Antimicrobial susceptibility of 840 clinical isolates of Haemophilus influenzae collected in four European countries in 2000-2001
2003; Elsevier BV; Volume: 9; Issue: 5 Linguagem: Inglês
10.1046/j.1469-0691.2003.00543.x
ISSN1469-0691
AutoresRenée S. Blosser-Middleton, D.F. Sahm, Clyde Thornsberry, Mark E. Jones, Patricia A. Hogan, Ian A. Critchley, James A. Karlowsky,
Tópico(s)Bacterial Infections and Vaccines
ResumoIn 2000-2001, 840 clinical isolates of Haemophilus influenzae were collected from laboratories in France, Germany, Italy and Spain (210 isolates/country). β-Lactamase production among the isolates varied considerably by country, ranging from 8.1 % in Germany to 34.8% in France. H. influenzae from patients ≤4 years old showed the highest prevalence of β-lactamase production (23.2%), compared with isolates from patients aged 5-17 years (17.8%) and ≥18 years (16.5%). All isolates were susceptible to amoxicillin-clavulanate, ciprofloxacin and levofloxacin; 99.6% and 98.9% of isolates were susceptible to azithromycin and cefuroxime, respectively. Among the macrolides tested, azithromycin (MIC90, 2 mg/L) was eight-fold more potent than clarithromycin (MIC90, 16 mg/L) and roxithromycin (MIC90, 16 mg/L). Despite variations in β-lactamase production between different countries, >99% of all isolates were susceptible to amoxicillin-clavulanate, ciprofloxacin, levofloxacin, and azithromycin. In 2000-2001, 840 clinical isolates of Haemophilus influenzae were collected from laboratories in France, Germany, Italy and Spain (210 isolates/country). β-Lactamase production among the isolates varied considerably by country, ranging from 8.1 % in Germany to 34.8% in France. H. influenzae from patients ≤4 years old showed the highest prevalence of β-lactamase production (23.2%), compared with isolates from patients aged 5-17 years (17.8%) and ≥18 years (16.5%). All isolates were susceptible to amoxicillin-clavulanate, ciprofloxacin and levofloxacin; 99.6% and 98.9% of isolates were susceptible to azithromycin and cefuroxime, respectively. Among the macrolides tested, azithromycin (MIC90, 2 mg/L) was eight-fold more potent than clarithromycin (MIC90, 16 mg/L) and roxithromycin (MIC90, 16 mg/L). Despite variations in β-lactamase production between different countries, >99% of all isolates were susceptible to amoxicillin-clavulanate, ciprofloxacin, levofloxacin, and azithromycin. Haemophilus influenzae is a common causative agent of otitis media in children and acute exacerbations of chronic bronchitis in the elderly, and is also isolated from patients with sinusitis and other community-acquired respiratory infections [1Berk SL Kalbfleisch JH The Alexander Project Collaborative Group. Antibiotic susceptibility patterns of community-acquired respiratory isolates of Moraxella catarrhalis in western Europe and in the USA.J Antimicrob Chemother. 1996; 38: 85-96Crossref PubMed Google Scholar, 2Credito KL Lin G Pankuch GA Bajaksouzian S Jacobs MR Appelbaum PC Susceptibilities of Haemophilus influenzae and Moraxella catarrhalis to ABT-773 compared to their susceptibilities to 11 other agents.Antimicrob Agents Chemother. 2001; 45: 67-72Crossref PubMed Scopus (41) Google Scholar]. β-Lactamase-mediated resistance in H. influenzae, encoded by TEM-1 and ROB-1, has been shown to account for most resistance to ampicillin, amoxicillin, and some oral cephalosporins [3Daum RS Murphey-Corb M Shapira E Dipp S Epidemiology of rob β-lactamase among ampicillin-resistant Haemophilus influenzae isolates in the United States.J Infect Dis. 1988; 157: 450-455Crossref PubMed Scopus (39) Google Scholar, 4Felmingham D Washington J The Alexander Project Group. Trends in the antimicrobial susceptibility of bacterial respiratory tract pathogens-findings of the Alexander Project 1992-1996.J Chemother. 