) for 201 penicillin- and tetracycline-susceptible and -resistant pneumococcal strains than tetracycline (< or = 0.06 to 128 microg/ml), minocycline (< or =0.06 to 16.0 microg/ml), or doxycycline (< or =0.06 to 32.0 microg/ml). GAR-936 was also bactericidal against 11 of 12 strains tested at the MIC after 24 h, with significant kill rates at earlier time points.

Antipneumococcal activities of gemifloxacin compared to those of nine other agents.

The activities of gemifloxacin compared to those of nine other agents was tested against a range of penicillin-susceptible and -resistant pneumococci by agar dilution, microdilution, time-kill, and post-antibiotic effect (PAE) methods. Against 64 penicillin-susceptible, 68 penicillin-intermediate, and 75 penicillin-resistant pneumococci (all quinolone susceptible), agar dilution MIC(50)s (MICs at which 50% of isolates are inhibited)/MIC(90)s (in micrograms per milliliter) were as follows: gemifloxacin, 0.03/0.06; ciprofloxacin, 1.0/4.0; levofloxacin, 1.0/2. 0; sparfloxacin, 0.5/1.0; grepafloxacin, 0.125/0.5; trovafloxacin, 0. 125/0.25; amoxicillin, 0.016/0.06 (penicillin-susceptible isolates), 0.125/1.0 (penicillin-intermediate isolates), and 2.0/4.0 (penicillin-resistant isolates); cefuroxime, 0.03/0.25 (penicillin-susceptible isolates), 0.5/2.0 (penicillin-intermediate isolates), and 8.0/16.0 (penicillin-resistant isolates); azithromycin, 0.125/0.5 (penicillin-susceptible isolates), 0. 125/>128.0 (penicillin-intermediate isolates), and 4.0/>128.0 (penicillin-resistant isolates); and clarithromycin, 0.03/0.06 (penicillin-susceptible isolates), 0.03/32.0 (penicillin-intermediate isolates), and 2.0/>128.0 (penicillin-resistant isolates). Against 28 strains with ciprofloxacin MICs of >/=8 microg/ml, gemifloxacin had the lowest MICs (0.03 to 1.0 microg/ml; MIC(90), 0.5 microg/ml), compared with MICs ranging between 0.25 and >32.0 microg/ml (MIC(90)s of 4.0 to >32.0 microg/ml) for other quinolones. Resistance in these 28 strains was associated with mutations in parC, gyrA, parE, and/or gyrB or efflux, with some strains having multiple resistance mechanisms. For 12 penicillin-susceptible and -resistant pneumococcal strains (2 quinolone resistant), time-kill results showed that levofloxacin at the MIC, gemifloxacin and sparfloxacin at two times the MIC, and ciprofloxacin, grepafloxacin, and trovafloxacin at four times the MIC were bactericidal for all strains after 24 h. Gemifloxacin was uniformly bactericidal after 24 h at

Antipneumococcal activity of MEN 10700, a new penem, compared with other compounds, by MIC and time-kill kinetics.

The antipneumococcal activity of MEN 10700 was compared with those of nine other compounds by MIC and time-kill kinetics. MIC90s (mg/L) of 202 penicillin-susceptible, -intermediate and -resistant pneumococci were: 0.06, 1.0 and 2.0 (MEN 10700); 0.06, 0.5-1.0 and 2.0 (amoxycillin +/- clavulanate); 0.06, 0.5 and 4.0 (cefotaxime); 0.125, 0.5 and 2.0 (cefepime); 0.016, 0.125 and 0.25 (imipenem); 0.03, 0.5 and 1.0 (meropenem); 2.0 (ciprofloxacin); 0.125, >64.0 and >64.0 (clarithromycin); and 0.5 (vancomycin). Time-kill kinetics showed that MEN 10700, at 4 x MIC, was bactericidal for all 12 isolates tested at 4 x MIC. Kinetics of other beta-lactams were similar to those of MEN 10700, relative to MICs. Ciprofloxacin, at 4 x MIC, was uniformly bactericidal after 24 h. Clarithromycin exhibited slow kill kinetics, after 24 h. Vancomycin was bactericidal against 11/12 isolates at 2 x MIC after 24 h.

Postantibiotic effects of gatifloxacin against gram-positive and -negative organisms.

Gatifloxacin pneumococcal, staphylococcal and enterococcal postantibiotic effects (PAEs) were 0.5 to 4.0 h, respectively. For Escherichia coli and Pseudomonas aeruginosa, PAEs were 2.2 to 4.8 h. Pneumococcal, staphylococcal, and enterococcal postantibiotic sub-MIC effects (PA-SMEs) (four times the MICs) were 3.7 to 8.6, 2.3 to 3.8, and 1.6 h, respectively, and E. coli and P. aeruginosa PA-SMEs were >/=9.6 and 4.4 h, respectively.

Susceptibilities of Streptococcus pneumoniae and Haemophilus influenzae to 10 oral antimicrobial agents based on pharmacodynamic parameters: 1997 U.S. Surveillance study.

