In vitro activities of 12 orally administered antimicrobial agents against four species of bacterial respiratory pathogens from U.S. Medical Centers in 1992 and 1993.

Clinical isolates of Haemophilus influenzae, Streptococcus pneumoniae, Streptococcus pyogenes, and Moraxella catarrhalis were gathered from 19 different clinical laboratories throughout the continental United States. The in vitro activities of 12 orally administered antimicrobial agents were compared by broth microdilution tests with 3,151 bacterial isolates. Among 890 H. influenzae isolates, 30% were capable of producing beta-lactamase enzymes (12 to 41% in different medical centers). Most of the 619 beta-lactamase-negative H. influenzae strains were susceptible to ampicillicin (MIC, < or = 1.0 micrograms/ml): 5 strains were intermediate in susceptibility (MIC, 2.0 micrograms/ml) and 1 strain was ampilicillin resistant (MIC, 4.0 micrograms/ml). Ninety-two percent of 698 M. catarrhalis strains were beta-lactamase positive. Of 799 S. pneumoniae isolates, 15% were intermediate in susceptibility to penicillin and 7% were resistant to penicillin. The prevalence of penicillin-susceptible pneumococci in different institutions ranged from 63 to 95%. Only 1% of 764 S. pyogenes isolates were resistant to the macrolides, but 5% of S. pneumoniae isolates were macrolide resistant. Only 71% of 58 penicillin-resistant S. pneumoniae isolates were erythromycin susceptible, whereas 97% of the 622 penicillin-susceptible strains were erythromycin susceptible. Penicillin-resistant pneumococci were also relatively resistant to the cephalosporins and amoxicillin. Penicillin-susceptible pneumococci were susceptible to amoxicillin-clavulanic acid (MIC for 90% of isolates tested [MIC90], < or = 0.12/0.06 microgram/ml), cefixime (MIC90, 0.25 microgram/ml), cefuroxime axetil (MIC90, < or = 0.5 microgram/ml), cefprozil (MIC90, < or = 0.5 micrograms/ml), cefaclor (MIC90, 0.5 microgram/ml), and loracarbef (MIC90, 1.0 microgram/ml). Most strains of the other species remained susceptible to the study drugs other than amoxicillin.

Performance of Haemophilus Test Media prepared with 12 different lots of Mueller-Hinton agar from four manufacturers.

Haemophilus Test Media (HTM) were prepared from 12 different lots of Mueller-Hinton agar. When tested with Haemophilus influenzae ATCC 49247, most lots were initially rejected because of small zones of inhibition for cefaclor, cefuroxime, and cefamandole disks, whereas five other drugs performed satisfactorily on the 11 lots that supported growth of the control strain. At the same time, tests of Escherichia coli ATCC 25922 and Staphylococcus aureus ATCC 25923 documented the acceptability of these agar media, with or without HTM supplements. The current control limits for cefaclor, cefuroxime, and cefamandole appear to be unrealistic. Because the beta-lactamase-negative, ampicillin-resistant control strain of H. influenzae (ATCC 49247) has altered penicillin-binding proteins, it is resistant to ampicillin and is probably also resistant to cefaclor, cefuroxime, and cefamandole. Consequently, media that produce no or very small zones of inhibition with those three drugs might be clinically correct and should not be rejected. One manufacturer provided three lots of Mueller-Hinton agar that gave unusually large zones of inhibition with all beta-lactams. The three other manufacturers provided eight Mueller-Hinton agars that were satisfactory for the preparation of HTM agar, provided that small zones of inhibition with cefaclor, cefuroxime, and cefamandole disks are accepted as the preferred result.

The prevalence of staphylococcal resistance to penicillinase-resistant penicillins. A retrospective and prospective national surveillance trial of isolates from 40 medical centers.

