Human plasma protein levels alter the in vitro antifungal activity of caspofungin: An explanation to the effect in critically ill?

BACKGROUND: Recent studies have shown low caspofungin concentrations in critically ill patients. In some patients, the therapeutic target, area under the total plasma concentration curve in relation to the minimal inhibition concentration (AUCtot /MIC), seems not to be achieved and therapeutic drug monitoring (TDM) has been proposed. Caspofungin is highly protein-bound and the effect of reduced plasma protein levels on pharmacodynamics has not been investigated. OBJECTIVES: Fungal killing activity of caspofungin in vitro was investigated under varying levels of human plasma protein. METHODS: Time-kill studies were performed with clinically relevant caspofungin concentrations of 1-9 mg/L on four blood isolates of C. glabrata, three susceptible and one strain with reduced susceptibility, in human plasma and plasma diluted to 50% and 25% using Ringer's acetate. RESULTS: Enhanced fungal killing of the three susceptible strains was observed in plasma with lower protein content (p < .001). AUCtot /MIC required for a 1 log10 CFU/ml kill at 24 h in 50% and 25% plasma was reduced with 36 + 12 and 80 + 9%, respectively. The maximum effect was seen at total caspofungin concentrations of 4-9 × MIC. For the strain with reduced susceptibility, growth was significantly decreased at lower protein levels. CONCLUSIONS: Reduced human plasma protein levels increase the antifungal activity of caspofungin in vitro, most likely by increasing the free concentration. Low plasma protein levels in critically ill patients with candidemia might explain a better response to caspofungin than expected from generally accepted target attainment and should be taken into consideration when assessing TDM based on total plasma concentrations.

Frequent emergence of porin-deficient subpopulations with reduced carbapenem susceptibility in ESBL-producing Escherichia coli during exposure to ertapenem in an in vitro pharmacokinetic model.

OBJECTIVES: Ertapenem resistance is increasing in Enterobacteriaceae. The production of extended-spectrum ß-lactamases (ESBLs) and reduced expression of outer membrane porins are major mechanisms of resistance in ertapenem-resistant Klebsiella pneumoniae. Less is known of ertapenem resistance in Escherichia coli. The aim of this study was to explore the impact of ESBL production in E. coli on the antibacterial activity of ertapenem. METHODS: Two E. coli strains, with and without ESBL production, were exposed to ertapenem in vitro for 48 h at concentrations simulating human pharmacokinetics with conventional and higher dosages. RESULTS: Isolates with non-susceptibility to ertapenem (MICs 0.75-1.5 mg/L) were detected after five of nine time-kill experiments with the ESBL-producing strain. All of these isolates had ompR mutations, which reduce the expression of outer membrane porins OmpF and OmpC. Higher dosage did not prevent selection of porin-deficient subpopulations. No mutants were detected after experiments with the non-ESBL-producing strain. Compared with other experiments, experiments with ompR mutants detected in endpoint samples showed significantly less bacterial killing after the second dose of ertapenem. Impaired antibacterial activity against E. coli with ESBL production and ompR mutation was also demonstrated in time-kill experiments with static antibiotic concentrations. CONCLUSIONS: The combination of ESBL production and porin loss in E. coli can result in reduced susceptibility to ertapenem. Porin-deficient subpopulations frequently emerged in ESBL-producing E. coli during exposure to ertapenem at concentrations simulating human pharmacokinetics. Inappropriate use of ertapenem should be avoided to minimize the risk of selection of ESBL-producing bacteria with reduced susceptibility to carbapenems.

Voriconazole-induced inhibition of the fungicidal activity of amphotericin B in Candida strains with reduced susceptibility to voriconazole: an effect not predicted by the MIC value alone.

