Synopsis

Antimicrobial Resistance with Streptococcus pneumoniae in the United States, 1997–98

Gary V. Doern, Angela B. Brueggemann, Holly Huynh, Elizabeth Wingert, and Paul Rhomberg
University of Iowa College of Medicine, Iowa City, Iowa, USA

Before 1990, most clinical isolates of Streptococcus pneumoniae in the United States were susceptible to a variety of antimicrobial drugs, including penicillin (1,2). In the early 1990s, however, antimicrobial resistance began to emerge (3-7), and ß-lactam resistance as a result of altered penicillin binding proteins was recognized (8-11). Resistance to other non-ß-lactam drugs, such as the macrolides, clindamycin, tetracycline, chloramphenicol, and trimethoprim/sulfamethoxazole (TMP/SMX), began to increase (4-6). Therapeutic failures in patients with pneumococcal infections treated with previously effective drugs were reported (12).

During November 1994 through April 1995, we assessed the prevalence of antimicrobial resistance with S. pneumoniae at 30 U.S. medical centers (4). Among 1,527 isolates of S. pneumoniae, 14.1% had intermediate resistance, and 9.5% were fully resistant to penicillin. Aggregate rates of intermediate and high penicillin resistance were 2.1% to 52.9%. In addition, high rates of resistance were noted with other antimicrobial drugs.

From November 1997 to April 1998, we surveyed 34 U.S. medical centers to assess changes in antimicrobial resistance rates with S. pneumoniae during the 3 years since the 1994-95 study. Twenty-four of these centers had also participated in the earlier investigation. Similar patient populations were sampled, and identical test methods were used. In addition, we assessed the relationship between various demographic factors and resistance and undertook a systematic analysis of multidrug resistance. Finally, macrolide and fluoroquinolone resistance was characterized at a molecular level.

The Study

From November 1, 1997, to April 30, 1998, 1,601 isolates of S. pneumoniae were recovered in 34 U.S. medical centers. All isolates included in this study were from consecutive patients. With the exception of specimens from the lower respiratory tract, all isolates were from normally sterile body sites (i.e., blood, cerebrospinal fluid, middle ear fluid, sinus aspirates, pericardial fluid, and pleural fluid). Isolates from lower respiratory tract specimens were included only if they were of clinical significance.

In the study centers, isolates were subcultured onto 5% sheep blood agar plates and incubated overnight at 35°C to 37°C in 5% to 7% CO₂. Colony growth was collected on a rayon swab and immediately immersed in a transport tube containing 12 ml of semisolid Ames transport medium with charcoal (Difco Laboratories, Detroit, MI). Transport tubes were then shipped overnight to the University of Iowa College of Medicine for additional analysis (Appendix). The recovery rate from this transport system was 100%. Twelve concentrations each of 23 antimicrobial drugs were tested against 1,601 isolates of S. pneumoniae.

Overall, 17.4% of isolates had intermediate and 12.1% had full resistance to penicillin (Table 1). Overall nonsusceptible rates with ceftriaxone and cefuroxime (intermediate plus fully resistant) were 14.9% and 23.3%, respectively. Because National Committee for Clinical Laboratory Standards (NCCLS)-approved breakpoints are lacking for the six other cephalosporins examined in this study (cefpodoxime, cefixime, ceftibuten, cefprozil, cefaclor, and loracarbef), rates of resistance were not determined for these drugs. However, when MIC values were compared, cefpodoxime was the most active.

Comparison of MIC values of the three macrolides we examined showed that clarithromycin was consistently twice as active as erythromycin, which in turn was consistently twice as active as azithromycin (Table 1). Overall rates of resistance, however, based on NCCLS breakpoints, which differ for these agents, were similar (18%-19%).

Compared with erythromycin as an indicator of macrolide activity, 302 (18.9%) isolates had MICs >1 µg/ml and thus were classified as resistant (Table 1). Among these, 217 (71.9%) had erythromycin MICs <32 µg/ml; the remaining 85 strains (28.1%) had erythromycin MICs >64 µg/ml. Of the 217 strains with erythromycin MICs <32, 214 had clindamycin MICs <0.25 and thus were categorized as clindamycin susceptible. Thirty-five of these strains, randomly selected, were examined by polymerase chain reaction (PCR) for the presence of ermAM and mefE genes. Of the 85 strains with erythromycin MICs >64 µg/ml, 83 had clindamycin MICs >8. Thirty-eight of these isolates, chosen randomly, were ermAM positive; 12 were also positive for mefE (Table 2). Five resistant strains were also characterized for the presence of macrolide resistance determinants (Table 2). Finally, the three isolates (Table 2) negative for both the ermAM and mefE genes were also negative by PCR for other known determinants of macrolide/lincosamide resistance in gram-positive bacteria (ermA, ermC, ereA, ereB, msrA, and linA genes).

Of the 1,601 isolates examined, one had intermediate resistance to trovafloxacin with an MIC 2 µg/ml; three strains (0.2%) were resistant, two with trovafloxacin MICs 4 µg/ml and one with a trovafloxacin MIC 8 µg/ml. The single strain with intermediate resistance had a ciprofloxacin MIC of 16 and an asp83tyr substitution in the C subunit of topoisomerase IV, as well as a ser84tyr substitution in the A subunit of DNA gyrase. The three trovafloxacin-resistant strains had ciprofloxacin MICs >32 µg/ml and a ser79phe substitution in the C subunit of topoisomerase, as well as a ser84phe substitution in the A subunit of DNA gyrase. None of these four strains had detectable mutations in the QRDR of par E.

Overall rates of resistance, expressed as the percentage of isolates intermediate or resistant, for selected other agents are described in Table 1: tetracycline, 13.2%, TMP/SMX, 31.1%, chloramphenicol, 7.2%, and rifampin, 0.2%. Linezolid was uniformly active over a narrow range of MICs (i.e., 0.12 to 2 µg/ml). Two strains among the 1,601 examined in this study were resistant to quinupristin/dalfopristin; one had an MIC 4 µg/ml and the other 8 µg/ml. Vancomycin was uniformly active against the 1,601 isolates of S. pneumoniae in this survey, with MICs <0.5 µg/ml.

The in vitro activity of all ß-lactams (penicillins, ß-lactamase inhibitor combinations and cephalosporins), macrolides, clindamycin, tetracycline, TMP/SMX, and chloramphenicol was lowest with high-level penicillin-resistant strains of S. pneumoniae and greatest with penicillin-susceptible isolates. This trend was not apparent with linezolid, trovafloxacin, rifampin, quinupristin/dalfopristin, or vancomycin.

The prevalence of resistance to selected agents was assessed according to the specimen source of isolates, the patient's age, and the health-care setting (Table 3). In general, the highest resistance rates for all antimicrobial drugs were observed in middle ear fluid and sinus aspirate isolates and in isolates from patients <5 years old and from patients seen in ambulatory-care settings.