In vitro bactericidal activity of ABT-773 and amoxicillin against erythromycin-susceptible and -resistant strains of Streptococcus pyogenes

Susan L. Pendlanda,*, Melinda M. Neuhauserb and Jennifer L. Prausea

a The University of Illinois at Chicago, College of Pharmacy, Department of Pharmacy Practice, Microbiology Research Laboratory, 833 South Wood Street, Chicago, IL 60612; b The University of Houston College of Pharmacy, Department of Clinical Sciences and Administration, Houston, TX, USA


    Abstract
 Top
 Abstract
 Introduction
 Materials and methods
 Results and discussion
 Acknowledgements
 References
 
The bactericidal activity of ABT-773 was compared with amoxicillin against 10 clinical isolates of S. pyogenes (six erythromycin susceptible and four erythromycin resistant). The MIC ranges (mg/L) were 0.004–0.25 of ABT-773 and 0.015–0.12 of amoxicillin. At 24 h, ABT-773 concentrations of 2 x MIC and 8 x MIC were bactericidal against three and six organisms, respectively. In comparison, amoxicillin was bactericidal against all 10 organisms at both test concentrations.


    Introduction
 Top
 Abstract
 Introduction
 Materials and methods
 Results and discussion
 Acknowledgements
 References
 
Streptococcus pyogenes (Group A streptococci) is the most common cause of bacterial pharyngitis. The organism is also frequently isolated in skin and skin-structure infections. Penicillin is generally considered the drug of choice for treatment of pharyngitis and other non-invasive streptococcal infections, with macrolides recommended as alternative agents in penicillin-allergic patients.1 While S. pyogenes remains uniformly susceptible to penicillin, there have been reports of increasing erythromycin resistance over the past decade.2

The ketolides, semisynthetic 14-membered ring macrolides, represent a new subclass of agents among the macrolide–lincosamide–streptogramin group. One of the newest agents, ABT-773, has demonstrated excellent in vitro activity against both erythromycin-susceptible and -resistant strains of S. pyogenes.3 While the MIC data look very promising, additional information is needed on the time–kill kinetics of these new agents. The purpose of this study was to compare the bactericidal activity of ABT-773 and amoxicillin against S. pyogenes in vitro.


    Materials and methods
 Top
 Abstract
 Introduction
 Materials and methods
 Results and discussion
 Acknowledgements
 References
 
MIC and time–kill assays were performed on 10 clinical isolates of S. pyogenes (six erythromycin susceptible and four erythromycin resistant). The erythromycin-susceptible strains were isolated from patients at the University of Illinois at Chicago Medical Center (Chicago, IL, USA). Erythromycin-resistant strains (two mefA and two ermTR) were kindly provided by Drs Michael Jacobs and Elizabeth Palavecino (Case Western Reserve University, Cleveland, OH, USA). The erythromycin-resistant isolates had been collected from patients at University hospitals in Cleveland. The control strain, S. pneumoniae ATCC 49619, was used for validation of MIC results.

ABT-773 (Abbott Laboratories, Abbott Park, IL, USA), amoxicillin and erythromycin (United States Pharmacopeia, Rockville, MD, USA) powders were prepared according to NCCLS guidelines or manufacturer's recommendations.4 Cation-supplemented Mueller–Hinton broth with 3% lysed horse blood (Remel, Lenexa, KS, USA) was the medium used for the MIC and time–kill assays. MICs were performed in duplicate following NCCLS guidelines using the broth microdilution method.4

The bactericidal activity of the antimicrobial agents was determined in duplicate using the time–kill method following NCCLS guidelines.5 Antimicrobial concentrations tested were 2 x and 8 x MIC. Sampling for colony counts was performed at 0, 2, 6 and 24 h. Antibiotic carryover was prevented by saline dilutions. Viable counts were determined after 24 h of incubation at 35°C with 5% CO2. The rate and extent of killing were determined by plotting colony counts (log10 cfu/mL) against time (h). Bactericidal activity was defined as a >=3 log10 decrease in cfu/mL, while bacteriostatic activity was defined as a <3 log10 decrease in cfu/mL. The lower limit of detection was 1.3 log10 cfu/mL.


    Results and discussion
 Top
 Abstract
 Introduction
 Materials and methods
 Results and discussion
 Acknowledgements
 References
 
Independently of erythromycin susceptibility, both ABT-773 and amoxicillin were highly active against all S. pyogenes isolates. The MIC ranges (mg/L) were 0.004–0.25 of ABT-773 and 0.015–0.12 of amoxicillin. Based upon the proposed breakpoints for ABT-773 against nonpneumococcal Streptococcus spp. (>=0.5 mg/L for susceptible, 1 mg/L for intermediate and >=2 mg/L for resistant), the 10 strains would be considered susceptible.6

The results of the time–kill experiments are given in Figures 1 and 2GoGo. At 24 h, ABT-773 8 x MIC demonstrated bactericidal activity against six strains (four erythromycin susceptible and two containing the mefA gene). Bactericidal activity was observed at 24 h against three strains at an ABT-773 concentration of 2 x MIC (one erythromycin susceptible and two containing the mefA gene). In comparison, amoxicillin at concentrations of 2 x and 8 x MIC was bactericidal against all 10 isolates at 24 h. Both the rate and extent of killing were less for ABT-773 than amoxicillin against the 10 isolates.