An antagonistic effect of voriconazole on the fungicidal activity of sequential doses of amphotericin B has previously been demonstrated in Candida albicans strains susceptible to voriconazole. Because treatment failure and the need to switch to other antifungals are expected to occur more often in infections that are caused by resistant strains, it was of interest to study whether the antagonistic effect was still seen in Candida strains with reduced susceptibility to voriconazole. With the hypothesis that antagonism will not occur in voriconazole-resistant strains, C. albicans strains with characterized mechanisms of resistance against voriconazole, as well as Candida glabrata and Candida krusei strains with differences in their degrees of susceptibility to voriconazole were exposed to voriconazole or amphotericin B alone, to both drugs simultaneously, or to voriconazole followed by amphotericin B in an in vitro kinetic model. Amphotericin B administered alone or simultaneously with voriconazole resulted in fungicidal activity. When amphotericin B was administered after voriconazole, its activity was reduced (median reduction, 61%; range, 9 to 94%). Levels of voriconazole-dependent inhibition of amphotericin B activity differed significantly among the strains but were not correlated with the MIC values (correlation coefficient, -0.19; P = 0.65). Inhibition was found in C. albicans strains with increases in CDR1 and CDR2 expression but not in the strain with an increase in MDR1 expression. In summary, decreased susceptibility to voriconazole does not abolish voriconazole-dependent inhibition of the fungicidal activity of amphotericin B in voriconazole-resistant Candida strains. The degree of interaction could not be predicted by the MIC value alone.

Posaconazole in human serum: a greater pharmacodynamic effect than predicted by the non-protein-bound serum concentration.

It is generally accepted that only the unbound fraction of a drug is pharmacologically active. Posaconazole is an antifungal agent with a protein binding of 98 to 99%. Taking into account the degree of protein binding, plasma levels in patients, and MIC levels of susceptible strains, it can be assumed that the free concentration of posaconazole sometimes will be too low to exert the expected antifungal effect. The aim was therefore to test the activity of posaconazole in serum in comparison with that of the calculated unbound concentrations in protein-free media. Significant differences (P < 0.05) from the serum control were found at serum concentrations of posaconazole of 1.0 and 0.10 mg/liter, with calculated free concentrations corresponding to 1× MIC and 0.1× MIC, respectively, against one Candida lusitaniae strain selected for proof of principle. In RPMI 1640, the corresponding calculated unbound concentration of 0.015 mg/liter resulted in a significant effect, whereas that of 0.0015 mg/liter did not. Also, against seven additional Candida strains tested, there was an effect of the low posaconazole concentration in serum, in contrast to the results in RPMI 1640. Fluconazole, a low-grade-protein-bound antifungal, was used for comparison at corresponding concentrations in serum and RPMI 1640. No effect was observed at the serum concentration, resulting in a calculated unbound concentration of 0.1× MIC. In summary, there was a substantially greater pharmacodynamic effect of posaconazole in human serum than could be predicted by the non-protein-bound serum concentration. A flux from serum protein-bound to fungal lanosterol 14α-demethylase-bound posaconazole is suggested.

Foreign travel is a major risk factor for colonization with Escherichia coli producing CTX-M-type extended-spectrum beta-lactamases: a prospective study with Swedish volunteers.

Foreign travel has been suggested to be a risk factor for the acquisition of extended-spectrum beta-lactamase (ESBL)-producing Enterobacteriaceae. To our knowledge, this has not previously been demonstrated in a prospective study. Healthy volunteers traveling outside Northern Europe were enrolled. Rectal swabs and data on potential travel-associated risk factors were collected before and after traveling. A total of 105 volunteers were enrolled. Four of them did not complete the study, and one participant carried ESBL-producing Escherichia coli before travel. Twenty-four of 100 participants with negative pretravel samples were colonized with ESBL-producing Escherichia coli after the trip. All strains produced CTX-M enzymes, mostly CTX-M-15, and some coproduced TEM or SHV enzymes. Coresistance to several antibiotic subclasses was common. Travel to India was associated with the highest risk for the acquisition of ESBLs (88%; n = 7). Gastroenteritis during the trip was an additional risk factor (P = 0.003). Five of 21 volunteers who completed the follow-up after 6 months had persistent colonization with ESBLs. This is the first prospective study demonstrating that international travel is a major risk factor for colonization with ESBL-producing Enterobacteriaceae. Considering the high acquisition rate of 24%, it is obvious that global efforts are needed to meet the emergence and spread of CTX-M enzymes and other antimicrobial resistances.

