a Microbiology Department, IDIBAPS, Hospital Clinic Universitari, c/Villarroel 170, 08036 Barcelona; b Microbiology Department, Hospital Gregorio Marañón, c/Doctor Esquerdo 46, 28007 Madrid; c Instituto Valenciano de Microbiología, Carretera de Bétera a San Antonio de Benageber Km 0.3, 46117 Valencia; d Medical Department, SmithKline Beecham Pharmaceuticals, c/Valle de la Fuenfría 3, 28034 Madrid, Spain
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Abstract |
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Introduction |
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We carried out a 1 year multicentre prospective study in order to assess the impact that geographical site, serotype, season of isolation and patient age might have on the antimicrobial susceptibility pattern of S. pneumoniae isolated from patients suffering from community-acquired respiratory infections in Spain.13
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Materials and methods |
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Between May 1996 and April 1997, all consecutive S. pneumoniae isolates obtained from patients suffering from upper or lower community-acquired respiratory tract infections (only one isolate from each patient) in 14 Spanish hospitals were collected.13
Each centre kept isolates at 70°C until used. Isolates were plated on enriched medium, incubated overnight at 3537°C and sent to a central laboratory (Instituto Valenciano de Microbiología, Valencia, Spain) along with epidemiological data such as age group of the patient (paediatric or adult), date of the sample, hospital, origin and source of the sample. Samples processed included middle ear exudate, pleural fluid, blood, sputum and bronchoscopy specimens.
Isolates were serotyped at the Instituto Valenciano de Microbiología and in the Servicio de Microbiología del Centro Nacional de Microbiología del Instituto de Salud Carlos III de Majadahonda (Madrid, Spain) using Quellung (Staten Seruminstitut, Copenhagen, Denmark) and dot blot analyses, respectively.14,15
Antibiotics used and antibiotic susceptibility studies
MICs were determined by a semiautomatic microdilution test (Sensititre, West Sussex, UK) following the recommended NCCLS guidelines,16 with the antibiotics most frequently used as empirical treatment for respiratory tract infections in Spain, namely penicillin, amoxycillin, amoxycillinclavulanic acid, cefixime, cefaclor, cefuroxime, cefotaxime, ceftriaxone, erythromycin, clarithromycin, azithromycin and ciprofloxacin. Wide antibiotic concentration ranges were chosen including a two-fold dilution higher and lower than the breakpoints of resistance and susceptibility to the antibiotics tested. The control strains used were Streptococcus pneumoniae ATCC 49619 and Staphylococcus aureus ATCC 29213.
The breakpoints used to calculate rates of resistance were 2 mg/L for penicillin, amoxycillin, amoxycillin clavulanic acid, cefotaxime, ceftriaxone, cefuroxime and azithromycin and
1 mg/L for clarithromycin and erythromycin.
Statistical methods
To compare the proportions obtained, a 2 test was used with YatesBonferroni adjustments (multiple comparisons) when necessary. Calculations were carried out with the software EpiInfo 6.02 (CDC, Atlanta, GA, USA).
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Results |
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Serotype 3 was, as expected, the most susceptible to antibiotics, in terms of both MIC90s (0.25 mg/L) and rates of resistance (<5%). Serotype 14 showed the highest rates of resistance (
50%) (Table I
). The prevalence of resistance to macrolides was
35% for all the main serotypes except serotypes 3, 9 and 11.
The non-typeable pneumococci showed penicillin and erythromycin resistance rates of 37.4% and 48.6%, respectively (Table I). In the 225 pneumococci of the less frequent serotypes, rates of resistance to penicillin and erythromycin were low (1.8% and 8.5%, respectively). The patterns of distribution of penicillin and erythromycin MICs differed among serotypes (Figures 1 and 2
).
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In paediatric isolates resistance to penicillin and cefuroxime was statistically significantly more prevalent than that from adults in internal medicine wards (50% and 37%, respectively, for penicillin; 62.4% and 45.6%, respectively, for cefuroxime; P < 0.05). There was no statistically significant difference between paediatric and adult isolates in terms of resistance to amoxycillin, amoxycillinclavulanic acid, third generation cephalosporins or macrolides (Table II).
Ninety (8%) of the samples came from the middle ear; of these, 69% were from children. The most frequent serotypes found in middle ear samples were again serotypes 6 (24.4%) and 19 (25.6%). In respiratory samples, other serotypes, serotype 6 and non-typeable isolates were most frequently found. Serotypes belonging to the category other and serotype 3 were the most common from blood cultures, accounting for up to 31% and 12.6% of the isolates, respectively. Serotype 11 (which is very similar to serotype 3 in its penicillin and erythromycin susceptibilities) represented 3.5% of blood pneumococci, so almost 50% of the blood isolates were in the susceptible category. Pneumococci isolated from middle ear fluids (n = 90) showed the highest prevalence of resistance, and those from blood cultures (n = 175) the lowest, with a statistical significance only for macrolides (P < 0.05) and not for ß-lactams.