In a retrospective survey of resistance to penicillinase-resistant penicillins (PRPs) in 152,076 clinical staphylococcal strains isolated in 40 United States Hospitals in 1985 and 1986, rates of resistance to oxacillin were found to be 11 and 13%, respectively, among Staphylococcus aureus isolates. The rates were approximately four times higher among coagulase-negative staphylococcal strains. In a prospective study of 1,408 wound or bacteremia isolates from the participant hospitals, oxacillin and methicillin agar screening, disk diffusion, and broth microdilution testing were conducted at a single reference laboratory. These tests yielded PRP resistance rates of 15% among S. aureus, 75% among S. epidermidis, and 48% among other coagulase-negative strains. No major changes in the distribution of resistance rates among hospitals or by hospital type were observed. Dilution susceptibility testing of several antimicrobial agents against PRP-resistant isolates and species-matched susceptible isolates from the same hospital showed that teicoplanin and vancomycin were the most active drugs (100% of S. aureus isolates were susceptible). Teicoplanin and vancomycin disk diffusion testing of all PRP-resistant staphylococcal strains also showed that these isolates were susceptible to the glycopeptides. However, agar dilution screening and broth microdilution tests revealed that several coagulase-negative strains, predominantly S. haemolyticus, had teicoplanin MICs greater than or equal to 8 micrograms/ml. S. haemolyticus isolates represented a very small number of the total stains tested. Teicoplanin and vancomycin were also the most active drugs when tested against older (1962-82) clinical PRP-resistant S. aureus strains from the reference laboratory collection. The methods found to be superior in detecting PRP-resistant strains were the oxacillin 6 micrograms/ml agar screening test in 4% NaCl-supplemented Mueller-Hinton agar and the 1 microgram oxacillin disk test. By reference laboratory standards, participant laboratories were incorrect in only 2.3% of species identifications and 4.5% of oxacillin-susceptibility determinations, indicating acceptable contemporary agreement and accuracy.

In vitro antimicrobial spectrum, occurrence of synergy, and recommendations for dilution susceptibility testing concentrations of the cefoperazone-sulbactam combination.

Broth microdilution tests and an antimicrobial interaction (synergy) studies using various combinations of cefoperazone and sulbactam were performed in an effort to determine the most appropriate in vitro dilution test system. The test results with cefoperazone and sulbactam were categorized as synergistic (complete or partial) for nearly 80% of the strains isolated from clinical trial patients. The results indicate that the cefoperazone-sulbactam fixed ratio (2:1) maximized the cefoperazone spectrum of activity and best approximated the parenteral formulation of the drug. The cefoperazone-sulbactam combination had a greater antimicrobial activity than did the other comparison beta-lactams, except for imipenem, tested against strains of the family Enterobacteriaceae. To be consistent with the National Committee for Clinical Laboratory Standards interpretive breakpoints for cefoperazone alone, the following MIC breakpoints should be applied to the combination (2:1 ratio): less than or equal to 16/8 micrograms/ml, susceptible; 32/16 micrograms/ml, moderately susceptible; and greater than or equal to 64/32 micrograms/ml, resistant.

Resistance to ten different fluoroquinolone antibiotics following in vitro exposures to nalidixic acid.

Ten different fluoroquinolone antibiotics were tested against selected Enterobacteriaceae. Against nalidixic acid-resistant clinical isolates, minimal inhibitory concentrations for all 10 drugs were eightfold to 32-fold greater than those against comparable nalidixic acid-susceptible strains. When the latter strains were serially exposed to increasing concentrations of nalidixic acid, susceptibility to all 10 fluoroquinolones decreased as resistance to nalidixic acid and cinoxacin was acquired in vitro.

Bactericidal activities of ten different fluoroquinolones against selected Enterobacteriaceae.

Ten fluoroquinolone compounds and two other 4-quinolones were tested in a microdilution system. Minimal inhibitory concentrations and minimal bactericidal concentrations were determined with 1 X 10(7) and 5 X 10(5) colony forming units per milliliter. All 12 compounds were bactericidal. Continued inhibition by drug carried over in the subcultured samples was documented and that represents a potential source of technical error that needs to be controlled when determining minimal bactericidal concentrations for these extremely potent drugs.

