Emergence of Erythromycin-Resistant, Clindamycin-Susceptible Streptococcus pyogenes Isolates in Madrid, Spain

In Spain, the frequency of resistance of group A streptococci (GAS) to erythromycin was very low, increasing from 0 to 0.7% in 1989 and 1990 (1, 7) to approximately 3% in 1991 to 1994 (2). The purpose of the present study was to determine the susceptibilities of recent isolates of GAS to macrolides and, in the case of resistant isolates, to ascertain their patterns of erythromycin resistance. We also studied the in vitro activities of penicillin G and clindamycin.

A total of 222 isolates of GAS were collected from April to August 1996 from outpatient samples in Madrid, Spain. A total of 203 isolates were throat swab samples and 19 were from other sources. Identification was done by biochemical tests according to standard criteria (8).

Penicillin G, erythromycin, azithromycin, miocamycin, and clindamycin were tested. The MICs were determined by the agar dilution method with Mueller-Hinton agar supplemented with 5% defibrinated horse blood. The plates were incubated overnight at 35°C in a 5% carbon dioxide atmosphere. The procedures for susceptibility testing were those recommended by the National Committee for Clinical Laboratory Standards (6). The MIC breakpoint for miocamycin resistance was >4 μg/ml, as defined by the Comité de l’Antibiogramme de la Société Française de Microbiologie (10).

Erythromycin resistance was classified according to the results of disk diffusion, with a double-disk test with erythromycin and clindamycin disks, as previously described (9), to test for inducible resistance. The MIC ranges and the MICs at which 50 and 90% of isolates were inhibited (MIC₅₀s and MIC₉₀s, respectively) for the 222 isolates of Streptococcus pyogenes are given in Table 1. Penicillin remained exquisitely active, with an MIC₉₀ of 0.03 μg/ml.

TABLE 1.

In vitro susceptibilities of 222 recent GAS strains to five antimicrobial agents

Antibiotic	MIC (μg/ml)a
	Range	50%	90%	S (%)
Penicillin G	≤0.008–0.06	0.015	0.03	100.0
Erythromycin	≤0.12–>64	≤0.12	16	82.4
Azithromycin	≤0.25–>16	≤0.25	>16	82.4
Miocamycin	≤0.25–>64	1	1	99.5
Clindamycin	≤0.12–>64	≤0.12	≤0.12	99.5

^a50 and 90%, MIC₅₀ and MIC₉₀, respectively. S, percent of strains susceptible.  

Thirty-nine (17.6%) of the isolates tested were resistant to erythromycin (MIC breakpoint, ≥1 μg/ml). In contrast, only 0.5% of the S. pyogenes isolates were resistant to clindamycin (MIC₉₀, ≤0.12 μg/ml). The resistance to both 14- and 15-membered macrolides tested was 17.6%, whereas the resistance to miocamycin, a 16-membered macrolide, was 0.5%. Thirty-seven (95%) of 39 erythromycin-resistant GAS were susceptible to clindamycin (MICs, ≤0.12 μg/ml) and miocamycin (MICs, 0.5 to 1 μg/ml). One isolate was resistant to erythromycin (MIC, >64 μg/ml), azithromycin (MIC, >16 μg/ml), miocamycin (MIC, >64 μg/ml), and clindamycin (MIC, >64 μg/ml), which indicates cross-resistance to macrolides, lincosamides, and streptogramin B-type antibiotics (MLS_B phenotype). Another erythromycin-resistant strain was susceptible to clindamycin but showed an inducible type of erythromycin resistance.

All erythromycin-resistant isolates were also resistant to the 15-membered macrolide azithromycin. However, the 16-membered macrolide miocamycin showed good activity against these isolates. The MICs for 37 of 39 erythromycin-resistant isolates were moderate (8 to 32 μg/ml), but the MICs of clindamycin were the same as those for the erythromycin-susceptible isolates (≤0.12 μg/ml); this pattern has been described with the M phenotype (11) and “novel resistance” (9). With this phenotype, susceptibility to clindamycin and streptogramin B antibiotics but resistance to 14- and 15-membered macrolides is seen (11). The mechanism of erythromycin resistance involves active efflux. The rate of isolation of strains with this novel phenotype among the erythromycin-resistant GAS of our work is even higher than that recently reported in the United States (75%) (11), although the incidence of erythromycin-resistant GAS is rare in this country (3). However, 50% of erythromycin-resistant GAS isolated in Japan (5) and in Taiwan (4) were also resistant to clindamycin and tetracycline.

In conclusion, our report documents the emergence of high rates of resistance to erythromycin in S. pyogenes in Spain. Strains with the M phenotype account for the majority of those isolates, which in a sample of this size represents a significant finding. Clindamycin remained active against these strains and might be useful for treatment of patients with clinical failure after phenoxymethyl penicillin treatment and for penicillin-allergic patients. We suggest that clinical laboratories review their local susceptibility data. If erythromycin resistance rates are high, we recommend that susceptibilities to both macrolides and clindamycin in S. pyogenes isolates be routinely tested.