Cross-protection against Marburg virus strains by using a live, attenuated recombinant vaccine

Abstract

Marburg virus (MARV) has been associated with sporadic episodes of hemorrhagic fever, including a recent highly publicized outbreak in Angola that produced severe disease and significant mortality in infected patients. MARV is also considered to have potential as a biological weapon. Recently, we reported the development of a promising attenuated, replication-competent vaccine against MARV based on recombinant vesicular stomatitis virus (VSV) expressing the glycoprotein of the Musoke strain of MARV (VSVDeltaG/MARVGP-Musoke). We used this vaccine to demonstrate complete protection of cynomolgus monkeys against a homologous MARV challenge. While these results are highly encouraging, an effective vaccine would need to confer protection against all relevant strains of MARV. Here, we evaluated the protective efficacy of the VSVDeltaG/MARVGP-Musoke vaccine against two heterologous MARV strains, the seemingly more pathogenic Angola strain and the more distantly related Ravn strain. In this study, seven cynomolgus monkeys were vaccinated with the VSVDeltaG/MARVGP-Musoke vector. Three of these animals were challenged with the Angola strain, three with the Ravn strain, and a single animal with the Musoke strain of MARV. Two animals served as controls and were each injected with a nonspecific VSV vector; these controls were challenged with the Angola and Ravn strains, respectively. Both controls succumbed to challenge by day 8. However, none of the specifically vaccinated animals showed any evidence of illness either from the vaccination or from the MARV challenges and all of these animals survived. These data suggest that the VSVDeltaG/MARVGP-Musoke vaccine should be sufficient to protect against all known MARV strains.

FIG. 1.

Phylogenetic tree analysis for the GPs of filoviruses. The amino acid sequences of filovirus GPs present in the protein database of GenBank were analyzed using MEGA version 3.1 (www.megasoftware.net). A neighbor-joining tree and 1,000 bootstrap replicates for branch points were prepared. The analysis shows that the GP of MARV-Angola is more closely related to that of MARV-Musoke than to that of MARV-Ravn and has substantial differences with the GPs of ZEBOV species. The numbers to the left of the strains show percentages of bootstrap values. With the exception of the two Durba strains, the assigned designations are from reference . CIEBOV, Ivory Coast ebolavirus; REBOV, Reston ebolavirus.

FIG. 2.

Vaccination and challenge of nonhuman primates. (A) Flow chart of the experimental design. Arrows indicate the days of sampling (blood and swabs). Days preceded by a minus indicate days prior to challenge. (B) Kaplan-Meier mortality chart of the MARV vaccine study. Open triangles, animal vaccinated with VSVΔG/EBOVGP and challenged with MARV-Ravn (control no. 1); X's, animal vaccinated with VSVΔG/EBOVGP and challenged with MARV-Angola (control no. 2); filled diamonds, animals vaccinated with VSVΔG/MARVGP and challenged with MARV-Ravn (subjects no. 1 to 3); open squares, animals vaccinated with VSVΔG/MARVGP-Musoke and challenged with MARV-Angola (subjects no. 4 to 6); open circles, animal vaccinated with VSVΔG/MARVGP-Musoke and challenged with MARV-Musoke (subject no. 7).

FIG. 3.

Viremia levels in nonhuman primates after vaccination and MARV challenge. (A) VSV viremia levels were determined after vaccination with VSVΔG/MARVGP-Musoke (subjects no. 1 to 7) or VSVΔG/ZEBOVGP (controls no. 1 and 2). (B) MARV viremia levels from plasma, determined at the indicated time points after challenge with MARV-Ravn, MARV-Angola, or MARV-Musoke. Viremias were determined by plaque assay.

FIG. 4.

Humoral immune responses to MARV before and after challenge in nonhuman primates. (A) IgG responses to MARV measured using an established ELISA (see Materials and Methods). Titers are presented as endpoint dilutions. (B) MARV neutralizing antibodies detected using a plaque reduction neutralization assay (PRNT₅₀) as described in Materials and Methods. Titers are presented as endpoint dilutions. Days preceded by a minus indicate days prior to challenge, and day 0 is the day of challenge.