Characterization of the inhibitory effect of voriconazole on the fungicidal activity of amphotericin B against Candida albicans in an in vitro kinetic model.

OBJECTIVES: The aim of the present investigation was to study and characterize the effect of voriconazole on the fungicidal activity of amphotericin B. METHODS: Four strains of Candida albicans susceptible to voriconazole were exposed to voriconazole and amphotericin B, either alone, simultaneously or sequentially in an in vitro kinetic model. Bolus doses resulting in voriconazole and amphotericin B concentrations of 0.005-5 and 2.5 mg/L, respectively, were administered. Antifungal-containing RPMI 1640 was eliminated and replaced by a fresh medium using a peristaltic pump, with a flow rate adjusted to obtain the desired half-lives. With two drugs tested, a computer-controlled dosing pump compensated for differences in the elimination rates. Using static time-kill methodology, one C. albicans strain was exposed to 5 mg/L voriconazole for varying durations followed by 2.5 mg/L amphotericin B after three repeated washes of voriconazole. RESULTS: Voriconazole and amphotericin B treatment alone resulted in fungistatic and fungicidal activities, respectively. Simultaneous administration of voriconazole and amphotericin B resulted in fungicidal activity, whereas only fungistatic activity was observed when repeated doses of amphotericin B were administered sequentially after voriconazole at 24-96 h. The inhibition of the fungicidal activity of amphotericin B was voriconazole dose-dependent, but seemed to be recovered once the voriconazole concentration fell below the MIC. The fungicidal activity was quickly regained after the removal of voriconazole, irrespective of the duration of voriconazole pre-exposure. CONCLUSIONS: Voriconazole inhibited the fungicidal effect of sequentially administered amphotericin B in a concentration- and time-dependent manner; the clinical significance of this needs further investigation.

[Cross allergy between penicillins and other beta lactam antibiotics--the risk is much less than previously thought]. / Korsallergi mellan penicilliner och övriga betalaktam-antibiotika - Risken är betydligt mindre än man tidigare trott.

Severe IgE-mediated allergic reactions to penicillins are rare but might be fatal. Because some studies demonstrated a high risk of cross-sensitivity to cephalosporins and carbapenems it has been recommended to avoid these antibiotics in patients with suspected hypersensitivity to penicillins. However, recent studies and analyses conclude that the risk of cross-reactivity was overestimated in the earlier studies and that it is in fact very low for parenteral cephalosporins and perhaps even negligible for carbapenems. The new knowledge has implications for the choice of therapy for bacterial infections in patients with a history of penicillin hypersensitivity, because alternative antibiotic regimens are often inferior to beta-lactam antibiotics. The aim of the present review is to present existing knowledge on cross-sensitivity between beta-lactams, as well as to discuss the management of patients with suspected allergic reactions to these antibiotics.

Pharmacodynamic effects of telavancin against methicillin-resistant and methicillin-susceptible Staphylococcus aureus strains in the presence of human albumin or serum and in an in vitro kinetic model.

Telavancin is a novel bactericidal lipoglycopeptide with multiple mechanisms of action against gram-positive pathogens. The aim of this study was to describe the dynamics of the antimicrobial effect of telavancin against two strains of Staphylococcus aureus (methicillin susceptible and methicillin resistant) in an in vitro kinetic model with simulated human pharmacokinetics. Also, static experiments were performed to determine the rate and extent of killing by telavancin in the presence and absence of human albumin and human serum. Experiments in broth and in nutrient-depleted medium were performed to study the rate and extent of killing by telavancin of bacteria in different growth phases. In the in vitro kinetic model regrowth was noted at 24 h for both strains when exposed to initial concentrations below 5 mg/liter. There was a >3-log(10) killing at all concentrations from 0.5x MIC and above at 24 h both in broth and in the presence of 40-g/liter human albumin. In contrast to the methicillin-susceptible strain, the methicillin-resistant strain in 40-g/liter human albumin showed a regrowth at concentrations of 0.5x MIC and 1x MIC at 24 h. At all the other concentrations >3-log(10) killing was seen at 24 h. Concordant results were seen in 50% human serum. At a target area under the curve/MIC ratio of 50 (corresponding to the human dose of 10 mg/kg of body weight, administered intravenously), >3-log(10) killing was observed at 6 to 8 h. Unlike most antibiotics, telavancin was able to kill both strains in a nongrowing phase.

