Laurie B. Hovde

Synergistic Activity of Colistin and Ceftazidime against Multiantibiotic-Resistant Pseudomonas aeruginosa in an In Vitro Pharmacodynamic Model

ABSTRACT Despite the marketing of a series of new antibiotics for antibiotic-resistant gram-positive bacteria, no new agents for multiple-antibiotic-resistant gram-negative infections will be available for quite some time. Clinicians will need to find more effective ways to utilize available agents. Colistin is an older but novel antibiotic that fell into disfavor with clinicians some time ago yet still retains a very favorable antibacterial spectrum, especially for Pseudomonas and Acinetobacter spp. Time-kill curves for two strains of multiantibiotic-resistant Pseudomonas aeruginosa were generated after exposure to colistin alone or in combination with ceftazidime or ciprofloxacin in an in vitro pharmacodynamic model. MICs of colistin, ceftazidime, ciprofloxacin, piperacillin-tazobactam, imipenem, and tobramycin were 0.125, ≥32, >4, >128/4, 16, and >16 mg/liter, respectively. Colistin showed rapid, apparently concentration-dependent bactericidal activity at concentrations between 3 and 200 mg/liter. We were unable to detect increased colistin activity at concentrations above 18 mg/liter due to extremely rapid killing. The combination of colistin and ceftazidime was synergistic (defined as at least a 2-log10 drop in CFU per milliliter from the count obtained with the more active agent) at 24 h. Adding ciprofloxacin to colistin did not enhance antibiotic activity. These data suggest that the antibacterial effect of colistin combined with ceftazidime can be maximized at a peak concentration of ≤18 mg/liter.

Increased Killing of Staphylococci and Streptococci by Daptomycin Compared with Cefazolin and Vancomycin in an In Vitro Peritoneal Dialysate Model

ABSTRACT Peritoneal dialysate fluid (PDF) is a bacteriostatic medium that compromises the antibacterial activity of cell wall-active agents. By use of an in vitro static model, methicillin-resistant Staphylococcus aureus (MRSA), methicillin-susceptible S. aureus (MSSA), methicillin-susceptible Staphylococcus epidermidis (MSSE), and Streptococcus sanguis were exposed to daptomycin at concentrations of 10, 30, and 100 mg/liter, cefazolin at 125 mg/liter, and vancomycin at 25 mg/liter in cation-adjusted Mueller-Hinton Broth or Todd Hewitt Broth (for S. sanguis) and PDF at pHs of 5.5 and 7.4. The pH had no effect on antibacterial activity. Neither cefazolin nor vancomycin produced a bactericidal or a bacteriostatic effect versus MRSA, MSSA, MSSE, or S. sanguis in PDF, while all concentrations of daptomycin were bactericidal against all organisms in PDF. Daptomycin did not exhibit concentration-dependent activity in PDF. Daptomycin appears to be a promising agent for use in peritoneal dialysis-associated peritonitis, producing bacterial kill to a greater extent and at a higher rate than cefazolin or vancomycin in PDF.

Levofloxacin plus Metronidazole Administered Once Daily versus Moxifloxacin Monotherapy against a Mixed Infection of Escherichia coli and Bacteroides fragilis in an In Vitro Pharmacodynamic Model

