One-step multiplex PCR assay for detecting Streptococcus pneumoniae serogroups/types covered by 13-valent...

The life-threatening illnesses caused by Streptococcus pneumoniae have been declined significantly after the use of pneumococcal conjugate vaccines. Continuous monitoring of the vaccine serogroups/types is necessary to follow the changing epidemiology of invasive pneumococcal diseases. Recently, the sequential multiplex PCR approach, which uses several different sets of reactions, has been commonly adopted for determining capsular serogroups/types of S. pneumoniae isolates. In our study, we focused on development of a one-step multiplex PCR assay detecting all 1, 3, 4, 5, 6A/B, 7F, 9V, 14, 18C, 19A, 19F and 23F serogroups/types targeted by PCV13. The content of multiplex PCR mix and the cycling conditions were optimized in a manner that allowed rapid and accurate serotyping of a pneumococcal isolate by performing only a single amplification reaction. In our study of 182 clinical isolates, the one-step multiplex PCR assay exhibited 100% sensitivity and specificity, suggesting that its utilization can significantly reduce the use of traditional antiserum method requiring expensive reagents.