Semimechanistic pharmacokinetic/pharmacodynamic model for assessment of activity of antibacterial agents from time-kill curve experiments.

Dosing of antibacterial agents is generally based on point estimates of the effect, even though bacteria exposed to antibiotics show complex kinetic behaviors. The use of the whole time course of the observed effects would be more advantageous. The aim of the present study was to develop a semimechanistic pharmacokinetic (PK)/pharmacodynamic (PD) model characterizing the events seen in a bacterial system when it is exposed to antibacterial agents with different mechanisms of action. Time-kill curve experiments were performed with a strain of Streptococcus pyogenes exposed to a wide range of concentrations of the following antibiotics: benzylpenicillin, cefuroxime, erythromycin, moxifloxacin, and vancomycin. Bacterial counts were monitored with frequent sampling during the experiment. A simultaneous fit of all data was accomplished. The degradation of the drugs was monitored and corrected for in the model, and a link model was used to account for an effect delay. In the final PK/PD model, the total bacterial population was divided into two subpopulations: one growing drug-susceptible population and one resting insusceptible population. The drug effect was included as an increase of the killing rate of bacteria in the susceptible state, according to a maximum-effect (E(max)) model. An internal model validation showed that the model was robust and had good predictability. In conclusion, for all drugs, the final PK/PD model successfully described bacterial growth and killing kinetics when the bacteria were exposed to different antibiotic concentrations. The semimechanistic model that was developed might, after further refinement, serve as a tool for the development of optimal dosing strategies for antibacterial agents.

Pharmacodynamic studies of amoxicillin against Streptococcus pneumoniae: comparison of a new pharmacokinetically enhanced formulation (2000 mg twice daily) with standard dosage regimens.

OBJECTIVES: To compare the pharmacodynamic effects of a pharmacokinetically enhanced formulation of amoxicillin 2000 mg twice daily, with amoxicillin 875 mg twice daily, 875 mg three times daily and 500 mg three times daily against Streptococcus pneumoniae with different susceptibility to amoxicillin in an in vitro kinetic model. METHODS: Strains of S. pneumoniae with amoxicillin MICs of 1, 2, 4 and 8 mg/L at an initial inoculum of approximately 10(5) cfu/mL were exposed to amoxicillin in an in vitro kinetic model simulating the human serum concentration-time profile of the pharmacokinetically enhanced formulation twice daily (C(max) 17 mg/L after 1.5 h). All isolates were also exposed to amoxicillin with concentration-time profiles correlating to the human dosage of 875 mg twice daily (C(max) 15 mg/L after 1 h), 875 mg three times daily and 500 mg (C(max) 8 mg/L after 1 h) three times daily with simulated half-life of 1 h. Repeated samples were taken regularly during 24 h and viable counts were carried out. RESULTS: Overall, the pharmacokinetically enhanced formulation was more effective at reducing bacterial counts than any of the other formulations evaluated. Eradication was achieved with the enhanced formulation for strains with a MIC of < or =2 mg/L, however, regrowth occurred with the other dosing regimens. In the experiments with the strain with a MIC of 4 mg/L, the enhanced formulation kept the bacterial counts < or =10(2) cfu/mL for at least 14 out of 24 h tested. In contrast, none of the other formulations reduced the bacterial counts down to < or =10(2) cfu/mL at any point. None of the regimens was able to eradicate the strain with an MIC of 8 mg/L, even though an initial substantial kill was noted with the enhanced formulation after both doses. The least effective dosage regimen for all strains was 875 mg twice daily.

