Skip to main page content

• HOME
• Current Issue
• Archive
• Alerts
• About ASM
• Contact us
• Tech Support
• Journals.ASM.org

This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Mascio, C. T. M. Articles by Silverman, J. A. Search for Related Content PubMed PubMed Citation Articles by Mascio, C. T. M. Articles by Silverman, J. A.

 Previous Article  |  Next Article 

Antimicrobial Agents and Chemotherapy, December 2007, p. 4255-4260, Vol. 51, No. 12
0066-4804/07/$08.00+0     doi:10.1128/AAC.00824-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Bactericidal Action of Daptomycin against Stationary-Phase and Nondividing Staphylococcus aureus Cells

Carmela T. M. Mascio,^* Jeff D. Alder, and Jared A. Silverman

Cubist Pharmaceuticals, Inc., Lexington, Massachusetts

Received 25 June 2007/ Returned for modification 27 July 2007/ Accepted 30 September 2007

Top ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 
Most antibiotics with bactericidal activity require that the bacteria be actively dividing to produce rapid killing. However, in many infections, such as endocarditis, prosthetic joint infections, and infected embedded catheters, the bacteria divide slowly or not at all. Daptomycin is a lipopeptide antibiotic with a distinct mechanism of action that targets the cytoplasmic membrane of gram-positive organisms, including Staphylococcus aureus. Daptomycin is rapidly bactericidal against exponentially growing bacteria (a 3-log reduction in 60 min). The objectives of this study were to determine if daptomycin is bactericidal against nondividing S. aureus and to quantify the extent of the bactericidal activity. In high-inoculum methicillin-sensitive S. aureus cultures in stationary phase (10¹⁰ CFU/ml), daptomycin displayed concentration-dependent bactericidal activity, requiring 32 µg/ml to achieve a 3-log reduction. In a study comparing several antibiotics at 100 µg/ml, daptomycin demonstrated faster bactericidal activity than nafcillin, ciprofloxacin, gentamicin, and vancomycin. In experiments where bacterial cell growth was halted by the metabolic inhibitor carbonyl cyanide m-chlorophenylhydrazone or erythromycin, daptomycin (10 µg/ml) achieved the bactericidal end point (a 3-log reduction) within 2 h. In contrast, ciprofloxacin (10 µg/ml) did not produce bactericidal activity. Daptomycin (2 µg/ml) remained bactericidal against cold-arrested S. aureus, which was protected from the actions of ciprofloxacin and nafcillin. The data presented here suggest that, in contrast to that of other classes of antibiotics, the bactericidal activity of daptomycin does not require cell division or active metabolism, most likely as a consequence of its direct action on the bacterial membrane.

Top ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 
Most in vitro studies examining the activity of antibiotics have been performed on rapidly dividing planktonic bacterial cultures supplemented with rich growth media. However, in infections, bacteria seldom encounter optimal conditions that allow logarithmic growth. Instead, long periods of limited or arrested growth are normal (20). Staphylococcus aureus is known to modulate gene expression to enhance endurance under less-than-optimal growth conditions and survive periods in stationary phase (27). It is likely that stationary-phase or nondividing bacteria are common in many persistent infections (e.g., endocarditis and osteomyelitis) and in biofilm-associated infections (e.g., on catheters, grafts, and foreign bodies) (11, 16). Therefore, antibiotics that are bactericidal under growth-arrested conditions may be beneficial.

The mechanism of action of many bactericidal antibiotics requires ongoing cell activity and cell division for the drugs' killing activity. Such drugs therefore display limited activity against nondividing cultures (4, 14, 15, 17). For example, beta-lactams require ongoing bacterial cell wall synthesis (30), and the action of some quinolones requires ongoing RNA and protein synthesis for bactericidal activity, while others require ongoing cell division (15). Rifampin is one of the few antibiotics identified as retaining bactericidal activity against nongrowing bacterial cultures (32).

Daptomycin is a lipopeptide antibiotic that displays rapid bactericidal activity in vitro against gram-positive pathogens, including streptococci, methicillin-resistant S. aureus, and vancomycin-resistant enterococci (5, 13, 18, 31). Daptomycin works by disrupting membrane function and causing leakage of essential potassium ions, ultimately leading to loss of membrane potential and cell death (26). Studies of daptomycin using artificial membranes have shown that daptomycin can act directly on the lipid bilayer in the absence of any bacterial protein or other cell surface component. Direct interaction with the bilayer suggests that daptomycin could act independently of growth phase or cellular metabolic activity (23, 24, 26). Bactericidal activity, in the absence of cell proliferation, may be valuable against infections involving slow-growing or growth-arrested gram-positive organisms.

The potency of daptomycin in comparison with other antibiotics was determined against stationary-phase S. aureus cultures under growth arrest by nutrient depletion, chemical treatment, and cold temperature.

(Portions of this work were previously presented [45th Intersci. Conf. Antimicrob. Agents Chemother., abstr. E-1743, 2005].)

