Serotype distribution and antimicrobial resistance of Streptococcus pneumoniae in Austria

Astrid Buxbaum1, Sabine Forsthuber2, Wolfgang Graninger2 and Apostolos Georgopoulos2,* on behalf of The Austrian Bacterial Surveillance Network{dagger}

1 Department of Internal Medicine IV, Division of Pulmology, and 2 Department of Internal Medicine I, Division of Infectious Diseases and Chemotherapy, University of Vienna, Währinger Gürtel 18–20, 1090 Vienna, Austria

Received 30 December 2003; returned 5 March 2004; revised 11 March 2004; accepted 28 March 2004


    Abstract
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Footnotes
 Acknowledgements
 References
 
Objectives: To determine the prevalence of antibiotic resistance and the distribution of serotypes among Streptococcus pneumoniae isolated in Austria.

Materials and methods: A total of 2367 strains of S. pneumoniae were collected in an Austrian-wide surveillance system between 1996 and 2002. Isolates were tested for their susceptibility to penicillin and clarithromycin and were serotyped by the capsular swelling method.

Results: An overall rise in penicillin resistance was observed from 4.9% in 1996 to 10.0% in 2002 (including both intermediate-resistant and resistant strains). A rise in clarithromycin resistance was also recorded in this period. The overall distribution of serogroups/types remained relatively stable, with 23, 19, 6 and 14 being the most frequent ones. Whereas in 1996 penicillin resistance was predominantly associated with serotype 23F, in 1998 and 2002, resistance was most frequently found in isolates of serogroup 9 and serotype 14, respectively. Coverage rates for currently available vaccines ranged from 57.4% (7-valent) to 72.4% (23-valent) of all serotyped strains.

Conclusions: This rise in pneumococcal resistance to penicillin and clarithromycin, and the change in distribution of serotypes in these resistant strains, indicates that ongoing surveillance programmes are warranted, in order to be able to formulate both effective vaccination strategies and optimal antibiotic therapies.

Keywords: serogroups , pneumococci , adults , vaccines


    Introduction
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Footnotes
 Acknowledgements
 References
 
Since the first description of a strain of Streptococcus pneumoniae with diminished penicillin susceptibility in 1967, resistance among pneumococci has spread worldwide. Furthermore, macrolide- and quinolone-resistant pneumococcal strains have been reported with increasing frequency, with macrolide-resistance rates up to 40%.1,2 Pneumococci are divided into >90 serogroups or serotypes on the basis of the immunochemistry of their capsular polysaccharides. Classically, some are particularly associated with invasive disease, whereas others are mainly associated with carriage and non-invasive disease.3 Also, pneumococcal strains that are resistant to penicillin appear to belong predominantly to a few selected serotypes, particularly 6B, 14, 19F and 23F. The emergence and spread of these often multiresistant strains have become a major concern worldwide and are seriously challenging current treatment strategies.4

Attention is now being focused on prevention, especially through widespread vaccination. Currently available vaccines cover 23 serotypes, which represent approximately 90% of the strains responsible for invasive disease, but are not immunogenic in young children. New conjugate vaccines, which are immunogenic in children <2 years of age, cover fewer serotypes (7-, 9- and 11-valent).5 Since pneumococci have the genetic capacity to switch serotype by horizontal transfer and recombination or other genetic events, knowledge of the frequency of these serotype exchanges is important to predict the long-term efficacy of new conjugate vaccines.

Continuous monitoring of both antimicrobial resistance and serotype distribution of S. pneumoniae remains therefore essential for the design of vaccination and antibiotic treatment plans. The Austrian Bacterial Surveillance Network was established in 1996 with the aim of providing nationwide data on bacterial epidemiology and antimicrobial resistance. This study describes the distribution of serotypes and the development of penicillin and clarithromycin resistance in S. pneumoniae in Austria from 1996 to 2002.


    Materials and methods
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Footnotes
 Acknowledgements
 References
 
Bacterial strains

S. pneumoniae strains isolated in the years 1996, 1998 and 2002 were collected at each participating centre in the Austrian Bacterial Surveillance Network and sent to the reference laboratory at the University Hospital of Vienna. In total, 26 centres participated in this study. Only one isolate per patient was accepted, thus excluding duplicates. Strains from patients under 18 years of age were not included in this study. Isolates were obtained from the respiratory tract, blood, CSF and other sites. In 1996, 1385 strains were collected and tested for their penicillin susceptibility, of which 385 were serotyped, while in 1998, 440 isolates were susceptibility tested and 161 serotyped, and in 2002, 542 isolates were susceptibility tested, and 193 were serotyped. When selecting strains for serotyping, all penicillin-intermediate and penicillin-resistant strains and a representative number of randomly selected penicillin-susceptible pneumococci (corresponding to the relative size of the participating hospital) were chosen each year.

