Key result
Random RT-PCR and high throughput sequencing successfully identified full-length genome sequences of poliovirus strains in vaccines, detecting strains comprising as little as 0.1% of sequence reads.
Random RT-PCR and high throughput sequencing provide high-resolution identification of poliovirus vaccine components, offering a robust alternative to serological methods for quality control.
May aid poliovirus vaccine quality control; leaves open replacement of serological methods pending validation.
BACKGROUND: Definitive identification of poliovirus strains in vaccines is essential for quality control, particularly where multiple wild-type and Sabin strains are produced in the same facility. Sequence-based identification provides the ultimate in identity testing and would offer several advantages over serological methods. METHODS: We employed random RT-PCR and high throughput sequencing to recover full-length genome sequences from monovalent and trivalent poliovirus vaccine products at various stages of the manufacturing process. RESULTS: All expected strains were detected in previously characterised products and the method permitted identification of strains comprising as little as 0.1% of sequence reads. Highly similar Mahoney and Sabin 1 strains were readily discriminated on the basis of specific variant positions. Analysis of a product known to contain incorrect strains demonstrated that the method correctly identified the contaminants. CONCLUSION: Random RT-PCR and shotgun sequencing provided high resolution identification of vaccine components. In addition to the recovery of full-length genome sequences, the method could also be easily adapted to the characterisation of minor variant frequencies and distinction of closely related products on the basis of distinguishing consensus and low frequency polymorphisms.
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Mee et al. (2015) studied Poliovirus vaccines. Random RT-PCR and high throughput sequencing was evaluated on Identification of poliovirus strains in vaccines. Random RT-PCR and high throughput sequencing successfully identified full-length genome sequences of poliovirus strains in vaccines, detecting strains comprising as little as 0.1% of sequence reads.
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