title: Random codon re-encoding induces stable reduction of replicative fitness of Chikungunya virus in primate and mosquito cells.
abstract: Large-scale codon re-encoding represents a powerful method of attenuating viruses to generate safe and cost-effective vaccines. In contrast to specific approaches of codon re-encoding which modify genome-scale properties, we evaluated the effects of random codon re-encoding on the re-emerging human pathogen Chikungunya virus (CHIKV), and assessed the stability of the resultant viruses during serial in cellulo passage. Using different combinations of three 1.4 kb randomly re-encoded regions located throughout the CHIKV genome six codon re-encoded viruses were obtained. Introducing a large number of slightly deleterious synonymous mutations reduced the replicative fitness of CHIKV in both primate and arthropod cells, demonstrating the impact of synonymous mutations on fitness. Decrease of replicative fitness correlated with the extent of re-encoding, an observation that may assist in the modulation of viral attenuation. The wild-type and two re-encoded viruses were passaged 50 times either in primate or insect cells, or in each cell line alternately. These viruses were analyzed using detailed fitness assays, complete genome sequences and the analysis of intra-population genetic diversity. The response to codon re-encoding and adaptation to culture conditions occurred simultaneously, resulting in significant replicative fitness increases for both re-encoded and wild type viruses. Importantly, however, the most re-encoded virus failed to recover its replicative fitness. Evolution of these viruses in response to codon re-encoding was largely characterized by the emergence of both synonymous and non-synonymous mutations, sometimes located in genomic regions other than those involving re-encoding, and multiple convergent and compensatory mutations. However, there was a striking absence of codon reversion (<0.4%). Finally, multiple mutations were rapidly fixed in primate cells, whereas mosquito cells acted as a brake on evolution. In conclusion, random codon re-encoding provides important information on the evolution and genetic stability of CHIKV viruses and could be exploited to develop a safe, live attenuated CHIKV vaccine.
authors: Aix Marseille Univ, IRD French Institute of Research for Development, EHESP French School of Public Health, UMR_D 190 Emergence des Pathologies Virales, Marseille, France. antoine.nougairede@univ-amu.fr
authors: Antoine
authors: A
authors: Nougairede
authors: Lauriane
authors: L
authors: De Fabritus
authors: Fabien
authors: F
authors: Aubry
authors: Ernest A
authors: EA
authors: Gould
authors: Edward C
authors: EC
authors: Holmes
authors: Xavier
authors: X
authors: de Lamballerie
mesh_terms: D000330:Aedes / Q000821:virology; D018354:Alphavirus Infections / Q000635:transmission / Q000821:virology; D019943:Amino Acid Substitution; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D065632:Chikungunya Fever; D002646:Chikungunya virus / Q000235:genetics* / Q000254:growth & development / Q000502:physiology; D002522:Chlorocebus aethiops; D003062:Codon / Q000235:genetics*; D016384:Consensus Sequence; D019143:Evolution, Molecular; D056084:Genetic Fitness; D014644:Genetic Variation; D016679:Genome, Viral / Q000235:genetics*; D006801:Humans; D009154:Mutation; D011323:Primates / Q000821:virology; D012367:RNA, Viral / Q000096:biosynthesis / Q000235:genetics; D017422:Sequence Analysis, DNA; D012692:Serial Passage; D014709:Vero Cells; D014764:Viral Proteins / Q000096:biosynthesis / Q000235:genetics; D014779:Virus Replication
chemical_list: D003062:Codon; D012367:RNA, Viral; D014764:Viral Proteins