Genomic evolution of vesicular stomatitis virus strains with differences in adaptability.

Virus strains with a history of repeated genetic bottlenecks frequently show a diminished ability to adapt compared to strains that do not have such a history. These differences in adaptability suggest differences in either the rate at which beneficial mutations are produced, the effects of beneficial mutations, or both. We tested these possibilities by subjecting four populations (two controls and two mutants with lower adaptabilities) to multiple replicas of a regimen of positive selection and then determining the fitnesses of the progeny through time and the changes in the consensus, full-length sequences of 56 genomes. We observed that at a given number of passages, the overall fitness gains observed for control populations were larger than fitness gains in mutant populations. However, these changes did not correlate with differences in the numbers of mutations accumulated in the two types of genomes. This result is consistent with beneficial mutations having a lower beneficial effect on mutant strains. Despite the overall fitness differences, some replicas of one mutant strain at passage 50 showed fitness increases similar to those observed for the wild type. We hypothesized that these evolved, high-fitness mutants may have a lower robustness than evolved, high-fitness controls. Robustness is the ability of a virus to avoid phenotypic changes in the face of mutation. We confirmed our hypothesis in mutation-accumulation experiments that showed a normalized fitness loss that was significantly larger in mutant bottlenecked populations than in control populations.

Antagonistic pleiotropy involving promoter sequences in a virus.

Selection of specialist genotypes, that is, populations with limited niche width, promotes the maintenance of diversity. Specialization to a particular environment may have a cost in other environments, including fitness tradeoffs. When the tradeoffs are the result of mutations that have a beneficial effect in the selective environment but a deleterious effect in other environments, we have antagonistic pleiotropy. Alternatively, tradeoffs can result from the fixation of mutations that are neutral in the selective environment but have a negative effect in other environments, and thus the tradeoff is due to mutation accumulation. We tested the mechanisms underlying the fitness tradeoffs observed during adaptation to persistent infection of vesicular stomatitis virus in insect cells by sequencing the full-length genomes of 12 strains with a history of replication in a single niche (acute mammalian infection or persistent insect infection) or in temporally heterogeneous niches and correlated genetic and fitness changes. Ecological theory predicts a correlation between the selective environment and the niche width of the evolved populations, such that adaptation to single niches should lead to the selection of specialists and niche cycling should result in the selection of generalists. Contrary to this expectation, adaptation to one of the single niches resulted in a generalist and adaptation to a heterogeneous environment led to the selection of a specialist. Only one-third of the mutations that accumulated during persistent infection had a fitness cost that could be explained in all cases by antagonistic pleiotropy. Mutations involved in fitness tradeoffs included changes in regulatory sequences, particularly at the 3' termini of the genomes, which contain the single promoter that controls viral transcription and replication.

Emergence of mammalian cell-adapted vesicular stomatitis virus from persistent infections of insect vector cells.

Arboviruses (arthropod-borne viruses) represent quintessential generalists, with the ability to infect and perform well in multiple hosts. However, antagonistic pleiotropy imposed a cost during the adaptation to persistent replication of vesicular stomatitis virus in sand fly cells and resulted in strains that initially replicated poorly in hamster cells, even when the virus was allowed to replicate periodically in the latter. Once a debilitated strain started replicating continuously in mammalian cells, fitness increased significantly. Fitness recovery did not entail back mutations or compensatory mutations, but instead, we observed the replacement of persistence-adapted genomes by mammalian cell-adapted strains with a full set of new, unrelated sequence changes. These mammalian cell-adapted genomes were present at low frequencies in the populations with a history of persistence for up to a year and quickly became dominant during mammalian infection, but coexistence was not stable in the long term. Periodic acute replication in mammalian cells likely contributed to extending the survival of minority genomes, but these genomes were also found in strictly persistent populations.