Molecular and Phenotypic Characterization of a Highly Evolved Type 2 Vaccine-Derived Poliovirus Isolated from Seawater in Brazil, 2014

Abstract

A type 2 vaccine-derived poliovirus (VDPV), differing from the Sabin 2 strain at 8.6% (78/903) of VP1 nucleotide positions, was isolated from seawater collected from a seaport in São Paulo State, Brazil. The P1/capsid region is related to the Sabin 2 strain, but sequences within the 5'-untranslated region and downstream of the P1 region were derived from recombination with other members of Human Enterovirus Species C (HEV-C). The two known attenuating mutations had reverted to wild-type (A481G in the 5'-UTR and Ile143Thr in VP1). The VDPV isolate had lost the temperature sensitive phenotype and had accumulated amino acid substitutions in neutralizing antigenic (NAg) sites 3a and 3b. The date of the initiating OPV dose, estimated from the number of synonymous substitutions in the capsid region, was approximately 8.5 years before seawater sampling, a finding consistent with a long time of virus replication and possible transmission among several individuals. Although no closely related type 2 VDPVs were detected in Brazil or elsewhere, this VDPV was found in an area with a mobile population, where conditions may favor both viral infection and spread. Environmental surveillance serves as an important tool for sensitive and early detection of circulating poliovirus in the final stages of global polio eradication.

Fig 1. Amino acid substitutions in the capsid protomer of isolate 44624.

VP1, VP2, VP3 and VP4 are represented as a 3-dimensional structured protomer. The image was generated using the software Swiss-PdbViewer [24], based on X-ray crystallographic analysis of type 2 poliovirus strain Lansing (Protein Data Bank accession number 1EAH.pdb) [23]. Colour codes: Substitutions at known antigenic sites, brown. Substitutions elsewhere, pink. The BC-loop of VP1 is not visible in this model.

Fig 2. Alignment of amino acids residues of neutralizing antigenic (NAg) sites for Sabin 2 (GenBank accession number AY184220) and isolate 44624.

Amino acid positions are numbered according to Sabin 2 NAg1 (VP1 88–106), NAg2 (VP2 163–169; VP2 268–270; VP1 220–225), NAg3a (VP3 54–61; VP3 70–74; VP1 286–291) and NA3b (VP2 71–73; VP3 75–79).

Fig 3. Schematic representation of the genome of recombinant aVDPV 44624, isolated in Brazil.

Non-vaccine sequences (HEV-C species) are represented in light blue. Highly mutated Sabin 2 regions are represented in gray.

Fig 4. Phylogenetic analysis of VP1 sequences of isolate 44624 and a set of type 2 VDPV isolated between 1998 and 2015.

Evolutionary distances were computed using Maximum Composite Likelihood method and Neighbor-joining tree. Consensus from 1000 bootstrap replicates is shown.

Fig 5. One-step growth curve analysis of isolate 44624 in comparison with Sabin 2 in RD cells.

Cells were infected at a MOI of 10 and incubated at 37 or 40°C. Total virus production at different times (0-12h) post-infection were determined by TCID 50 assays on RD cells. Each point represents the mean + standard deviation of virus titers from three different experiments.