FIG 10.

Evaluation of the in vivo relevance of the Fcγ receptor in protection against ma-MARV in a murine challenge model. (A) Experimental timeline for immunization of BALB/c mice and the immunization groups included in the study. Groups of wild-type or Fcγ knockout mice (n = 12 per group) were immunized intramuscularly in the gastrocnemius muscle with the indicated treatments. Mice receiving 2 doses of adjuvanted inactivated vaccine (groups 6 and 7) were primed 61 days before challenge (day −61) and boosted at both 54 and 40 days before challenge (day −54 and day −40, respectively). All other groups of mice were primed 40 days before challenge (day −40). (B) Survival of BALB/c mice challenged intraperitoneally (i.p.) with a lethal dose of ma-MARV (1,000 PFU). Statistical significance was performed using the log-rank (Mantel-Cox) test for comparison of survival curves (**, P < 0.01). (C) Prechallenge humoral response in pooled mouse sera from each vaccine group after completion of the immunization schedule (day 0). The left and right panels show MARV GP- and RABV G-specific antibody titers (bar graph of EC₅₀ values), respectively, compared to the positive-control monoclonal antibody titers (gray bars). The D’Agostino and Pearson normality test was performed to test for the normal distribution in each data set. Statistical significance for both MARV GP and RABV G antibody titers was performed using the nonparametric Kruskal-Wallis test and Dunn’s multiple-comparison test (*, P < 0.05; **, P < 0.01; n.s., not significant). (D) Postchallenge humoral response in pooled sera from survivor mice in the indicated vaccine groups after challenge with ma-MARV (day 28, necropsy). (Left and middle) MARV GP- and RABV G-specific antibody titers, respectively, are represented by the EC₅₀ values of the ELISA curves compared to those for positive-control monoclonal antibodies (gray bars). The D’Agostino and Pearson normality test was performed to test for the normal distribution in each data set. Tests for the statistical significance for MARV GP antibody titers were performed using the nonparametric Kruskal-Wallis test and Dunn’s multiple-comparison test (n.s., not significant). (Right) Bar graph of MARV GP-specific IgG1 and IgG2 isotype ELISA OD₄₉₀ readings at the lowest antibody dilution (1:50) in postchallenge sera (day 28) for survivors in the indicated vaccine groups. (E) Pooled sera from mice from the indicated vaccine groups were analyzed in an in vitro pseudotyped lentivirus luciferase assay to determine the titers of both prechallenge (day 0) and postchallenge (day 28) MARV GP- and RABV G-neutralizing antibodies compared to those of a positive-control monoclonal antibody known to neutralize either MARV or RABV pseudotyped virus in vitro (gray lines). Graphs are representative of average data from three independent experiments. The gray horizontal lines indicate the threshold for a 50% reduction in infection.