Lethal Mutagenesis of Foot-and-Mouth Disease Virus Involves Shifts in Sequence Space

Abstract

Lethal mutagenesis or virus transition into error catastrophe is an antiviral strategy that aims at extinguishing a virus by increasing the viral mutation rates during replication. The molecular basis of lethal mutagenesis is largely unknown. Previous studies showed that a critical substitution in the foot-and-mouth disease virus (FMDV) polymerase was sufficient to allow the virus to escape extinction through modulation of the transition types induced by the purine nucleoside analogue ribavirin. This substitution was not detected in mutant spectra of FMDV populations that had not replicated in the presence of ribavirin, using standard molecular cloning and nucleotide sequencing. Here we selectively amplify and analyze low-melting-temperature cDNA duplexes copied from FMDV genome populations passaged in the absence or presence of ribovirin Hypermutated genomes with high frequencies of A and U were present in both ribavirin -treated and untreated populations, but the major effect of ribavirin mutagenesis was to accelerate the occurrence of AU-rich mutant clouds during the early replication rounds of the virus. The standard FMDV quasispecies passaged in the absence of ribavirin included the salient transition-modulating, ribavirin resistance mutation, whose frequency increased in populations treated with ribavirin. Thus, even nonmutagenized FMDV quasispecies include a deep, mutationally biased portion of sequence space, in support of the view that the virus replicates close to the error threshold for maintenance of genetic information.

Extinction of Hepatitis C Virus by Ribavirin in Hepatoma Cells Involves Lethal Mutagenesis

“…Statistically significant increases of nucleotide diversity were also observed for increasing Rib concentrations (NS5A p<0.004; NS5B p<0.04 and p<0.005; Permutation test) (Table 1). Ribavirin induces preferentially G→A and C→U transitions in HCV and other RNA viruses [35], [56], [57], [77], [102]. While the percentage of transitions relative to the total number of mutations did not vary as a result of Rib treatment (79.1%, 75.6% and 79.4% in the absence and presence of 50 and 100 μM Rib, respectively), the corresponding percentage of [(G→A)+(C→U)] transitions increased from 32.5% in the absence of Rib to 51.7% and 57.7% in the presence of 50 and 100 μM Rib, respectively, reflected also in an increase of the [(G→A)+(C→U)]/[(A→G)]+[(U→C)] ratio (Fig.…”

“…Neither the increase in mutation frequency nor that of G→A and C→U transitions reached statistical significance (A.M. Ortega-Prieto et al, unpublished results). (ii) Rib can produce transient expansions of mutant spectrum complexity [102], [138], so that a mutational activity might be missed depending on the time of sampling of the mutant spectrum. (iii) If quantifications of template HCV RNA are not performed prior to RT-PCR amplifications intended for molecular cloning and Sanger sequencing or for UDPS, the repertoire of genomes screened might not reflect the repertoire present in the natural sample.…”

Mutagenesis-Mediated Virus Extinction: Virus-Dependent Effect of Viral Load on Sensitivity to Lethal Defection

“…Mutagenic nucleoside analogues are currently being investigated as antiviral agents because they can promote RNA virus extinction through lethal mutagenesis [[52], [50], [76], [51], [30], [33], [77], [78], [53], [73], [79]–[81]; reviewed in [11], [12], [9]]. Examination of the events that accompany FU-mediated extinction of LCMV [31], [33], [27], [34], [30], [32] led to the proposal of lethal defection as a mechanism of virus extinction [30].…”

Getting to Know Viral Evolutionary Strategies: Towards the Next Generation of Quasispecies Models

“…Mutagenic treatments enlarge the region of sequence space explored by a virus (Ojosnegros et al 2008) due to an increase in the diversity of the population. Examination of the FMDV mutant spectrum in the presence of ribavirin showed an increase in the frequency of A/G and C/U transitions, causing a displacement towards regions of sequence space where the virus was more prone to experience deleterious mutations (Perales et al 2011c). A ribavirin-resistant mutant isolated in that same virus restored the normal transition pattern without affecting the average mutation frequency or the incorporation of ribavirin by the replicase (Agudo et al 2010).…”

“…Differences in the viral mutation rates have also been observed across hosts or cell type infected (Pita et al 2007). Actually, lethal mutagenesis studies yield many examples of the variety of responses elicited from RNA viruses subjected to artificial increases in the error rate: fitness decreases and extinction (Perales et al 2011a;Domingo et al 2012), genomic shifts in sequence space (Perales et al 2011c), alterations of the network of interactions within the mutant spectrum (Grande-Pérez et al 2005) or selection of resistant mutants (Arias et al 2008;Arribas et al 2011;Agudo et al 2010;Pfeiffer and Kirkegaard 2003;Iranzo et al 2011). The evolutionary history of a viral population eventually determines the region of the sequence space it occupies (its genomic diversity), a fact that affects genetic robustness, viral plasticity and the effect of new mutations Graci et al 2012).…”