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Antimicrobial susceptibility profile among β-haemolytic Streptococcus spp. collected in the SENTRY antimicrobial surveillance program—North America, 2001
Diagnostic Microbiology and Infectious Disease 46 (2003) 291–294
www.elsevier.com/locate/diagmicrobio
Notes
Antimicrobial susceptibility profile among -haemolytic Streptococcus spp. collected in the SENTRY Antimicrobial Surveillance Program— North America, 2001 Douglas J. Biedenbacha,*, Jennifer M. Stephena, Ronald N. Jonesa,b a
The JONES Group/JMI Laboratories, North Liberty, IA, USA b Tufts University School of Medicine, Boston, MA, USA
Received 10 December 2002; received in revised form 13 March 2003
Abstract The surveillance of antimicrobial resistance among -hemolytic Streptococcus spp. is an important health care concern due to the serious nature of the diseases that are caused by these pathogens and the emerging antimicrobial resistances. National and international studies have documented diverse rates of resistance to macrolide and lincosamide agents. The SENTRY Antimicrobial Surveillance Program evaluated the resistance rates of -hemolytic Streptococcus spp. to several drug classes including erythromycin and clindamycin during the 2001 study year. Twenty-five medical centers in North America contributed 787 isolates from serogroups A (SGA; 397 strains), B (SGB; 318 strains), G (SGG; 45 strains), C (SGC; 19 strains), and F (SGF; eight strains). Isolates were tested at a reference center by broth microdilution using NCCLS methods. A subset of isolates were characterized by molecular methods to determine the presence of erm and mef resistance mechanisms. All isolates were susceptible to -lactams, linezolid, vancomycin, chloramphenicol, quinupristin/dalfopristin, and fluoroquinolones. Garenoxacin (MIC90, 0.06 g/ml) was the most potent fluoroquinolone tested. Tetracycline was inactive against SGB (14.8% susceptible) compared to SGC, SGG, SGF and SGA (48.9-85.6% susceptible). Resistance to erythromycin and clindamycin was highest among SGB isolates with M-phenotypes (mef) representing the majority (59.1%) of strains. Among isolates randomly selected for genotypic characterization, eight strains had mef (A), 16 had erm (A) [subclass erm (TR)], seven had erm (B) and one strain had erm (A) [subclass erm (TR)] and mef (A) present. These data when compared to previous SENTRY Program analysis showed macrolide resistance has remained stable over the last five years and M-phenotypes are the most prevalent expression of MLSB resistance in North America. © 2003 Elsevier Inc. All rights reserved.
1. Introduction Disease caused by -hemolytic streptococci remains an important global health concern and requires structured surveillance programs to detect the isolation frequency of more virulent clones and emerging antimicrobial-resistant strains. These pathogens are responsible for acute and chronic disease including respiratory tract infections, endocarditis, sepsis, meningitis, bone and joint infections, pylonephritis, necrotizing facitis and delayed sequelae including glomerulonephritis, rheumatic fever and permanent neurologic problems (Ruoff et al., 1999). Current practice guidelines for the management of pharyngitis caused by Streptococcus pyogenes (serogroup A) includes the use of erythromycin as * Corresponding author. Tel.: ⫹1-319-665-3370; fax: ⫹1-319-6653371. E-mail address: [email protected] (D. Biedenbach). 0732-8893/03/$ – see front matter © 2003 Elsevier Inc. All rights reserved. doi:10.1016/S0732-8893(03)00065-8
an alternative to penicillin when indicated and clindamycin for persons with multiple, recurrent episodes (Bisino et al., 2002). Macrolide or lincosamide therapy is also a recommended treatment option for S. agalactiae (serogroup B) infection or for prophylaxis when streptococcal colonization among pregnant women is suspected (Schuchat, 2001). The high transmissibility of -hemolytic streptococci including resistant clones and the association of increased macrolide usage, may play significant roles in the variable resistance rates that have been reported during the last decade (Avanzini et al., 2000). It has been well documented that the frequency of macrolide-lincosamide-streptograminB-resistance (MLSB phenotypes) among streptococci varies considerably between countries (Avanzini et al., 2000; Critchley et al., 2001; Gordon et al., 2002; Perez-Trallero et al., 2001; Weiss et al., 2001). Global results from the SENTRY Antimicrobial Surveillance Program determined that the overall resistance
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rate among -hemolytic streptococci was 13.8% for erythromycin and 5.3% for clindamycin during 1997-2000 (Gordon et al., 2002). This same study showed that susceptibility to erythromycin was significantly lower in North America (81.4%) when compared to the Asia-Pacific (89.1%), Europe (90.3%) and Latin America (97.3%). It was also noted that drug efflux, consistent with the M-phenotype (resistant to erythromycin and susceptible to clindamycin) was more prevalent in North American medical centers compared to Europe or the Asia-Pacific, which referred more strains having potential ribosomal modification with a constitutive resistance (erm-type). A large study sample of S. pyogenes isolated in the United States during 1999 showed that macrolide resistance was 6.2%, with the highest rates reported in the western region of the country (Critchley et al., 2001). Supportive North American data from Canada during 1998 showed similar macrolide resistance rates (4.6%) with regional variation (Weiss et al., 2001). In Europe, studies frequently document the decreased susceptibility to macrolides among S. pyogenes. A study conducted in Spain reported 20.4% macrolide resistance rates during 1998-99 dominated (90%) by M-phenotypes (Perez-Trallero et al., 2001). Studies in Italy documented an increasing erythromycin resistance trend beginning in 1983 which surpassed 50% in 1996 and 1997, during which time constitutive resistance phenotypes were most prevalent (Avanzini et al., 2000). Erythromycinresistant S. pyogenes isolates in central and Eastern European countries were detected at a 12.3% rate, with a predominance