Evaluation of a proprietary broth medium for microdilution susceptibility testing of nutritionally fastidious bacteria.

For microdilution susceptibility tests with nutritionally fastidious microorganisms, a new clear broth medium developed at Micro-Media Systems, Inc., Potomac, Md., was evaluated in a three-laboratory collaborative study. Replicate tests were performed with 80 isolates (51 Streptococcus spp., 27 Haemophilus influenzae isolates, and 2 Neisseria meningitidis isolates) against 15 antimicrobial agents. In standard 100-microliters volumes, results of tests in the new broth medium were comparable to those in the reference medium (Mueller-Hinton broth with 2 to 3% lysed horse blood), but MICs were somewhat easier to read in the new broth medium. Results of similar tests in smaller panels, containing 40 microliters in each well, were less satisfactory; i.e., growth failures and poorly defined endpoints were more commonly encountered. With drugs other than erythromycin or clindamycin, the 40-microliters panels provided MICs which compared favorably with those obtained by standard reference methods.

The anti-microbial activity, beta-lactamase stability, and disk diffusion susceptibility testing of carumonam (RO 17-2301, AMA-1080), a new monobactam.

Carumonam, a new monobactam, was found to have an anti-microbial spectrum similar to aztreonam. Its spectrum includes Enterobacteriaceae, Haemophilus influenzae, pathogenic Neisseria species, Pseudomonas aeruginosa, and some streptococci. Staphylococcus species, enterococci, and many other nonenteric gram-negative bacilli were not inhibited. Enterobacteriaceae resistant to cefoperazone (minimum inhibitory concentrations [MICs] greater than or equal to 32 mg/L) were more likely inhibited by carumonam (52% at less than or equal to 8.0 mg/L) than aztreonam (39%) or ceftazidime (35%). Dilution test methods on agar or in Mueller-Hinton broth produced similar results. Carumonam minimum bactericidal concentrations were usually the same or one dilution above the MIC. Carumonam and aztreonam were very stable to most chromosomal (P99, K1, K14) and plasmid-mediated beta-lactamases (TEM, OXA, PSE). The Klebsiella oxytoca enzymes hydrolyzed aztreonam at rates greater than or equal to fivefold higher than carumonam but at a rate less than 1% that of cephaloridine. The aztreonam MICs for these Klebsiella stains were greater than or equal to 32 mg/L, but the hydrolysis rates do not fully explain the high-grade resistance to aztreonam. In vitro susceptibility tests with 30-micrograms carumonam disks were found to be very predictive. Similar regression statistics were observed for aztreonam and cefotaxime. Recommendations for carumonam susceptibility testing are susceptible greater than or equal to 21 mm (less than or equal to 8.0 mg/L) and resistant less than or equal to 14 mm (greater than or equal to 32 mg/L). Cross-resistance analysis favors the independent testing of carumonam or aztreonam against gram-negative species other than Enterobacteriaceae and P. aeruginosa.

Antistaphylococcal activity of ceforanide and cefonicid in the presence of human serum.

Tests with 52 strains of Staphylococcus aureus compared ceforanide and cefonicid. Addition of 50% human serum to the test system reduced the bacteriostatic and bactericidal activities of cefonicid, but ceforanide was not affected as greatly.

Cefoperazone and cefoperazone-sulbactam susceptibility tests with anaerobic bacteria by the thioglycolate disk elution method.

Tests were performed with 104 anaerobic microorganisms to evaluate the thioglycolate disk elution technique for the detection of resistance to cefoperazone and cefoperazone-sulbactam. An unacceptably high false-resistance rate and a poor reproducibility record make the disk elution procedure unsatisfactory for routine testing of this drug or combination of drugs.