Postantibiotic, postantibiotic sub-MIC, and subinhibitory effects of PGE-9509924, ciprofloxacin, and levofloxacin.

Postantibiotic effects (PAEs), postantibiotic sub-MIC effects, and sub-MIC effects of the new nonfluoroquinolone PGE-9509924, ciprofloxacin, and levofloxacin against gram-positive and gram-negative strains were investigated. In comparison to ciprofloxacin and levofloxacin, PGE-9509924 exerted very similar PAEs against all strains except for both strains of Streptococcus pneumoniae, where longer PAEs were found for PGE-9509924. All three investigated quinolones showed no minimal PAEs against Pseudomonas aeruginosa.

Bactericidal effects of levofloxacin in comparison with those of ciprofloxacin and sparfloxacin.

OBJECTIVE: To investigate and compare the in vitro activity of levofloxacin with the activities of ciprofloxacin and sparfloxacin. METHODS: The following experiments were performed: (1) comparative studies of the rate of killing by the three quinolones of different strains of Streptococcus pneumoniae at a concentration corresponding to the 1-h serum level following a 500-mg dose in humans; (2) comparative studies of the rate of killing by levofloxacin and ciprofloxacin of different strains of Staphylococcus aureus, Escherichia coli and Pseudomonas aeruginosa at the same concentrations as above; (3) comparative studies of the rate of killing by levofloxacin at four different concentrations of reference and clinical strains of Streptococcus pneumoniae, Staphylococcus aureus, E. coli and P. aeruginosa. RESULTS: Levofloxacin exhibited statistically significantly higher bactericidal activity than sparfloxacin after 2 and/or 3 h against all strains of Streptococcus pneumoniae. Compared to ciprofloxacin, levofloxacin showed a statistically significantly higher bactericidal activity after 2 and/or 3 h against all strains of Streptococcus pneumoniae except the one resistant to both penicillin and cefotaxime. No differences in killing rate between levofloxacin and ciprofloxacin were seen against Staphylococcus aureus, E. coli and P. aeruginosa, with almost complete killing after 3 h of the P. aeruginosa strains and after 6 h for the E. coli strains. No concentration-dependent killing was seen at concentrations above 4xMIC of levofloxacin against Staphyloccus aureus, E. coli and P. aeruginosa. CONCLUSION: Levofloxacin was shown to be active against both Gram-positive and Gram-negative bacteria. In terms of MIC values, ciprofloxacin was the most active drug against the Gram-negative organisms, and sparfloxacin against the strains of Streptococcus pneumoniae, but levofloxacin exhibited a similar or even better bactericidal activity against the investigated strains compared with the other two fluoroquinolones when killing curves were compared.

Comparative in vitro pharmacodynamics of BO-2727, meropenem and imipenem against Gram-positive and Gram-negative bacteria.

OBJECTIVE: To investigate and compare the in vitro pharmacodynamics of three carbapenems: imipenem, meropenem and BO-2727. METHODS: The following studies were performed: (1) comparative studies of the rate of killing of the three carbapenems of reference strains of Gram-positive and Gram-negative bacteria at a concentration corresponding to the 1-h serum level following 500 mg intravenously in humans; (2) comparative studies of the rate of killing of BO-2727, meropenem and imipenem at different antibiotic concentrations of reference strains of Gram-positive and Gram-negative bacteria; (3) comparative studies of the rate of killing of BO-2727, meropenem and imipenem of bacteria which are phenotypically tolerant; (4) studies of the postantibiotic effect of BO-2727 using viable counts and optical density; (5) studies of the postantibiotic sub-MIC effect (PA SME) of BO-2727 using optical density. RESULTS: No difference in killing rate was noted between the three carbapenems, and there was no concentration-dependent killing of the Gram-negative strains after 6 h. A pronounced paradoxical effect was seen against Staphylococcus aureus. All three antibiotics were able to kill phenotypically tolerant bacteria. Only very short or no postantibiotic effect of BO-2727 was found against the investigated strains. Very long PA SMEs were noted for the Gram-negative strains, although there was a pronounced variation for the different strains of Pseudomonas aeruginosa. CONCLUSION: There was no significant difference between the studied carbapenems in their pharmacodynamic properties. All three antibiotics acted similarly to other beta-lactam antibiotics.

