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.2023 Mar 30;97(3):e0160122.
doi: 10.1128/jvi.01601-22. Epub 2023 Mar 8.

Comparative Efficacy of Mayaro Virus-Like Particle Vaccines Produced in Insect or Mammalian Cells

Affiliations

Comparative Efficacy of Mayaro Virus-Like Particle Vaccines Produced in Insect or Mammalian Cells

Sandra R Abbo et al. J Virol..

Abstract

Mayaro virus (MAYV) is a mosquito-transmitted alphavirus that causes often debilitating rheumatic disease in tropical Central and South America. There are currently no licensed vaccines or antiviral drugs available for MAYV disease. Here, we generated Mayaro virus-like particles (VLPs) using the scalable baculovirus-insect cell expression system. High-level secretion of MAYV VLPs in the culture fluid of Sf9 insect cells was achieved, and particles with a diameter of 64 to 70 nm were obtained after purification. We characterize a C57BL/6J adult wild-type mouse model of MAYV infection and disease and used this model to compare the immunogenicity of VLPs from insect cells with that of VLPs produced in mammalian cells. Mice received two intramuscular immunizations with 1 μg of nonadjuvanted MAYV VLPs. Potent neutralizing antibody responses were generated against the vaccine strain, BeH407, with comparable activity seen against a contemporary 2018 isolate from Brazil (BR-18), whereas neutralizing activity against chikungunya virus was marginal. Sequencing of BR-18 illustrated that this virus segregates with genotype D isolates, whereas MAYV BeH407 belongs to genotype L. The mammalian cell-derived VLPs induced higher mean neutralizing antibody titers than those produced in insect cells. Both VLP vaccines completely protected adult wild-type mice against viremia, myositis, tendonitis, and joint inflammation after MAYV challenge.IMPORTANCE Mayaro virus (MAYV) is associated with acute rheumatic disease that can be debilitating and can evolve into months of chronic arthralgia. MAYV is believed to have the potential to emerge as a tropical public health threat, especially if it develops the ability to be efficiently transmitted by urban mosquito vectors, such as Aedes aegypti and/or Aedes albopictus. Here, we describe a scalable virus-like particle vaccine against MAYV that induced neutralizing antibodies against a historical and a contemporary isolate of MAYV and protected mice against infection and disease, providing a potential new intervention for MAYV epidemic preparedness.

Keywords: Mayaro virus; baculovirus; mouse model; vaccine; virus-like particle.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