Minimal inhibitory concentrations (MICs)

MICs were determined using the NCCLS broth microdilution method with Mueller–Hinton broth, supplemented with 5% lysed horse blood.6,7 Standard quality control strains (S. pneumoniae ATCC 49619 and 33400) were included in each run. Isolates were tested against penicillin G and clarithromycin. The susceptibility and resistance percentages of the isolates were determined using the breakpoints recommended in the 2003 NCCLS guidelines.

Serotyping

The strains were serotyped by the Quellung reaction with the Pneumotest antisera panel (Staten Serum Institut, Copenhagen, Denmark).


    Results
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Footnotes
 Acknowledgements
 References
 
Minimal inhibitory concentrations

Susceptibilities of the pneumococci to penicillin and clarithromycin are presented in Table 1. There were no distinct regional differences between the participating centres regarding susceptibility status of the isolates collected between 1996 and 2002 (data not shown). Penicillin-intermediate and -resistant isolates comprised 4.9% (2.9% and 2.0%, respectively) of all isolates in 1996; the prevalence of clarithromycin-resistance was also comparatively low at 3.2%. In 1998, a rise in penicillin resistance to 9.9% was recorded, accompanied by an increase in clarithromycin resistance, to 7.6%. This trend was also observed in 2002, with both penicillin and clarithromycin resistance having increased to about 10%. The percentage of strains resistant to both penicillin and clarithromycin increased from 27% in 1996 to 63% in 2002 (data not shown).


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Table 1. Susceptibility of 2367 pneumococci to penicillin and clarithromycin

 
Serotyping

The overall distribution of pneumococcal serotypes was consistent among the participating centres (data not shown). Between 1996 and 2002, this distribution remained stable with serogroups 19 (13.9%), 23 (11.9%), 6 (10.4%), 14 (9.2%), 3 (8.1%) and 9 (7.8%) being predominant. In 1996, penicillin resistance was predominantly associated with serogroups 23 (33.2%), 19 (21.6%), 15 (13.7) and 6 (9.8%). In 1998, serogroups 9 (32.4%), 19 (11%), 14 (10.7) and 23 (10.7) were most frequent, while in 2002, serogroups 14 (32.5%), 6 (18%), 19 (13.5%) and 23 (9%) were predominant. Predominant serogroups of clarithromycin-resistant pneumococci were 19 (1996), 9 (1998) and 14 (2002). Serogroup 14 was also observed in the majority of strains resistant to both penicillin and clarithromycin (data not shown).

Vaccine coverage

The proportions of pneumococcal isolates covered by the 7-, 9- and 11-valent conjugate vaccines were 57.4%, 59.2% and 70.6%, respectively, when including all pneumococci regardless of their penicillin susceptibility. The 23-valent vaccine was able to cover 72.4% of all isolates. Percentages for penicillin-intermediate and -resistant isolates were comparable (Table 2).


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Table 2. Proportion of pneumococcal isolates covered by vaccines

 

    Discussion
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Footnotes
 Acknowledgements
 References
 
Over the past two decades, the emergence of pneumococci with decreased susceptibility to ß-lactam and non-ß-lactam antibiotics has emphasized the need for new therapeutic agents.8 Besides the 90 capsular serotypes already recognized, serotype distribution varies annually, and with geographical area, disease and age. Because of these considerations, the Austrian Bacterial Surveillance Network was set up in 1996 to promote the surveillance of bacterial infections and to provide nationwide bacteriological surveillance data.

In 1996, the start of the surveillance programme, 4.9% of all tested pneumococci proved to be intermediate or resistant to penicillin and 3.2% to clarithromycin. The prevalence of penicillin-resistant pneumococci obtained in this study was surprisingly low when compared with the data reported from other central European countries like Hungary.5 This has been attributed to the strict Austrian antibiotic policy which advocates high-dose treatment regimens, and to the moderate consumption of ß-lactams by the Austrian community. In 1998, a dramatic rise in penicillin resistance was recorded with a doubling of the rate to nearly 10%, accompanied by an increase in clarithromycin resistance. These findings were confirmed in 2002, when pneumococcal resistance to both penicillin and clarithromycin crossed the 10% threshold for the first time. Also notable was the increase in strains with intermediate penicillin resistance with MIC values close to high-level resistance.

Penicillin-non-susceptible pneumococci frequently belong to serogroups 6, 9, 14 and 23.5 These serogroups have also been associated with carriage. In Austria, a change in serotype distribution was recorded during the observation period. Whereas in 1996, serotype 23F was predominantly associated with penicillin resistance, in 1998 serogroup 9 and in 2002 serogroup 14 were predominant among penicillin-non-susceptible strains. At the same time, serogroups predominant among clarithromycin-resistant pneumococci changed from 19 (1996) to 9 (1998) and then to 14 (2002). Serotype 14 seems to be mainly associated with penicillin-intermediate resistant strains at the border of high-level resistance.

