In vitro activity of ertapenem against selected respiratory pathogens

A. Marchese*, L. Gualco, A. M. Schito, E. A. Debbia and G. C. Schito

Sezione di Microbiologia del Di.S.C.A.T., University of Genoa, Largo R.Benzi 10, 16132 Genoa, Italy

Received 14 May 2004; returned 4 July 2004; revised 31 August 2004; accepted 3 September 2004


    Abstract
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
 References
 
Objectives: The in vitro activity of ertapenem was evaluated in comparison to 21 selected agents against a large collection of recently isolated respiratory tract pathogens including: 180 Streptococcus pneumoniae, 100 Streptococcus pyogenes, 70 Haemophilus influenzae, 70 Moraxella catarrhalis, 100 methicillin-susceptible Staphylococcus aureus and 30 Klebsiella pneumoniae. Additional in vitro tests (time–kill curves with ertapenem alone and in combination with four other agents) for S. pneumoniae were carried out.

Methods: MIC determinations and time–kill curves were carried out following the procedures suggested by the NCCLS.

Results: According to NCCLS susceptibility breakpoints, ertapenem was comparable to the most potent compounds tested for all pathogens studied. Ertapenem was 100% active against penicillin-susceptible and -intermediate S. pneumoniae and against 60% of penicillin-resistant strains. Time–kill tests at 4x MIC confirmed a pronounced bactericidal potency of ertapenem against these organisms. Interactions of ertapenem with several other agents against pneumococci resulted in clear synergic interactions (98.4%). Indifference was extremely rare and antagonism was not observed. All S. pyogenes strains tested were inhibited by ertapenem, irrespective of their macrolide resistance phenotypes. Ertapenem was also fully active against H. influenzae (100% susceptible) and M. catarrhalis (MIC90 0.015–0.03 mg/L) even when capable of synthesizing ß-lactamases. Methicillin-susceptible S. aureus and K. pneumoniae, including extended-spectrum ß-lactamase-producing strains, were 100% susceptible to ertapenem.

Conclusions: Our results indicate that ertapenem has a suitable spectrum of activity against organisms encountered in community-acquired bacterial respiratory tract infections.

Keywords: carbapenems , time–kill , interactions , bactericidal activity , Streptococcus pneumoniae


    Introduction
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
 References
 
Respiratory tract infections are a major cause of morbidity and mortality worldwide.1 Therapeutic choices considered in most international guidelines include ß-lactams, macrolides and quinolones.2 However, during the past decade, antibiotic resistance among pathogens causing these infections has increased at an alarming rate, particularly penicillin and macrolide resistance.3 Alternative agents are therefore needed.

This study was designed to assess the activity of ertapenem, a new parenteral non-antipseudomonal carbapenem, against a large collection of recently isolated respiratory pathogens (550 strains). In addition, bactericidal activity of ertapenem against Streptococcus pneumoniae was determined by time–kill curve experiments. Similarly, the outcome of combining ertapenem with several other antimicrobial agents commonly employed for antipneumococcal therapy was determined.


    Materials and methods
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
 References
 
The following organisms, isolated from patients suffering from respiratory tract infections during 2001 in Italy, were studied: 180 S. pneumoniae (100 penicillin-susceptible, 50 penicillin-intermediate and 30 penicillin-resistant), 100 Streptococcus pyogenes, including 60 with well-characterized erythromycin resistance phenotypes (20 isolates each possessing the following phenotype: constitutive MLSB, inducible MLSB and M), 70 Haemophilus influenzae (20 ß-lactamase positive), 70 Moraxella catarrhalis (50 ß-lactamase positive), 100 methicillin-susceptible Staphylococcus aureus and 30 Klebsiella pneumoniae, including 10 extended-spectrum ß-lactamase (ESBL) producer strains.

Ertapenem was provided by Merck Sharp and Dohme (Rome, Italy). The comparator molecules were obtained from commercial sources (Sigma–Aldrich, Milan, Italy) or from their respective manufacturers.

