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Antimicrob Agents Chemother. 2006 Dec; 50(12): 4229–4230.
Published online 2006 Oct 16. doi:  10.1128/AAC.00943-06
PMCID: PMC1693999

Clonal Spread of Macrolide- and Tetracycline-Resistant [erm(A) tet(O)] emm77 Streptococcus pyogenes Isolates in Italy and Norway

Claudio Palmieri and Manuela Vecchi
Institute of Microbiology and Biomedical Sciences
Polytechnic University of Marche Medical School
60131 Ancona, Italy1
Pia Littauer and Arnfinn Sundsfjord
Reference Centre for Detection of Antimicrobial Resistance
Department of Microbiology and Virology
University Hospital of North-Norway and University of Tromsø
N-9037 Tromsø, Norway2
Pietro E. Varaldo and Bruna Facinelli*
Institute of Microbiology and Biomedical Sciences
Polytechnic University of Marche Medical School
60131 Ancona, Italy3

Over the last 15 years, a general increase in macrolide resistance in Streptococcus pyogenes has been observed in many parts of the world (1). In Europe, its prevalence is higher in Mediterranean countries and lower in Scandinavia. In particular, whereas an overall incidence around 43%, the highest rate recorded after a Japanese epidemic in the 1970s (10), was reported in a nationwide survey in Italy (17), the resistance rate remains low (<5%) in Norway (8). An association between erythromycin resistance and cell invasiveness in S. pyogenes has been documented in Italy (3).

Erythromycin-resistant (ER) S. pyogenes isolates are phenotypically and genotypically heterogeneous (5, 7). An efflux mechanism encoded by different mef genes causes resistance to 14- and 15-membered macrolides (M phenotype). In contrast, ribosomal methylation causes coresistance to macrolide-lincosamide-streptogramin B (MLS) antibiotics, which is expressed constitutively or inducibly (cMLS or iMLS-A phenotype, respectively) when encoded by the erm(B) gene and inducibly (iMLS-B or iMLS-C phenotype, depending on high- or low-level resistance) when encoded by the erm(A) gene.

Population structure analysis of ER S. pyogenes isolates (carrying the internalization-associated gene prtF1) collected in Italy in the late 1990s (17) and subsequently investigated for cell invasiveness (3) revealed the predominance of a limited number of clones with different combinations of resistance and virulence genes (14). The most widespread of three clones, comprising about half the isolates (herein named IMC-77), is characterized by inducible macrolide resistance [erm(A)/iMLS-B], emm77 type, high cell invasion efficiency, and ability to persist in cultured respiratory cells (13). In a parallel study, the same strains were also found to carry the tetracycline resistance gene tet(O) linked to erm(A) (4). A similar investigation, using different typing approaches, of ER strains collected in Norway from 1993 to 2002 (8) revealed four clonal complexes comprising ca. 75% of ER S. pyogenes isolates; of them, the second in frequency had phenotypic and genotypic characteristics [erm(A)/iMLS, emm77, tet(O)] which closely resemble those of IMC-77.

We recently detected prtF1 in several ER S. pyogenes isolates collected in Norway in 2003 (unpublished results). Of these, three erm(A) emm77 strains (K5-27, from vagina; K7-45, from throat; and K9-31, from abscess) were selected for the present study and compared to SP1900, an Italian throat isolate (from 1997) representing IMC-77, by use of complementary typing approaches. The macrolide resistance phenotype and genotype (5), tet(O) (4) and prtF1 (3) genes, cell invasion efficiency (3) and intracellular persistence (13), emm and RD2 typing (14), SmaI macrorestriction and pulsed-field gel electrophoresis (PFGE) analysis (12), and multilocus sequence typing (2) were determined as described previously. Besides the same erythromycin resistance genotype/phenotype [erm(A)/iMLS-B; MIC, >128 μg/ml] and emm type (emm77), the four strains shared identical tetracycline MICs (32 μg/ml) and resistance genotype [tet(O)], virulence traits (prtF1 [RD2 type d] and high cell invasion efficiency and persistence), and sequence type (ST369). Compared with the PFGE profile of the Italian strain, the three Norwegian strains shared a two-band difference, resulting from the disappearance of a ca.-280-kb fragment and the appearance of a new one of ca. 250 kb (Fig. (Fig.1),1), consistent with a close relatedness (16) to IMC-77.

FIG. 1.
PFGE patterns of SmaI-digested genomic DNA of four S. pyogenes strains. Lane 1, Italian strain SP1900; lanes 2, 3, and 4, Norwegian strains K5-27, K7-45, and K9-31, respectively; lane M, molecular size marker (low-range PFG marker; New England Biolabs, ...

Littauer et al. (8) argued that ER S. pyogenes in Norway may result from either introduction of resistant strains or local selection in internationally disseminated susceptible clones. The present findings suggest that the former hypothesis is likely to apply to ST369, also considering the far greater prevalence of ER S. pyogenes isolates in Italy than in Norway. The ability of prtF1-positive ER S. pyogenes isolates to escape β-lactams because of intracellular location and macrolides because of resistance could have conferred a selective advantage. Interestingly, ST369 is a single locus variant of ST63, which was first described to occur in the United States (2) and subsequently in Europe both in ER strains [erm(A)/iMLS, erm(B)/iMLS, and mef(A)/M] and in susceptible strains from different sources, all belonging to emm77 (6, 8, 11, 15). Thus, the dissemination of ST369 might also depend on the emm77 type, which belongs to the emm pattern E (no obvious preference for tissue site infection) and confers high colonization adaptability (9).


This work was supported in part by a grant from the Italian Ministry of Education, University, and Research.


Published ahead of print on 16 October 2006.


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