You are engineering an attenuated viral vaccine and must minimize reversion risk. Which multi-pronged genetic strategy best balances retention of limited replication and reduced chance of reversion to pathogenicity?

• A. Introduce a single point mutation in a highly conserved virulence gene that can easily revert but is expected to be stable without adding any additional safeguards because one change is sufficient.
• B. Engineer the virus to integrate into host genomic DNA to ensure persistence without replication, which simultaneously reduces reversion risk but increases oncogenic potential and is therefore always preferred.
• C. Remove all antigenic proteins and rely entirely on synthetic adjuvants to mimic infection, eliminating replication entirely but maintaining all features of live vaccines.
• D. Introduce multiple independent attenuating mutations in separate essential virulence loci (e.g., delete virulence gene A, introduce point mutation in polymerase that reduces replication rate, and modify regulatory element controlling a second virulence factor), so simultaneous reversion is extremely unlikely while low-level replication persists.

Answer: (D)