MICs of ertapenem and of the other antimicrobial agents tested were determined using the microdilution method and interpreted according to the breakpoints suggested by the National Committee for Clinical Laboratory Standards.4

The antibacterial activity of ertapenem against pneumococci was assessed by carrying out time–kill curves following the NCCLS (1999) recommendations.5 Ertapenem (at a concentration corresponding to four times the MIC) was tested against 15 isolates of S. pneumoniae, characterized by the following phenotypes: three penicillin-susceptible (and susceptible to erythromycin, tetracycline, co-trimoxazole and chloramphenicol), three penicillin-intermediate (and susceptible to erythromycin, tetracycline, co-trimoxazole and/or chloramphenicol), three penicillin-resistant (and susceptible to erythromycin, tetracycline, co-trimoxazole and chloramphenicol), three erythromycin-resistant (and susceptible to penicillin, co-trimoxazole and chloramphenicol) and three multi-resistant strains (simultaneously penicillin-, co-trimoxazole- and chloramphenicol-resistant). Time–kill tests to determine the activity of ertapenem combined with other agents (vancomycin, clarithromycin, rifampicin and levofloxacin), at half their respective MIC against the same S. pneumoniae isolates detailed above, were carried out as described by the NCCLS.5


    Results
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
 References
 
MIC figures for all 550 strains studied are presented in Table 1.


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Table 1. In vitro activity of ertapenem and other selected comparative antimicrobial agents

 
Against penicillin-susceptible S. pneumoniae ertapenem, all the other ß-lactams, levofloxacin, vancomycin and rifampicin were fully active. Ertapenem also demonstrated an excellent activity (all strains susceptible) against penicillin-intermediate pneumococci. Towards high-level penicillin-resistant S. pneumoniae, a considerable proportion of the strains (60%) were susceptible to ertapenem. The activity of the compound was comparable to that of amoxicillin and co-amoxiclav (63.3%), but lower than that of cefotaxime (73.3%) and ceftriaxone (70.0%). As expected, against fully penicillin-resistant pneumococci, the most potent antimicrobial agents were non-ß-lactam agents such as levofloxacin and vancomycin (100% efficacy).

Time–kill curves at 4x MIC showed that after 6 h, ertapenem behaved as a bacteriostatic agent (90–99% killing) against 15 S. pneumoniae strains irrespective of their resistance traits. However, prolonged incubation gave rise to a consistent bactericidal activity against all S. pneumoniae strains, with 99.9% killing after 24 h of exposure (Figure 1).



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Figure 1. Bactericidal activity of ertapenem (4x MIC) against penicillin-susceptible (a), penicillin-intermediate (b), penicillin-resistant (c), erythromycin-resistant (d) and multi-resistant (e) S. pneumoniae strains. Controls untreated (filled squares) and treated strains (filled circles). The horizontal broken line indicates the limit of the assay. Each value represents the mean of nine experiments (three for each strain) ± S.D.

 
When ertapenem was tested in combination with other agents against 15 S. pneumoniae showing different phenotypes, synergy was the prevalent response obtained with all strains and in all tests (59/60). In no instance was antagonism observed. In particular, ertapenem in combination with vancomycin, levofloxacin and rifampicin constantly gave rise to synergic interactions (45/45 tests). With clarithromycin, against a macrolide-resistant S. pneumoniae indifference was observed.

Against erythromycin-susceptible S. pyogenes strains, ertapenem was the most potent agent tested together with other ß-lactams, levofloxacin, vancomycin, chloramphenicol and macrolides (100% susceptible strains), followed by tetracycline (87.5%). Similar results were obtained, macrolides excluded, for erythromycin-resistant strains, irrespective of the mechanism of resistance.

All antimicrobial agents studied, with the exception of ampicillin for ß-lactamase-producing strains, were highly active, in terms of MIC90, against M. catarrhalis. Ertapenem, meropenem, co-amoxiclav and rifampicin showed the lowest MIC90 for ß-lactamase-negative M. catarrhalis (0.015 mg/L), while against the ß-lactamase-positive organisms the MIC90 of ertapenem was two-fold higher (0.03 mg/L).

