In vitro evolution of herpes simplex virus 1 (HSV-1) reveals selection for syncytia and other minor variants in cell culture

Abstract The large dsDNA virus herpes simplex virus 1 (HSV-1) is considered to be genetically stable, yet it can rapidly evolve in response to strong selective pressures such as antiviral treatment. Deep sequencing has revealed that clinical and laboratory isolates of this virus exist as populations that contain a mixture of minor alleles or variants, similar to many RNA viruses. The classic virology approach of plaque purifying virus creates a genetically homogenous population, but it is not clear how closely this represents the mixed virus populations found in nature. We sought to study the evolution of mixed versus highly purified HSV-1 populations in controlled cell culture conditions, to examine the impact of this genetic diversity on evolution. We found that a mixed population of HSV-1 acquired more genetic diversity and underwent a more dramatic phenotypic shift than a plaque-purified population, producing a viral population that was almost entirely syncytial after just ten passages. At the genomic level, adaptation and genetic diversification occurred at the level of minor alleles or variants in the viral population. Certain genetic variants in the mixed viral population appeared to be positively selected in cell culture, and this shift was also observed in clinical samples during their first passages in vitro. In contrast, the plaque-purified viral population did not appear to change substantially in phenotype or overall quantity of minor allele diversity. These data indicate that HSV-1 is capable of evolving rapidly in a given environment, and that this evolution is facilitated by diversity in the viral population.