Establishing a small animal model for evaluating protective immunity against mumps virus

Abstract

Although mumps vaccines have been used for several decades, protective immune correlates have not been defined. Recently, mumps outbreaks have occurred in vaccinated populations. To better understand the causes of the outbreaks and to develop means to control outbreaks in mumps vaccine immunized populations, defining protective immune correlates will be critical. Unfortunately, no small animal model for assessing mumps immunity exists. In this study, we evaluated use of type I interferon (IFN) alpha/beta receptor knockout mice (IFN-α/βR-/-) for such a model. We found these mice to be susceptible to mumps virus administered intranasally and intracranially. Passive transfer of purified IgG from immunized mice protected naïve mice from mumps virus infection, confirming the role of antibody in protection and demonstrating the potential for this model to evaluate mumps immunity.

Fig 1. Susceptibility of IFN-α/βR^−/− mice to rMuV-RLuc by various inoculation routes.

IFN-α/βR^−/− mice were infected with (A) 6x10⁶ PFU rMuV-RLuc in 200μL s.c., (B) 6x10⁶ PFU in 200μL i.p., (C) 1000 PFU in 20μL i.c., (D) 3x10⁶ PFU in 100μL i.n., or mock infected with PBS in the respective volume for each route. Lungs, spleens, and brains were harvested from mice 4, 7, and 10 dpi (s.c., i.p., and i.n. routes) or 2, 4, and 6 dpi (i.c. route). Tissues were homogenized and renilla luciferase activity was measured as described in materials and methods. ANOVA and Tukey multiple comparison tests were used to calculate P values.

Fig 2. Lung viral titers of IFN-α/βR^−/− and C57BL/6 mice challenged with MuV-IA.

(A) IFN-α/βR^−/− and (B) wild-type C57BL/6 mice were challenged i.n. with 1x10⁶ PFU MuV-IA in 100μL. Lungs were harvested on 2, 3, and 4 dpi. Tissues were weighed and homogenized and viral titers were determined by plaque assay in duplicate. The limit of detection of 5.8 PFU/g tissue is represented by the dashed line.

Fig 3. Reduced lung viral titers of challenged IFN-α/βR^−/− mice that received passive transfer of serum from MuV-IA-immunized mice.

BALB/c mice were immunized i.n. with 1x10⁶ PFU of MuV in a volume of 100 μL or mock immunized with 100 μL PBS. Mice were boosted at 3 and 6 wpi, and serum was collected and pooled at 8 wpi. Naïve IFN-α/βR^−/− mice received passive transfer of 300 μL i.p. of inactived serum from MuV-IA immunized mice (50 μL and 150 μL groups received serum diluted in PBS for 300 μL total volume), or 300 μL i.p. inactivated serum from PBS immunized mice. Mice were challenged 24 hours post passive transfer i.n. with 1x10⁶ PFU of MuV-IA in a volume of 100 μL. Lungs were collected and homogenized 2 dpi and viral titers were determined by plaque assay in duplicate. The limit of detection of 5.9 PFU/g tissue is represented by the dashed line. ANOVA and Tukey multiple comparison tests were used to calculate P values.

Fig 4. Reduced lung viral titers of challenged IFN-α/βR^−/− mice that received passive transfer of serum eluate and purified IgG from MuV-IA-immunized mice.

BALB/c mice were immunized i.n. with 1x10⁶ PFU of MuV-IA in a volume of 100 μL. Mice were boosted at 3 and 5 wpi, and serum was collected and pooled at 8 wpi. IgG was purified by affinity chromatography as described in materials and methods and dialyzed in PBS. Column eluate (referred to as serum eluate) was collected during IgG purification. Naïve IFN-α/βR^−/− mice received passive transfer i.p. of 1 mL PBS, serum eluate, or purified IgG. Mice were challenged 24 hours post passive transfer i.n. with 1x10⁶ PFU of MuV-IA in a volume of 100 μL. Lungs were collected and homogenized 2 dpi and viral titers were determined by plaque assay in duplicate. The limit of detection of 5.4 PFU/g tissue is represented by the dashed line. ANOVA and Tukey multiple comparison tests were used to calculate P values.