All ß-lactamase-negative H. influenzae isolates were susceptible to ertapenem, other ß-lactams, quinolones, azithromycin and rifampicin, whereas 96% of these isolates were susceptible to tetracycline, 88% to clarithromycin and co-trimoxazole and 86% to chloramphenicol. On the other hand, 100% of ß-lactamase-positive ampicillin-resistant strains were susceptible to ertapenem and all other antibiotics, excluding tetracycline (80%) and co-trimoxazole (90%).

Against methicillin-susceptible S. aureus, ertapenem, imipenem, the cephalosporins, co-amoxiclav, vancomycin and levofloxacin were active against 100% of the isolates followed by ciprofloxacin (99%), rifampicin (99%), chloramphenicol (98%), amikacin (98%), tetracycline (97%), co-trimoxazole (96%), gentamicin (91%), clindamycin (71%), clarithromycin (69%), azithromycin (69%), ampicillin (18%) and penicillin (18%).

Ertapenem was active against all K. pneumoniae, including ESBL-producers. With respect to these last microorganisms, ertapenem, imipenem, meropenem, co-amoxiclav and amikacin activity (all strains susceptible) exceeded that of tetracycline (80%), ciprofloxacin (30%) and co-trimoxazole (30%) and, as expected, extended-spectrum parenteral cephalosporins (0%).


    Discussion
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 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
 References
 
Ertapenem is a novel, non-antipseudomonal, long-acting, parenteral 1-ßmethyl-carbapenem licensed in the EU and in the USA for the treatment of different conditions, including intra-abdominal infections and community-acquired pneumonia.6 The results of this study support previous findings indicating that ertapenem exhibits a broad antimicrobial activity against contemporary respiratory pathogens independent of their resistance patterns.7,8 Ertapenem is stable against hydrolysis by a variety of ß-lactamases6 and, consistent with this feature, in this study it has been shown to retain full activity against ß-lactamase-positive H. influenzae and M. catarrhalis, and ESBL-producing K. pneumoniae.

Ertapenem, like other carbapenems, is efficacious against methicillin-susceptible staphylococci, but remains inactive against methicillin-resistant strains. Contrary to imipenem, ertapenem was fully active not only against penicillin-susceptible but also penicillin-intermediate S. pneumoniae. Ertapenem MIC90s were two-fold higher than those of imipenem, but in terms of percentage of susceptible strains, ertapenem is consistently more active than imipenem against pneumococci fully refractory to penicillin (60% versus 46.6%). On the contrary, ertapenem showed MIC90 values lower than those of cefotaxime and ceftriaxone, but since the NCCLS interpretative guidelines for non-meningeal isolates were adopted in this paper, the percentages of strains classified as susceptible (73.3% and 70.0%), rendered the third-generation injectable cephalosporins the most active ß-lactam agents tested. On the other hand, recent results from clinical trials with ertapenem versus ceftriaxone for the treatment of pneumococcal pneumonia, although on a limited number of patients, showed comparable clinical and bacteriological responses even for organisms for which penicillin MICs were ≥2 mg/L.9

A further confirmation of the good antipneumococcal activity of ertapenem has been provided here through the killing kinetics approach. Ertapenem was bactericidal against all pneumococci studied, irrespective of their penicillin or macrolide susceptibility phenotypes. The killing rates of ertapenem at 4x MIC were similar to those observed by other authors at 2x MIC.8 Increasing ertapenem concentrations did not therefore significantly influence the extent of killing, as expected due to ß-lactam general pharmacodynamics.

Given its characteristics, ertapenem has been approved in combination with a macrolide for initial empirical therapy of suspected community-acquired bacterial pneumonia in immunocompetent hospitalized adults.2 The fact that synergy often occurs when ertapenem interacts with other agents (clarithromycin, levofloxacin, rifampicin and vancomycin) suggested for the treatment of community-acquired pneumonia2 (CAP) even against penicillin-resistant pneumococci is a new finding, supporting its recent approval for such usage.

Although, at present, ertapenem has been licensed only for CAP, it can be predicted that its usage might be extended to other respiratory infections such as severe acute exacerbation of chronic bronchitis because of its appropriate spectrum.

