Comparative in vitro activity of telithromycin (HMR 3647), three macrolides, amoxycillin, cefdinir and levofloxacin against Gram-positive clinical isolates in Japan

Hiroki Okamoto*, Shuichi Miyazaki, Kazuhiro Tateda, Yoshikazu Ishii and Keizo Yamaguchi

Department of Microbiology, Toho University School of Medicine, Omori-Nishi, Ota-ku, Tokyo, Japan


    Abstract
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 Abstract
 Introduction
 Materials and methods
 Results and discussion
 References
 
The in vitro activities (MIC and MBC) of telithromycin (HMR 3647) against clinical isolates in Japan were investigated in comparison with those of erythromycin A, clarithromycin, azithromycin, amoxycillin, cefdinir and levofloxacin. Telithromycin was more potent than the reference compounds against erythromycin A-susceptible or resistant Streptococcus pneumoniae isolates possessing either mef (mefA or mefE) genes or the ermB gene. Against erythromycin A-susceptible or inducibly resistant Staphylococcus aureus and erythromycin A-susceptible and intermediate Enterococcus faecalis, telithromycin was highly active.


    Introduction
 Top
 Abstract
 Introduction
 Materials and methods
 Results and discussion
 References
 
Telithromycin (HMR 3647) is a new ketolide antibacterial agent synthesized by chemical modification of erythromycin A. Many investigations have demonstrated that telithromycin shows good in vitro antibacterial activity against Gram-positive bacteria, including erythromycin A-resistant isolates of Streptococcus pneumoniae, an organism important in the aetiology of community-acquired respiratory tract infections.1,2

A recent survey shows that the rate of pneumococcal resistance to erythromycin A is high (36.3%) in Japan.3 Therefore, telithromycin might find a useful role in the treatment of respiratory tract infections in Japan.

In this study, we investigated the in vitro inhibitory (MIC) and bactericidal (MBC) activity of telithromycin against Gram-positive clinical isolates in Japan. Isolates were classified on the basis of their susceptibility to erythromycin A, or the phenotype or genotype of resistance to erythromycin A. The activity of telithromycin was compared with that of erythromycin A, clarithromycin, azithromycin, amoxycillin, cefdinir and levofloxacin.


    Materials and methods
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 Abstract
 Introduction
 Materials and methods
 Results and discussion
 References
 
Antimicrobial agents

Antimicrobial agents were kindly provided by the sources indicated: telithromycin (Hoechst Marion Roussel Co., Ltd, Tokyo, Japan); erythromycin A (Shionogi Pharmaceutical Co., Ltd, Osaka, Japan); clarithromycin (Taisho Pharmaceutical Co., Ltd, Tokyo, Japan); clindamycin (Upjohn Co., Ltd, Tokyo, Japan); azithromycin (Pfizer Laboratories, Groton, CT, USA); amoxycillin (Sigma Chemical, St Louis, MO, USA); cefdinir (Fujisawa Pharmaceutical Co., Osaka, Japan); levofloxacin (Daiichi Pharmaceutical Co. Ltd, Tokyo, Japan).

Bacterial isolates

The bacterial isolates used in this study were isolated in Toho University Hospital from 1994–98.

MIC and MBC determination

The MICs were determined by the broth microdilution method4 using cation-adjusted Mueller–Hinton broth (Difco Laboratories, Detroit, MI, USA) for staphylococci. For streptococci and enterococci the broth was supplemented with 5% lysed horse blood plus 5 g/L yeast extract (Oxoid, Hampshire, UK) and 15 mg/L NAD (Sigma Chemical). The breakpoints for susceptibility classification were those specified by the NCCLS.5 Resistant strains of Staphylococcus aureus were subdivided into inducible and constitutive phenotypes on the basis of the agar diffusion test with discs for erythromycin and clindamycin as described by Hamilton-Miller.6

MBC was determined by subculture of 10 µL of broth from each well without visible growth on brain–heart infusion agar (Difco) plates for S. aureus, and blood agar plates for S. pneumoniae and Enterococcus faecalis, and was defined as the lowest concentration of each drug that exhibited >99.9% reduction in growth.

