Marlene Spohr

In vitro Antibiotic Susceptibility of Legionellaceae: Search for Alternative Antimicrobial Drugs

Reference strains of Legionella pneumophila (serotypes 1-4, and 6), Legionella micdadei , and Legionella bozemanii proved susceptible (agar dilution test, buffered charcoal yeast extract agar) to erythromycin, rifampin, augmentin (amoxycillin plus clavulanic acid), cefotaxime, cefoxitin, enoxacin , fusidic acid, and norfloxacin; cefamandole was less active. The strains varied in susceptibility to fosfomycin.

Pseudomonas aeruginosa: in vitro susceptibility to antimicrobial drugs, single and combined, with and without defibrinated human blood.

Twelve clinical isolates of Pseudomonas aeruginosa of distinct pyocin type varied in susceptibility to 14 of 17 antimicrobial drugs. The 2 x MIC concentrations of 16 antimicrobial drugs combined with 55% (v/v) of fresh, defibrinated human blood yielded additive effects. Additive effects were noted with blood plus the MIC concentrations of all drugs tested except cefoperazone, gentamicin, and netilmicin. Blood combined with subinhibitory (1/2 MIC) concentrations of aztreonam, ceftazidime, ciprofloxacin, fleroxacin, imipenem, and tobramycin, respectively, yielded additive effects; indifferent effects were observed with the remaining 10 blood plus 1/2 MIC drug combinations. The following drug combinations additively augmented the antibacterial activity of 65% (v/v) of human blood against two selected isolates of P. aeruginosa: tobramycin (1 microgram/ml) plus the MIC or 2 x MIC concentrations of azlocillin, aztreonam, ceftazidime, ciprofloxacin, imipenem, norfloxacin, ofloxacin, piperacillin, and ticarcillin, respectively. Imipenem (8 micrograms/ml) combined with ceftazidime, cefoperazone, and piperacillin, but not aztreonam, enhanced the bactericidal activity of human blood. Rifampin (2 micrograms/ml) plus tobramycin (0.5-1 microgram/ml) combined with 8 or 16 micrograms/ml of azlocillin, aztreonam, cefoperazone, ceftazidime, imipenem, and piperacillin, respectively, enhanced blood-mediated killing of three representative multiple-drug-resistant P. aeruginosa isolates. Additional effective triple-drug combinations with human blood were rifampin + tobramycin + polymyxin B, rifampin + ciprofloxacin + imipenem, and rifampin + amikacin + imipenem. Ciprofloxacin (2 micrograms/ml) was the most potent intraphagocytic bactericidal drug of 16 tested agents (greater than or equal to 2 x MBC concentrations) against P. aeruginosa control strain ATCC 27853.

Active immunization of NMRI mice against serratia marcescens

Phenol-hot water lipopolysaccharide (LPS) extracts of Serratia marcescens strains CDC O3:H1, CDC O6:H3, NEW CDC O14:H12, and SH 186 (serotype O6/O14:H12) significantly protected NMRI mice against intraperitoneal challenge with the more mouse virulent homologous strains; overall, there was moderate cross-protection against the minority of heterologous challenge strains. Trichloroacetic acid LPS extracts and K-antigen extracts of strains NEW CDC O14:H12 and SH 186 also proved protective antigens. The purified metalloproteases of strains SH 186 and SF 178 (serotype O6/O14:H12) effected active murine immunization. Neither active nor passive immunization of NMRI mice with E. coli Rc mutant J5 afforded significant protection against various challenge strains of S. marcescens.

Fosfomycin: Interpretation of Inhibition Zones Obtained with the Bauer-Kirby Agar Disk Diffusion Susceptibility Test

A total of 430 strains representing diverse gram-positive and gram-negative bacterial species were examined for susceptibility to fosfomycin utilizing the Bauer-Kirby agar disk diffusion and the Steers’ agar dilution methods. Tentative criteria are proposed for the interpretation of the diameters of resultant inhibition zones (50 μg fosfomycin disks plus 50 μg glucose-6-phosphate) on Mueller-Hinton agar, based on MIC breakpoints of d 16 μg/ml = sensitive (S), 32–64 μg/ml = intermediate susceptible (I), and ≥ 128 μg/ml = resistant (R). For staphylococci: ≥ 20 mm = S; 14–19 mm = I; ≤ 13 mm = R; for Enterobacteriaceae and Pseudomonas aeruginosa: ≥ 16 mm = S; 11-15 mm = I; ≤ 10mm = R.