BMY28142, cefbuperazone (T-1982), and Sch 34343. Antimicrobial activity against 94 anaerobes compared to seven other antimicrobial agents.

Three new beta-lactams were evaluated against 94 anaerobic strains representing 15 species using a Wilkins-Chalgren broth microdilution method. The penems, Sch 29482 and Sch 34343, were most active with all minimum inhibitory concentrations (MICs) at less than or equal to 4.0 micrograms/ml and MIC90s of less than or equal to 0.25 micrograms/ml. BMY 28142 had a more limited antianaerobic activity against Bacteroides fragilis with a MIC50 and MIC90 of 32 and 128 micrograms/ml, respectively. Cefbuperazone (T-1982) had low B. fragilis MICs (MIC90, 8.0 micrograms/ml), but potentially resistant range MIC90 results for the other species in the B. fragilis group and Clostridium species.

Feasibility study of disk diffusion susceptibility tests with Mueller-Hinton broth solidified with Gelrite, an agar substitute.

Feasibility studies were done to determine whether a new agar substitute, Gelrite gellan gum, could be used to prepare a solid Mueller-Hinton medium for disk diffusion susceptibility tests. Mueller-Hinton broth was combined with 0.43% of the gellan gum and 0.75% KCI. The resulting medium had performance characteristics similar to those of Mueller-Hinton agars; however, zones on the gellan gum media tended to be a little larger. Significant differences among Mueller-Hinton broths and among Mueller-Hinton agars from different manufacturers were documented: zones on different lots of the gellan gum were more consistent. The Mueller-Hinton broth-gellan gum medium appears to represent a satisfactory alternative to agar media. However, because somewhat larger zones were seen on the gellan gum plates, further study will be needed to develop quality control limits and interpretive zone size standards for tests on this new medium.

Determination of susceptibility of anaerobic bacteria to cefotetan and cefoxitin by the thioglycolate disk elution method.

The in vitro activities of two cephamycins, cefotetan and cefoxitin, against 107 anaerobic bacteria were evaluated. The aerobically incubated thioglycolate disk elution technique of Kurzynski et al. (Antimicrob. Agents Chemother. 10:727-732, 1976) was also evaluated to establish the appropriate number of 30-micrograms disks to be added to each tube, assuming that strains with MICs of less than or equal to 16 micrograms/ml are susceptible. Optimal predictive values were obtained when two tubes were prepared for each antimicrobial agent, one with two 30-micrograms disks and the other with three 30-micrograms disks. After 48 h of incubation, resistant strains grew in both tubes and susceptible strains provided no growth or growth less than or equal to 50% of that in the control broth. Growth in one tube but not in the other was considered an equivocal test result indicating the need for additional tests; 5 to 7% of our strains gave equivocal results. Reproducibility studies confirmed that broth microdilution tests were more reproducible than disk elution susceptibility tests.

Unpredictable interactions between cefotetan and penicillins or aminoglycosides.

The bacteriostatic and bactericidal activity of gentamicin, amikacin, piperacillin, azlocillin, ampicillin, benzyl penicillin, and oxacillin against 85 selected isolates was measured alone and in the presence of subinhibitory concentrations of cefotetan. A potential for antagonistic interactions between cefotetan and acylamino penicillins (piperacillin greater than azlocillin greater than ampicillin) were observed with some strains; synergy was suggested with other isolates, however, Bacteriostatic and bactericidal activity of the two aminoglycosides was enhanced by cefotetan with most gram-negative bacilli, but bactericidal activity was significantly diminished with 4 of 65 strains. Piperacillin and cefotetan were clearly antagonistic with 11 of 44 strains and synergistic with 4 of 44 strains of gram-negative bacilli: the remaining 21 strains gave off scale end points and could not be analyzed in that way.

The activity of cefotaxime and desacetylcefotaxime alone and in combination against anaerobes and staphylococci.