of erm (A) [subclass erm (TR)] phenotypes (60.5%) during 1999-2000 (Nagai et al., 2002). The increasing isolation of macrolide-resistant S. agalactiae has also become alarming, and it has been suggested that MLSB agents are no longer reliable empiric or preventative treatment options (Murdoch and Reller, 2001). Studies have documented that the macrolide resistance rates for invasive S. agalactiae isolates during 1997-99 was 8-14.3% and 25.4% in Canada and the United States, respectively (Andrews et al., 2000; de Azavedo et al., 2001). Investigators in Taiwan have reported erythromycin-resistance that is at a stable, very high rate of 46% during 1997-2000, which was predominately (85.5%) constitutive resistance phenotypes (Hsueh et al., 2001). The cited studies above are a testament to the regional diversity of resistant phenotypes of -hemolytic Streptococcus spp. In 2001, the SENTRY Program initiated an objective to determine the antimicrobial resistance patterns of -hemolytic streptococci isolated from all monitored medical centers worldwide. This report will focus on serogroup A (SGA), B (SGB), C (SGC), G (SGG) and F (SGF) isolates from the North American sample and determine the prevalence and diversity of macrolide-resistant phenotypes. A selected sample of 32 strains isolated in the United States and Canada had molecular characterization performed to determine the responsible resistance mechanism(s). The activity of several other drug classes and the potencies of
newer generation fluoroquinolones, including gatifloxacin and garenoxacin (formerly BMS 284756), will also be presented. During 2001, the SENTRY Program participants were requested to forward to the regional monitor (North Liberty, IA), twenty-five isolates of -hemolytic Streptococcus spp. from clinically significant infections. Acceptable samples from which -hemolytic streptococci were to be isolated included throat swabs, high quality sputum samples, cutaneous exudate (cellulitis, pyoderma and wound infections), urine, blood and cerebral spinal fluid. Twenty-five sites in North America including twenty in the United States and five in Canada complied with the protocol. The sample of organisms was geographically diverse and included medical centers from sixteen states in the Northeast (6), Midwest (4), South (5) and West (5) regions of the United States, and five provinces in Canada. A total of 787 isolates were sent by overnight courier which were viable and grew adequately in test media to determine minimum inhibitory concentrations (MICs) for thirty-one antimicrobial agents (thirteen presented in this study), including -lactams, MLSB compounds, fluoroquinolones, tetracycline, linezolid, chloramphenicol and vancomycin. The majority of -hemolytic streptococci isolates were SGA (397 strains) and SGB (318 strains), with smaller collections of SGG (45 strains), SGC (19 strains) and SGF (8 strains) isolates. The species identification was determined locally. After overnight incubation, isolates were examined for hemolysis of blood agar and colony morphology appropriate for the Lancefield group, including colony size and the zone size of -hemolysis. The regional monitor confirmed the isolate identification using bacitracin disks, CAMP factor test and the Vitek System (BioMerieux, Hazelwood, MO) when local identification was either questionable or not provided (i.e., -hemolytic Streptococcus spp., NOS). Patient demographic data were collected which included the age and sex of patient and the source of the infection. After a second subculture, several colonies were selected and suspended into Mueller-Hinton broth to an inoculum density equivalent to a 0.5 McFarland standard. The suspension was further diluted (5 ⫻ 105 CFU/ml) into MuellerHinton with 3-5% lysed horse blood supplement and dispensed (100l) into dry-form microdilution panels using an autoinoculating device (TREK Diagnostics Systems, Cleveland, OH). The panels were incubated in 35°C ambient air overnight and MICs were read visually after 20-24 h. All methods and interpretation of results conformed to National Committee for Clinical Laboratory Standards (NCCLS 2003 and 2003). Routine quality control was performed using American Type Culture Collection Strains (ATCC) including S. pneumoniae ATCC 49619, Hemophilus influenzae ATCC 49247 and Staphylococcus aureus ATCC 29213. A subset of isolates were molecularly characterized to determine the presence of erm and mef genes by multi-
D.J. Biedenbach et al. / Diagnostic Microbiology and Infectious Disease 46 (2003) 291–294
293
Table 1 Activity and susceptibility ratesa for 13 antimicrobial agents tested against five serogroups of -haemolytic streptococci isolated in North America (SENTRY Antimicrobial Surveillance Program, 2001) Antimicrobial agent
Activity by organism tested: Serogroup A (397)
Erythromycin Clindamycin Tetracycline Chloramphenicol Penicillin Cefepime Ceftriaxone Quinupristin/Dalfopristin Linezolid Vancomycin Garenoxacinb Gatifloxacin Levofloxacin a b
Serogroup B (318)
Serogroup G (45)
Serogroup C (19)
Serogroup F (8)
MIC50/90
% Sa
MIC50/90
% Sa
MIC50/90
% Sa
MIC50/90
% Sa
MIC50/90
% Sa
ⱕ0.06/ⱕ0.06 ⱕ0.06/ⱕ0.06 ⱕ4/⬎8 ⱕ2/4 ⱕ0.015/ⱕ0.015 ⱕ0.12/ⱕ0.12 ⱕ0.25/ⱕ0.25 0.12/0.25 1/1 0.25/0.5 0.06/0.12 0.25/0.25 0.5/0.5
91.4 99.2 85.6 100.0 100.0 100.0 100.0 100.0 100.0 100.0 100.0 100.0 100.0
ⱕ0.06/8 ⱕ0.06/⬎8 ⬎8/⬎8 ⱕ2/ⱕ2 0.03/0.06 ⱕ0.12/ⱕ0.12 ⱕ0.25/ⱕ0.25 0.5/0.5 1/1 0.5/0.5 0.06/0.06 0.25/0.25 0.5/1
69.8 88.1 14.8 100.0 100.0 100.0 100.0 100.0 100.0 100.0 100.0 100.0 100.0
ⱕ0.06/2 ⱕ0.06/ⱕ0.06 8/⬎8 ⱕ2/4 ⱕ0.015/0.03 ⱕ0.12/ⱕ0.12 ⱕ0.25/ⱕ0.25 0.25/0.5 1/1 0.25/0.5 0.06/0.06 0.25/0.25 0.5/0.5
84.4 97.8 48.9 100.0 100.0 100.0 100.0 100.0 100.0 100.0 100.0 100.0 100.0
ⱕ0.06/ⱕ0.06 ⱕ0.06/ⱕ0.06 ⱕ4/⬎8 ⱕ2/4 0.03/0.06 ⱕ0.12/0.5 ⱕ0.25/ⱕ0.25 0.5/0.5 1/1 0.5/0.5 ⱕ0.03/0.06 0.12/0.25 0.25/0.5
94.7 100.0 57.9 100.0 100.0 100.0 100.0 100.0 100.0 100.0 100.0 100.0 100.0
ⱕ0.06/⫺ ⱕ0.06/⫺ ⱕ4/⫺ ⱕ2/⫺ 0.03/⫺ 0.5/⫺ ⱕ0.25/⫺ 1/⫺ 1/⫺ 1/⫺ ⱕ0.03/⫺ 0.12/⫺ 0.25/⫺
100.0 100.0 75.0 100.0 100.0 100.0 100.0 100.0 100.0 100.0 100.0 100.0 100.0
S ⫽ susceptible, susceptibility criteria per NCCLS (2002). Garenoxacin susceptibility (ⱕ1 g/ml) was defined the same as gatifloxacin [NCCLS, 2002] for comparison purposes only.