ABSTRACT Moxifloxacin has been suggested as an option for monotherapy of intra-abdominal infections. Recent data support the use of a once-daily metronidazole regimen. The purpose of this study was to investigate the activity of levofloxacin (750 mg every 24 h [q24h]) plus metronidazole (1,500 mg q24h) compared with that of moxifloxacin (400 mg q24h) monotherapy in a mixed-infection model. By using an in vitro pharmacodynamic model in duplicate, Escherichia coli and Bacteroides fragilis were exposed to peak concentrations of 8.5 mg of levofloxacin/liter q24h, 32 mg of metronidazole/liter q24h, and 2 mg for moxifloxacin/liter q24h for 24 h. The activities of levofloxacin, metronidazole, moxifloxacin, and levofloxacin plus metronidazole were evaluated against E. coli, B. fragilis, and E. coli plus B. fragilis. The targeted half-lives of levofloxacin, metronidazole, and moxifloxacin were 8, 8, and 12 h, respectively. Time-kill curves were analyzed for time to 3-log killing, slope, and regrowth. Pre- and postexposure MICs were determined. The preexposure levofloxacin, metronidazole, and moxifloxacin MICs for E. coli and B. fragilis were 0.5 and 1, >64 and 0.5, and 1 and 0.25 mg/liter, respectively. Levofloxacin and moxifloxacin achieved a 3-log killing against E. coli and B. fragilis in all experiments, as did metronidazole against B. fragilis. Metronidazole did not decrease the starting inoculum of E. coli. The area under the concentration-time curve/MIC ratios for E. coli and B. fragilis were 171.7 and 85.9, respectively, for levofloxacin and 26 and 103.9, respectively, for moxifloxacin. Levofloxacin plus metronidazole exhibited the fastest rates of killing. The levofloxacin and moxifloxacin MICs for B. fragilis increased 8- to 16-fold after the organism was exposed to moxifloxacin. No other changes in the postexposure MICs were found. Levofloxacin plus metronidazole administered once daily exhibited activity similar to that of moxifloxacin against the mixed E. coli and B. fragilis infection. A once-daily regimen of levofloxacin plus metronidazole looks promising for the treatment of intra-abdominal infections.

Colistin Methanesulfonate against Multidrug-Resistant Acinetobacter baumannii in an In Vitro Pharmacodynamic Model

ABSTRACT Using an in vitro pharmacodynamic model, a multidrug-resistant strain of Acinetobacter baumannii was exposed to colistin methanesulfonate alone and in combination with ceftazidime. Pre- and postexposure colistin sulfate MICs were determined. A single daily dose of colistin methanesulfonate combined with continuous-infusion ceftazidime prevented regrowth and postexposure MIC increases.

Mutant Prevention Concentrations of ABT-492, Levofloxacin, Moxifloxacin, and Gatifloxacin against Three Common Respiratory Pathogens

ABSTRACT The purpose of this study was to compare the mutant prevention concentration (MPC) of ABT-492 to those of levofloxacin, moxifloxacin, and gatifloxacin against Streptococcus pneumoniae, Haemophilus influenzae, and Moraxella catarrhalis. The fluoroquinolones had comparable mutation selection windows, which is the ratio of MPC/MIC, for all isolates.

Comparative Pharmacodynamics of Three Newer Fluoroquinolones versus Six Strains of Staphylococci in an In Vitro Model under Aerobic and Anaerobic Conditions

ABSTRACT Six strains of staphylococci were exposed to levofloxacin, moxifloxacin, or trovafloxacin in an in vitro pharmacodynamic model under both aerobic and anaerobic conditions. Each agent demonstrated a rapid 3-log10 kill versus susceptible isolates regardless of condition. Against clinical isolates with reduced susceptibility, regrowth occurred by 24 h and was frequently associated with further increases in MICs.

Enhanced detection of hydrogen sulfide generated in cell culture using an agar trap method

Lack of reliable methods to accurately measure hydrogen sulfide (H(2)S) produced in vitro has impeded research on the physiology of this gaseous mediator. Current in vitro methods involve measurement of H(2)S in cell culture media following incubation with H(2)S-releasing compounds. However, this method is inaccurate because H(2)S gas has a short life and thus evades detection. To overcome this, we have adapted a method that employs a modified agar layer to instantly trap H(2)S, allowing measurement of H(2)S accumulated with time. The amount of H(2)S trapped in the agar is quantified using an in situ methylene blue assay. We were able to detect H(2)S produced from sodium hydrogen sulfide (NaHS) added at concentrations as low as 10 μM. Following a 24-h incubation of endothelial-like or vascular smooth muscle cells with 50 μM NaHS, we were able to recover twice more H(2)S than conventional methods. When H(2)S-releasing compounds L-cysteine and N-acetylcysteine were added to the cell culture, the amount of H(2)S increased in a concentration-, time-, and cell line-dependent manner. In conclusion, we have developed an improved method to quantify H(2)S generated in vitro. This method could be used to screen compounds to identify potential H(2)S donors and inhibitors for therapeutic use.