In Austria, one of the currently available vaccines is the 23-valent polysaccharide vaccine that is used in the USA and Europe. The coverage rate of this vaccine in the years 1996–2002 was approximately 70% and remained stable within this period. The newly licensed 7-valent conjugate vaccine specifically targets serotypes commonly associated with penicillin resistance that infect infants and young children. Coverage for this new vaccine was slightly lower than for the 23-valent vaccine. The 9-valent and 11-valent vaccines did not markedly increase the coverage rate for penicillin-non-susceptible strains. These figures are in keeping with findings of other European and US studies and indicate that the new conjugate vaccines would be suitable formulations for use in Austria.9

Some authors argue that future widespread vaccination programmes will exert significant evolutionary pressure on the bacteria, resulting in the replacement of serotypes covered by the vaccine by strains carrying novel capsular types and perhaps leading to the spread of resistance genes to strains with capsules not covered by the vaccine.10 Nevertheless, increasing antibiotic resistance of S. pneumoniae and the speed of the spread of resistant clones strongly supports the need to consider vaccination as the main measure in fighting pneumococcal infections. The long-term solution for control of the spread of pneumococcal resistance to antibiotics can only be prevention.11 Given the potential disparity between the results of local studies and those of multinational studies, intermediate studies such as national multicentre studies like the Austrian Bacterial Surveillance Network seem mandatory if relevant data are to be obtained. These data will be vital not only in tracking the epidemiology of pneumococcal serotypes and the emergence of drug-resistant strains, but also in assessing the impact of measures such as vaccination programmes, and ensuring that vaccines are based on serotypes found in strains causing disease.


    Acknowledgements
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Footnotes
 Acknowledgements
 References
 
We would like to thank W. Schmidt and K. Stich for excellent technical assistance. This study was partially supported by grant ‘Jubiläumsfondprojekt Nr. 7024’ from the Austrian National Bank.

The Austrian Bacterial Surveillance Network comprises: A. Hirschl, D. Wolf, W. Ulrich, U. Setinek-Liszka (Vienna); F. Allerberger (Innsbruck); W. Sixl, A. Bojatzis (Graz); M. Müller (Salzburg); E. Grund (Klagenfurt); G. Alpi (Villach); G. Leitner (Leoben); W. Pflanzl (Oberwart); W. Aichinger (Wels); J. Feichtinger (Steyr); H. Gogl (Vöcklabruck); G. Brinninger (Ried); W. Öhlinger (Krems); L. Gerstner (Mistelbach); W. Stiglbauer (Wr.Neustadt); M. Drlicek (Linz).


    Footnotes
 
* Corresponding author. Tel: +43-1-40400-5139; Fax: +43-1-40400-5200; Email: apostolos.georgopoulos{at}akh-wien.ac.at

{dagger} Members of The Austrian Bacterial Surveillance Network are listed in the Acknowledgements. Back


    References
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 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Footnotes
 Acknowledgements
 References
 
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6 . National Committee for Clinical Laboratory Standards (2003). Methods for Dilution Antimicrobial Tests for Bacteria that Grow Aerobically: Approved Standard M7-A6 NCCLS, Wayne, PA, USA.

7 . National Committee for Clinical Laboratory Standards (2003). Performance Standards for Antimicrobial Susceptibility Testing: Supplemental Tables, M100-S13/M7 NCCLS, Wayne, PA, USA.

8 . McGee, L., Goldsmith, C. E. & Klugman, K. P. (2002). Fluoroquinolone resistance among clinical isolates of Streptococcus pneumoniae belonging to international multiresistant clones. Journal of Antimicrobial Chemotherapy 49, 173–6.[Abstract/Free Full Text]

9 . Eskolja, J., Kilpi, T. & Palmu, A. (2001). Efficacy of a pneumococcal conjugate vaccine against acute otitis media. New England Journal of Medicine 344, 403–9.[Abstract/Free Full Text]

10 . Mbelle, N., Hubner, R. E., Wasas, A. D. et al. (1999). Immunogenicity and impact on nasopharyngeal carriage of a nonavalent pneumococcal conjugate vaccine. Journal of Infectious Diseases 180, 1171–6.[CrossRef][ISI][Medline]

11 . Marco, F., Bouza, E., Garcia-de-Lomas, J. et al. (2000). Streptococcus pneumoniae in community-acquired respiratory tract infections in Spain: the impact of serotype and geographical, seasonal and clinical factors on its susceptibility to the most commonly prescribed antibiotics. Journal of Antimicrobial Chemotherapy 46, 557–64.[Abstract/Free Full Text]