1999; 11: 5-21Crossref PubMed Google Scholar, 5Livermore DM β-Lactamases in laboratory and clinical practice.Clin Microbiol Rev. 1995; 8: 557-584Crossref PubMed Google Scholar]. As a result, aminopenicillins, in the absence of a β-lactamase inhibitor, are not used as first-line therapies for community-acquired respiratory tract infections in some countries. The prevalence of β-lactamase production among H. influenzae strains has been shown to vary among European countries [6Sahm DF Jones ME Hickey ML Diakun DR Mani SV Thornsberry C Resistance surveillance of Streptococcus pneumoniae, Haemophilus influenzae and Moraxella catarrhalis isolated in Asia and Europe, 1997-1998.J Antimicrob Chemother. 2000; 45: 457-466Crossref PubMed Scopus (178) Google Scholar]. Differences in empirical prescription guidelines and the availability of antimicrobial agents may contribute to differences in resistance in certain countries. This report summarizes data from a surveillance study of H. influenzae isolates collected from clinical laboratories in four European countries in 2000-2001. During 2000-2001, 840 isolates of H. influenzae were prospectively collected from patient specimens at 20 hospital laboratories in France, Germany, Italy and Spain (from five laboratories in each country). The location of the participating laboratories was representative of the various geographic regions in each of the countries. Patient demographic information, including age, specimen source (blood, cerebrospinal fluid, respiratory, other) and patient location (inpatient, outpatient) was collected for each isolate. In total, 220 (26.2%) isolates were collected from patients ≤4 years old, 101 (12.0%) from patients 5-17 years old, 460 (54.7%) from patients ≥18 years old, and 59 (7.0%) from patients of unknown age. Isolates were submitted to our central laboratory (Focus Technologies, Herndon, VA, USA) and subcultured on chocolate agar. The identity of each isolate was confirmed by using standard laboratory methods [7Campos JM Haemophilus.in: Murray PR Baron EF Pfaller MA Tenover FC Yolken RH Manual of clinical microbiology. American Society for Microbiology Press, Washington, DC1999: 539-560Google Scholar], and each isolate was tested for the production of β-lactamase by use of the chromogenic substrate nitrocefin (BBL Dry Slide Nitrocefin, Becton Dickinson, Sparks, MD, USA). Antimicrobial susceptibility to ampicillin, amoxicillin-clavulanate, cefaclor, cefuroxime, azithromycin, clarithromycin, roxithromycin, ciprofloxacin and levofloxacin was tested by broth microdilution according to the National Committee for Clinical Laboratory Standards (NCCLS) guidelines with the use of frozen microdilution panels (TREK Diagnostics, Westlake, OH, USA). Minimum inhibitory concentrations (MICs) for H. influenzae were interpreted using the NCCLS M100-S11 recommendations, where available [8National Committee for Clinical Laboratory Standards.Performance standards for antimicrobial susceptibility testing. NCCLS, Wayne, PA2001Google Scholar]. The NCCLS-recommended quality control strains H. influenzae ATCC 49247 and H. influenzae ATCC 49766 were used throughout testing. Of the 840 H. influenzae isolates collected, 18.8% (158/840) were β-lactamase positive and ampicillin resistant (France, 34.8%; Germany, 8.1%; Italy, 8.6%; Spain 23.8%) (Table 1). All of the isolates tested were susceptible to amoxicillin-clavulanate, ciprofloxacin and levofloxacin. Cefuroxime susceptibility was >96% in each country (Table 1).Table 1Susceptibility of H. influenzae to antimicrobial agents according to β-lactamase statusFrancea137 β-lactamase-negative and 73 β-lactamase-positive isolates collected.Germanyb193 β-lactamase-negative and 17 β-lactamase-positive isolates collected.Italyc192 β-lactamase-negative and 18 β-lactamase-positive isolates collected.Spaind160 β-lactamase-negative and 50 β-lactamase-positive isolates collected.MIC90 (mg/L)%SeS = susceptible; I = intermediate; R = resistant.%IeS = susceptible; I = intermediate; R = resistant.