Another potential role of ertapenem, as suggested by other authors, is in outpatient therapy.6 This last therapeutic approach could be of particular interest in Italy, a country where third-generation injectable cephalosporins have been vastly employed. In view of this peculiar situation, it has been speculated that the relatively low prevalence of resistant S. pneumoniae circulating in our country during the 1990s might have been related, at least in part, to aggressive intramuscular therapy.10 This hypothesis is supported by the recent increase in overall S. pneumoniae resistance in our country concomitant to governmental directions that limit the prescription of third-generation injectable cephalosporins in the community.

Outpatient intravenous therapy because of its clear pharmacokinetic advantages over the intramuscular route, may thus represent, if correctly and judiciously adopted, a new strategy in the struggle to achieve higher clinical success and eradication rates as well as providing the pharmacoeconomic advantages that stem from avoiding the costs of hospitalization.

Since the widespread use of carbapenems may promote the dissemination of resistant strains, it is mandatory to stress that prudent prescribing practices will be fundamental in maintaining the clinical utility of these ‘last resort’ antimicrobial agents.


    Acknowledgements
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
 References
 
This study was partially supported by Merck Sharp and Dohme (Rome, Italy).


    Footnotes
 
* Corresponding author. Tel: +39-010-3537655; Fax: +39-010-3537698; Email: anna.marchese{at}unige.it


    References
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 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
 References
 
1 . Fauci, A. S. (2001). Infectious diseases: considerations for the 21st century. Clinical Infectious Diseases 32, 675–85.[CrossRef][ISI][Medline]

2 . Mandell, L. A., Bartlett, J. G., Dowell, S. F. et al. (2003). Update of practice guidelines for the management of community-acquired pneumonia in immunocompetent adults. Clinical Infectious Diseases 37, 1405–33.[CrossRef][ISI][Medline]

3 . Felmingham, D. (2002). Evolving resistance patterns in community-acquired respiratory tract pathogens: first results from the PROTEKT global surveillance study. Prospective Resistant Organism Tracking and Epidemiology for the Ketolide Telithromycin. Journal of Infection 44, Suppl. A, 3–10.[ISI][Medline]

4 . National Committee for Clinical Laboratory Standards. (2004). Methods for Dilution Antimicrobial Susceptibility Tests for Bacteria that Grow Aerobically—Sixth Edition: Approved Standard M7-A6 and M100-S14. NCCLS, Wayne, PA, USA.

5 . National Committee for Clinical Laboratory Standards. (1999). Methods for Determining Bactericidal Activity of Antimicrobial Agents: Approved Guideline M26-A. NCCLS, Wayne, PA, USA.

6 . Livermore, D. M., Sefton, A. M. & Scott, G. M. (2003). Properties and potential of ertapenem. Journal of Antimicrobial Chemotherapy 52, 331–44.[Abstract/Free Full Text]

7 . Jones, R. N. (2001). In vitro evaluation of ertapenem (MK-0826), a long-acting carbapenem, tested against selected resistant strains. Journal of Chemotherapy 13, 363–76.[ISI][Medline]

8 . Pankuch, G. A., Davies, T. A., Jacobs, M. R. et al. (2002). Antipneumococcal activity of ertapenem (MK-0826) compared to those of other agents. Antimicrobial Agents and Chemotherapy 46, 42–6.[Abstract/Free Full Text]

9 . Ortiz-Ruiz, G., Vetter, N., Isaacs, R. et al. (2004). Ertapenem versus ceftriaxone for the treatment of community-acquired pneumonia in adults: combined analysis of two multicentre randomized, double-blind studies. Journal of Antimicrobial Chemotherapy 53, Suppl. S2, ii59–66.[Abstract/Free Full Text]

10 . Marchese, A., Mannelli, S., Tonoli, E. et al. (2001). Prevalence of antimicrobial resistance in Streptococcus pneumoniae circulating in Italy: results of the Italian Epidemiological Observatory Survey (1997–1999). Microbial Drug Resistance 7, 277–87.[CrossRef][ISI][Medline]