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Antimicrobial Agents and Chemotherapy, November 2001, p. 3226-3228, Vol. 45, No. 11
Microbiology Department, IDIBAPS,
Hospital Cliníc, Barcelona1;
Instituto Valenciano de Microbiología,
Valencia2; Medical Department,
GlaxoSmithKline, Tres Cantos, Madrid3;
Microbiology Department, Hospital General Universitario
Gregorio Marañón, Madrid4; and
Microbiology Department, Residencia Marqués de
Valdecilla, Santander,5 Spain
Received 28 December 2000/Returned for modification 25 June
2001/Accepted 8 August 2001
A The aim of the present study is to
describe the current susceptibility patterns of Haemophilus
influenzae to first-line antibiotics used for respiratory tract
infections, as well as to describe the prevalence of the new phenotypes
of antibiotic resistance in Spain.
A total of 1,730 consecutive clinical isolates of H. influenzae obtained from community-acquired respiratory tract
infections, collected between November 1998 and October 1999, were
included in this prospective surveillance study involving 17 hospitals selected on the basis of geographic location. Strains were sent to a
central laboratory (Instituto Valenciano de Microbiología, Valencia, Spain) to confirm the identification by colony morphology, Gram staining, growth in chocolate but not in blood agar, catalase test, and X and V factor requirements. In addition, a H. influenzae ATCC 49247, H. influenzae ATCC
49766, and Escherichia coli ATCC 35218 were used as quality
control strains following NCCLS recommendations (7). BLNAR
strains were defined as those strains failing to detect A total of 1,534 samples (88.6%) were collected from the lower
respiratory tract (sputum and bronchoalveolar lavage), 26 (1.5%) were
from hemoculture, and 170 (9.8%) were from middle ear exudate. The
vast majority of isolates (97.3%) were not typeable, whereas serogroups b, d, e, and f represented just 1.2, 0.2, 0.2, and 1%,
respectively. The different production of Prevalence of susceptibility, and MICs for 50 and 90% of strains
(MIC50 and MIC90) are shown in Table
1. According to NCCLS breakpoints, all
antibiotics tested exhibited susceptibility rates equal to or higher
than 99% except for ampicillin (70%), clarithromycin (89%), and
cefaclor (91%). The antibiotics with lowest intrinsic potency (as
measured by MIC90) happened to be penicillin, amoxicillin, ampicillin, cefaclor, erythromycin, and clarithromycin, all of them
displaying an MIC90 of
0066-4804/01/$04.00+0 DOI: 10.1128/AAC.45.11.3226-3228.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
Antimicrobial Susceptibilities of 1,730 Haemophilus influenzae Respiratory Tract Isolates in Spain
in 1998-1999
ABSTRACT
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-lactamase prevalence of 23% was found among 1,730 Haemophilus influenzae isolates. Ampicillin
susceptibility was 70%, and 12% of -lactamase-negative strains
presented diminished susceptibility to ampicillin (BLNAR phenotype).
Susceptibility of 90% was found for cefaclor and clarithromycin,
whereas it was nearly 100% for cefotaxime, cefixime, azithromycin, and
cefuroxime. Ciprofloxacin-resistant (0.1%) and -lactamase-positive
amoxicillin/clavulanate-resistant (BLPACR) phenotypes (0.1%) are
anecdotal so far.
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-lactamase test (nitrocefin; Becton Dickinson) was performed (9).
Susceptibility testing was performed following the National Committee
for Clinical Laboratory Standards (NCCLS) 1999 guidelines using a
semiautomated microdilution (7) method (Sensititre; Trek
Diagnostics Inc., Westlake, Ohio) against antimicrobials commonly used
as empiric therapy of respiratory tract infections in Spain. These were
penicillin, amoxicillin, ampicillin, amoxicillin-clavulanate, cefaclor,
cefuroxime, cefixime, cefotaxime, erythromycin, clarithromycin,
azithromycin, and ciprofloxacin.
-lactamase
production and presenting with either a high (
4 µg/ml) or
intermediate (2 µg/ml) ampicillin MIC (3). On the other
hand, -lactamase-positive amoxicillin-clavulanate-resistant (BLPACR)
strains were defined as those with an MIC to amoxicillin-clavulanate of
8 µg/ml that rendered positive with the nitrocefin disk test (3). Differences between -lactamase-positive and
-lactamase-negative strains with respect to the prevalence of
susceptibility were calculated using comparison of proportions with the
chi square test. Estimation of 95% confidence intervals (95%CI) for
population proportions was made by the exact binomial method. Epi-Info
version 6.04 was used for statistical calculations.
-lactamase in hemoculture (30.7%) in comparison with middle ear samples (23.5%) and lower respiratory tract samples (22.8%) did not show a significant
statistical difference.
8 µg/ml. The remaining
antibiotics for which there are NCCLS breakpoints presented a far
better intrinsic potency, and as expected, third-generation
cephalosporins and ciprofloxacin showed the lowest MIC90.
TABLE 1.
MIC50, MIC90, range of MICs, and
susceptibility of 1,730 clinical isolates of H. influenzaea
Nearly one of every four isolates (399 of 1,730) was a -lactamase
producer (23%; 95%CI, 21 to 25%), a figure very much like that found
in our previous surveillance study (25.7%) (4), but
significantly lower than that reported in Spain by other national (10) and multinational (11) studies, around
30 to 35%. Considering geographical differences in our 17 centers,
-lactamase production ranged from 15.8 to 30.4%, but no statistical
significance was reached. The -lactamase production status of the
isolates did not influence their susceptibility to the antibiotics
tested except for ampicillin (9.7% for -lactamase positive versus
87.9% for -lactamase negative; P < 0.0001) and
cefaclor (70.4% for -lactamase positive versus 97.7% for
-lactamase negative; P < 0.0001). Cefaclor was the
sole oral cephalosporin for which -lactamase production influenced
the prevalence of susceptibility.