%ReS = susceptible; I = intermediate; R = resistant.MIC90 (mg/L)%S%I%RMIC90 (mg/L)%S%I%RMIC90 (mg/L)%S%I%RAmpicillinAll1664.80.534.8191.908.1190.51.08.61675.70.523.8β-Lactamase-negative0.599.30.700.5100000.599.01.000.599.40.60β-Lactamase-positive1600100160010016001001600100Amoxicillin-clavulanateAll2100–fNCCLS breakpoints unavailable for categorical interpretation as susceptible, intermediate, or resistant.01100–01100–02100–0β-Lactamase-negative1100–00.5100–00.5100–01100–0β-Lactamase-positive2100–02100–02100–04100–0CefaclorAll3272.913.313.81687.110.52.41685.210.54.33271.911.416.7β-Lactamase-negative891.27.31.5890.77.81.61688.58.92.61683.18.18.8β-Lactamase-positive6438.424.737.03247.141.211.83250.027.822.26436.022.042.0CefuroximeAll210000299.50.50299.50.50496.73.30β-Lactamase-negative210000299.50.50299.50.50495.64.40β-Lactamase-positive410000410000110000210000AzithromycinAll299.5gOne isolate was azithromycin non-susceptible (MIC ≥ 8 mg/L).––2100––2100––299.0hTwo isolates were azithromycin non-susceptible (MICs ≥ 8 mg/L).––β-Lactamase-negative299.3––2100––2100––298.8––β-Lactamase-positive2100––2100––2100––2100––ClarithromycinAll1681.917.60.51682.916.70.51676.722.41.01677.621.41.0β-Lactamase-negative1686.113.10.71683.416.10.51677.122.40.5168019.40.6β-Lactamase-positive1674.026.001676.523.501672.222.25.6167028.02.0RoxithromycinAll16–––16–––16–––16–––β-Lactamase-negative16–––16–––16–––16–––β-Lactamase-positive16–––16–––16–––16–––CiprofloxacinAll0.008100––0.015100––0.008100––0.015100––β-Lactamase-negative0.008100––0.015100––0.008100––0.015100––β-Lactamase-positive0.015100––0.008100––0.015100––0.015100––LevofloxacinAll0.015100––0.015100––0.015100––0.015100––β-Lactamase-negative0.015100––0.015100––0.015100––0.015100––β-Lactamase-positive0.015100––0.015100––0.015100––0.015100––a 137 β-lactamase-negative and 73 β-lactamase-positive isolates collected.b 193 β-lactamase-negative and 17 β-lactamase-positive isolates collected.c 192 β-lactamase-negative and 18 β-lactamase-positive isolates collected.d 160 β-lactamase-negative and 50 β-lactamase-positive isolates collected.e S = susceptible; I = intermediate; R = resistant.f NCCLS breakpoints unavailable for categorical interpretation as susceptible, intermediate, or resistant.g One isolate was azithromycin non-susceptible (MIC ≥ 8 mg/L).h Two isolates were azithromycin non-susceptible (MICs ≥ 8 mg/L). Open table in a new tab Overall, H. influenzae isolates collected from patients ≤4 years old had the highest prevalence of β-lactamase production (23.2%). β-Lactamase production was >5% lower among isolates collected from patients aged 5-17 years (17.8%) and ≥18 years (16.5%) than among patients ≤4 years old; this pattern was observed in all countries. Macrolide activity was unaffected by β-lactamase production, as MIC distributions were similar for both β-lactamase-negative and β-lactamase-positive isolates (Figure 1). In all countries, azithromycin (MIC90, 2 mg/L; ≥99% susceptible) was more potent than clarithromycin (MIC90, 16 mg/L; 64 mg/L (confirmed by repeat testing), while clarithromycin and roxithromycin MICs were consistently higher (16, 32 and >128 mg/L for both agents; data not shown). The prevalence of β-lactamase production reported here (18.8%) is similar to that previously reported in other studies of H. influenzae in Europe [6Sahm DF Jones ME Hickey ML Diakun DR Mani SV Thornsberry C Resistance surveillance of Streptococcus pneumoniae, Haemophilus influenzae and Moraxella catarrhalis isolated in Asia and Europe, 1997-1998.J Antimicrob Chemother. 2000; 45: 457-466Crossref PubMed Scopus (178) Google Scholar, 9Biedenbach DJ Jones RN Pfaller MA The SENTRY Participants Group (Americas and Europe). Activity of BMS284756 against 2,681 recent clinical isolates of Haemophilus influenzae and Moraxella catarrhalis: report from The SENTRY Antimicrobial Surveillance Program (2000) in Europe, Canada and the United States.Diagn Microbiol Infect Dis. 2001; 39: 245-250Abstract Full Text Full Text PDF PubMed Scopus (25) Google Scholar, 10Schito GC Debbia EA Marchese A The evolving threat of antibiotic resistance in Europe: new data from the Alexander Project.J Antimicrob Chemother. 