Inactivated Wild-Type Chikungunya Virus

In previous studies, the researchers found that a novel chikungunya virus particle (ΔC-CHIKV) that completely lacked the capsid was still infectious in BHK-21 cells and demonstrated its potential as a live attenuated vaccine candidate. potential. However, ΔC-CHIKV has low yield and cannot be propagated in the vaccine production cell line Vero, which is not practical for commercial vaccine development. In this study, the researchers not only successfully propagated viral particles in Vero cells, but also significantly improved their production by constructing chimeric VEEV-ΔC-CHIKV and mass-passaging in Vero cells. Mechanistically, the high yield of VEEV-ΔC-CHIKV is due to adaptive mutations (especially mutations in the envelope protein) that improve the packaging efficiency of viral RNA. Similar to ΔC-CHIKV, the passaged VEEV-ΔC-CHIKV is safe, highly immunogenic, and effective. Only one immunization can protect mice from CHIKV attack. These studies demonstrate that utilizing CHIKV infectious capsidless viral particles as vaccine candidates is a practical strategy for developing alphavirus vaccines.

Previous studies have found that ΔC-CHIKV produces a large number of empty particles and a higher ratio of total viral particles to infectious particles compared with WT CHIKV. Here, the researchers found that the particle-to-PFU ratio of P50 VEEV-ΔC-CHIKV was much lower than that of P0 VEEV-ΔC-CHIKV, because 104 PFU contained a similar amount of envelope protein as 10 PFU P0 VEEV-ΔC-CHIKV (Figure 1A). Next, VEEV-ΔC-CHIKV in P0 and P50 were collected for sucrose gradient purification. Each fraction (fractions 1 to 16) was recovered from top to bottom and subjected to WB assay to quantify viral proteins. As shown in Figures 1B and C, infectious particles and empty particles were located in fractions 7-11 and fractions 1-3, respectively. The majority of P50 VEEV-ΔC-CHIKV was concentrated in fractions 7–11, whereas P0 particles were distributed in fractions 1–13 in a smear pattern, confirming that increased viral assembly efficiency was responsible for the higher viral titers of passaged VEEV-ΔC-CHIKV.

Figure 1. Passaged VEEV-ΔC-CHIKV produces more infectious viral particles. (A) Western blot analysis of CHIKV E1 expression in CHIKV-WT and VEEV-ΔC-CHIKV at different viral loads at P0, P10, and P50 using a CHIKV E1 polyclonal antibody. (B) Western blot analysis of sucrose density gradient fractions of VEEV-ΔC-CHIKV at P0 and P50 produced in Vero cells. (C) The intensity of the viral E1 protein band in each fraction was analyzed using ImageJ software. (Zhang Y N, et al., 2022)