Top ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 
Bacterial strain and culture conditions. Methicillin-sensitive S. aureus strain ATCC 29213 was used for all of the studies presented here. Bacterial cultures were incubated overnight at 37°C with shaking (200 rpm) in Mueller-Hinton broth supplemented with 50-mg/liter free Ca²⁺ (MHBc) and diluted 1:1,000 into fresh MHBc. To prepare exponential-phase cultures, these diluted cultures were grown in MHBc for 2 h prior to experimentation. To prepare stationary-phase cultures, diluted cultures were grown for 17 to 20 h prior to testing.

Susceptibility testing. MIC determinations were performed in MHBc following Clinical and Laboratory Standards Institute (formerly the National Committee for Clinical Laboratory Standards) guidelines for broth microdilution MIC assays, except that cultures were incubated at 37°C with shaking at 200 rpm.

Time-kill studies. Bacterial suspension cultures in MHBc were treated with various concentrations of different antibiotics. Time-kill studies were performed at 37°C with shaking at 200 rpm, unless otherwise indicated. Culture aliquots (100 µl) were removed at the times shown in the figures, serially diluted in MHB or phosphate-buffered saline (pH 7.4 to 7.6), and then plated on tryptic soy agar fortified with 5% sheep blood and incubated overnight in a 35 to 37°C ambient-air incubator. Cell viability was assessed by enumerating the CFU per milliliter. The lower limit of detection for these studies was 10³ CFU/ml, and bactericidal activity was defined as a 3-log reduction with antibiotic treatment compared with the untreated control at the start of each assay. Undiluted and 1:10-diluted samples were not titrated for the number of CFU per milliliter in order to reduce the effect of drug carryover and avoid misrepresentation of the effect of each compound during the course of the time-kill study.

Time-kill studies of exponential- and stationary-phase S. aureus. Exponential-phase S. aureus cultures were prepared in MHBc as detailed and then treated with daptomycin at 2 µg/ml. Separately, high cell density stationary-phase cultures of S. aureus in MHBc were treated with various concentrations of daptomycin (0, 8, 16, 32, 64, and 128 µg/ml). On the basis of the data generated from the concentration range-finding study, the bactericidal actions of daptomycin and comparator antibiotics were evaluated. Stationary-phase cultures were separately treated with daptomycin, nafcillin, ciprofloxacin, gentamicin, or vancomycin at 100 µg/ml in MHBc. Cell density (the number of CFU per milliliter) was determined at the indicated time points.

Time-kill study of nutrient-depleted, growth-arrested S. aureus. The bactericidal activity of daptomycin against exponentially growing, concentrated exponentially growing, stationary, and diluted stationary-phase S. aureus was determined in nutrient-depleted (or spent) MHBc medium (depMHBc). depMHBc was prepared by adding S. aureus to 400 ml of MHBc and incubating it at 37°C for 46.5 h. Afterward, the cultures were centrifuged (4,648 x g) for 10 min at 4°C to collect whole bacterial cells. The supernatant was removed and passed through a 0.45-µm-pore-size filter (Nalgene, Rochester, NY). The calcium concentration in the filtered supernatant was measured with the QuantiChrom Calcium Assay Kit (BioAssay Systems, Hayward, CA) and adjusted to 50 mg/liter.

Stationary-phase cells were prepared as detailed earlier and then centrifuged (6,588 x g) for 10 min at room temperature, washed with depMHBc, centrifuged again, and suspended to equal the initial volume in depMHBc. Exponentially growing, concentrated exponentially growing, stationary-phase, and diluted stationary-phase S. aureus cells were prepared in depMHBc by standardizing the optical density (OD₆₀₀) to 3.75 for concentrated exponentially growing and stationary-phase cells and 0.1 for exponentially growing and diluted stationary-phase cells. Bacterial cultures were treated with daptomycin at 10 or 100 µg/ml and assayed at the indicated time points. During each assay, depMHBc sterility was confirmed by incubating a blank sample during the time-kill study.

Time-kill study of chemically growth-arrested S. aureus. Exponentially growing cells were grown to an OD₆₀₀ of approximately 0.3. Cultures were divided into three equal portions and left untreated (control) or treated with 10 µM carbonyl cyanide m-chlorophenylhydrazone (CCCP) or erythromycin at 2.5 µg/ml. Samples were incubated for 1 h at 37°C while shaking (200 rpm) to allow growth arrest. After the 1-h pretreatment, parallel experiments were performed to test the activity of daptomycin or ciprofloxacin at 10 µg/ml on growth-arrested cultures. Bacterial counts were assayed at various time points.

Time-kill study of S. aureus cultures growth arrested by cold temperature. Exponentially growing S. aureus cells were chilled in an ice bath for 1 h while shaking (200 rpm) prior to treatment with a concentration of ciprofloxacin, daptomycin, or nafcillin equal to four times the MIC (2 µg/ml). Samples were incubated on ice (0°C) with shaking (200 rpm) during the time course experiment. After 24 h under these conditions, samples were returned to 37°C for 2 h. Numbers of CFU per milliliter were assayed at various time points.

