Purpose Penicillin-resistant invasive Streptococcus pneumoniae emerged in the 1990s and occurred subsequently with increasing frequency in Huntington, WV. We investigated the evolution of penicillin resistance during 23 years by tracking changes in the penicillin-binding protein gene 2b (pbp2b) among all invasive strains of capsular types 6, 9, 14, 19, and 23, the types likely to become resistant, to assess whether penicillin resistance evolved from other streptococci or was imported into the community.
Methods We tested all consecutive invasive strains of S. pneumoniae isolated from children and adults admitted to hospital between 1983 and 2005 for penicillin susceptibility by E-test and capsular type by Quellung procedures. For genomic studies of the pbp2b gene, DNA was extracted from 289 strains, including 48 resistant (PRSP), 67 intermediate (PISP), 174 susceptible (PSSP), for restriction fragment length polymorphism (RFLP) using restriction enzyme HaeIII. A representative strain of each RFLP pattern was sequenced using internal primers and the assembled sequences were compared by multiple sequence alignment of DNA and amino acid sequences for homology.
Results PSSP occurred every year and belonged to five RFLP patterns; 167 of 174 PSSP belonged to two RFLP patterns of nearly identical genomic sequences and identical amino acid sequences, remained stable throughout the study period, and were closely related to the reference strain R6. PRSP occurred almost every year starting in 1994, with the first one related to Spain 23F-1, belonged to five other RFLP patterns, and exhibited distinctly different genomic sequences from each other but were closely related to PRSP first identified in other countries. PISP occurred in all years except four and belonged to 15 RFLP patterns. By RFLP patterns, they all showed different genomic sequences from each other and from R6. The sequences of PISP reflected a spectrum between PSSP and PRSP.
Conclusions These findings suggest that the PSSP persisted unchanged in the community throughout the study period. Unlike the PSSP, the PRSP appeared to have been imported into the community because all were closely related to PRSP previously identified in other countries, making it unlikely that antibiotic pressure in our community influenced the development of PRSP. However, it may have influenced the development of some PISP because only some PISP appear to have been imported based on a comparison of their genomic sequences with sequences registered in Genbank.
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