plex rapid cycle PCR with microwell-format probe hybridization (Farrell et al., 2001). The age of the patients from which cultures were obtained ranged from one month to 97 years old with a mode of 15 and 39 years for patients infected by SGA and SGB, respectively, and 31 years for the other three serogroups combined. The distribution of male and female patients was nearly even for all serogroups except for patients infected by SGB (females represented 69.8%). The significant sources of infection due to SGA were throat (80.3%), wound (11.8%), sterile body sites (3.3%) and ear, eye and nasal sources (1.8%). SGB isolates were cultured primarily from urine (36.3%), wound (24.5%), genital tract and placenta (19.3%), sterile body sites (7.8%) and pulmonary sources (5.6%). The remaining serogroups were isolated mainly from throat (39.8%), wound (32.5%), pulmonary sources (16.9%) and blood culture (7.2%). The majority of the isolates were cultured from outpatients, particularly SGA (88.1%) ⬎ SGB (68.1%) ⬎ SGC, SGG and SGF (59.7%, combined). The antimicrobial potencies and the susceptibilities of -hemolytic streptococci separated by serogroup are shown in Table 1. All isolates regardless of serogroup were susceptible to penicillin, cephalosporins, quinupristin/dalfopristin (Q/D), linezolid, vancomycin, chloramphenicol and the three tested fluoroquinolones. The potency of Q/D and vancomycin (MIC90, 0.25-0.5 g/ml) was slightly greater than linezolid (MIC90, 1 g/ml). The rank order of fluoroquinolone potency was garenoxacin (MIC90, 0.06-0.12 g/ ml) ⬎ gatifloxacin (MIC90, 0.25 g/ml) ⬎ levofloxacin (MIC90, 0.5-1 g/ml). Tetracycline was not active against the SGB isolates (14.8% susceptible) and only marginally active against SGG, SGC and SGF (48.9-75.0% susceptible). A fairly high percentage (85.6%) of SGA isolates from
North America remained susceptible to tetracycline. Susceptibility to erythromycin was lowest among the SGB (69.8%) and SGG (84.4%) isolates compared to SGA (91.4%), SGC (94.7%) and SGF (100%). All SGC and SGF isolates were susceptible to clindamycin and resistance was rarely detected among SGA and SGG isolates (0.8-2.2% susceptible) but was higher for SGB (11.4%). Among the 25 centers monitored during 2001 in North America, 16 sites forwarded isolates of SGA and/or SGB streptococci that were resistant to erythromycin. Among the 35 SGA strains with macrolide resistance (MIC, ⱖ 1 g/ ml), regional variation with regards to the prevalence or phenotypic patterns of erythromycin and clindamycin resistance was not evident. Thirty-one (91.4%) of the SGA isolates were of the mef-phenotype. Most isolates with macrolide resistance did not cause invasive disease with the highest number of strains from throat (60.0%) and wound (30.0%) cultures. The overall rate of M-phenotypes (mef) among the SGB isolates was 59.1%, concluding that constitutive resistance was much higher for this species compared to the other serogroup streptococci in North America. It should be noted that two medical centers were overly compliant and sent the majority of the SGB isolates in the collection resulting in a slight over-representation. The data in Table 2 shows the genotypic analysis for the three most prevalent serogroups with erythromycin-resistant isolates. Thirty-two selected isolates were tested by PCR methods to determine the mechanism(s) that were responsible for macrolide-resistant patterns. A total of 9, 18 and 5 isolates of SGA, SGB and SGG, respectively, were selected from the United States and Canada. Three isolates of SGA were confirmed as mef (A), 5 were erm (A) [subclass erm (TR)] and one isolate was erm (B). Among the SGB isolates, 8 were erm (A) [subclass erm (TR)], 6 were erm (B), 3 were
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Table 2 Molecular characterization of MLSB resistances among 32 -haemolytic streptococci isolated in the SENTRY Antimicrobial Surveillance Program in North America (2001) Organism
Serogroup A Serogroup B Serogroup G
Region (no. tested)
Resistance mechanism erm (A) [subclass erm (TR)]
erm (B)
mef (A)
erm (A) [subclass erm (TR)]⫹mef (A)
USA (7) Canada (2) USA (17) Canada (1) USA (3) Canada (2)
4 1 8 0 3 0
0 1 5 1 0 0
3 0 3 0 0 2
0 0 1 0 0 0
mef (A) and one isolate from the United States had both erm (A) [subclass erm (TR)] and mef (A) genes. The SGG isolates in the United States were all erm (A) [subclass erm (TR)] and isolates from Canada were both mef (A). All isolates with erm (B) were resistant to erythromycin, azithromycin and clindamycin. Although penicillin remains an effective treatment for infections caused by -hemolytic Streptococcus spp., drug tolerance and clinical therapeutic failures have been reported (Gillespie, 1998). Macrolides and lincosamides have been frequently used to circumvent patient -lactam allergy, and as empiric and preventative therapies for the treatment of -hemolytic Streptococcus spp. This report indicates that during 2001 in North America, macrolide resistance was relatively common particularly among S. agalactiae (SGB) and that M-phenotypes dominate the population of macrolide-resistant streptococcal strains. It was also noted that tetracycline resistance is species dependent. No fluoroquinolone-resistance was detected during the monitored year and the potencies of newer fluoroquinolones (gatifloxacin and garenoxacin) were found to be superior overall. Continual monitoring of antimicrobial resistance among -hemolytic Streptococcus spp. appears to be a necessary objective to provide the medical community with data regarding the resistance mechanisms that are most common to their local or regional environments. This is particularly important for macrolide resistance which can vary quite significantly between communities and regions within a nation.