Reagents. Daptomycin was provided by Cubist Pharmaceuticals (Lexington, MA). Other antibiotics, including erythromycin, gentamicin, nafcillin, and vancomycin, were purchased from Sigma (St. Louis, MO), while ciprofloxacin was purchased from U.S. Biological (Swampscott, MA). CCCP and calcium chloride were obtained from Sigma (St. Louis, MO). Culture media, including blood agar plates and MHB, were obtained from bioMérieux (Lombard, IL) and Difco Becton Dickinson (Sparks, MD), respectively.

Top ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 
Susceptibility testing. MIC determinations were performed in MHBc (9). The MICs of the antibiotics used in these studies, determined under growth conditions that would support exponential growth as defined by the Clinical and Laboratory Standards Institute, were as follows: ciprofloxacin, 0.5 µg/ml; daptomycin, 0.5 µg/ml; gentamicin, 0.25 µg/ml; nafcillin, 0.125 µg/ml; vancomycin, 0.5 µg/ml.

Bactericidal activity of daptomycin against exponentially growing S. aureus. As a basis for comparison with experiments in growth-arrested cultures, the effects of daptomycin were evaluated in exponentially growing cultures of S. aureus. Daptomycin, at a concentration equal to four times the MIC, was added to cultures to determine the effects on cell viability compared with untreated control cells. At this concentration, daptomycin (2 µg/ml) exhibited rapid bactericidal activity in active cultures of S. aureus (as defined by a 3-log reduction in the number of CFU per milliliter) within 60 min (Fig. 1). Untreated bacterial cells divided rapidly, leading to a 20-fold cell density increase (from 5 x 10⁷ to 1 x 10⁹ CFU/ml) during the 120-min-long experiment (Fig. 1).

View larger version (12K): [in this window] [in a new window]

FIG. 1. Daptomycin (2 µg/ml) exhibits rapid bactericidal activity against exponential-phase cultures of methicillin-sensitive S. aureus. DAP, daptomycin.

In order to determine whether daptomycin also had bactericidal activity against stationary-phase organisms, bacteria were grown for at least 17 h to create high inoculum density cultures (1 x 10¹⁰ CFU/ml) and then subjected to increasing concentrations of daptomycin in MHBc. In this experiment, S. aureus cells appeared to be in stationary phase because untreated cells did not proliferate, as shown by the flat viability curve (Fig. 2). However, in cultures treated with daptomycin, concentration-dependent cell killing was measured. In comparison to exponentially growing S. aureus, stationary-phase cultures required higher concentrations of daptomycin (32 µg/ml) to reach the bactericidal end point after 24 h of treatment. This altered concentration response to daptomycin may be due to an altered bacterial physiological (growth) state, culture density, or both.

View larger version (21K): [in this window] [in a new window]

FIG. 2. Daptomycin displays concentration-dependent bactericidal activity against high inoculum density stationary-phase cultures of S. aureus. Prior to exposure to daptomycin, cultures were incubated for >17 h to generate high cell density stationary-phase samples.

Bactericidal activities of daptomycin and comparator antibiotics against stationary-phase S. aureus. The previous experiments demonstrated that higher concentrations of daptomycin were required for bactericidal activity against stationary-phase S. aureus cells compared with exponentially growing cells. Therefore, daptomycin at 100 µg/ml and similarly high concentrations of comparator antibiotics were examined for activity in high inoculum density stationary-phase cultures in MHBc. Stationary-phase S. aureus cultures were left untreated or were treated separately with ciprofloxacin, daptomycin, gentamicin, nafcillin, or vancomycin at 100 µg/ml. Under these growth conditions, daptomycin and gentamicin retained bactericidal activity against stationary-phase cultures (Fig. 3). Daptomycin treatment resulted in an immediate and progressive reduction in viability, demonstrating a 3-log reduction in the number of CFU per milliliter after 8 h. Gentamicin required a longer time (24 h) to achieve a bactericidal effect; no change in viability was seen for the first 4 h of exposure. In contrast, under these conditions, ciprofloxacin, nafcillin, and vancomycin did not achieve the bactericidal end point. At the 24-h time point, cultures treated with these three drugs had cell densities equal to those of untreated control cultures (Fig. 3).

View larger version (19K): [in this window] [in a new window]

FIG. 3. Effect of exposing stationary-phase S. aureus cultures to either daptomycin or a comparator antibiotic at 100 µg/ml. DAP, daptomycin; NAF, nafcillin; CIP, ciprofloxacin; GEN, gentamicin; VAN, vancomycin.