DNA isolation and detection of ermB and mef (mefA or mefE) genes

Genomic DNA was extracted according to the instructions provided with SepaGene (Sanko junyaku Co., Tokyo, Japan). Macrolide-resistance genes were detected by the PCR method as described by Sutcliffe et al.7 Strain 3585 (ermB) and strain 02J1175 (mefE), kindly provided by Dr Joyce de Azavedo, University of Toronto, were used as positive controls. It has been reported that the mefA and mefE genes mediate resistance by excretion of the drug, while ermB gene resistance is due to the presence of a 50S ribosome subunit mutation.7,8 A further distinction of mef genes into mefA and mefE was not possible with the methodology used.


    Results and discussion
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 Abstract
 Introduction
 Materials and methods
 Results and discussion
 References
 
The 100 isolates of S. pneumoniae included 32 erythromycin A-susceptible isolates and 67 erythromycin Aresistant isolates (Table IGo). In addition, the erythromycin A-resistant isolates included two isolates that possessed both the mef and ermB resistance genes, 32 isolates that possessed only the mef genes and 33 isolates that possessed only the ermB gene. The MIC90 of telithromycin was 0.125 mg/L for isolates carrying either of these resistance genes, although this value was 16 times higher than that (0.008 mg/L) for the erythromycin A-susceptible isolates. The MIC90s of erythromycin A, clarithromycin and azithromycin were 2–4 mg/L for erythromycin A-resistant S. pneumoniae with only the mef genes and >128 mg/L for S. pneumoniae with only the ermB gene. The potent activity of telithromycin compared with that of the macrolides tested could be related either to the affinity of the drug for methylated ribosome and/or to a lack of capacity to produce inducible resistance.9 Nishijima et al.10 reported recently that almost all ermB+-mediated resistance was inducible and resistance in mef+ isolates was non-inducible. Therefore, the activity of telithromycin against ermB+ isolates might be associated with its poor activity as an inducer, and against mef+ isolates due to the high affinity of telithromycin for the target ribosome. The MIC90s of amoxycillin, cefdinir and levofloxacin for the erythromycin A-resistant S. pneumoniae isolates were 8–128 times higher than that of telithromycin.


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Table I. Antibacterial activity of telithromycin and reference compounds against clinical isolates
 
Erythromycin A-resistant isolates of Streptococcus pyogenes and Streptococcus agalactiae were not available for this study (Table IGo). Telithromycin showed excellent antibacterial activity against both isolates, with MIC90s of 0.008–0.016 mg/L and these activities were 2–16 times greater than those of the reference compounds tested.

Telithromycin had excellent antibacterial activity against erythromycin A-susceptible and inducible erythromycin A-resistant S. aureus with an MIC90 of 0.125 mg/L (Table IGo). These activities were greater than those of the reference compounds. However, telithromycin, in common with the macrolides tested, exhibited little activity against the S. aureus isolates with constitutive erythromycin A resistance. Against Staphylococcus epidermidis, the antibacterial activity of telithromycin was similar to that against S. aureus (data not shown).

Telithromycin was much more potent against enterococcal isolates than the reference compounds tested (Table IGo). In particular, the MIC90 of telithromycin was <=0.063 mg/L for erythromycin A-susceptible and intermediate E. faecalis. In addition, telithromycin showed activity, even against erythromycin A-resistant E. faecalis, which was >=32 times greater than those of the macrolides tested. Telithromycin was the most potent antibacterial agent against E. faecium among the compounds tested.

Telithromycin showed bactericidal activity against not only erythromycin A-susceptible isolates of S. pneumoniae, but also erythromycin A-resistant isolates that possessed only the mef genes. The MBC90/MIC90 ratio for those isolates was 1 (Table IIGo). However, against erythromycin A-resistant S. pneumoniae isolates that possessed the ermB gene, the MBC90/MIC90 ratio of telithromycin was 4. This result suggests that 50S ribosome mutation rather than excretion of the drug might influence the bactericidal activity of this compound.


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Table II. Bactericidal activities of telithromycin and reference compounds against clinical isolates
 
MBC90/MIC90 ratios of telithromycin for erythromycin A-susceptible and for inducible erythromycin A-resistant S. aureus were 16 and 8, respectively (Table IIGo). Against the E. faecalis isolates, telithromycin showed an MBC90/MIC90 ratio of 4 for the erythromycin A-intermediate isolates, whereas the ratio for the erythromycin A-resistant isolates was 32. These differences in antibacterial and/or bactericidal activities of telithromycin against Gram-positive cocci according to the species might be influenced by the binding of telithromycin to the target site. Amoxycillin, cefdinir and levofloxacin showed bactericidal activity against bacterial isolates tested in this study.