Concerning -lactamase-negative isolates, 87 (6.5%) and 74 (5.5%)
strains were found to be intermediate and resistant to ampicillin, respectively, and were therefore categorized as BLNAR. This means that 9.3% of the whole H. influenzae population of the
study were BLNAR (95% CI, 8 to 10.7%), suggesting an increase
compared with previous reports in Spain of around 5%
(3,10). Again, no significant differences in the
prevalence of BLNAR strains were seen on a geographical basis.
Macrolide nonsusceptibility was around 1% for azithromycin and 10% for clarithromycin, most of this being nonsusceptibility categorized as intermediate resistance. In any case, clarithromycin was significantly less active than azithromycin in terms of MIC90 values (16 µg/ml versus 2 µg/ml).
Regarding the extremely rare ciprofloxacin resistance phenotype, we found only two isolates (0.1%) with MICs of 2 and 4 µg/ml, which is in accordance with other authors (1). Likewise, only two isolates (0.1%) were found to be BLPACR, which is less than reported elsewhere (3).
H. influenzae is one of the most prevalent isolates in
community-acquired respiratory tract infections. Multicenter
surveillances have had key importance in ascertaining -lactamase
production (2,4,5,8,10), decreasing susceptibility to
certain macrolides (4), and increasing intrinsic
resistance to ampicillin mediated by altered penicillin-binding
proteins (PBP) (6), the so-called
-lactamase-negative ampicillin resistance (BLNAR). Likewise,
surveys have revealed novel resistance phenotypes as very rare
ciprofloxacin-resistant (1) and
-lactamase-positive and amoxicillin-clavulanate-resistant
(BLPACR) isolates (3).
Our survey shows that -lactamase production in Spain (23%) is
homogeneously distributed and seems to be decreasing compared with
other reports, but ampicillin susceptibility hardly changes (69.9%;
95% CI, 67.7 to 72%). However, the prevalence of the BLNAR phenotype
seems to be increasing (9.3%), and sharp differences in the activity
of macrolides are also evident. Both BLPACR and ciprofloxacin-resistant
phenotypes constitute to date just anecdotal events (0.1%), but they
must be monitored. Regarding BLNAR strains and following NCCLS, the
question is open as to what extent ampicillin-intermediate strains
should be considered resistant to the remaining -lactams antibiotics
within the family.
Differences in susceptibility patterns between different surveillances over time due to the increasing prevalence of certain phenotypes of resistance or the emergence of new ones make national multicenter surveys a must. Most of the multicenter surveillances of resistance carried out so far have as a main criticism that they make their estimations based either on a relatively small number of isolates from a single country or on a large number of isolates from many different countries. Both attitudes cannot but lead to a somewhat inadequate representation of the complex reality of the resistance dynamics for a given country. That is why extensive national multicenter surveillances provide more reliable information, ensuring a better estimation for the country whose specific information on resistance is sought.
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FOOTNOTES |
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* Corresponding author. Mailing address: Medical Department, GlaxoSmithKline, C/Severo Ochoa, 2, 28760 Tres Cantos, Madrid, Spain. Phone: 34 91 807 5912. Fax: 34 91 807 0596. E-mail: lorenzo.aguilar-alfaro{at}gsk.com.
Members of the Spanish Surveillance Group for Respiratory Pathogens
are: E. Cercenado, Hospital Gregorio Marañón, Madrid; T. Jiménez de Anta, Hospital Clínic, Barcelona; A. García-Perea, S. García, and M. Güeni, Hospital
La Paz, Madrid; E. Pérez-Trallero and J. Larruskain, Complejo
Hospitalario Donostia, San Sebastián; J. Barrón and L. López, Hospital de Cruces, Baracaldo; C. Rubio and C. García, Hospital Clínico Universitario, Zaragoza;
J. A. García-Rodríguez (Study Coordinator) and I. Trujillano, Hospital Clínico Universitario, Salamanca; J. Ruiz and E. Simarro, Hospital Virgen de la Arrixaca, Murcia; C. García-Riestra, B. Regueiro, A. Jato, and M. Prieto, Hospital
Clínico Universitario, Santiago de Compostela; J. M. Nogueira, Hospital Dr. Peset, Valencia; C. Latorre and A. Gené,
Hospital Sant Joan de Deu, Barcelona; M. de la Rosa, Hospital Virgen de
las Nieves, Granada; E. Perea and L. Martínez, Hospital Virgen
de la Macarena, Sevilla; A. M. Martín and F. Cañas, Hospital Insular, Las Palmas; M. Casal and A. Ibarra, Hospital Reina Sofía, Córdoba; D. Romero and M. González, Hospital Nuestra Señora de Alarcos, Ciudad Real;
F. Baquero (Study Coordinator), Hospital Ramón y Cajal, Madrid;
J. J. Granizo, Fundación Jiménez Díaz,
Madrid; L. López and C. Gimeno, Instituto Valenciano de Microbiología, Valencia; and R. Dal-Ré,
GlaxoSmithKline, Tres Cantos, Madrid, Spain.
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Antimicrobial Agents and Chemotherapy, November 2001, p. 3226-3228, Vol. 45, No. 11
0066-4804/01/$04.00+0 DOI: 10.1128/AAC.45.11.3226-3228.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
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