Effects of S. aureus physiology on the bactericidal activity of daptomycin. The previous experiment demonstrates that daptomycin retains bactericidal activity against stationary-phase cultures but that elevated levels of the antibiotic are required to achieve the 3-log reduction end point. The requirement for higher drug concentrations could be due to the well-documented inoculum effect (29) or to changes in cellular physiology in stationary-phase cultures. To address this, exponential- and stationary-phase bacterial cultures were prepared at high cell densities (5 x 10⁹ CFU/ml) for treatment with a low (10 µg/ml) or high (100 µg/ml) concentration of daptomycin. The lower daptomycin concentration had a minimal effect on high inoculum density stationary-phase S. aureus over the 24-h experiment, while the same concentration was bactericidal against high-density exponentially growing cells over the same time period (Fig. 4A). The high daptomycin concentration (100 µg/ml) had a very rapid bactericidal effect on high-density exponentially growing cells (a >3-log decrease in cell density in 1 h), but in stationary-phase high-density cells, the same concentration was bactericidal only after a prolonged treatment period (Fig. 4A).

View larger version (23K): [in this window] [in a new window]

FIG. 4. (A) Effect of cell physiology on the dose response and bactericidal activity of daptomycin against concentrated exponential-phase (Expo.) and stationary-phase (Stat.) S. aureus. (B) Log reductions of dilute and concentrated stationary-phase cultures. Cells were suspended in depMHBc and treated with either a high (100-µg/ml) or a low (10-µg/ml) concentration of daptomycin (DAP).

To test if altered cell physiology mediates reduced daptomycin susceptibility in stationary-phase S. aureus, a culture dilution experiment was performed. Stationary-phase cells were standardized to an OD₆₀₀ of 3.75 and then diluted in depMHBc to an OD₆₀₀ of 0.1. Diluted cultures were treated with a low (10 µg/ml) or high (100 µg/ml) concentration of daptomycin and compared with concentrated stationary-phase cells. The lower daptomycin concentration (10 µg/ml) had no bactericidal effect on concentrated stationary-phase cells over 24 h; however, the diluted cell cultures had some susceptibility (a >1-log decrease in the number of CFU per milliliter) to the lower daptomycin concentration (Fig. 4B). In contrast, the higher daptomycin concentration (100 µg/ml) was bactericidal in both concentrated and diluted S. aureus cultures in stationary phase, although the bactericidal effect was more pronounced in diluted cultures. Taken together, these data suggest that under these experimental conditions, S. aureus cell physiology influences daptomycin susceptibility more than inoculum density does.

Effects of daptomycin and ciprofloxacin on metabolically arrested S. aureus. To determine the effect of artificial metabolic arrest on antibiotic susceptibility, exponentially growing cells were treated with a proton ionophore (CCCP) to abrogate cellular energy production or a protein synthesis inhibitor (erythromycin) prior to antibiotic treatment. Untreated control cultures grew almost 3 log units during the 4-h experiment (Fig. 5A and B), while treatment with CCCP or erythromycin stopped the proliferation of bacteria over 24 h (data not shown). Control cultures treated with daptomycin or ciprofloxacin at 10 µg/ml were effectively killed (Fig. 5A and B). Daptomycin also demonstrated a bactericidal effect against S. aureus in cultures growth arrested by both CCCP and erythromycin (Fig. 5A). However, CCCP- and erythromycin-mediated metabolic arrest protected S. aureus from the bactericidal action of ciprofloxacin (Fig. 5B).

View larger version (22K): [in this window] [in a new window]

FIG. 5. Effect of daptomycin on chemically arrested S. aureus. Exponentially growing cells were chemically arrested by a 1-h pretreatment with CCCP (a proton ionophore) or erythromycin (a protein synthesis inhibitor). Cells were treated with either daptomycin (A) or ciprofloxacin (B) at 10 µg/ml. CIP, ciprofloxacin; DAP, daptomycin; ERY, erythromycin.

Effects of daptomycin, ciprofloxacin, and nafcillin on S. aureus cultures growth arrested by cold temperature. The effects of antibiotics on S. aureus cultures growth arrested by cold temperature were also examined. Exponentially growing cultures were chilled in an ice bath prior to treatment with ciprofloxacin, daptomycin, or nafcillin at 2 µg/ml. Time-kill curves demonstrate that daptomycin remained bactericidal against cold-arrested S. aureus, while cold arrest protected the cells against ciprofloxacin or nafcillin (Fig. 6). After 24 h on ice, cultures were incubated at 37°C and each of the three antibiotics was active and effectively demonstrated activity (Fig. 6).

View larger version (16K): [in this window] [in a new window]

FIG. 6. Bactericidal activities of daptomycin, ciprofloxacin, and nafcillin (each at 2 µg/ml) against S. aureus that has been growth arrested by ice bath submersion. After 24 h, bacteria were warmed to 37°C. DAP, daptomycin; CIP, ciprofloxacin; NAF, nafcillin.

Top ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 
Daptomycin is a cyclic lipopeptide antibacterial agent that exhibits rapid, concentration-dependent in vitro bactericidal activity against clinically significant strains of gram-positive pathogens, including streptococci, methicillin-resistant S. aureus, and vancomycin-resistant enterococci (5, 13, 18, 31). In the present study, the observed bactericidal activity against stationary-phase S. aureus ATCC 29213 is remarkable since most other bactericidal agents interfere with cell division or processes necessary for cell division and typically have little activity against nondividing cells (30).