These data, in general, show close agreement with the results that have already been reported for in vitro antibacterial activity of telithromycin against clinical isolates in various geographical regions of the world.1,2

In this study, we confirmed that telithromycin was a very useful antimicrobial agent for the treatment of community-acquired respiratory tract infections in Japan, because this compound was active against erythromycin A-resistant S. pneumoniae, an organism that is frequently detected in Japan, posing serious therapeutic difficulties.


    Notes
 
* Correspondence address. Aventis Pharma Ltd Pharmacology, Lead Optimization, 1-3-2 Minamidai, Kawagoe, Saitama 350-1165, Japan. Tel: +81-492-43-2421; Fax: +81-492-43-4002; E-mail: hiroki.okamoto{at}aventis.com Back


    References
 Top
 Abstract
 Introduction
 Materials and methods
 Results and discussion
 References
 
1 . Boswell, F. J., Andrews, J. M., Ashby, J. P., Fogarty, C., Brenwald, N. P. & Wise, R. (1998). The in-vitro activity of HMR 3647, a new ketolide antimicrobial agent. Journal of Antimicrobial Chemotherapy 42, 703–9.[Abstract]

2 . Hamilton-Miller, J. M. & Shah, S. (1998). Comparative in-vitro activity of ketolide HMR 3647 and four macrolides against Gram-positive cocci of known erythromycin susceptibility status. Journal of Antimicrobial Chemotherapy 41, 649–53.[Abstract]

3 . Okamoto, H., Tateda K., Ishii Y., Matsumoto T., Kashitani, F., Miyazaki S. et al. (1999). High frequency of macrolide resistance and distribution of mefE and ermB genes in clinical isolates of Streptococcus pneumoniae in Japan. In Program and Abstracts of the Thirty-Ninth Interscience Conference on Antimicrobial Agents and Chemotherapy, San Francisco, 1999. Abstract 1222, p. 158. American Society for Microbiology, Washington, DC.

4 . Japan Society of Chemotherapy. (1993). Method for the determination of minimum inhibitory concentration (MIC) of aerobic bacteria by microdilution method. Chemotherapy (Tokyo) 41, 183–9

5 . National Committee for Clinical Laboratory Standards. (2000). Methods for Dilution Antimicrobial Susceptibility Tests for Bacteria that Grow Aerobically—Approved Standard. Fifth Edition M7-A5. NCCLS, Villanova, PA.

6 . Hamilton-Miller, J. M. (1992). In-vitro activities of 14-, 15- and 16-membered macrolides against Gram-positive cocci. Journal of Antimicrobial Chemotherapy 29, 141–7.[Abstract]

7 . Sutcliffe, J., Grebe, T., Tait-Kamradt, A. & Wondrack, L. (1996). Detection of erythromycin-resistant determinants by PCR. Antimicrobial Agents and Chemotherapy 40, 2562–6.[Abstract]

8 . Oster, P., Zanchi, A., Cresti, S., Lattanzi, M., Montagnani, F., Cellesi, C. et al. (1999). Patterns of macrolide resistance determinants among community-acquired Streptococcus pneumoniae isolates over a 5-year period of decreased macrolide susceptibility rates. Antimicrobial Agents and Chemotherapy 43, 2510–2.[Abstract/Free Full Text]

9 . Rosato, A., Vicarini, H., Bonnefoy, A., Chantot, J. F. & Leclercq, R. (1998). A new ketolide, HMR 3004, active against streptococci inducibly resistant to erythromycin. Antimicrobial Agents and Chemotherapy 42, 1392–6.[Abstract/Free Full Text]

10 . Nishijima, T., Saito, Y., Aoki, A., Toriya, M., Toyonaga, Y. & Fujii, R. (1999). Distribution of mefE and ermB genes in macrolide-resistant strains of Streptococcus pneumoniae and their variable susceptibility to various antibiotics. Journal of Antimicrobial Chemotherapy 43, 637–43.[Abstract/Free Full Text]

Received 1 February 2000; returned 22 May 2000; revised 15 June 2000; accepted 7 August 2000