Comparative disk and broth dilution susceptibility test results with ticarcillin and timentin against Pseudomonas aeruginosa and Pseudomonas maltophilia.

A total of 500 clinical isolates of Pseudomonas aeruginosa and 14 isolates of Pseudomonas maltophilia were examined for susceptibility to ticarcillin and timentin (ticarcillin plus clavulanic acid). Clavulanic acid enhanced the activity of ticarcillin against only 34 of 500 isolates of P. aeruginosa. Among 23 isolates of P. aeruginosa, that had yielded divergent disk diffusion results, there were 3 major discrepancies (3 = ticarcillin-resistant by disk diffusion, but susceptible by broth dilution test criteria) and 1 very major discrepancy (timentin-susceptible by disk diffusion, but resistant by broth dilution test criteria). Clavulanic acid enhanced the activity of ticarcillin against 12 of 14 isolates of P. maltophilia; there were 2 major discrepancies (ticarcillin resistance in terms of disk diffusion, but susceptibility according to ticarcillin MICs). Consequently, it was decided to continue employing timentin disks alone, rather than add ticarcillin disks in our Bauer-Kirby test battery for Pseudomonadaceae.

Streptococcus faecalis: in vitro susceptibility to antimicrobial drugs, single and combined, with and without defibrinated human blood.

Ampicillin, fusidic acid, gentamicin, imipenem, mezlocillin, ofloxacin, penicillin G, piperacillin, and vancomycin were examined for inhibitory and bactericidal activity in various broth media against 7 clinical isolates of Streptococcus faecalis. On a weight-for-weight basis, ampicillin, imipenem, mezlocillin, and ofloxacin proved to be more efficacious. All enterococcal isolates were resistant against gentamicin; fusidic acid and vancomycin lacked bactericidal activity. The combinations of either ampicillin, imipenem, mezlocillin, ofloxacin, piperacillin, or vancomycin with a subinhibitory concentration (4 micrograms/ml) of gentamicin, with or without added 65% (v/v) fresh defibrinated human blood, respectively, yielded additive effects against all enterococcal isolates. The addition of fresh human blood failed to enhance the antienterococcal activity of 4 micrograms/ml of gentamicin; in contrast, addition of 65% (v/v) fresh or heat-inactivated (56 degrees C, 30 min) normal rabbit, bovine, and human sera augmented the activity of gentamicin, an effect that was ablated through the addition of either 0.005 M DTT or 0.01 M MgCl2 + 0.01 M EGTA + 0.01 M CaCl2, supplements known to antagonize human serum beta-lysin, but not lysozyme activity.

Susceptibility of Clostridium perfringens Type A to 23 Antimicrobial Drugs

A total of 106 strains of Clostridium perfringens type A, that had been isolated very recently from faecal cultures of healthy adults, were examined for susceptibility to 23 antimicrobial drugs (agar dilution method). All strains were susceptible to ampicillin, cefazolin, cefotaxime, cefotetan, cefoxitin, ceftriaxone, ciprofloxacin, fusidic acid, imipenem, metronidazole, mezlocillin, ofloxacin, penicillin G, piperacillin, teicoplanin, and vancomycin. Teicoplanin and imipenem were the most active, whereas fosfomycin and tetracycline were the least active drugs. Three of the 106 strains revealed multiple drug resistance against clindamycin, erythromycin, josamycin, tetracycline, and, in one instance, against chloramphenicol.

Gentamicin- and Methicillin-Resistant, Clinical Isolates of Staphylococcus aureus: Comparative in vitro and in vivo Efficacy of Alternative Antimicrobial Drugs