The susceptibilities of Streptococcus pneumoniae (1,476 strains) and untypeable Haemophilus influenzae (1,676 strains) to various oral beta-lactam, macrolide-azalide, and fluoroquinolone antimicrobial agents were determined by broth microdilution. Organisms were isolated from specimens obtained from outpatients in six geographic regions of the United States. MIC data were interpreted according to pharmacodynamically derived breakpoints applicable to the oral agents tested. Among H. influenzae strains, 41.6% were beta-lactamase positive. Virtually all H. influenzae strains were susceptible to amoxicillin-clavulanate (98%), cefixime (100%), and ciprofloxacin (100%), while 78% were susceptible to cefuroxime, 57% were susceptible to amoxicillin, 14% were susceptible to cefprozil, 9% were susceptible to loracarbef, 2% were susceptible to cefaclor, and 0% were susceptible to azithromycin and clarithromycin. Among S. pneumoniae isolates, 49.6% were penicillin susceptible, 17.9% were intermediate, and 32.5% were penicillin resistant, with penicillin MICs for 50 and 90% of the isolates tested of 0.12 and 4 microg/ml, respectively. Overall, 94% of S. pneumoniae isolates were susceptible to amoxicillin and amoxicillin-clavulanate, 69% were susceptible to azithromycin and clarithromycin, 63% were susceptible to cefprozil and cefuroxime, 52% were susceptible to cefixime, 22% were susceptible to cefaclor, and 11% were susceptible to loracarbef. Although ciprofloxacin has marginal activity against S. pneumoniae, no high-level fluoroquinolone-resistant strains were found. Significant cross-resistance was found between penicillin and macrolides-azalides among S. pneumoniae isolates, with 5% of the penicillin-susceptible strains being macrolide-azalide resistant, compared with 37% of the intermediate isolates and 66% of the resistant isolates. Resistance was highest in S. pneumoniae isolates from patients younger than 10 years of age, middle ear and paranasal sinus specimens, and the southern half of the United States. With the continuing rise in resistance, judicious use of oral antimicrobial agents is necessary in all age groups.

In vitro development of resistance to five quinolones and amoxicillin-clavulanate in Streptococcus pneumoniae.

The ability of 50 sequential subcultures in subinhibitory concentrations of ciprofloxacin, levofloxacin, grepafloxacin, sparfloxacin, trovafloxacin, and amoxicillin-clavulanate to select for resistance was studied for six penicillin-susceptible and four penicillin-intermediate pneumococci. Subculturing in ciprofloxacin, grepafloxacin, levofloxacin, and sparfloxacin led to selection of mutants requiring increased MICs for all 10 strains, with MICs rising from (i) 0.5 to 4.0 to (ii) 4.0 to 32.0 microgram/ml after 7 to 12 passages for ciprofloxacin, from (i) 0.06 to 0.25 to (ii) 0.5 to 8.0 microgram/ml after 5 to 23 passages for grepafloxacin, from (i) 0.5 to 1.0 to (ii) 4.0 to 64 microgram/ml after 14 to 49 passages for levofloxacin, and from (i) 0.125 to 0.25 to (ii) 1.0 to 16.0 microgram/ml after 8 to 26 passages for sparfloxacin. Subculturing in trovafloxacin led to increased MICs for eight strains, with MICs rising from (i) 0.06 to 0.125 to (ii) 0.5 to 8.0 microgram/ml after 6 to 28 passages. Subculturing in amoxicillin-clavulanate led to raised MICs for only one strain, with the MIC rising from 0.015 to 0. 125 microgram/ml after 24 passages. Double mutations in both ParC and GyrA led to high-level quinolone resistance when ParC mutations were at S79. Trovafloxacin MICs were 1 to 2 microgram/ml in double mutants with ParC mutations at positions other than S79 (e.g., D83). Mutations in ParE (at D435, R447, and E474) and GyrB (at S405, D406, and D435) were found in four and six mutants, respectively. In the presence of reserpine, 29 mutants had lower ciprofloxacin MICs (2 to 16 times lower), 8 mutants had lower levofloxacin MICs (2 times), and one mutant had a lower trovafloxacin MIC (2 times), suggesting the involvement of an efflux mechanism. In contrast to the case for quinolones, subculturing in the presence of amoxicillin-clavulanate did not select for resistance to this drug.

In vitro selection of resistance to four beta-lactams and azithromycin in Streptococcus pneumoniae.

Selection of resistance to amoxicillin (with or without clavulanate), cefaclor, cefuroxime, and azithromycin among six penicillin G- and azithromycin-susceptible pneumococcal strains and among four strains with intermediate penicillin sensitivities (azithromycin MICs, 0.125 to 4 microg/ml) was studied by performing 50 sequential subcultures in medium with sub-MICs of these antimicrobial agents. For only one of the six penicillin-susceptible strains did subculturing in medium with amoxicillin (with or without clavulanate) lead to an increased MIC, with the MIC rising from 0.008 to 0.125 microg/ml. Five of the six penicillin-susceptible strains showed increased azithromycin MICs (0.5 to >256.0 microg/ml) after 17 to 45 subcultures. Subculturing in medium with cefaclor did not affect the cefaclor MICs of three strains but and led to increased cefaclor MICs (from 0.5 to 2.0 to 4.0 microg/ml) for three of the six strains, with MICs of other beta-lactams rising 1 to 3 twofold dilutions. Subculturing in cefuroxime led to increased cefuroxime MICs (from 0.03 to 0.06 microg/ml to 0.125 to 0.5 microg/ml) for all six strains without significantly altering the MICs of other beta-lactams, except for one strain, which developed an increased cefaclor MIC. Subculturing in azithromycin did not affect beta-lactam MICs. Subculturing of the four strains with decreased penicillin susceptibility in amoxicillin (with or without clavulanate) or cefuroxime did not select for beta-lactam resistance. Subculturing of one strain in cefaclor led to an increase in MIC from 0.5 to 2.0 microg/ml after 19 passages. In contrast to strains that were initially azithromycin susceptible, which required >10 subcultures for resistance selection, three of four strains with azithromycin MICs of 0.125 to 4.0 microg/ml showed increased MICs after 7 to 13 passages, with the MICs increasing to 16 to 32 microg/ml. All azithromycin-resistant strains were clarithromycin resistant. With the exception of strains that contained mefE at the onset, no strains that developed resistance to azithromycin contained ermB or mefE, genes that have been found in macrolide-resistant pneumococci obtained from clinic patients.