Direct detection of Sabin poliovirus vaccine strains in stool specimens offirst-dose vaccinees by a sensitive reverse transcription PCR method

A multiplex reverse transcription-PCR method was optimized to monitor the d uration of excretion of Sabin poliovirus strains in stools of vaccinees fol lowing administration of the first dose of the trivalent oral vaccine. The assay detected approximately 1 50% tissue culture infective dose of each po liovirus serotype spiked into cell culture media. Although PCR inhibitors w ere frequently encountered in the stool specimens, a 1:20 dilution of the e xtracted RNA was sufficient to obtain a positive PCR result. Analysis of 19 5 stool specimens collected from 26 vaccinees showed that poliovirus types 1, 2, and 3 were identified more frequently by PCR than by tissue culture i solation. The percentages of specimens positive by PCR for poliovirus types 1, 2, and 3 were 67.2, 82.6, and 53.8, respectively. In contrast, the cult ure method identified types 1, 2, and 3 virus in 55.4, 64.1, and 27.7% of t he samples, respectively. Poliovirus type 2 excretion was detected by PCR i n practically all of the oral poliovirus vaccine recipients for 4 to 8 week s following vaccination. In contrast, excretion of type 1 and 3 viruses was more variable, with a range of 1 to 8 weeks. Shedding of type 3 virus ceas ed in similar to 70% of vaccinees within a week after immunization. In addi tion to an enhanced sensitivity for the detection of poliovirus, this PCR m ethod permits the direct characterization of virus in stool specimens witho ut further passage in culture, which may select for genetic variants that m ay not accurately reflect the virus composition in the original specimen.