Six representative clinical isolates of gentamicin- and methicillin-resistant (GRMR) Staphylococcus aureus, constituting phage groups II and III, were susceptible only to amikacin, cefamandole, clindamycin, fosfomycin, fusidic acid, netilmicin, nitrofurantoin, trimethoprim-sulfamethoxazole, and vancomycin (Bauer-Kirby test). With few exceptions, the minimal bactericidal concentrations of the beta-lactam and aminoglycoside antibiotics tested, fusidic acid, and irregularly those of vancomycin, but not those of fosfomycin and rifampin, exceeded minimal inhibitory concentrations values by at least 8-fold. In vitro, combinations of rifampin with cefamandole, cefazolin, cefotaxime, erythromycin, fosfomycin, fusidic acid, netilmicin, and vancomycin yielded indifferent effects. The GRMR S. aureus strains were refractory against 50, 65, and 80 vol% of fresh defibrinated blood; following reduction of viable counts at 2 h (greater than or equal to 90%), rebound growth invariably occurred within 4 h after exposure. Combined human blood (55 vol%)-antibiotic assays revealed rifampin as the most effective drug, followed by vancomycin, fusidic acid, and cefamandole, in that order. Blood plus cefotaxime, cefazolin, or netilmicin yielded indifferent effects; fosfomycin failed in vitro. In terms of speed of recovery and survival data (chi 2 test), cyclophosphamide-pretreated, i.e., leukopenic NMRI mice responded best to chemotherapy with rifampin, followed by vancomycin, cefamandole, netilmicin, and fosfomycin, in that order; cefazolin yielded variable results.

In vitro Activity of Three Fluoroquinolones against Enterococci and Pseudomonas aeruginosa: Modified Categorization System for Discrepant Broth Dilution and Agar Disk Diffusion Test Results

The fluoroquinolones ciprofloxacin, norfloxacin, and ofloxacin were examined in vitro against 103 enterococcal isolates and 138 isolates of Pseudomonas aeruginosa, in an attempt to determine which respective proposed interpretive criteria (minimal inhibitory concentration (MIC) versus diameter of agar disk diffusion inhibition zones) correlated best, i.e., resulted in fewer discrepant results. A previously proposed system for grouping discrepant results (very major, major, and minor discrepancies) of Barry and co-workers was modified to comprise 6 categories (very major, major, minor, slight, minimal, and negligible); this expanded system rendered the encountered discrepant in vitro test results more transparent. Overall, the currently employed interpretive criteria for norfloxacin (MICs versus 10-micrograms disks) resulted in fewer discrepancies with the above two groups of bacterial isolates than those proposed for ciprofloxacin and ofloxacin (MICs versus 5-micrograms disks, respectively).

Gentamicin- and methicillin-resistant Staphylococcus aureus: in vitro susceptibility to antimicrobial drugs

Thirteen nosocomially significant, gentamicin- and methicillin-resistant (GRMR) Staphylococcus aureus isolates, all of phage group III/M (lysotype 42E/47/53/54/75/77/83A/84/85/94/96), were uniformly resistant against augmentin, erythromycin, fosfomycin, gentamicin, methicillin, oxacillin, penicillin G, tetracycline, and tobramycin, but differed in susceptibility to cefamandole, ciprofloxacin, clindamycin, imipenem, josamycin, the synthetic chinolone Ro 23-6240, and ofloxacin. All isolates were susceptible to chloramphenicol, coumermycin, fusidic acid, novobiocin, rifampin, teicoplanin, trimethoprim-sulfamethoxazole (cotrimoxazole), and vancomycin. One isolate was of intermediate susceptibility to netilmicin. On a weight-for-weight basis, the 7 most active drugs were rifampin, coumermycin, cotrimoxazole, novobiocin, teicoplanin, fusidic acid, and vancomycin (in decreasing order) in terms of minimal inhibitory concentrations. With regard to minimal bactericidal concentrations, coumermycin, rifampin, vancomycin, teicoplanin, cotrimoxazole, ofloxacin, and ciprofloxacin (in decreasing order) were the 7 most potent antimicrobial drugs. Freshly defibrinated human blood [65% (v/v)] combined with chloramphenicol and rifampin, respectively, resulted in a weak additive effect (time kill curves). Indifferent effects were observed following combination of blood with ciprofloxacin, cotrimoxazole, coumermycin, fusidic acid, imipenem, netilmicin, novobiocin, ofloxacin, compound Ro 23-6240, teicoplanin, and vancomycin. Rifampin combined with novobiocin, teicoplanin, and vancomycin, respectively, in the presence of 65% (v/v) human blood, resulted in an additive effect. Combinations of rifampin with 9 other antimicrobial drugs in blood yielded essentially indifferent effects.