The mechanism of action of daptomycin has not been fully elucidated, despite 20 years of study. Early efforts (3) suggested that the drug inhibited an early step in peptidoglycan synthesis, although no particular reaction was identified. Other authors (7) have suggested that daptomycin targets lipoteichoic acid biosynthesis. Subsequently, it has been shown that daptomycin becomes inserted into the bacterial plasma membrane in a calcium-dependent fashion, leading to membrane depolarization, release of intracellular potassium ions, and rapid cell death (1, 26). Since depolarization leads to a loss of transport and other metabolic activity, inhibition of lipoteichoic acid and peptidoglycan biosynthesis is a consequence of the primary mechanism (2, 22).

The ability of daptomycin to kill bacteria whose metabolism has been arrested by chemical treatment (dissipation of proton motive force by CCCP or inhibition of protein synthesis by erythromycin), cold, or nutrient depletion further supports the idea that inhibition of macromolecular synthesis is a secondary component of the bactericidal mechanism of action. Recent studies with artificial membranes (19, 26, 28) have suggested that daptomycin insertion causes positive-curvature strain of the membrane, which would ultimately result in cell leakage (28). Although this process has not been demonstrated to occur in vivo, it is consistent with the ability of daptomycin to kill growth-arrested bacteria.

In the present study, antibiotics with different mechanisms of action were selected as comparators. Ciprofloxacin blocks DNA replication, and nafcillin and vancomycin perturb cell wall synthesis, while gentamicin binds to the 30S ribosomal subunit, causing bacterial mRNA to be misread (30). As would be expected, ciprofloxacin, nafcillin, and vancomycin were not bactericidal against stationary-phase S. aureus cultures in all of the growth arrest models tested. Interestingly, gentamicin was bactericidal in stationary-phase cells at the 24-h time point. This may be a result of gentamicin's ability to disrupt the cell membrane as it is transported into the cell prior to acting on bacterial ribosomes (6).

It is well established that a high inoculum density can deter the effectiveness of many antibiotics (29). In the experiments presented here, inoculum effects were observed but appeared to have less of an impact on the effectiveness of daptomycin than did cell physiology or the growth state. The reason for this result is unknown. It is possible, however, that positive-curvature strain in the membrane is increased during bacterial cell division, resulting in more rapid bactericidal activity. In some S. aureus strains, the bactericidal activity of oleic acid has differential effects, depending on the phase of growth, that appear to coincide with changes in membrane fluidity (33).

A previous study by Lamp et al. had shown that in the presence of human sera, daptomycin was bactericidal in stationary-phase and exponentially growing isolates in vitro and exhibited statistically significantly faster killing rates than vancomycin against both methicillin-sensitive and methicillin-resistant strains (25). In each case, daptomycin and vancomycin were more effective against exponentially growing cultures than against stationary-phase cultures (25). In the present study, daptomycin had bactericidal activity against stationary-phase S. aureus at concentrations (32 µg/ml) approximately 15 times that which is effective against exponentially growing cells (2 µg/ml). High concentrations of daptomycin (minimum bactericidal concentration, 32 µg/ml) have shown bactericidal activity against adherent S. epidermidis (21) in an adherent-cell biofilm model. Interestingly, many of the genes characterized as assisting in bacterial survival in stationary phase, such as Clp homologues, are expressed by bacteria within biofilms (8); these similarities in physiology may translate into similarities in susceptibility.

In conclusion, the data suggest that the bactericidal activity of daptomycin in vitro does not require cell division or active metabolism, consistent with the proposed mechanism of disrupting membrane function. The activity of daptomycin, at high concentrations, against nongrowing cells might also indicate the alternative possibility that daptomycin possesses two mechanisms of action—one active against exponential-phase cultures at low concentrations (near the standard MIC) and one active against stationary-phase cells and observed only at high concentrations of the drug. We have no evidence that would support such a dual mechanism, but the possibility cannot be ruled out by the studies reported here.

The demonstration of daptomycin's bactericidal effect in stationary-phase bacteria may have implications for the additional in vivo use of this drug. Daptomycin was recently approved by the Food and Drug Administration for the treatment of S. aureus bacteremia and right-sided infective endocarditis (10, 12). The limitation of these daptomycin experiments is similar to that of many other in vitro studies; experimental conditions do not necessarily reflect those in human infections. However, these studies are valuable in gaining further understanding regarding the mechanism of its action. The present study suggests that further investigations of daptomycin in infection models involving stationary or slowly growing gram-positive bacteria would be of great interest.

 
We thank the other members of the Discovery Microbiology group (past and present) and the Drug Evaluation group of Cubist Pharmaceuticals, Inc., for their support, scientific discussion, and assistance with all aspects of this research.

We are all Cubist employees and own stock options.