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Figure 1. Time–kill curves for six erythromycin-susceptible strains (a–f) of S. pyogenes. Control, {diamondsuit}; ABT-773 8 x MIC, {blacksquare}; ABT-773 2 x MIC, {square}; amoxicillin 8 x MIC, •; amoxicillin 2 x MIC, {circ}; lower limit of detection, - - - .

 


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Figure 2. Time–kill curves for four erythromycin-resistant strains (a–d) of S. pyogenes. Control, {diamondsuit}; ABT-773 8 x MIC, {blacksquare}; ABT-773 2 x MIC, {square}; amoxicillin 8 x MIC, •; amoxicillin 2 x MIC, {circ}; lower limit of detection, - - - .

 
Limited data are available on the time–kill kinetics of ketolides against S. pyogenes.7,8 Odenholt et al.7 studied the bactericidal activity of telithromycin against one erythromycin-susceptible and two erythromycin-resistant strains. Similar to our results, the authors noted a slow rate of killing (<2 log10 cfu/mL after 12 h) against the three isolates. The telithromycin concentration (0.6 mg/L) utilized in their study was reported to correspond to a 2 h free, unbound serum level following a 800 mg dose. Boswell et al.8 also performed time–kill studies with telithromycin against three isolates of S. pyogenes. At concentrations of 10 x MIC, bactericidal activity was reported with telithromycin in 2/3 strains at 24 h. Similar results were obtained in our study with 8 x MIC ABT-773. Organisms exposed to concentrations of telithromycin 2 x MIC demonstrated increased growth (1.51–4.17 log10 cfu/mL), whereas in our study, ABT-773 2 x MIC demonstrated bacteriostatic activity against seven isolates and bactericidal activity against three isolates.

In conclusion, ABT-773 demonstrated in vitro activity against both erythromycin-susceptible and -resistant strains of S. pyogenes. Further in vitro and in vivo studies are needed to define further the role of ABT-773 in infections due to S. pyogenes.


    Acknowledgements
 Top
 Abstract
 Introduction
 Materials and methods
 Results and discussion
 Acknowledgements
 References
 
This work was supported by a grant from Abbott Laboratories. This work was presented in part at the poster session of The 22nd International Congress of Chemotherapy, Amsterdam, The Netherlands, 2001.


    Notes
 
* Corresponding author. Tel: +1-312-996-8639; Fax: +1-312-413-1797; E-mail: pendland{at}uic.edu Back


    References
 Top
 Abstract
 Introduction
 Materials and methods
 Results and discussion
 Acknowledgements
 References
 
1 . Bisno, A. L., Gerber, M. A., Gwaltney, Jr. J. M., Kaplan, E. L. & Schwartz, R. H. (1997). Diagnosis and management of group A streptococcal pharyngitis: a practice guideline. Infectious Diseases Society of America. Clinical Infectious Diseases 25, 574–83.[ISI][Medline]

2 . Betriu, C., Casado, M. C., Gómez, M., Sanchez, A., Palau, M. L. & Picazo, J. J. (1999). Incidence of erythromycin resistance in Streptococcus pyogenes: a 10-year study. Diagnostic Microbiology and Infectious Disease 33, 255–6.[ISI][Medline]

3 . Nilius, A. M., Bui, M. H., Almer, L., Hensey-Rudloff, D., Beyer, J., Ma, Z. et al. (2001). Comparative in vitro activity of ABT-773, a novel antibacterial ketolide. Antimicrobial Agents and Chemotherapy 45, 2163–8.[Abstract/Free Full Text]

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

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

6 . Stone, G., Nilius, A., Hensey, D., Almer, L., Beyer, J. & Flamm, R. (2000). Development of tentative interpretive criteria for ABT-773, a novel ketolide antibacterial agent. In Programs and Abstracts of the Fortieth Interscience Conference on Antimicrobial Agents and Chemotherapy, Toronto, Canada, 2000. Abstract 2164, p. 181. American Society for Microbiology, Washington, DC.

7 . Odenholt, I., Lowdin, E. & Cars, O. (2001). Pharmacodynamics of telithromycin in vitro against respiratory tract pathogens. Antimicrobial Agents and Chemotherapy 45, 23–9.[Abstract/Free Full Text]

8 . Boswell, F. J., Andrews, J. M. & Wise, R. (1998). Pharmacodynamic properties of HMR 3647, a novel ketolide, on respiratory pathogens, enterococci, and Bacteroides fragilis demonstrated by studies of time–kill kinetics and postantibiotic effect. Journal of Antimicrobial Chemotherapy 41, 149–53.[Abstract]

Received 3 August 2001; returned 19 November 2001; revised 4 January 2002; accepted 14 January 2002





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