Molecular epidemiology of penicillin-susceptible non-beta-lactam-resistant Streptococcus pneumoniae isolates from Greek children

Abstract

A total of 128 Streptococcus pneumoniae isolates that were susceptible to penicillin but resistant to non-beta-lactam agents were isolated from young carriers in Greece and analyzed by antibiotic susceptibility testing, serotyping, restriction fragment end labeling (RFEL), and antibiotic resistance genotyping. The serotypes 6A/B (49%), 14 (14%), 19A/F (11%), 11A (9%), 23A/F (4%), 15B/C (2%), and 21 (2%) were most prevalent in this collection. Of the isolates, 65% were erythromycin resistant, while the remaining isolates were tetracycline and/or trimethoprim-sulfamethoxazole resistant. Fifty-nine distinct RFEL types were identified. Twenty different RFEL clusters, harboring 2 to 19 strains each, accounted for 76% of all strains. Confirmatory multilocus sequence typing analysis of the genetic clusters showed the presence of three international clones (Tennessee(23F)-4, England(14)-9, and Greece(6B)-22) representing 30% of the isolates. The erm(B) gene was present in 70% of the erythromycin-resistant isolates, whereas 18 and 8% contained the mef(A) and mef(E) genes, respectively. The pneumococci representing erm(B), erm(A), and mef genes belonged to distinct genetic clusters. In total, 45% of all isolates were tetracycline resistant. Ninety-six percent of these isolates contained the tet(M) gene. In conclusion, penicillin-susceptible pneumococci resistant to non-beta-lactams are a genetically heterogeneous group displaying a variety of genotypes, resistance markers, and serotypes. This suggests that multiple genetic events lead to non-beta-lactam-resistant pneumococci in Greece. Importantly, most of these genotypes are capable of disseminating within the community.

FIG.1.

Dendrogram of the 59 RFEL types observed among the 128 Greek S. pneumoniae nasopharyngeal isolates. The molecular sizes of reference bands (in bases [b]), serotypes, PBP types, RFEL types, MLST types, number of isolates per RFEL type, clusters, and cluster codes are depicted. One or two strains per RFEL clusters were analyzed by MLST. Day care center isolates from the 1995 to 1996 study (asterisks) and day care center isolates from the 1997 to 1999 study (degree symbols) are indicated. If more than one day care center isolate belongs to a specific genotype, the total number is displayed in parentheses. Two new strains with new MLST types (new¹ and new²) are indicated. These two strains have been submitted to the MLST database. nt, nontypeable.

FIG. 2.

Genetic relatedness of the 15 MLST sequence types (ST) observed within the 20 RFEL clusters.