Geographical variations in the susceptibility to penicillin or erythromycin are shown in Table IV. Strains from the Hospital Insular de Las Palmas (Canary Islands) and those from the Hospital Xeral de Santiago de Compostela (Galicia) were more susceptible to antibiotics (
31% of strains showing high resistance to any antibiotic). Strains from the Hospital Santa Creu i Sant Pau de Barcelona and from the Hospital Basurto de Bilbao exhibited the highest rates of resistance.
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Discussion |
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The awareness that respiratory isolates differ from meningeal isolates influences choice of appropriate breakpoints, given that for respiratory tract infections the outcome is the same whether the strains are susceptible or intermediately resistant to penicillin.17 This is acknowledged in the newly published NCCLS breakpoints for amoxycillin, amoxycillinclavulanate, cefuroxime and oral cephalosporins.18 Our isolates were obtained before current NCCLS amendments were published, and breakpoints for parenteral cephalosporins remain unchanged (and are even lower than those of oral ß-lactams) so, to maintain consistency in the comparison of results, we have used the former breakpoints to categorize susceptibility. Using the new breakpoints for full resistance, (8 and
4 mg/L for amoxycillin (alone or with clavulanate) and oral cephalosporins, respectively) throughout the pneumococcal population, resistance rates of 4.85% for amoxycillin (alone or with clavulanate) and 34.86% for oral cefuroxime are obtained.
The distribution by serotype differs slightly from that found in other studies carried out in Spain.2,8 Some of the serotypes, such as serotypes 3, 6 and 9, retain their epidemiological importance but the prevalence of others, such as serotypes 1, 5, 8 and 7, which in previous studies were >5%,2,8 did not exceed 2% in this study, although the different geographical provenance of the samples in the different studies should be borne in mind.
Serotypes 6, 19 and 23 were the most common in both groups, although with different prevalence rates (13.6%, 11.5% and 9.9%, respectively, for adults, and 24.0%, 19.2% and 13.6%, respectively, for children). No single serotype showed a clear predominance in adults, but three serotypes made up 55% of the total in children, in agreement with a previous report.3
The association between the resistance to penicillin and serotype is well known. In this study, serotype 3 was particularly susceptible, as expected, to all the antibiotics tested, with resistance rates of <5%. The rates of resistance among the other more prevalent serotypes were higher, although without a homogeneous pattern: serotypes 6 and 9 both had penicillin resistance rates of >50%, but differed in their resistance to erythromycin (68% and 12%, respectively). In serotype 19, the rate of resistance to penicillin and erythromycin was 33% and 42%, respectively. Serotypes 14 and 23 both showed a high prevalence of resistance to oral ß-lactams and macrolides (above 40%). In the light of these results, it seems that the widely held opinion that resistance to penicillin is associated with resistance to other antibiotics may require modification, as our data suggest that this may only be true for some serotypes (serotypes 9 and 15).
The differences between participating hospitals in the prevalence of resistance might result from differences in the populations studied, as the proportion of paediatric samples differed between centres, and to environmental factors, such as local differences in consumption of antibiotics. Nevertheless, the huge sample size together with the large number of hospitals involved and the short period of time (1 year) during which the study was conducted might be expected to have outweighed bias. The wide range in the number of isolates submitted from each hospital should also be noted; obviously, the fewer isolates per centre the higher the potential bias in reporting resistance.
Although statistically significant seasonality has already been reported for penicillin resistance,13 this has not been reported for oral or parenteral ß-lactams. One of the most interesting findings in this study is the existence of a seasonal pattern of resistance to oral ß-lactams, and, to a much lesser extent, parenteral third generation cephalosporins. This phenomenon may be a result of the seasonal variability of isolation of serotypes over the 12 months of study.
Our population MIC data for the different serotypes show the existence of different pneumococcal subpopulations on the basis of their susceptibility to penicillin and erythromycin. This adds to the multifactorial complexity of penicillin or erythromycin resistance in S. pneumoniae.
Whether penicillin susceptibility is of clinical importance in the treatment of respiratory infections is controversial, but it seems that at least for pneumococcal strains with a penicillin MIC 2 mg/L, penicillin can be safely used in treating pneumonia. Nevertheless, clinical resistance is not the same as loss of susceptibility. It is the slow and progressive shift in MIC for a given bacterial population (loss of susceptibility) that allows clinically susceptible strains to accumulate progressive mutations that would ultimately render them clinically resistant.
If practical and rational antibiotic policies are to be developed with the aim of helping doctors to prescribe more suitable empirical antibiotics, then national multicentre surveillance studies that take into account and integrate regional data seem more useful than multinational studies that can only evaluate nationally collected raw data, which may differ between countries. Evidently, regional studies would be even more accurate than national surveillance, but results would also be more difficult to generalize (accuracy does not mean usefulness). Given the disparity between the extreme accuracy of local studies and the extreme generalization of results in multinational studies, intermediate surveillance studies such as national multicentre projects seem mandatory if relevant susceptibility results are sought for the empirical antibiotic treatment of any infectious condition.
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Acknowledgments |
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Notes |
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Members of the Spanish Surveillance Group for Respiratory Tract Pathogens are listed in the Acknowledgements.
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References |
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Received 28 January 2000; returned 24 April 2000; revised 8 May 2000; accepted 9 June 2000