Postantibiotic and sub-MIC effects of benzylpenicillin against Streptococcus pneumoniae with different susceptibilities for penicillin.

BACKGROUND: The purpose of the study was to examine whether penicillin-susceptible and nonsusceptible strains of Streptococcus pneumoniae exhibited different pharmacodynamic responses to benzylpenicillin. METHODS: The postantibiotic effects (PAEs) and the postantibiotic sub-MIC effects (PA SMEs) were investigated by optical density against strains of S. pneumoniae with different susceptibilities to benzylpenicillin. To validate the data, the PAE and PA SME of one susceptible and one resistant strain were also tested with the viable count method. The post-MIC effects (PMEs) were studied in an in vitro kinetic model, simulating human pharmacokinetics with a half-life of 1 h and a time above MIC of approximately 20% of 24 h. RESULTS: There were no differences with respect to the PAEs, PA SMEs and PMEs of benzylpenicillin for the various strains of S. pneumoniae, irrespective of their susceptibility to penicillin. For both some of the susceptible and resistant strains investigated, longer PA SMEs at 0.2 and 0.3 x MIC were noted, indicating that these parameters might be more dependent on the type of strain rather than on the susceptibility status. CONCLUSION: No differences in the pharmacodynamic response after similar drug exposure were seen for S. pneumoniae strains with different penicillin susceptibility.

Pharmacodynamics of moxifloxacin against Streptococcus pyogenes in an in vitro kinetic model.

The aim of the present study was to investigate the pharmacodynamics of moxifloxacin against strains of Streptococcus pyogenes with different susceptibilities to erythromycin by using an in vitro kinetic model simulating human pharmacokinetics of moxifloxacin at oral doses of 400 and 200 mg, respectively. When the different strains of S. pyogenes were exposed to the higher dose, the number of bacteria was reduced below the detection limit after 12 h and no regrowth was noted during the following 12 h. At the lower dose there was regrowth of the strains with constitutive and inducible erythromycin resistance of the MLS(B) phenotype. Replication assays of the regrowing bacteria indicated that the failure of moxifloxacin to kill the MLS(B) strains at the lower dose was likely caused by the emergence of preexisting resistant subpopulations. Thus, the present study indicates that the presently used 400-mg dose seems to have an advantage over the lower dose in that the risk for selection of resistant subpopulations is minimized.

Suboptimal antibiotic dosage as a risk factor for selection of penicillin-resistant Streptococcus pneumoniae: in vitro kinetic model.

Optimizing pharmacokinetic/pharmacodynamic indices of antibiotics to obtain clinical and microbiological efficacy is essential, but dosing regimens must also be tailored to minimize the risk for emergence of resistance. The aim of the present study was to investigate whether certain concentrations of benzylpenicillin are critical for the selection of resistant subpopulations. A mixed culture of Streptococcus pneumoniae containing ca. 90% susceptible (MIC = 0.031 mg/liter), 9% intermediate (MIC = 0.25 mg/liter), and 1% resistant (MIC = 8 mg/liter) was studied in an in vitro kinetic model. The time that concentrations exceeded the MIC (T>MIC) for the three strains in the culture was varied by different initial concentrations of benzylpenicillin. Samples for viable counts were withdrawn at different times during 24 h and seeded on blood agar plates and on selective antibiotic-containing plates. The T>MIC varied from 46 to 100% for the susceptible strain, from 6 to 100% for the intermediate strain, and from 0 to 48% for the resistant strain. Our study, which may mimic the clinical situation with carriage of a mixed population of S. pneumoniae with different antibiotic susceptibilities, has shown that selection of resistant bacteria may easily occur if dosing regimens are only targeted toward fully susceptible strains.