Calibration of Multiple Poliovirus Molecular Clocks Covering an Extended Evolutionary Range

Abstract

We have calibrated five different molecular clocks for circulating poliovirus based upon the rates of fixation of total substitutions (K t ), synonymous substitutions (K s ), synonymous transitions (A s ), synonymous transversions (B s ), and nonsynonymous substitutions (K a ) into the P1/capsid region (2,643 nucleotides). Rates were determined over a 10-year period by analysis of sequences of 31 wild poliovirus type 1 isolates representing a well-defined phylogeny derived from a common imported ancestor. Similar rates were obtained by linear regression, the maximum likelihood/single-rate dated-tip method, and Bayesian inference. . Nonsynonymous substitutions at all P1/capsid sites, including the neutralizing antigenic sites, appeared to be constrained by purifying selection. Simulation of the evolution of third-codon positions suggested that saturation of synonymous transitions would be evident at 10 years and complete at ϳ65 years of independent transmission. Saturation of synonymous transversions was predicted to be minimal at 20 years and incomplete at 100 years. The rapid evolution of the K t , K s , and A s clocks can be used to estimate the dates of divergence of closely related viruses, whereas the slower B s and K a clocks may be used to explore deeper evolutionary relationships within and across poliovirus genotypes.Poliovirus is one of the most rapidly evolving viruses known (17,31,38,52,58). Estimates of the rates of total nucleotide substitution into poliovirus capsid regions average ϳ10 Ϫ2 substitutions per site per year (31, 39, 52-54, 89, 90). The rates appear to be similar across the three poliovirus serotypes and for both circulating polioviruses and polioviruses associated with chronic infections. This very rapid rate of genomic evolution has facilitated high-resolution molecular epidemiologic studies, permitting the unambiguous identification of the sources of imported viruses (10, 41) and the resolution of separate lineages during outbreaks (39,43,74,75), during endemic transmission (23,52,90), during prolonged poliovirus replication in immunodeficient patients (9,31,53,89), and from environmental surveillance (23). However, the rapid accumulation of nucleotide substitutions, most of which are synonymous transitions, obscures deeper evolutionary relationships (46,70).Underlying the rapid pace of poliovirus genomic evolution are the high rates of base misincorporation (in the range of 10 Ϫ5 to 10 Ϫ3 per base per replication) by the poliovirus RNAdependent RNA polymerase (2,15,19,81,82,84). These high mutation rates are attributable to the absence of 3Ј35Ј exonuclease proofreading mechanisms for the viral RNA polymerases (19), although other mechanisms may also be involved (17, 18). The strong transition bias of the poliovirus polymerase (2, 46) is reflected in the pattern of fixation nucleotide substitutions into poliovirus genomes.In this report, we have calibrated five different molecular clocks based upon the rates of fixation of total substitutions (K t ), synonymous substitutions (K s ),...

Genus-Specific Substitution Rate Variability among Picornaviruses

“…These higher rates are evinced despite a wide range of mean rates; there was more variability among mean rates of enterovirus evolution than among those of isolates from five other genera (the standard deviation for enterovirus rates was four times [VP1] and two times [3D pol ] greater than for the other genera). Our novel and collected VP1 rates are similar to published rates of the entire P1 structural region (15,29,30), and our 3D pol rates echo that of an adjacent nonstructural region of a veterinary enterovirus (29). The striking similarity between evolutionary rates of structural and nonstructural picornavirus genes has not previously been discussed in the literature, but for each virus the rates were remarkably consistent with the estimated rates for both regions.…”

“…The genus Enterovirus is also by far the largest and most diverse genus in the family (2), which has allowed for more rate analyses of enteroviruses than of other genera. While all enterovirus substitution rates are of serotypes and most other available substitution rates are of species, the substitution rates of an enterovirus species should closely resemble the average of its serotype rates (30). However, our analyses showed that slower-evolving nonenterovirus species have much longer TMRCAs, which means they have had more time to become saturated at synonymous positions.…”

“…Due to their error-prone RNA-dependent RNA polymerases (RdRp), ssRNA viruses are known for high mutation rates, which are on the order of 10 Ϫ5 to 10 Ϫ3 mutations per nucleotide per replication event (10,12,25,30,57). In the absence of selection, the per-year substitution rate is solely a function of the per-replication mutation rate, so higher mutation rates can directly translate into higher substitution rates.…”

Limits of variation, specific infectivity, and genome packaging of massively recoded poliovirus genomes

“…To improve the interpretation of the data in Fig. 3F, we added Table S3, which depicts the amount of synonymous changes from PV1(M) as a percentage of total nucleotides in the coding domain P1, P2, and P3 (36). Whereas the number of natural synonymous changes in the PVs and CAV20 is large, the changes acquired in P(1+2+3) Max In PV-infected cells, icosahedral "procapsids" (also known as "empty capsids") (35,44) are formed that, remarkably, are void of viral or cellular RNAs.…”

Characterization of a rare natural intertypic type 2/type 3 penta-recombinant vaccine-derived poliovirus isolated from a child with acute flaccid paralysis

“…The date of the initiating OPV dose for the AFP case patient described in this study was estimated from the K s (synonymous substitutions per synonymous site) and K t (total substitutions per site) values by assuming evolution rates of 0.032 synonymous substitutions per synonymous site per year and 0.011 total substitutions per site per year (Jorba et al, 2008).…”