Cefotaxime (CTX) and desacetylcefotaxime (des-CTX) alone and in combination were tested against anaerobic bacteria collected from clinical infections from several geographically diverse medical centers. The CTX minimum inhibitory concentration (MIC) inhibiting 50% of tested Bacteroides fragilis strains was in the moderately susceptible range (32 micrograms/ml), but when placed in combination with des-CTX it had a potency compatible to cefoxitin (MIC50, 8.0 micrograms/ml). Other B. fragilis group species (B. distasonis and B. vulgatus) were also susceptible to CTX and des-CTX alone at the MIC50 level. MIC90 statistics for CTX, cefoxitin, and ticarcillin were generally in the resistant range. Synergy studies showed that 80% of tested anaerobes were synergistically killed by the combination of CTX and des-CTX. Most of these strains had their synergy occur at drug levels that could be achieved in vivo. A large number of the B. thetaiotaomicron strains must be considered resistant to the combination because of the very high levels of des-CTX required to produce synergistic killing. Other drugs routinely used for anaerobic infections (clindamycin, chloramphenicol, and metronidazole) also had elevated B. thetaiotaomicron MICs. Endemic difference in susceptibility to the beta-lactam drugs were observed, especially the CTX-des-CTX combination. The combination and other beta-lactams were most usable for strains isolated from the Portland metropolitan area, were moderately active against those from Cleveland, and were rarely usable on Bacteroides isolates at Northwestern in Chicago. Laboratories are urged to monitor the cephalosporin and semisynthetic penicillin in vitro efficacy and not rely on published statistics. Staphylococcus aureus strains were susceptible to CTX alone, but were even more susceptible (two- to fourfold reduction in MICs) when used in combination with des-CTX. These data show CTX to be the most active antistaphylococcal compound among the new cephalosporins and to be comparable to cefamandole and cefuroxime, but superior to the anaerobe-active cefoxitin.

Cefotaxime, cephalothin, and cephapirin: antimicrobial activity and synergy studies of cephalosporins with significant in vivo desacetyl metabolite concentrations.

The desacetyl metabolites of cefotaxime, cephalothin, and cephapirin were 5-55% as active as the parent drug, depending upon the bacterial species tested. Synergy or partial synergy was demonstrated in 64% of 25 strains of Enterobacteriaceae and Staphylococcus aureus tested against cephalothin/desacetylcephalothin and cephapirin/desacetylcephapirin combinations. Some species variations were identified that influenced synergy rates, particularly among the S. aureus strains (for example, highest rates for cephapirin).

Antimicrobial activity of amikacin combinations against Enterobacteriaceae moderately susceptible to third-generation cephalosporins.

Enterobacteriaceae strains having elevated minimal inhibitory concentrations (greater than or equal to 2.0 to less than or equal to 32 micrograms/ml) of cefoperazone, cefotaxime, ceftazidime, and moxalactam were synergistically inhibited by amikacin combinations (54.1 to 69.6% occurrence). Indifference was rare (8.1% for moxalactam), and true antagonistic interactions were not observed. Strains resistant or susceptible to these new cephalosporins were also synergistically inhibited by the addition of amikacin, reducing resistant cephalosporin minimal inhibitory concentrations to clinically achievable levels.

Antibacterial activity of fortimicin A compared with those of five other aminoglycosides, and factors affecting susceptibility tests.

Fortimicin A, a pseudodisaccharide aminoglycoside, was found to have broad-spectrum activity against most clinically important aerobic and facultatively anaerobic bacteria, except Pseudomonas aeruginosa, some other Pseudomonas species, and streptococci. It was comparable to amikacin in its level of activity (minimum inhibitory concentrations) and spectrum of activity (except for the lack of activity on P. aeruginosa). Fortimicin A was bactericidal and was affected by cations when tested against P. aeruginosa. Minimum inhibitory concentrations were affected by the inoculum used in the susceptibility test. The drug was resistant to most aminoglycoside-inactivating enzymes, but probably is not active against permeability mutants.