References Andrews, J. I., Diekema, D. J., Hunter, S. K., Rhomberg, P. R., Pfaller, M. A., Jones, R. N., & Doern, G. V. (2000). Group B streptococci causing neonatal blood stream infection: Antimicrobial susceptibility and serotyping. Results from the SENTRY centers in the Western Hemisphere. Am J Obst Gyn 4, 859 – 862. Avanzini, C., Bosio, K., Volpe, G., Dotti, G., & Savoia, D. (2000). Streptococcus pyogenes collected in Torino (northwest Italy) between 1983 and 1998: Survey of macrolide resistance and trend of genotype by RAPD. Microbial Drug Resistance 6, 289 –295.
Bisno, A. L., Gerber, M. A., Gwaltney, J. M., Kaplan, E. L., & Schwartz, R. H. (2002). Practice guidelines for the diagnosis and management of group A streptococcal pharyngitis. Clin Infect Dis 35, 113–125. Critchley, I. A., Sahm, D. F., Thornsberry, C., Blosser-Middleton, R. S., Jones, M. E., & Karlowsky, J. A. (2001). Antimicrobial susceptibilities of Streptococcus pyogenes isolated from respiratory and skin and soft tissue and infections: United States LIBRA surveillance data from 1999. Diagn Microbiol Infect Dis 42, 129 –135. De Azavedo, J. C., McGavin, M., Duncan, C., Low, D. E., & McGeer, A (2001). Prevalence and mechanisms of macrolide resistance in invasive and noninvasive group B Streptococcus isolates from Ontario, Canada. Antimicrob Agents Chemother 45, 3504 –3508. Farrell, D. J., Morrisey, I., Bakker, G., & Felmingham, D. (2001). Detection of macrolide resistance mechanisms in Streptococcus pneumoniae and Streptococcus pyogenes using a multiplex rapid cycle PCR with microwell-format probe hybridization. J Antimicrob Chemother 48, 541–544. Gillespie, S. H. (1998). Failure of penicillin in Streptococcus pyogenes pharyngeal infection. Lancet 352, 1954 –1956. Gordon, K. A., Beach, M. L., Biedenbach, D. J., Jones, R. N., Rhomberg, P. R., & Mutnick, A. H. (2002). Antimicrobial susceptibility patterns of -haemolytic and viridans group streptococci: Report from the SENTRY Antimicrobial Surveillance Program (1997-2000). Diagn Microbiol Infect Dis 43, 157–162. Hsueh, P.-R., Teng, L.-J., Lee, L.-N., Ho, S.-W., Yang, P.-C., & Luh, K.-T. (2001). High incidence of erythromycin resistance among clinical isolates of Streptococcus agalactiae in Taiwan. Antimicrob Agents Chemother 45, 3205–3208. Leclercq, R. (2002). Mechanisms of resistance to macrolides and lincosamides: Nature of the resistance elements and their clinical implications. Clin Infect Dis 34, 482– 492. Murdoch, D. R., & Reller, L. B. (2001). Antimicrobial susceptibilities of group B streptococci isolated from patients with invasive disease: 10 year perspective. Antimicrob Agents Chemother 45, 3623–3624. Nagai, K., Appelbaum, P. C., Davies, T. A., Kelly, L. M., Hoellman, D. B., Andrasevic, A. T., Drukalska, L., Hryniewicz, W., Jacobs, M. R., Kolman, J., Miciuleviciene, J., Pana, M., Setchanova, L., Thege, M. K., Hupkova, H., Trupl, J., & Urbaskova, P. (2002). Susceptibility to telithromycin in 1,011 Streptococcus pyogenes isoaltes from 10 central and eastern European countries. Antimicrob Agents Chemother 46, 546–549. National Committee for Clinical Laboratory Standards (NCCLS) (2003). Methods for dilution antimicrobial susceptibility tests for bacteria that grow aerobically—Fifth Edition: Approved standard M7-A6. Wayne, PA: NCCLS. National Committee for Clinical Laboratory Standards (NCCLS) (2003). Performance standards for antimicrobial susceptibility testing. Supplemental tables, M100-S13. Wayne, PA: NCCLS. Perez-Trallero, E., Fernandez-Mazarrasa, C., Garcia-Rey, C., Bouza, E., Aguilar, L., Garcia-de-Lomas, J., Baquero, F., & The Spanish Surveillance Group for Respiratory Pathogens (2001). Antimicrobial susceptibilities of 1,684 Streptococcus pneumoniae and 2,039 Streptococcus pyogenes isolates and their ecological relationships: Results of a 1-year (1998-1999) multicenter surveillance study in Spain. Antimicrob Agents Chemother 45, 3334 –3340. Ruoff, K. L. (1999). Streptococcus. In: P. R. Murray, E. J. Baron, M. A. Pfaller, C. Tenover, & R. H. Yolken (Eds), Manual of clinical microbiology, 7th ed. Washington, DC: American Society for Microbiology, pp. 283–296. Schuchat, A. (2001). Group B streptococcal disease: From trials and tribulations to triumph and trepidation. Clin Infect Dis 33, 751–756. Weiss, K., De Azavedo, J., Restieri, C., Galarneau, L. A., Gourdeau, M., Harvey, P., Paradis, J. F., Salim, K., & Low, D. E. (2001). Phenotypic and genotypic characterization of macrolide-resistant group a Streptococcus strains in the province of Quebec, Canada. J Antimicrob Chemother 47, 345–348.