 
* Corresponding author. Mailing address: Cubist Pharmaceuticals, Inc., 65 Hayden Avenue, Lexington, MA 02421. Phone: (781) 860-8525. Fax: (781) 861-1164. E-mail: carmela.mascio{at}cubist.com

Published ahead of print on 8 October 2007.

Top ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 

1
1. Alborn, W. E., Jr., N. E. Allen, and D. A. Preston. 1991. Daptomycin disrupts membrane potential in growing Staphylococcus aureus. Antimicrob. Agents Chemother. 35:2282-2287.[Abstract/Free Full Text]
2
2. Allen, N. E., W. E. Alborn, Jr., and J. N. Hobbs, Jr. 1991. Inhibition of membrane potential-dependent amino acid transport by daptomycin. Antimicrob. Agents Chemother. 35:2639-2642.[Abstract/Free Full Text]
3
3. Allen, N. E., J. N. Hobbs, and W. E. Alborn, Jr. 1987. Inhibition of peptidoglycan biosynthesis in gram-positive bacteria by LY146032. Antimicrob. Agents Chemother. 31:1093-1099.[Abstract/Free Full Text]
4
4. Anderl, J. N., J. Zahller, F. Roe, and P. S. Stewart. 2003. Role of nutrient limitation and stationary-phase existence in Klebsiella pneumoniae biofilm resistance to ampicillin and ciprofloxacin. Antimicrob. Agents Chemother. 47:1251-1256.[Abstract/Free Full Text]
5
5. Barry, A. L., P. C. Fuchs, and S. D. Brown. 2001. In vitro activities of daptomycin against 2,789 clinical isolates from 11 North American medical centers. Antimicrob. Agents Chemother. 45:1919-1922.[Abstract/Free Full Text]
6
6. Bryan, L. E., and S. Kwan. 1983. Roles of ribosomal binding, membrane potential, and electron transport in bacterial uptake of streptomycin and gentamicin. Antimicrob. Agents Chemother. 23:835-845.[Abstract/Free Full Text]
7
7. Canepari, P., M. Boaretti, M. M. Lleo, and G. Satta. 1990. Lipoteichoic acid as a new target for activity of antibiotics: mode of action of daptomycin (LY146032). Antimicrob. Agents Chemother. 34:1220-1226.[Abstract/Free Full Text]
8
8. Chatterjee, I., P. Becker, M. Grundmeier, M. Bischoff, G. A. Somerville, G. Peters, B. Sinha, N. Harraghy, R. A. Proctor, and M. Herrmann. 2005. Staphylococcus aureus ClpC is required for stress resistance, aconitase activity, growth recovery, and death. J. Bacteriol. 187:4488-4496.[Abstract/Free Full Text]
9
9. Clinical and Laboratory Standards Institute. 2005. Performance standards for antimicrobial susceptibility testing. Fifteenth informational supplement, vol. 25, no. 1. Clinical and Laboratory Standards Institute, Wayne, PA.
10
10. Cubist Pharmaceuticals. 2006. Cubicin (daptomycin for injection) prescribing information. Cubist Pharmaceuticals, Lexington, MA.
11
11. Donlan, R. M., and J. W. Costerton. 2002. Biofilms: survival mechanisms of clinically relevant microorganisms. Clin. Microbiol. Rev. 15:167-193.[Abstract/Free Full Text]
12
12. Fowler, V. G., Jr., H. W. Boucher, R. Corey, E. Abrutyn, A. W. Karchmer, M. E. Rupp, D. P. Levine, H. F. Chambers, F. P. Tally, G. A. Vigliani, C. H. Cabell, A. S. Link, I. Demeyer, S. G. Filler, M. Zervos, P. Cook, J. Parsonnet, J. M. Bernstein, C. S. Price, G. N. Forrest, G. Fatkenheuer, M. Gareca, S. J. Rehm, H. R. Brodt, A. Tice, and S. E. Cosgrove. 2006. Daptomycin versus standard therapy for bacteremia and endocarditis caused by Staphylococcus aureus. N. Engl. J. Med. 355:653-665.[Abstract/Free Full Text]
13
13. Fuchs, P. C., A. L. Barry, and S. D. Brown. 2002. In vitro bactericidal activity of daptomycin against staphylococci. J. Antimicrob. Chemother. 49:467-470.[Abstract/Free Full Text]
14
14. Fux, C. A., S. Wilson, and P. Stoodley. 2004. Detachment characteristics and oxacillin resistance of Staphylococcus aureus biofilm emboli in an in vitro catheter infection model. J. Bacteriol. 186:4486-4491.[Abstract/Free Full Text]
15
15. Gradelski, E., B. Kolek, D. Bonner, and J. Fung-Tomc. 2002. Bactericidal mechanism of gatifloxacin compared with other quinolones. J. Antimicrob. Chemother. 49:185-188.[Abstract/Free Full Text]
16
16. Hall-Stoodley, L., J. W. Costerton, and P. Stoodley. 2004. Bacterial biofilms: from the natural environment to infectious diseases. Nat. Rev. Microbiol. 2:95-108.[CrossRef][Medline]
17
