Of the 12 serotypes present, the majority of isolates were not contained in PCV7 (the 7-valent pneumococcal conjugate vaccine), including 1 isolate that was serotype 1, 12 isolates that were serotype 2, 58 isolates that were serotype 5, 7 isolates that were serotype 7F, and 1 isolate that was serotype 12F. To determine whether clonal dissemination of the predominant serotypes had taken place, genotyping was performed on 100 S. pneumoniae isolates by using two methods: pulsed-field gel electrophoresis (PFGE) of SmaI-digested genomic DNA, and the Bacterial Barcodes repetitive-element PCR (rep-PCR) method. Criteria for delineating rep-PCR genotypes were established such that isolates of different serotypes were generally not grouped together. The two methods were equally discriminatory within a given pneumococcal serotype. PFGE separated the isolates into 15 genotypes and 7 subtypes; rep-PCR separated isolates into 15 genotypes and 6 subtypes. Using either method, isolates within serotypes 2, 5, and 7 formed three large, separate clusters containing 1 genotype each. Both methods further distinguished related subtypes within serotypes 2 and 5. Interestingly, one of the PFGE subtypes of serotype 5 is indistinguishable from the Columbia 5 -19 clone circulating in Latin America since 1994. The data support that serotypes 2 and 5 were likely to be the result of dissemination of particular clones, some of which are responsible for invasive disease over a broad population range.