Development of a Quantitative One-Step RT-PCR Method for the Detection of Sabin 2 Virus Contamination in a Novel Oral Poliovirus Vaccine Type 2

摘要

To control circulating vaccine-derived type 2 poliovirus outbreaks, a more genetically stable novel Oral Poliovirus Vaccine type 2 (nOPV2) was developed by targeted modifications of Sabin 2 genome. Since the use of OPV2 made of Sabin 2 strain has been stopped, it is important to exclude the possibility that batches of nOPV2 are contaminated with Sabin 2 virus. Here, we report the development of a simple quantitative one-step reverse-transcription polymerase chain reaction assay for the detection and quantitation of Sabin 2 virus in the presence of overwhelming amounts of nOPV2 strain. The method is specific and linear within 8 log10 range even in the presence of relevant amounts of nOPV2 virus. It is sensitive, with a lower limit of detection of 0.2 CCID50/mL (an equivalent of 198 genome copies per mL), and generates reproducible results. This assay can be used for quality control and lot release of the nOPV2.