17. Herbert, D., C. N. Paramasivan, P. Venkatesan, G. Kubendiran, R. Prabhakar, and D. A. Mitchison. 1996. Bactericidal action of ofloxacin, sulbactam-ampicillin, rifampin, and isoniazid on logarithmic- and stationary-phase cultures of Mycobacterium tuberculosis. Antimicrob. Agents Chemother. 40:2296-2299.[Abstract]
18
18. Hermsen, E. D., L. B. Hovde, J. R. Hotchkiss, and J. C. Rotschafer. 2003. Increased killing of staphylococci and streptococci by daptomycin compared with cefazolin and vancomycin in an in vitro peritoneal dialysate model. Antimicrob. Agents Chemother. 47:3764-3767.[Abstract/Free Full Text]
19
19. Jung, D., A. Rozek, M. Okon, and R. E. Hancock. 2004. Structural transitions as determinants of the action of the calcium-dependent antibiotic daptomycin. Chem. Biol. 11:949-957.[CrossRef][Medline]
20
20. Kolter, R., D. A. Siegele, and A. Tormo. 1993. The stationary phase of the bacterial life cycle. Annu. Rev. Microbiol. 47:855-874.[CrossRef][Medline]
21
21. Labthavikul, P., P. J. Petersen, and P. A. Bradford. 2003. In vitro activity of tigecycline against Staphylococcus epidermidis growing in an adherent-cell biofilm model. Antimicrob. Agents Chemother. 47:3967-3969.[Abstract/Free Full Text]
22
22. Laganas, V., J. Alder, and J. A. Silverman. 2003. In vitro bactericidal activities of daptomycin against Staphylococcus aureus and Enterococcus faecalis are not mediated by inhibition of lipoteichoic acid biosynthesis. Antimicrob. Agents Chemother. 47:2682-2684.[Abstract/Free Full Text]
23
23. Lakey, J. H., and E. J. Lea. 1986. The role of acyl chain character and other determinants on the bilayer activity of A21978C an acidic lipopeptide antibiotic. Biochim. Biophys. Acta 859:219-226.[Medline]
24
24. Lakey, J. H., and M. Ptak. 1988. Fluorescence indicates a calcium-dependent interaction between the lipopeptide antibiotic LY146032 and phospholipid membranes. Biochemistry 27:4639-4645.[CrossRef][Medline]
25
25. Lamp, K. C., M. J. Rybak, E. M. Bailey, and G. W. Kaatz. 1992. In vitro pharmacodynamic effects of concentration, pH, and growth phase on serum bactericidal activities of daptomycin and vancomycin. Antimicrob. Agents Chemother. 36:2709-2714.[Abstract/Free Full Text]
26
26. Silverman, J. A., N. G. Perlmutter, and H. M. Shapiro. 2003. Correlation of daptomycin bactericidal activity and membrane depolarization in Staphylococcus aureus. Antimicrob. Agents Chemother. 47:2538-2544.[Abstract/Free Full Text]
27
27. Somerville, G. A., M. S. Chaussee, C. I. Morgan, J. R. Fitzgerald, D. W. Dorward, L. J. Reitzer, and J. M. Musser. 2002. Staphylococcus aureus aconitase inactivation unexpectedly inhibits exponential-phase growth and enhances stationary-phase survival. Infect. Immun. 70:6373-6382.[Abstract/Free Full Text]
28
28. Straus, S. K., and R. E. Hancock. 2006. Mode of action of the new antibiotic for gram-positive pathogens daptomycin: comparison with cationic antimicrobial peptides and lipopeptides. Biochim. Biophys. Acta 1758:1215-1223.[Medline]
29
29. Tedesco, K., and M. J. Rybak. 2003. Impact of high inoculum Staphylococcus aureus on the activities of nafcillin (NAF), vancomycin (VAN), linezolid (LZD), gentamicin (GEN), and daptomycin (DAP) in an in vitro pharmacodynamic model (IVPD), abstr. A-1151. In 43rd Intersci. Conf. Antimicrob. Agents Chemother. American Society for Microbiology, Washington, D.C.
30
30. Tenover, F. C. 2006. Mechanisms of antimicrobial resistance in bacteria. Am. J. Med. 119:S3-S10.[Medline]
31
31. Wise, R., J. M. Andrews, and J. P. Ashby. 2001. Activity of daptomycin against gram-positive pathogens: a comparison with other agents and the determination of a tentative breakpoint. J. Antimicrob. Chemother. 48:563-567.[Abstract/Free Full Text]
32
32. Xie, Z., N. Siddiqi, and E. J. Rubin. 2005. Differential antibiotic susceptibilities of starved Mycobacterium tuberculosis isolates. Antimicrob. Agents Chemother. 49:4778-4780.[Abstract/Free Full Text]
33
33. Xiong, Z., S. Ge, N. R. Chamberlain, and F. A. Kapral. 1993. Growth cycle-induced changes in sensitivity of Staphylococcus aureus to bactericidal lipids from abscesses. J. Med. Microbiol. 39:58-63.[Abstract/Free Full Text]