FIG 1
FIG 1
MAYV VLP production using insect cells and recombinant baculoviruses. (A) Schematic overview of the MAYV structural cassette expressed in insect cells. The molecular weight of each protein is shown in kilodaltons. Autocatalytic, host furin, and host signalase cleavage sites are indicated. (B) BACe56/MAYV-infected Sf9 insect cells and mock-infected Sf9 insect cells at 4 days postinfection. Black arrow indicates dense nuclear body, which presumably consisted of accumulated MAYV core-like particles. (C and D) MAYV structural protein expression in Sf9 cells was analyzed at 4 days postinfection by western blotting using antiserum derived from a MAYV-infected mouse (C) or anti-CHIKV capsid antibody 5.5D11 (D). (E) Detection of MAYV structural proteins in purified WUR VLP fraction from Sf9 insect cells and dilution series of TNAC MAYV VLPs from HEK293 human cells. (F) Transmission electron microscopy photo of purified WUR MAYV VLPs. Black arrows indicate MAYV VLPs; white arrow indicates baculovirus. (G) Size distribution of WUR MAYV VLPs based on diameter measurements of 107 VLPs.
FIG 2
FIG 2
MAYV BeH407 in adult wild-type C57BL/6J mice. (A) Viremia in adult female C57BL/6J mice (6 to 24 weeks old) infected with 104, 105, or 106 TCID50 or mock-infected with PBS, withn = 4 to 6 mice per group. (B) Percentage increase in foot height × width (relative to day 0) for mice infected as described for panel A, withn = 8 to 12 feet from 4 to 6 mice per group per time point. (C) Photographs showing examples of feet on day 6 postinfection, illustrating foot swelling in MAYV-infected groups compared to the mock-infected control group. H&E staining of muscle (M), tendon (T), synovial space (SS), and subcutaneous edema (*) in foot sections from 6- to 24-week-old C57BL/6J wild-type female mice infected as described for panel A. Black ovals indicate some of the areas containing inflammatory infiltrates in the muscles. Inflammatory infiltrates near and in tendon areas are visible, as well as inflammatory infiltrates near joint tissues. (D) Ratio of nuclear (purple) to nonnuclear (red) staining of H&E-stained foot sections (a measure of leukocyte infiltration). Data from 4 to 6 feet from 2 to 3 mice per group, with 3 sections scanned per foot and values averaged to produce one value for each foot. Statistical analysis used the Kolmogorov-Smirnov test. Multiple test correction was not applied. Error bars indicate one standard error of the mean.
FIG 3
FIG 3
MAYV VLP vaccination and challenge with MAYV BeH407 in adult C57BL/6J mice. (A) Timeline of vaccination with two 1-μg doses of nonadjuvanted MAYV VLPs, with a single 1-μg dose of adjuvanted MAYV VLPs, or with two doses of RPMI 1640 medium (negative control), then antibody measurements after bleeds, and disease determinations of viremia and foot swelling following MAYV BeH407 challenge. (B) MAYV BeH407 endpoint IgG ELISA titers after 1 or 2 vaccinations of female 6 -to 8-week-old C57BL/6J mice with nonadjuvanted MAYV VLPs or RPMI control, or 1 vaccination with MAYV VLPs with adjuvant. Lines among the dots indicate the mean ELISA titers, and error bars show the standard errors of the means. Dashed line represents the limit of detection (1:30 serum dilution). Statistical analysis used the Kolmogorov-Smirnov test. Multiple test correction was not applied. (C) MAYV BeH407 50% neutralization titers after 1 or 2 vaccinations with nonadjuvanted MAYV VLPs or RPMI control, or 1 vaccination with MAYV VLPs with adjuvant. Lines among the dots indicate the mean neutralization titers, and error bars show the standard errors of the means. Dashed line represents the limit of detection (1:10 serum dilution). Statistical analysis was with the Kolmogorov-Smirnov test. Multiple test correction was not applied. (D to F) Comparison of neutralization titers at week 8 against MAYV BeH407 (D), MAYV BR-18 (E), and CHIKV (Reunion isolate) (F). (G) MAYV BeH407 viremia postchallenge in mice vaccinated twice with nonadjuvanted MAYV VLPs or RPMI, or vaccinated once with MAYV VLPs with adjuvant (n = 5 to 6 per group). The limit of detection for each mouse was 102 TCID50/mL, with means from 5/6 mice plotted. Statistical analysis was with the Kolmogorov-Smirnov test. Multiple test correction was not applied. (H) Percentage increase in foot height × width (relative to day 0) for C57BL/6J mice vaccinated as described for panel G, withn = 6 to 12 feet from 3 to 6 mice per group per time point. Statistical analysis was with thet test.
FIG 4
FIG 4
Histopathology of MAYV VLP-vaccinated adult C57BL/6J mice after challenge with MAYV BeH407. (A) Photographs of mouse feet at 6 days postchallenge, illustrating swelling on day 6 in the RPMI-vaccinated, infected, and WUR VLP with adjuvant groups. H&E staining of tissues in foot sections from RPMI-vaccinated or VLP-vaccinated, uninfected, and MAYV-infected adult C57BL/6J mice is shown. Black ovals indicate some of the areas containing inflammatory infiltrates in the muscles (M). Inflammatory infiltrates near tendons (T) as well as inflammatory infiltrates near joint tissues (#) were also seen. Subcutaneous edema is shown with an asterisk. Hemorrhage is indicated by black arrows. These images are representative of a larger number analyzed. (B) Ratio of nuclear (purple) to nonnuclear (red) staining of H&E-stained foot sections (n = 2 to 3 mice, 4 to 6 feet per group, 3 sections per foot; values were averaged to produce one value for each foot). Statistical analysis used the Kolmogorov-Smirnov test. Multiple test correction was not applied.
FIG 5
FIG 5
Sequencing of the contemporary isolate, BR-18. (A, top) The E1/E2 amino acids that make contact with the receptor MXRA8 are identical for the two MAYV isolates, showing 60% amino acid identity with CHIKV (Reunion Island isolate). (Bottom) The crystal structure of CHIKV E1/E2 dimer, with the receptor contact residues that differed between MAYV and CHIKV colored as above. (B, top) Differences between the two MAYV isolates in E1/E2 (these amino acids are believed not to be involved in interaction with the receptor). (Bottom) The crystal structure of CHIKV E1/E2 dimer, with the residues that differed between the two isolates colored as above.
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