Optimization and Deoptimization of Codons in SARS‐CoV‐2 and Related Implications for Vaccine Development

The spread of coronavirus disease 2019 (COVID‐19), caused by severe respiratory syndrome coronavirus 2 (SARS‐CoV‐2), has progressed into a global pandemic. To date, thousands of genetic variants have been identified among SARS‐CoV‐2 isolates collected from patients. Sequence analysis reveals that the codon adaptation index (CAI) values of viral sequences have decreased over time but with occasional fluctuations. Through evolution modeling, it is found that this phenomenon may result from the virus's mutation preference during transmission. Using dual‐luciferase assays, it is further discovered that the deoptimization of codons in the viral sequence may weaken protein expression during virus evolution, indicating that codon usage may play an important role in virus fitness. Finally, given the importance of codon usage in protein expression and particularly for mRNA vaccines, it is designed several codon‐optimized Omicron BA.2.12.1, BA.4/5, and XBB.1.5 spike mRNA vaccine candidates and experimentally validated their high levels of expression. This study highlights the importance of codon usage in virus evolution and provides guidelines for codon optimization in mRNA and DNA vaccine development.