Microbiologic effectiveness of time- or concentration-based dosing strategies in Streptococcus pneumoniae

This in vitro study evaluated the pharmacodynamic performance of levofloxacin using different dosing strategies against both a levofloxacin-sensitive (MIC = 1 mg/liter) and -resistant (MIC = 16 mg/liter) strain of Streptococcus pneumoniae. The strain was genotypically characterized by a mutation in gyrA and two mutations in parE; resistance was shown not to be efflux-mediated. The purpose of this study was to determine if simulated levofloxacin dosing strategies focused either on time or concentration would affect microbiologic outcome. Differing peak concentration/MIC ratios (1,2, and 10), T>MIC (3.6,9.6,15.6, and 24 h corresponding to 15, 40, 65, and 100% of the 24-h dosing interval), and AUC/MIC ratios (13-180) were generated by varying dosing strategies. Initial bacterial inocula were decreased by 99.9% in each experiment conducted. Despite the wide variation in exposure levels, in terms of AUC/MIC, Cp-max/MIC, and T>MIC, the kill portions of the bacterial density curves were super-imposable between all permutations of antibiotic exposure. However, there appeared to be an AUC/MIC breakpoint (35-40) defining bacterial regrowth. Over a 10-fold concentration range, levofloxacin appeared to kill S. pneumoniae in a concentration-independent fashion. When given in concentrations suitable to achieve specified pharmacodynamic endpoints (AUC/MIC >/=35), levofloxacin demonstrated the ability to eradicate both a levofloxacin-resistant and levofloxacin-sensitive strain of S. pneumoniae in the in vitro model.

In Vitro Evaluation of the Activities of Telavancin, Cefazolin, and Vancomycin against Methicillin-Susceptible and Methicillin-Resistant Staphylococcus aureus in Peritoneal Dialysate

ABSTRACT This study compared the ability of telavancin to the ability of cefazolin and vancomycin to eliminate staphylococci from peritoneal dialysis fluid by using a static in vitro model to simulate the conditions of peritoneal dialysis. The results showed that telavancin exhibited statistically significantly better kill (P < 0.05) against both methicillin-susceptible and methicillin-resistant Staphylococcus aureus.

Pharmacodynamics of Pulse Dosing versus Standard Dosing: In Vitro Metronidazole Activity against Bacteroides fragilis and Bacteroides thetaiotaomicron

ABSTRACT Pulse dosing is a novel approach to dosing that produces escalating antibiotic levels early in the dosing interval followed by a prolonged dose-free period. Antibiotic is frontloaded by means of four sequential bolus injections, after which antibiotic levels are allowed to diminish until the next dose. This study compares standard thrice-daily dosing and pulse dosing of metronidazole against Bacteroides spp. in an in vitro model. Two American Type Culture Collection Bacteroides fragilis isolates (metronidazole MIC for each organism = 1 mg/liter) were exposed to metronidazole for 48 or 96 h. Human pharmacokinetics were simulated for an oral 500-mg dose given every 8 h (maximum concentration of drug [Cmax] = 12 mg/liter; half-life = 8 h; area under the curve [AUC] = 294 mg · h/liter) and for pulse dosing. Pulses, each producing an increase in metronidazole concentration of 9 mg/liter, were administered at times 0, 2, 4, and 6 h of each 24-h cycle, with a targeted half-life of 8 h (AUC = 347 mg · h/liter). A metronidazole-resistant B. fragilis strain (metronidazole MIC = 32 mg/liter) was exposed to both dosing regimens and, additionally, to a regimen of 1,500 mg administered once daily (Cmax = 36 mg/liter; AUC = 364 mg · h/liter). Furthermore, regimens against one B. fragilis isolate and one B. thetaiotaomicron isolate corresponding to one-fourth and one-eighth of the thrice-daily and pulse dosing regimens, mimicking peak metronidazole concentrations achieved in abscesses, were simulated in 48-h experiments (metronidazole MIC = 1 mg/liter). Time-kill curves were generated for each experiment and analyzed for bactericidal activity, defined as a bacterial burden reduction ≥ 3 log10 CFU/ml. The results of paired (Wilcoxon matched-pair signed-rank test) and nonpaired (Mann-Whitney test) statistical analyses conducted on time to 3 log10 kill data and area under the kill curve data from each of the thrice-daily dosing experiments versus each of the pulse dosing experiments were considered not significant for a given isolate-dosing regimen combination. The thrice-daily dosing, pulse dosing, and once-daily dosing regimens all exhibited bactericidal activity. Metronidazole administered in standard or pulse dosing fashion was highly active against both susceptible and resistant strains of Bacteroides spp.