www.elsevier.com/locate/diagmicrobio
Notes
Antimicrobial susceptibility profile among -haemolytic Streptococcus spp. collected in the SENTRY Antimicrobial Surveillance Program— North America, 2001 Douglas J. Biedenbacha,*, Jennifer M. Stephena, Ronald N. Jonesa,b a
The JONES Group/JMI Laboratories, North Liberty, IA, USA b Tufts University School of Medicine, Boston, MA, USA
Received 10 December 2002; received in revised form 13 March 2003
Abstract The surveillance of antimicrobial resistance among -hemolytic Streptococcus spp. is an important health care concern due to the serious nature of the diseases that are caused by these pathogens and the emerging antimicrobial resistances. National and international studies have documented diverse rates of resistance to macrolide and lincosamide agents. The SENTRY Antimicrobial Surveillance Program evaluated the resistance rates of -hemolytic Streptococcus spp. to several drug classes including erythromycin and clindamycin during the 2001 study year. Twenty-five medical centers in North America contributed 787 isolates from serogroups A (SGA; 397 strains), B (SGB; 318 strains), G (SGG; 45 strains), C (SGC; 19 strains), and F (SGF; eight strains). Isolates were tested at a reference center by broth microdilution using NCCLS methods. A subset of isolates were characterized by molecular methods to determine the presence of erm and mef resistance mechanisms. All isolates were susceptible to -lactams, linezolid, vancomycin, chloramphenicol, quinupristin/dalfopristin, and fluoroquinolones. Garenoxacin (MIC90, 0.06 g/ml) was the most potent fluoroquinolone tested. Tetracycline was inactive against SGB (14.8% susceptible) compared to SGC, SGG, SGF and SGA (48.9-85.6% susceptible). Resistance to erythromycin and clindamycin was highest among SGB isolates with M-phenotypes (mef) representing the majority (59.1%) of strains. Among isolates randomly selected for genotypic characterization, eight strains had mef (A), 16 had erm (A) [subclass erm (TR)], seven had erm (B) and one strain had erm (A) [subclass erm (TR)] and mef (A) present. These data when compared to previous SENTRY Program analysis showed macrolide resistance has remained stable over the last five years and M-phenotypes are the most prevalent expression of MLSB resistance in North America. © 2003 Elsevier Inc. All rights reserved.
1. Introduction Disease caused by -hemolytic streptococci remains an important global health concern and requires structured surveillance programs to detect the isolation frequency of more virulent clones and emerging antimicrobial-resistant strains. These pathogens are responsible for acute and chronic disease including respiratory tract infections, endocarditis, sepsis, meningitis, bone and joint infections, pylonephritis, necrotizing facitis and delayed sequelae including glomerulonephritis, rheumatic fever and permanent neurologic problems (Ruoff et al., 1999). Current practice guidelines for the management of pharyngitis caused by Streptococcus pyogenes (serogroup A) includes the use of erythromycin as * Corresponding author. Tel.: ⫹1-319-665-3370; fax: ⫹1-319-6653371. E-mail address: [email protected] (D. Biedenbach). 0732-8893/03/$ – see front matter © 2003 Elsevier Inc. All rights reserved. doi:10.1016/S0732-8893(03)00065-8
an alternative to penicillin when indicated and clindamycin for persons with multiple, recurrent episodes (Bisino et al., 2002). Macrolide or lincosamide therapy is also a recommended treatment option for S. agalactiae (serogroup B) infection or for prophylaxis when streptococcal colonization among pregnant women is suspected (Schuchat, 2001). The high transmissibility of -hemolytic streptococci including resistant clones and the association of increased macrolide usage, may play significant roles in the variable resistance rates that have been reported during the last decade (Avanzini et al., 2000). It has been well documented that the frequency of macrolide-lincosamide-streptograminB-resistance (MLSB phenotypes) among streptococci varies considerably between countries (Avanzini et al., 2000; Critchley et al., 2001; Gordon et al., 2002; Perez-Trallero et al., 2001; Weiss et al., 2001). Global results from the SENTRY Antimicrobial Surveillance Program determined that the overall resistance
292
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rate among -hemolytic streptococci was 13.8% for erythromycin and 5.3% for clindamycin during 1997-2000 (Gordon et al., 2002). This same study showed that susceptibility to erythromycin was significantly lower in North America (81.4%) when compared to the Asia-Pacific (89.1%), Europe (90.3%) and Latin America (97.3%). It was also noted that drug efflux, consistent with the M-phenotype (resistant to erythromycin and susceptible to clindamycin) was more prevalent in North American medical centers compared to Europe or the Asia-Pacific, which referred more strains having potential ribosomal modification with a constitutive resistance (erm-type). A large study sample of S. pyogenes isolated in the United States during 1999 showed that macrolide resistance was 6.2%, with the highest rates reported in the western region of the country (Critchley et al., 2001). Supportive North American data from Canada during 1998 showed similar macrolide resistance rates (4.6%) with regional variation (Weiss et al., 2001). In Europe, studies frequently document the decreased susceptibility to macrolides among S. pyogenes. A study conducted in Spain reported 20.4% macrolide resistance rates during 1998-99 dominated (90%) by M-phenotypes (Perez-Trallero et al., 2001). Studies in Italy documented an increasing erythromycin resistance trend beginning in 1983 which surpassed 50% in 1996 and 1997, during which time constitutive resistance phenotypes were most prevalent (Avanzini et al., 2000). Erythromycinresistant S. pyogenes isolates in central and Eastern European countries were detected at a 12.3% rate, with a predominance of erm (A) [subclass erm (TR)] phenotypes (60.5%) during 1999-2000 (Nagai et al., 2002). The increasing isolation