Evolutionary Transition toward Defective RNAs That Are Infectious by Complementation

Abstract

Passage of foot-and-mouth disease virus (FMDV) in cell culture resulted in the generation of defective RNAs that were infectious by complementation. Deletions (of nucleotides 417, 999, and 1017) mapped in the L proteinase and capsid protein-coding regions. Cell killing followed two-hit kinetics, defective genomes were encapsidated into separate viral particles, and individual viral plaques contained defective genomes with no detectable standard FMDV RNA. Infection in the absence of standard FMDV RNA was achieved by cotransfection of susceptible cells with transcripts produced in vitro from plasmids encoding the defective genomes. These results document the first step of an evolutionary transition toward genome segmentation of an unsegmented RNA virus and provide an experimental system to compare rates of RNA progeny production and resistance to enhanced mutagenesis of a segmented genome versus its unsegmented counterpart.Mutation, recombination, and ensuing phenotypic modifications in response to selective pressures can be readily observed and quantitated with RNA viruses both in cell culture and in vivo within modest time periods. This has rendered viruses suitable experimental systems for studies of basic Darwinian processes of genetic modification, competition, and selection or random drift as agents of genome diversification and biological adaptation (reviewed in references 1, 8, and 12). However, some major evolutionary transitions such as RNA genome segmentation have never been observed in the laboratory, yet they have probably occurred, given the hundreds of animal, plant, bacterial, and fungal viruses with segmented RNA genomes that have been described, including picornavirus-like plant RNA viruses (43). We have carried out extensive studies on the population dynamics of the important animal picornavirus foot-and-mouth disease virus (FMDV), including that of phenotypic evolution upon long-term serial cytopathic infections of BHK-21 cells (2). Unexpectedly, after more than 200 serial infections of viral population C-S8c1 of FMDV (a clone of serotype C [reviewed in reference 33]), defective genomes became dominant in the population. Defective interfering (DI) particles are frequently produced upon passage of RNA viruses at a high multiplicity of infection (MOI). DI particles are deletion mutants that require the presence of helper virus for replication and can interfere with the replication of the standard infectious virus (14-16, 29, 32, 35). The results described here show that defective genomes which individually could not cause cytopathology could nevertheless complement each other to produce progeny and kill cells in the absence of standard virus. The results provide, to our knowledge, the first description of defective RNA genomes occurring during viral replication that can be stably maintained by complementation upon passage at a high MOI in the absence of standard infectious virus. Therefore, the results provide experimental evidence of the initial step of an evolutionary transition towards genome ...

Evolutionary transitions during RNA virus experimental evolution

“…The advantages of RNA genome segmentation have been discussed in the past and they range from better resisting the deleterious effect of high mutation rates [22], bolstering evolvability by segment reassortment during mixed infections [22] and selection of shorter and faster replicators, [23,24] to optimizing the co-regulation of the expression of linked genes [25,26]. García-Arriaza et al [27,28] have documented the first step of an evolutionary transition toward genome segmentation of an unsegmented RNA virus, the foot-and-mouth disease virus (FMDV). A clone of FMDV was serially passaged at high multiplicity of infection (MOI) in cell culture for 143 passages.…”

Error threshold in RNA quasispecies models with complementation

“…Henceforth, the phenotype of a genome may not reflect its genotype, a situation which in the case of viruses is known as phenotypic mixing and hiding [17][18][19][20]. Moreover, a substantial amount of evidences, gathered with viruses as different as the murine cytomegalovirus [21], foot-and-mouth disease virus [22] or tobacco mosaic virus [23], give support to the fact that deletion mutants can be replicated in cells coinfected with the full-genome helper viruses.…”

“…The mechanisms ensuring the survival and persistence of DIPs are not entirely clear, as they behave as hyperparasites and usually get involve in an arms race with the full virus [33][34][35]. It is possible that the emergence of genomes shortened by deletions might confer an advantage in terms of replication speed compared with the full virus [22]. Furthermore, there is also evidence of a stronger form of interference whereby the DIPs genome competes more successfully for the viral replicative machinery [34,36].…”

Pathways to extinction: beyond the error threshold

“…Parasites are unable to produce themselves the correct products to ensure their viability, but are often able to use in trans the products correctly encoded by other types. The extreme version of complementation is represented by fragmented genomes encapsidated in different particles, a situation where co-infection becomes a must for viability (García-Arriaza et al 2004;S. Ojosnegros et al 2010, unpublished results).…”