Twenty-Eight Years of Poliovirus Replication in an Immunodeficient Individual: Impact on the Global Polio Eradication Initiative

Abstract

There are currently huge efforts by the World Health Organization and partners to complete global polio eradication. With the significant decline in poliomyelitis cases due to wild poliovirus in recent years, rare cases related to the use of live-attenuated oral polio vaccine assume greater importance. Poliovirus strains in the oral vaccine are known to quickly revert to neurovirulent phenotype following replication in humans after immunisation. These strains can transmit from person to person leading to poliomyelitis outbreaks and can replicate for long periods of time in immunodeficient individuals leading to paralysis or chronic infection, with currently no effective treatment to stop excretion from these patients. Here, we describe an individual who has been excreting type 2 vaccine-derived poliovirus for twenty eight years as estimated by the molecular clock established with VP1 capsid gene nucleotide sequences of serial isolates. This represents by far the longest period of excretion described from such a patient who is the only identified individual known to be excreting highly evolved vaccine-derived poliovirus at present. Using a range of in vivo and in vitro assays we show that the viruses are very virulent, antigenically drifted and excreted at high titre suggesting that such chronic excreters pose an obvious risk to the eradication programme. Our results in virus neutralization assays with human sera and immunisation-challenge experiments using transgenic mice expressing the human poliovirus receptor indicate that while maintaining high immunisation coverage will likely confer protection against paralytic disease caused by these viruses, significant changes in immunisation strategies might be required to effectively stop their occurrence and potential widespread transmission. Eventually, new stable live-attenuated polio vaccines with no risk of reversion might be required to respond to any poliovirus isolation in the post-eradication era.

Fig 1. Sequence analysis of iVDPV strains.

Neighbour-joining tree representing phylogenetic relationships through the entire capsid coding sequence (2637 nt) between iVDPV isolates from the case study (shown as a number that corresponds to the date of isolation in the format ddmmyy), Sabin 2 vaccine strain and other type 2 VDPV and wild polioviruses. EMBL Data Library accession numbers for published capsid sequences are shown in the tree. Numbers at nodes indicate the percentage of 1000 bootstrap pseudoreplicates supporting the cluster. The sequence of PV1-Mahoney reference strain was introduced as an outgroup for the correct rooting of the tree. Isolates from the patient are labelled with blue circles on the tree, other iVDPV isolates are indicated in yellow, cVDPVs in red, VDPVs found in sewage samples in green and wild polioviruses in purple.

Fig 2. Neurovirulence of iVDPV strains.

The graphic represents 50% Paralytic Doses (PD₅₀) values (with 95% Confidence Intervals) for selected poliovirus strains determined in Tg21-bx mice using the Probit method. Isolates from the patient are shown underlined.

Fig 3. Antigenic structure of iVDPV strains.

Radial diagrams representing the reactivity of poliovirus strains with Sabin 2-specific monoclonal antibodies. The results are shown as OD values at 492nm obtained in ELISA assays and expressed as normalised values relative to those obtained with antibody 1102 which reacted with all poliovirus strains. These values denote the average of two duplicate assays. Monoclonal antibodies used in the assay in the order 1 to 14 shown in the graph (from the top and clockwise), with antigenic site specificity shown in brackets, were: 969 (site 1), 435 (1), 433 (1), 434 (1), 436 (1), 1231 (2a), 1247 (2a), 1269 (2a), 1037 (2b), 1050 (3a), 1102 (3b), 1103 (3b), 1121 (3b) and 1051 (3b). Sabin 2 vaccine virus, iVDPV isolates from the patient (160198, 190100, 080503, 071108 and 171012), cVDPV strain MAD029 and wild strains (EGY42, EGY52, VEN59, MOR78 and KUW80) were used in the assays.

Fig 4. Location of mutations in iVDPV isolate 160198.

Molecular surface diagram of the three-dimensional structure of type 2 wild poliovirus strain Lansing viewed from the outside of the virion [25]. A pentameric unit is represented. The virus particle consists of 60 protomers. Each protomer contains a single copy of VP1, VP2, VP3, and VP4 arranged in icosahedral symmetry. The location of mutations found in known antigenic sites of iVDPV isolate 160198 with respect to Sabin 2 vaccine strain are shown in red, other amino acid changes from Sabin 2 are displayed in cyan. The image was generated using PyMOL Molecular Graphics System, Version 1.7.0.3 software (Schrödinger, LLC).

Fig 5. Neutralization of poliovirus strains by sera from fully immunised humans.

The graphs represent comparison of neutralization titres in 40 sera from UK adults against iVDPV isolate 160198 versus MEF-1 (A) or versus Sabin 2 (B) vaccine strains in cell culture assays. The values are expressed as reciprocals (Log2) of the highest dilution of serum that protected 50% of the cell cultures determined by the Karber formula.

Fig 6. Survival curve analysis of Tg21-bx mice immunised with IPV and challenged with paralytic doses of poliovirus.

Survival curves showing animals protected against paralysis caused by MEF-1 (A) or iVDPV strain 171012 (B) following immunisation with conventional (cIPV) products are shown with opened symbols and continuous lines; results for Sabin IPV (sIPV) products are shown as dashed lines and closed symbols; and survival data for mice injected with diluent (control) are shown as dotted lines and inverted triangles. All three cIPV products showed 100% protection against challenge with both viruses with the exception of cIPV-C that protected 7 out of the 8 animals used in the test. sIPV-A vaccine also protected all immunised animals against challenge with the MEF-1 strain.