of macrolide-resistant S. agalactiae has also become alarming, and it has been suggested that MLSB agents are no longer reliable empiric or preventative treatment options (Murdoch and Reller, 2001). Studies have documented that the macrolide resistance rates for invasive S. agalactiae isolates during 1997-99 was 8-14.3% and 25.4% in Canada and the United States, respectively (Andrews et al., 2000; de Azavedo et al., 2001). Investigators in Taiwan have reported erythromycin-resistance that is at a stable, very high rate of 46% during 1997-2000, which was predominately (85.5%) constitutive resistance phenotypes (Hsueh et al., 2001). The cited studies above are a testament to the regional diversity of resistant phenotypes of -hemolytic Streptococcus spp. In 2001, the SENTRY Program initiated an objective to determine the antimicrobial resistance patterns of -hemolytic streptococci isolated from all monitored medical centers worldwide. This report will focus on serogroup A (SGA), B (SGB), C (SGC), G (SGG) and F (SGF) isolates from the North American sample and determine the prevalence and diversity of macrolide-resistant phenotypes. A selected sample of 32 strains isolated in the United States and Canada had molecular characterization performed to determine the responsible resistance mechanism(s). The activity of several other drug classes and the potencies of
newer generation fluoroquinolones, including gatifloxacin and garenoxacin (formerly BMS 284756), will also be presented. During 2001, the SENTRY Program participants were requested to forward to the regional monitor (North Liberty, IA), twenty-five isolates of -hemolytic Streptococcus spp. from clinically significant infections. Acceptable samples from which -hemolytic streptococci were to be isolated included throat swabs, high quality sputum samples, cutaneous exudate (cellulitis, pyoderma and wound infections), urine, blood and cerebral spinal fluid. Twenty-five sites in North America including twenty in the United States and five in Canada complied with the protocol. The sample of organisms was geographically diverse and included medical centers from sixteen states in the Northeast (6), Midwest (4), South (5) and West (5) regions of the United States, and five provinces in Canada. A total of 787 isolates were sent by overnight courier which were viable and grew adequately in test media to determine minimum inhibitory concentrations (MICs) for thirty-one antimicrobial agents (thirteen presented in this study), including -lactams, MLSB compounds, fluoroquinolones, tetracycline, linezolid, chloramphenicol and vancomycin. The majority of -hemolytic streptococci isolates were SGA (397 strains) and SGB (318 strains), with smaller collections of SGG (45 strains), SGC (19 strains) and SGF (8 strains) isolates. The species identification was determined locally. After overnight incubation, isolates were examined for hemolysis of blood agar and colony morphology appropriate for the Lancefield group, including colony size and the zone size of -hemolysis. The regional monitor confirmed the isolate identification using bacitracin disks, CAMP factor test and the Vitek System (BioMerieux, Hazelwood, MO) when local identification was either questionable or not provided (i.e., -hemolytic Streptococcus spp., NOS). Patient demographic data were collected which included the age and sex of patient and the source of the infection. After a second subculture, several colonies were selected and suspended into Mueller-Hinton broth to an inoculum density equivalent to a 0.5 McFarland standard. The suspension was further diluted (5 ⫻ 105 CFU/ml) into MuellerHinton with 3-5% lysed horse blood supplement and dispensed (100l) into dry-form microdilution panels using an autoinoculating device (TREK Diagnostics Systems, Cleveland, OH). The panels were incubated in 35°C ambient air overnight and MICs were read visually after 20-24 h. All methods and interpretation of results conformed to National Committee for Clinical Laboratory Standards (NCCLS 2003 and 2003). Routine quality control was performed using American Type Culture Collection Strains (ATCC) including S. pneumoniae ATCC 49619, Hemophilus influenzae ATCC 49247 and Staphylococcus aureus ATCC 29213. A subset of isolates were molecularly characterized to determine the presence of erm and mef genes by multi-
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Table 1 Activity and susceptibility ratesa for 13 antimicrobial agents tested against five serogroups of -haemolytic streptococci isolated in North America (SENTRY Antimicrobial Surveillance Program, 2001) Antimicrobial agent
Activity by organism tested: Serogroup A (397)
Erythromycin Clindamycin Tetracycline Chloramphenicol Penicillin Cefepime Ceftriaxone Quinupristin/Dalfopristin Linezolid Vancomycin Garenoxacinb Gatifloxacin Levofloxacin a b
Serogroup B (318)
Serogroup G (45)
Serogroup C (19)
Serogroup F (8)
MIC50/90
% Sa
MIC50/90
% Sa
MIC50/90
% Sa
MIC50/90
% Sa
MIC50/90
% Sa
ⱕ0.06/ⱕ0.06 ⱕ0.06/ⱕ0.06 ⱕ4/⬎8 ⱕ2/4 ⱕ0.015/ⱕ0.015 ⱕ0.12/ⱕ0.12 ⱕ0.25/ⱕ0.25 0.12/0.25 1/1 0.25/0.5 0.06/0.12 0.25/0.25 0.5/0.5
91.4 99.2 85.6 100.0 100.0 100.0 100.0 100.0 100.0 100.0 100.0 100.0 100.0
ⱕ0.06/8 ⱕ0.06/⬎8 ⬎8/⬎8 ⱕ2/ⱕ2 0.03/0.06 ⱕ0.12/ⱕ0.12 ⱕ0.25/ⱕ0.25 0.5/0.5 1/1 0.5/0.5 0.06/0.06 0.25/0.25 0.5/1
69.8 88.1 14.8 100.0 100.0 100.0 100.0 100.0 100.0 100.0 100.0 100.0 100.0
ⱕ0.06/2 ⱕ0.06/ⱕ0.06 8/⬎8 ⱕ2/4 ⱕ0.015/0.03 ⱕ0.12/ⱕ0.12 ⱕ0.25/ⱕ0.25 0.25/0.5 1/1 0.25/0.5 0.06/0.06 0.25/0.25 0.5/0.5
84.4 97.8 48.9 100.0 100.0 100.0 100.0 100.0 100.0 100.0 100.0 100.0 100.0
ⱕ0.06/ⱕ0.06 ⱕ0.06/ⱕ0.06 ⱕ4/⬎8 ⱕ2/4 0.03/0.06 ⱕ0.12/0.5 ⱕ0.25/ⱕ0.25 0.5/0.5 1/1 0.5/0.5 ⱕ0.03/0.06 0.12/0.25 0.25/0.5
94.7 100.0 57.9 100.0 100.0 100.0 100.0 100.0 100.0 100.0 100.0 100.0 100.0
ⱕ0.06/⫺ ⱕ0.06/⫺ ⱕ4/⫺ ⱕ2/⫺ 0.03/⫺ 0.5/⫺ ⱕ0.25/⫺ 1/⫺ 1/⫺ 1/⫺ ⱕ0.03/⫺ 0.12/⫺ 0.25/⫺
100.0 100.0 75.0 100.0 100.0 100.0 100.0 100.0 100.0 100.0 100.0 100.0 100.0
S ⫽ susceptible, susceptibility criteria per NCCLS (2002). Garenoxacin susceptibility (ⱕ1 g/ml) was defined the same as gatifloxacin [NCCLS, 2002] for comparison purposes only.
