Molecular characterization of penicillin-resistant Streptococcus pneumoniae isolates causing respiratory disease in the United States

Three hundred twenty-eight (328) penicillin-resistant Streptococcus pneumon iae isolates collected in 39 states of the United States between October, 1 996, and March, 1997, from (mostly adult) patients with respiratory disease were characterized by microbiological, serological, and molecular fingerpr inting techniques, including determination of chromosomal macrorestriction pattern with pulsed-field gel electrophoresis (BFGE) and hybridization with DNA probes specific for various antibiotic resistance genes. The overwhelm ing majority of the isolates were in five serogroups (23, 6, 19, 9, 14). Al l isolates had penicillin MIC values of at least 2 mu g/ml, but the collect ion also included isolates with MIC values as high as 16 mu g/ml. Virtually all isolates (96.6%) were resistant to trimethoprim/sulfamethoxazole (SXT) and many isolates were also resistant to chloramphenicol (43%), tetracycli ne (55%), and erythromycin (65%). Resistance to levofloxacin was extremely rare. The molecular fingerprinting methods showed that a surprisingly large proportion (167 out of 328, or 50.9%) of the isolates belonged to two inte rnational epidemic clones of S. pneumoniae: clone A (127, or 38.7%) with pr operties indistinguishable from that of the 23F multiresistant "Spanish/USA " clone widely spread in Europe, Asia, Latin America, and South Africa, and clone B (40, or 12.2%) belonging to the "French" serogroup 9/14 clone wide ly spread in Europe and South America. Virtually all members of clone A wer e also resistant to chloramphenicol (cat+), tetracycline (tetM+), and SXT, and about 75% were also resistant to erythromycin (mefE+ or ermB+). Close t o 30% (39 out of 127) of the clone A isolates expressed anomalous serotypes (primarily serotypes 19 and 14, and nontypable) and most likely represente d spontaneous capsular transformants. Most of the 40 isolates (35/40) belon ging to clone B expressed serotype 9, with five of the isolates expressing serotypes 14 or 19, or were nontypable. All members of this clone were resi stant to penicillin and SXT with only occasional isolates showing resistanc e to macrolides, tetracycline, and chloramphenicol. The combination of micr obiological tests and DNA hybridizations also allowed the identification of unusual strains, for instance, isolates that reacted with the tetM or mefE DNA probes without showing phenotypic antibiotic resistance, an isolate sh owing phenotypic macrolide resistance without hybridizing with either the e rmB or mefE DNA probes, or isolates that hybridized with both of these DNA probes. In addition to clones A and B, another large portion of the S. pneu moniae isolates (112 of 328, or 34.1%) was represented by eight clusters, e ach with a unique PFGE type. These clusters, together with the clone A and clone B isolates, made up 85% of all the penicillin-resistant isolates iden tified in this survey in the United States. Both international clones and t he unique clusters showed wide geographic dispersal: Clone A was present in 30 of the 39 states and clone B in 18. The data suggest that the major mod e of spread of penicillin-resistant pneumococci in the United States is by clonal expansion and that the most significant components (clones A and B) have been imported into the United States from abroad.