The aims of this study were to develop modified sequential multiplex PCR (SM-PCR) primer sets and to evaluate their ability and efficiency for serotype determination. We selected target serotypes for SM-PCR testing according to serotype prevalence as reported in Asian publications. The modified SM-PCR consisted of 6 groups of PCR reactions, and each reaction was performed using 5 primer pairs. We evaluated the efficiency and performance of this modified multiplex PCR using 378 pneumococcal strains by comparing the findings with the results of the Quellung reaction. A total of 30 primer pairs were used in a consecutive set of 6 reactions. All results were concordant with those of the Quellung reaction and there was no cross-reactivity to unintended serotypes. We could identify the final serotypes of 370 isolates (97.9%). The coverage rates of modified SM-PCR were 42.6%, 65.9%, and 79.4% in reactions1, 2, and 3, respectively. The modified SM-PCR showed acceptable performance for detecting pneumococcal serotypes and can serve as useful alternative to the Quellung reaction.
Keywords: Streptococcus pneumoniae; sequential multiplex PCR; serotype.