Research Article: Zika (PRVABC59) Infection Is Associated with T cell Infiltration and Neurodegeneration in CNS of Immunocompetent Neonatal C57Bl/6 Mice

Date Published: November 17, 2016

Publisher: Public Library of Science

Author(s): Mohanraj Manangeeswaran, Derek D. C. Ireland, Daniela Verthelyi, Robyn S Klein.


The recent spread of Zika virus (ZIKV) and its association with increased rates of Guillain Barre and other neurological disorders as well as congenital defects that include microcephaly has created an urgent need to develop animal models to examine the pathogenesis of the disease and explore the efficacy of potential therapeutics and vaccines. Recently developed infection models for ZIKV utilize mice defective in interferon responses. In this study we establish and characterize a new model of peripheral ZIKV infection using immunocompetent neonatal C57BL/6 mice and compare its clinical progression, virus distribution, immune response, and neuropathology with that of C57BL/6-IFNAR KO mice. We show that while ZIKV infected IFNAR KO mice develop bilateral hind limb paralysis and die 5–6 days post-infection (dpi), immunocompetent B6 WT mice develop signs of neurological disease including unsteady gait, kinetic tremors, severe ataxia and seizures by 13 dpi that subside gradually over 2 weeks. Immunohistochemistry show viral antigen predominantly in cerebellum at the peak of the disease in both models. However, whereas IFNAR KO mice showed infiltration by neutrophils and macrophages and higher expression of IL-1, IL-6 and Cox2, B6 WT mice show a cellular infiltration in the CNS composed predominantly of T cells, particularly CD8+ T cells, and increased mRNA expression levels of IFNg, GzmB and Prf1 at peak of disease. Lastly, the CNS of B6 WT mice shows evidence of neurodegeneration predominantly in the cerebellum that are less prominent in mice lacking the IFN response possibly due to the difference in cellular infiltrates and rapid progression of the disease in that model. The development of the B6 WT model of ZIKV infection will provide insight into the immunopathology of the virus and facilitate assessments of possible therapeutics and vaccines.

Partial Text

Zika virus (ZIKV) is an emerging mosquito-borne pathogen that belongs to the Flavivirus genus of the Flaviviridae family, which includes globally relevant arthropod-transmitted human pathogens such as dengue (DENV), yellow fever (YFV), West Nile (WNV), Japanese encephalitis (JEV), and tick-borne encephalitis viruses. The first strain of ZIKV (MR 766) was isolated in 1947 from a febrile sentinel rhesus monkey in the Zika forest near Entebbe, Uganda after the virus underwent intracerebral passage in Swiss albino mice[1]. For the next 50 years infections with ZIKV were reported sporadically in different regions of Africa and Asia, but were associated with mild symptoms consisting of skin rashes, conjunctivitis, fever and headaches[2]. In 2007 ZIKV started spreading west, first with an outbreak in Island of Yap where it infected over 70% of the population, followed in 2013 by an outbreak in French Polynesia[3]. This last outbreak was associated with a sharp increase in cases of Guillain Barre Syndrome (GBS), an autoimmune disease characterized by weakening and even paralysis of the limbs and face [4,5]. In 2015 Zika spread to South and Central America, infecting thousands of people in Brazil and Colombia, where it associated with an increase in GBS rates as well as a significant increase in severe fetal abnormalities that include spontaneous abortion, stillbirth, hydrocephaly, microcephaly, and placental insufficiency[6–9]. The temporal association of the viral outbreak and increased incidence of GBS and birth defects did not necessarily imply causality, however recent studies showing infection of the CNS in utero as well as infections of human neural progenitor and neural stem cells leading to cell cycle arrest and death [10,11] lend credence to the causality of ZIKV in microcephaly [12,13].

ZIKV belongs to the Flavivirus genus that includes several etiological agents of viral encephalitis, the most significant being Japanese encephalitis virus, West Nile virus, and tick-borne encephalitis virus. As with other flaviviruses, the majority of infected individuals will not develop disease, but a minority will develop a severe illness with a significant chance of permanent neurological damage, congenital malformations, or death. The factors that determine this are likely numerous, involving complex interactions between virus and host that are yet to be uncovered. Animal models can help us understand the pathophysiology of the virus, identify therapeutic targets, and explore the safety and efficacy of new therapeutics and vaccines. This study shows that neonatal B6 WT mice challenged with ZIKV develop a slow onset non-lethal encephalitis that is characterized by unsteady gait, kinetic tremors, severe ataxia, loss of balance and seizures. The virus localizes to the CNS where it elicits a strong IFN response, T cell infiltration with increased expression of RNA coding for Ifng, granzymeB, perforin1 and Il-2. In addition, these mice show evidence of neurodegeneration in particular affecting the Purkinje and granular cell layers of the cerebellum as evidenced by Fluoro-Jade C staining. Our data suggests that innate and adaptive responses can limit viral expansion but may also play a role in pathological changes in the CNS.




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