2000; 46: 3-9Crossref PubMed Scopus (133) Google Scholar]. The 2000 SENTRY study reported a β-lactamase prevalence of 12.6% [9Biedenbach DJ Jones RN Pfaller MA The SENTRY Participants Group (Americas and Europe). Activity of BMS284756 against 2,681 recent clinical isolates of Haemophilus influenzae and Moraxella catarrhalis: report from The SENTRY Antimicrobial Surveillance Program (2000) in Europe, Canada and the United States.Diagn Microbiol Infect Dis. 2001; 39: 245-250Abstract Full Text Full Text PDF PubMed Scopus (25) Google Scholar]. The 1998 Alexander Project, which included France, Germany and Italy among the 14 countries studied, reported the prevalence of β-lactamase production to be 11.6% [10Schito GC Debbia EA Marchese A The evolving threat of antibiotic resistance in Europe: new data from the Alexander Project.J Antimicrob Chemother. 2000; 46: 3-9Crossref PubMed Scopus (133) Google Scholar]. A 1997-98 European surveillance study performed by Sahm et al reported β-lactamase production among H. influenzae isolates collected from five countries to be 19.4% [10Schito GC Debbia EA Marchese A The evolving threat of antibiotic resistance in Europe: new data from the Alexander Project.J Antimicrob Chemother. 2000; 46: 3-9Crossref PubMed Scopus (133) Google Scholar]. On a per country basis, Sahm et al reported β-lactamase frequencies similar to those observed here: France, 27.6%; Germany, 5.7%; Italy, 7.7%; and Spain, 32.0% [10Schito GC Debbia EA Marchese A The evolving threat of antibiotic resistance in Europe: new data from the Alexander Project.J Antimicrob Chemother. 2000; 46: 3-9Crossref PubMed Scopus (133) Google Scholar]. In the USA, the frequency of β-lactamase production has been reported to be as high as 33% [11Thornsberry C Ogilvie PT Holley Jr HP Sahm DF Survey of susceptibilities of Streptococcus pneumoniae, Haemophilus influenzae, and Moraxella catarrhalis isolates to 26 antimicrobial agents: a prospective US study.Antimicrob Agents Chemother. 1999; 43: 2612-2623PubMed Google Scholar], which is similar to the frequencies observed in France and Spain. While overall frequencies of β-lactamase production in the USA have reached a plateau [12Thornsberry C Ogilvie P Kahn J Mauriz Y the Laboratory Investigator Group.Surveillance of antimicrobial resistance in Streptococcus pneumoniae, Haemophilus influenzae, and Moraxella catarrhalis in the United States in 1996-1997 respiratory season.Diagn Microbiol Infect Dis. 1997; 29: 249-257Abstract Full Text PDF PubMed Scopus (281) Google Scholar, 13Doern GV Brueggemann AB Pierce G Holley Jr, HP Rauch A Antibiotic resistance among clinical isolates of Haemophilus influenzae in the United States in 1994 and 1995 and detection of betalactamase-positive strains resistant to amoxicillin-clavulanate: results of a national multicenter surveillance study.Antimicrob Agents Chemother. 1997; 41: 292-297PubMed Google Scholar], the frequency detected here was slightly higher than in previous years, suggesting that, in some European countries, β-lactamase production may be increasing. Historically, azithromycin has been shown to be more active than erythromycin against Gram-negative organisms, due to the positive charge created by a methyl-substituted nitrogen in the 15-membered macrolide ring [14Farmer S Li ZS Hancock RE Influence of outer membrane mutations on susceptibility of Escherichia coli to the dibasic macrolide azithromycin.J Antimicrob Chemother. 1992; 29: 27-33Crossref PubMed Scopus (45) Google Scholar, 15Retsema J Girard A Schelkly W et al.Spectrum and mode of action of azithromycin (CP-62,993), a new 15-membered-ring macrolide with improved potency against gram-negative organisms.Antimicrob Agents Chemother. 