---

Antimicrobial Agents and Chemotherapy, December 2007, p. 4255-4260, Vol. 51, No. 12
0066-4804/07/$08.00+0     doi:10.1128/AAC.00824-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

This article has been cited by other articles:

• Ooi, N., Miller, K., Randall, C., Rhys-Williams, W., Love, W., Chopra, I. (2009). XF-70 and XF-73, novel antibacterial agents active against slow-growing and non-dividing cultures of Staphylococcus aureus including biofilms. J Antimicrob Chemother 0: dkp409v1-dkp409 [Abstract] [Full Text]  
• Murillo, O., Garrigos, C., Pachon, M. E., Euba, G., Verdaguer, R., Cabellos, C., Cabo, J., Gudiol, F., Ariza, J. (2009). Efficacy of High Doses of Daptomycin versus Alternative Therapies against Experimental Foreign-Body Infection by Methicillin-Resistant Staphylococcus aureus. Antimicrob. Agents Chemother. 53: 4252-4257 [Abstract] [Full Text]  
• Weiss, E. C., Zielinska, A., Beenken, K. E., Spencer, H. J., Daily, S. J., Smeltzer, M. S. (2009). Impact of sarA on Daptomycin Susceptibility of Staphylococcus aureus Biofilms In Vivo. Antimicrob. Agents Chemother. 53: 4096-4102 [Abstract] [Full Text]  
• Wu, M., von Eiff, C., Al Laham, N., Tsuji, B. T. (2009). Vancomycin and Daptomycin Pharmacodynamics Differ against a Site-Directed Staphylococcus epidermidis Mutant Displaying the Small-Colony-Variant Phenotype. Antimicrob. Agents Chemother. 53: 3992-3995 [Abstract] [Full Text]  
• Weiss, E. C., Spencer, H. J., Daily, S. J., Weiss, B. D., Smeltzer, M. S. (2009). Impact of sarA on Antibiotic Susceptibility of Staphylococcus aureus in a Catheter-Associated In Vitro Model of Biofilm Formation. Antimicrob. Agents Chemother. 53: 2475-2482 [Abstract] [Full Text]  
• Begic, D., von Eiff, C., Tsuji, B. T. (2009). Daptomycin pharmacodynamics against Staphylococcus aureus hemB mutants displaying the small colony variant phenotype. J Antimicrob Chemother 63: 977-981 [Abstract] [Full Text]  
• Hachmann, A.-B., Angert, E. R., Helmann, J. D. (2009). Genetic Analysis of Factors Affecting Susceptibility of Bacillus subtilis to Daptomycin. Antimicrob. Agents Chemother. 53: 1598-1609 [Abstract] [Full Text]  
• Belley, A., Neesham-Grenon, E., McKay, G., Arhin, F. F., Harris, R., Beveridge, T., Parr, T. R. Jr., Moeck, G. (2009). Oritavancin Kills Stationary-Phase and Biofilm Staphylococcus aureus Cells In Vitro. Antimicrob. Agents Chemother. 53: 918-925 [Abstract] [Full Text]  
• Camargo, I. L. B. d. C., Neoh, H.-M., Cui, L., Hiramatsu, K. (2008). Serial Daptomycin Selection Generates Daptomycin-Nonsusceptible Staphylococcus aureus Strains with a Heterogeneous Vancomycin-Intermediate Phenotype. Antimicrob. Agents Chemother. 52: 4289-4299 [Abstract] [Full Text]  
• Hawkey, P. M. (2008). Pre-clinical experience with daptomycin. J Antimicrob Chemother 62: iii7-iii14 [Abstract] [Full Text]  
• Livermore, D. M. (2008). Future directions with daptomycin. J Antimicrob Chemother 62: iii41-iii49 [Abstract] [Full Text]  
• Cotroneo, N., Harris, R., Perlmutter, N., Beveridge, T., Silverman, J. A. (2008). Daptomycin Exerts Bactericidal Activity without Lysis of Staphylococcus aureus. Antimicrob. Agents Chemother. 52: 2223-2225 [Abstract] [Full Text]  
• Baltch, A. L., Ritz, W. J., Bopp, L. H., Michelsen, P., Smith, R. P. (2008). Activities of Daptomycin and Comparative Antimicrobials, Singly and in Combination, against Extracellular and Intracellular Staphylococcus aureus and Its Stable Small-Colony Variant in Human Monocyte-Derived Macrophages and in Broth. Antimicrob. Agents Chemother. 52: 1829-1833 [Abstract] [Full Text]  

This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Mascio, C. T. M. Articles by Silverman, J. A. Search for Related Content PubMed PubMed Citation Articles by Mascio, C. T. M. Articles by Silverman, J. A.