plex rapid cycle PCR with microwell-format probe hybridization (Farrell et al., 2001). The age of the patients from which cultures were obtained ranged from one month to 97 years old with a mode of 15 and 39 years for patients infected by SGA and SGB, respectively, and 31 years for the other three serogroups combined. The distribution of male and female patients was nearly even for all serogroups except for patients infected by SGB (females represented 69.8%). The significant sources of infection due to SGA were throat (80.3%), wound (11.8%), sterile body sites (3.3%) and ear, eye and nasal sources (1.8%). SGB isolates were cultured primarily from urine (36.3%), wound (24.5%), genital tract and placenta (19.3%), sterile body sites (7.8%) and pulmonary sources (5.6%). The remaining serogroups were isolated mainly from throat (39.8%), wound (32.5%), pulmonary sources (16.9%) and blood culture (7.2%). The majority of the isolates were cultured from outpatients, particularly SGA (88.1%) ⬎ SGB (68.1%) ⬎ SGC, SGG and SGF (59.7%, combined). The antimicrobial potencies and the susceptibilities of -hemolytic streptococci separated by serogroup are shown in Table 1. All isolates regardless of serogroup were susceptible to penicillin, cephalosporins, quinupristin/dalfopristin (Q/D), linezolid, vancomycin, chloramphenicol and the three tested fluoroquinolones. The potency of Q/D and vancomycin (MIC90, 0.25-0.5 g/ml) was slightly greater than linezolid (MIC90, 1 g/ml). The rank order of fluoroquinolone potency was garenoxacin (MIC90, 0.06-0.12 g/ ml) ⬎ gatifloxacin (MIC90, 0.25 g/ml) ⬎ levofloxacin (MIC90, 0.5-1 g/ml). Tetracycline was not active against the SGB isolates (14.8% susceptible) and only marginally active against SGG, SGC and SGF (48.9-75.0% susceptible). A fairly high percentage (85.6%) of SGA isolates from
North America remained susceptible to tetracycline. Susceptibility to erythromycin was lowest among the SGB (69.8%) and SGG (84.4%) isolates compared to SGA (91.4%), SGC (94.7%) and SGF (100%). All SGC and SGF isolates were susceptible to clindamycin and resistance was rarely detected among SGA and SGG isolates (0.8-2.2% susceptible) but was higher for SGB (11.4%). Among the 25 centers monitored during 2001 in North America, 16 sites forwarded isolates of SGA and/or SGB streptococci that were resistant to erythromycin. Among the 35 SGA strains with macrolide resistance (MIC, ⱖ 1 g/ ml), regional variation with regards to the prevalence or phenotypic patterns of erythromycin and clindamycin resistance was not evident. Thirty-one (91.4%) of the SGA isolates were of the mef-phenotype. Most isolates with macrolide resistance did not cause invasive disease with the highest number of strains from throat (60.0%) and wound (30.0%) cultures. The overall rate of M-phenotypes (mef) among the SGB isolates was 59.1%, concluding that constitutive resistance was much higher for this species compared to the other serogroup streptococci in North America. It should be noted that two medical centers were overly compliant and sent the majority of the SGB isolates in the collection resulting in a slight over-representation. The data in Table 2 shows the genotypic analysis for the three most prevalent serogroups with erythromycin-resistant isolates. Thirty-two selected isolates were tested by PCR methods to determine the mechanism(s) that were responsible for macrolide-resistant patterns. A total of 9, 18 and 5 isolates of SGA, SGB and SGG, respectively, were selected from the United States and Canada. Three isolates of SGA were confirmed as mef (A), 5 were erm (A) [subclass erm (TR)] and one isolate was erm (B). Among the SGB isolates, 8 were erm (A) [subclass erm (TR)], 6 were erm (B), 3 were
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Table 2 Molecular characterization of MLSB resistances among 32 -haemolytic streptococci isolated in the SENTRY Antimicrobial Surveillance Program in North America (2001) Organism
Serogroup A Serogroup B Serogroup G
Region (no. tested)
Resistance mechanism erm (A) [subclass erm (TR)]
erm (B)
mef (A)
erm (A) [subclass erm (TR)]⫹mef (A)
USA (7) Canada (2) USA (17) Canada (1) USA (3) Canada (2)
4 1 8 0 3 0
0 1 5 1 0 0
3 0 3 0 0 2
0 0 1 0 0 0
mef (A) and one isolate from the United States had both erm (A) [subclass erm (TR)] and mef (A) genes. The SGG isolates in the United States were all erm (A) [subclass erm (TR)] and isolates from Canada were both mef (A). All isolates with erm (B) were resistant to erythromycin, azithromycin and clindamycin. Although penicillin remains an effective treatment for infections caused by -hemolytic Streptococcus spp., drug tolerance and clinical therapeutic failures have been reported (Gillespie, 1998). Macrolides and lincosamides have been frequently used to circumvent patient -lactam allergy, and as empiric and preventative therapies for the treatment of -hemolytic Streptococcus spp. This report indicates that during 2001 in North America, macrolide resistance was relatively common particularly among S. agalactiae (SGB) and that M-phenotypes dominate the population of macrolide-resistant streptococcal strains. It was also noted that tetracycline resistance is species dependent. No fluoroquinolone-resistance was detected during the monitored year and the potencies of newer fluoroquinolones (gatifloxacin and garenoxacin) were found to be superior overall. Continual monitoring of antimicrobial resistance among -hemolytic Streptococcus spp. appears to be a necessary objective to provide the medical community with data regarding the resistance mechanisms that are most common to their local or regional environments. This is particularly important for macrolide resistance which can vary quite significantly between communities and regions within a nation.