1987; 31: 1939-1947Crossref PubMed Scopus (418) Google Scholar]. Based on the MIC90 in this study, azithromycin (2 mg/L in all countries) was more potent than both clarithromycin (16 mg/L in all countries) and roxithromycin (16 mg/L in all countries). Macrolide potency was unaffected by β-lactamase production. The MIC90s reported here for azithromycin were identical to those reported in the 1997-98 European surveillance by Sahm et al [6Sahm DF Jones ME Hickey ML Diakun DR Mani SV Thornsberry C Resistance surveillance of Streptococcus pneumoniae, Haemophilus influenzae and Moraxella catarrhalis isolated in Asia and Europe, 1997-1998.J Antimicrob Chemother. 2000; 45: 457-466Crossref PubMed Scopus (178) Google Scholar]; however, the clarithromycin MIC90 of 16 mg/L was one doubling dilution higher than this study, but identical to that reported in the 2000 SENTRY study [9Biedenbach DJ Jones RN Pfaller MA The SENTRY Participants Group (Americas and Europe). Activity of BMS284756 against 2,681 recent clinical isolates of Haemophilus influenzae and Moraxella catarrhalis: report from The SENTRY Antimicrobial Surveillance Program (2000) in Europe, Canada and the United States.Diagn Microbiol Infect Dis. 2001; 39: 245-250Abstract Full Text Full Text PDF PubMed Scopus (25) Google Scholar]. The MIC90 of roxithromycin was one dilution lower (16 mg/L) than reported in previous studies [2Credito KL Lin G Pankuch GA Bajaksouzian S Jacobs MR Appelbaum PC Susceptibilities of Haemophilus influenzae and Moraxella catarrhalis to ABT-773 compared to their susceptibilities to 11 other agents.Antimicrob Agents Chemother. 2001; 45: 67-72Crossref PubMed Scopus (41) Google Scholar, 16Wootton M Bowker KE Janowska A Holt HA MacGowan AP In-vitro activity of HMR 3647 against Streptococcus pneumoniae, Haemophilus influenzae, Moraxella catarrhalis and β-haemolytic streptococci.J Antimicrob Chemother. 1999; 44: 445-453Crossref PubMed Scopus (41) Google Scholar]. The MIC90S of both azithromycin and clarithromycin were identical to those reported in the USA during the 1990s [13Doern GV Brueggemann AB Pierce G Holley Jr, HP Rauch A Antibiotic resistance among clinical isolates of Haemophilus influenzae in the United States in 1994 and 1995 and detection of betalactamase-positive strains resistant to amoxicillin-clavulanate: results of a national multicenter surveillance study.Antimicrob Agents Chemother. 1997; 41: 292-297PubMed Google Scholar, 17Barry AL Pfaller MA Fuchs PC Packer RR In vitro activities of 12 orally administered antimicrobial agents against four species of bacterial respiratory pathogens from US medical centers in 1992 and 1993.Antimicrob Agents Chemother. 1994; 38: 2419-2425Crossref PubMed Scopus (171) Google Scholar, 18Hogan PA Sheehan DJ Macrolide susceptibility and β-lactamase production among Haemophilus influenzae isolates in the United States, 1996-1997.Antimicrob Agents Chemother. 1998; 42: 3313-3314PubMed Google Scholar], suggesting that the overall potencies of the macrolides against clinical isolates of H. influenzae have not changed in recent years. Three azithromycin-non-susceptible H. influenzae isolates were collected, one of which demonstrated an azithromycin MIC of >64 mg/L and clarithromycin and roxithromycin MICs of >128 mg/L. These unusually high MICs were also reported in the 1992-97 Alexander Project [19Felmingham D, Grüneberg RN, the Alexander Project Group.The Alexander Project 1996-1997: latest susceptibility data from this international study of bacterial pathogens from community-acquired lower respiratory tract infections.J Antimicrob Chemother. 2000; 45: 191-203Crossref PubMed Scopus (394) Google Scholar], as two isolates collected in 1997 were found to have azithromycin MICs ≥32 mg/L. Further work is necessary to determine the mechanism of resistance for these types of isolate. Overall, >99% of the H. influenzae isolates tested were susceptible to amoxicillin-clavulanate, ciprofloxacin, levofloxacin and azithromycin, despite variations in β-lactamase production among countries. This work was supported by Pfizer, New York, NY, USA. We thank David R. Diakun of Focus Technologies, Herndon, Pfizer, Inc., VA, USA for providing technical support in preparation of the manuscript.
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