Development of a small animal peripheral challenge model of Japanese encephalitis virus using interferon deficient AG129 mice and the SA14-14-2 vaccine virus strain

Abstract

Japanese encephalitis virus (JEV) is the most common cause of viral encephalitis in Asia, and it is increasingly a global public health concern due to its recent geographic expansion. While commercial vaccines are available and used in some endemic countries, JEV continues to be a public health problem, with 50,000 cases reported annually. Research with virulent JEV in mouse models to develop new methods of prevention and treatment is restricted to BSL-3 containment facilities, confining these studies to investigators with access to these facilities. We have developed an adult small animal peripheral challenge model using interferon-deficient AG129 mice and the JEV live-attenuated vaccine SA14-14-2, thus requiring only BSL-2 containment. A low dose of virus (10PFU/0.1ml) induced 100% morbidity in infected mice. Increased body temperatures measured by implantable temperature transponders correlated with an increase in infectious virus and viral RNA in serum, spleen and brain as well as an increase in pro-inflammatory markers measured by a 58-biomarker multi-analyte profile (MAP) constructed during the course of infection. In the future, the MAP measurements can be used as a baseline for comparison in order to better assess the inhibition of disease progression by other prophylactic and therapeutic agents. The use of the AG129/JEV SA14-14-2 animal model makes vaccine and therapeutic studies feasible for laboratories with limited biocontainment facilities.

Keywords: AG129 mouse model; Flavivirus; Flavivirus pathogenesis; Interferon-deficient mice; JEV SA14-14-2 vaccine strain; Japanese encephalitis virus; Viral encephalitis.

Figure 1. Dose-dependent virulence of JEV SA14-14-2 in AG129 mice

Mice (n=12) were inoculated i.p. with 10 (◊), 1(□) and 0.1 (○) PFU of JEV SA14-14-2.

Figure 2. Body temperatures of uninfected and JEV SA14-14-2 infected AG12 mice

Panel A: Mean of observable body temperatures of infected (solid line) and uninfected (dashed line) mice. Panel B: Estimated mean body temperatures from model with mouse-specific and diurnal effects removed for infected (solid) and uninfected (dashed) mice. Panel C: Difference in estimated mean body temperatures (infected minus uninfected) with 95% confidence bound (dashed lines).

Figure 3. Infection virus titers in various tissues of AG129 mice infected with JEV SA14-14-2 expressed as PFU/gram (or ml)

Panels: A, serum; B, spleen; C, brain. Plaque assays of tissue samples taken at 6 different time points during infection were performed in duplicate.

Figure 4. Virus RNA in various tissues of AG129 mice infected with JEV SA14-14-2 expressed as genomic copies/gram (or ml)

Panels: A, serum; B, spleen; C, brain. Virus RNA titers of tissue samples taken at 6 different time points during infection were measured by quantitative RT-PCR. Each assay was performed in triplicate.