References Andrews, J. I., Diekema, D. J., Hunter, S. K., Rhomberg, P. R., Pfaller, M. A., Jones, R. N., & Doern, G. V. (2000). Group B streptococci causing neonatal blood stream infection: Antimicrobial susceptibility and serotyping. Results from the SENTRY centers in the Western Hemisphere. Am J Obst Gyn 4, 859 – 862. Avanzini, C., Bosio, K., Volpe, G., Dotti, G., & Savoia, D. (2000). Streptococcus pyogenes collected in Torino (northwest Italy) between 1983 and 1998: Survey of macrolide resistance and trend of genotype by RAPD. Microbial Drug Resistance 6, 289 –295.
Bisno, A. L., Gerber, M. A., Gwaltney, J. M., Kaplan, E. L., & Schwartz, R. H. (2002). Practice guidelines for the diagnosis and management of group A streptococcal pharyngitis. Clin Infect Dis 35, 113–125. Critchley, I. A., Sahm, D. F., Thornsberry, C., Blosser-Middleton, R. S., Jones, M. E., & Karlowsky, J. A. (2001). Antimicrobial susceptibilities of Streptococcus pyogenes isolated from respiratory and skin and soft tissue and infections: United States LIBRA surveillance data from 1999. Diagn Microbiol Infect Dis 42, 129 –135. De Azavedo, J. C., McGavin, M., Duncan, C., Low, D. E., & McGeer, A (2001). Prevalence and mechanisms of macrolide resistance in invasive and noninvasive group B Streptococcus isolates from Ontario, Canada. Antimicrob Agents Chemother 45, 3504 –3508. Farrell, D. J., Morrisey, I., Bakker, G., & Felmingham, D. (2001). Detection of macrolide resistance mechanisms in Streptococcus pneumoniae and Streptococcus pyogenes using a multiplex rapid cycle PCR with microwell-format probe hybridization. J Antimicrob Chemother 48, 541–544. Gillespie, S. H. (1998). Failure of penicillin in Streptococcus pyogenes pharyngeal infection. Lancet 352, 1954 –1956. Gordon, K. A., Beach, M. L., Biedenbach, D. J., Jones, R. N., Rhomberg, P. R., & Mutnick, A. H. (2002). Antimicrobial susceptibility patterns of -haemolytic and viridans group streptococci: Report from the SENTRY Antimicrobial Surveillance Program (1997-2000). Diagn Microbiol Infect Dis 43, 157–162. Hsueh, P.-R., Teng, L.-J., Lee, L.-N., Ho, S.-W., Yang, P.-C., & Luh, K.-T. (2001). High incidence of erythromycin resistance among clinical isolates of Streptococcus agalactiae in Taiwan. Antimicrob Agents Chemother 45, 3205–3208. Leclercq, R. (2002). Mechanisms of resistance to macrolides and lincosamides: Nature of the resistance elements and their clinical implications. Clin Infect Dis 34, 482– 492. Murdoch, D. R., & Reller, L. B. (2001). Antimicrobial susceptibilities of group B streptococci isolated from patients with invasive disease: 10 year perspective. Antimicrob Agents Chemother 45, 3623–3624. Nagai, K., Appelbaum, P. C., Davies, T. A., Kelly, L. M., Hoellman, D. B., Andrasevic, A. T., Drukalska, L., Hryniewicz, W., Jacobs, M. R., Kolman, J., Miciuleviciene, J., Pana, M., Setchanova, L., Thege, M. K., Hupkova, H., Trupl, J., & Urbaskova, P. (2002). Susceptibility to telithromycin in 1,011 Streptococcus pyogenes isoaltes from 10 central and eastern European countries. Antimicrob Agents Chemother 46, 546–549. National Committee for Clinical Laboratory Standards (NCCLS) (2003). Methods for dilution antimicrobial susceptibility tests for bacteria that grow aerobically—Fifth Edition: Approved standard M7-A6. Wayne, PA: NCCLS. National Committee for Clinical Laboratory Standards (NCCLS) (2003). Performance standards for antimicrobial susceptibility testing. Supplemental tables, M100-S13. Wayne, PA: NCCLS. Perez-Trallero, E., Fernandez-Mazarrasa, C., Garcia-Rey, C., Bouza, E., Aguilar, L., Garcia-de-Lomas, J., Baquero, F., & The Spanish Surveillance Group for Respiratory Pathogens (2001). Antimicrobial susceptibilities of 1,684 Streptococcus pneumoniae and 2,039 Streptococcus pyogenes isolates and their ecological relationships: Results of a 1-year (1998-1999) multicenter surveillance study in Spain. Antimicrob Agents Chemother 45, 3334 –3340. Ruoff, K. L. (1999). Streptococcus. In: P. R. Murray, E. J. Baron, M. A. Pfaller, C. Tenover, & R. H. Yolken (Eds), Manual of clinical microbiology, 7th ed. Washington, DC: American Society for Microbiology, pp. 283–296. Schuchat, A. (2001). Group B streptococcal disease: From trials and tribulations to triumph and trepidation. Clin Infect Dis 33, 751–756. Weiss, K., De Azavedo, J., Restieri, C., Galarneau, L. A., Gourdeau, M., Harvey, P., Paradis, J. F., Salim, K., & Low, D. E. (2001). Phenotypic and genotypic characterization of macrolide-resistant group a Streptococcus strains in the province of Quebec, Canada. J Antimicrob Chemother 47, 345–348.