Detection of Emerging Vaccine-Related Polioviruses by Deep Sequencing

Abstract

Oral poliovirus vaccine can mutate to regain neurovirulence. To date, evaluation of these mutations has been performed primarily on culture-enriched isolates by using conventional Sanger sequencing. We therefore developed a culture-independent, deep-sequencing method targeting the 5′ untranslated region (UTR) and P1 genomic region to characterize vaccine-related poliovirus variants. Error analysis of the deep-sequencing method demonstrated reliable detection of poliovirus mutations at levels of <1%, depending on read depth. Sequencing of viral nucleic acids from the stool of vaccinated, asymptomatic children and their close contacts collected during a prospective cohort study in Veracruz, Mexico, revealed no vaccine-derived polioviruses. This was expected given that the longest duration between sequenced sample collection and the end of the most recent national immunization week was 66 days. However, we identified many low-level variants (<5%) distributed across the 5′ UTR and P1 genomic region in all three Sabin serotypes, as well as vaccine-related viruses with multiple canonical mutations associated with phenotypic reversion present at high levels (>90%). These results suggest that monitoring emerging vaccine-related poliovirus variants by deep sequencing may aid in the poliovirus endgame and efforts to ensure global polio eradication.

Assessing the individual risk of fecal poliovirus shedding among vaccinated and non-vaccinated subjects following national health weeks in Mexico

“…In a previous study [18], we have described that the presence of mutant variants and variants per sample was quite low in a subset of the samples we collected; we therefore consider that it is unlikely that there were any VDPVs among our samples.…”

Whole Genome Sequencing of Enteroviruses Species A to D by High-Throughput Sequencing: Application for Viral Mixtures

“…Research has shown the importance of analyzing the complete genome for enteroviruses ( Oberste et al, 2004 ; Joffret et al, 2012 ) because nucleotidic differences and inter and intra-typic recombination events in non-structural regions differentiate types and lineages ( Lukashev et al, 2003 ; Simmonds, 2006 ; Combelas et al, 2011 ). The amplification and sequencing of whole genomic sequences of strains belonging to EV-A, -B, and -C species using generic and specific primers were successfully performed ( Tan et al, 2015 ; Bessaud et al, 2016 ; Montmayeur et al, 2017 ; Sahoo et al, 2017 ). One particular study aimed to isolate polioviruses using random amplification and NGS to better optimize current protocols for whole genome sequencing and identification of a variety of vaccine-derived polioviruses ( Montmayeur et al, 2017 ).…”

“…Next generation sequencing (NGS) methods offer a new powerful sequencing tool for the identification and characterization of enteroviruses. This has been successfully used to sequence partial or whole genome sequences of poliovirus and of enteroviruses species C ( Bessaud et al, 2016 ; Montmayeur et al, 2017 ; Sahoo et al, 2017 ). Additionally, a generic assay for whole genome amplification and deep sequencing of enterovirus A71 was published ( Tan et al, 2015 ).…”

Protocol Paper: Oral Poliovirus Vaccine Transmissibility in Communities After Cessation of Routine Oral Poliovirus Vaccine Immunization

“…Finally, genetic sequencing using the Sanger technique was performed on these OPV-positive samples to rule out the possibility of false positives due to other enteroviruses [ 4 ].…”