Abstract:
:Intrinsic genetic instability of RNA viruses may lead to the accumulation of revertants during manufacture of live viral vaccines, requiring rigorous quality control to ensure vaccine safety. Each lot of oral poliovirus vaccine (OPV) is tested for neurovirulence in animals and also for the presence of neurovirulent revertants. Mutant analysis by PCR and restriction enzyme cleavage (MAPREC) is used to measure the frequency of neurovirulent mutations at the 5' untranslated region (UTR) of the viral genome that correlate with the level of neurovirulence determined by the monkey neurovirulence test. However, MAPREC can only monitor mutations at a few genomic loci and miss mutations at other sites that could adversely affect vaccine quality. Here we propose to use massively parallel sequencing (MPS) for sensitive detection and quantification of all mutations in the entire genome of attenuated viruses. Analysis of vaccine samples and reference preparations demonstrated a perfect agreement with MAPREC results. Quantitative MPS analysis of validated reference preparations tested by MAPREC produced identical results, suggesting that the method could take advantage of the existing reference materials and be used as a replacement for the MAPREC procedure in lot release of OPV. Patterns of mutations present at a low level in vaccine preparations were characteristic of seed viruses used for their manufacture and could be used for identification of individual batches. This approach may represent the ultimate tool for monitoring genetic consistency of live viral vaccines.
journal_name
Proc Natl Acad Sci U S Aauthors
Neverov A,Chumakov Kdoi
10.1073/pnas.1012537107subject
Has Abstractpub_date
2010-11-16 00:00:00pages
20063-8issue
46eissn
0027-8424issn
1091-6490pii
1012537107journal_volume
107pub_type
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