Characterization and pathogenesis of aerosolized eastern equine encephalitis in the common marmoset (Callithrix jacchus)

Abstract

BackgroundLicensed antiviral therapeutics and vaccines to protect against eastern equine encephalitis virus (EEEV) in humans currently do not exist. Animal models that faithfully recapitulate the clinical characteristics of human EEEV encephalitic disease, including fever, drowsiness, anorexia, and neurological signs such as seizures, are needed to satisfy requirements of the Food and Drug Administration (FDA) for clinical product licensing under the Animal Rule.MethodsIn an effort to meet this requirement, we estimated the median lethal dose and described the pathogenesis of aerosolized EEEV in the common marmoset (Callithrix jacchus). Five marmosets were exposed to aerosolized EEEV FL93-939 in doses ranging from 2.4 × 101 PFU to 7.95 × 105 PFU.ResultsThe median lethal dose was estimated to be 2.05 × 102 PFU. Lethality was observed as early as day 4 post-exposure in the highest-dosed marmoset but animals at lower inhaled doses had a protracted disease course where humane study endpoint was not met until as late as day 19 post-exposure. Clinical signs were observed as early as 3 to 4 days post-exposure, including fever, ruffled fur, decreased grooming, and leukocytosis. Clinical signs increased in severity as disease progressed to include decreased body weight, subdued behavior, tremors, and lack of balance. Fever was observed as early as day 2–3 post-exposure in the highest dose groups and hypothermia was observed in several cases as animals became moribund. Infectious virus was found in several key tissues, including brain, liver, kidney, and several lymph nodes. Clinical hematology results included early neutrophilia, lymphopenia, and thrombocytopenia. Key pathological changes included meningoencephalitis and retinitis. Immunohistochemical staining for viral antigen was positive in the brain, retina, and lymph nodes. More intense and widespread IHC labeling occurred with increased aerosol dose.ConclusionWe have estimated the medial lethal dose of aerosolized EEEV and described the pathology of clinical disease in the marmoset model. The results demonstrate that the marmoset is an animal model suitable for emulation of human EEEV disease in the development of medical countermeasures.Electronic supplementary materialThe online version of this article (doi:10.1186/s12985-017-0687-7) contains supplementary material, which is available to authorized users.

Aerosol infection of Balb/c mice with eastern equine encephalitis virus; susceptibility and lethality

“…Virus distribution and load was comparable with Honnold et al I, [7] in the brain, although unsurprisingly a generally higher viral load was observed in the lung, spleen and blood of mice exposed to 100 × LD 50 EEEV FL93–939 than described here. Exposure of marmosets to EEEV by the aerosol route culminates in a very similar disease course, where clinical signs were observed from 3 to 4 days post challenge, and included ruffled fur, lack of grooming and progressively, loss of balance, subdued behaviours, tremors and death in animals exposed to high doses of the virus (> 1 × 10 3 pfu inhaled dose) [12]. Unlike rodent models, the higher the challenge dose of virus the lower the MTTD and the earlier severe clinical signs were observed in marmosets, with a dose-dependent MTTD range of 4–19 days for animals that succumbed to disease.…”

Eastern Equine Encephalitis Virus Rapidly Infects and Disseminates in the Brain and Spinal Cord of Infected Cynomolgus Macaques Following Aerosol Challenge

“…In these previous studies, virus was localized almost exclusively in the brain and was readily detected in the frontal cortex, corpus striatum, thalamus, hippocampus, mesencephalon, pons, medulla oblongata, and cerebellum [11][12][13][14]. In contrast, EEEV could not be detected in the heart, liver, lung, spleen, and kidney of guinea pigs and marmosets [11,14].…”

“…The dissemination of EEEV following aerosol infection has not been investigated in previous macaque studies; however, it has been examined in mice, guinea pigs, and marmosets [11][12][13][14]. In these previous studies, virus was localized almost exclusively in the brain and was readily detected in the frontal cortex, corpus striatum, thalamus, hippocampus, mesencephalon, pons, medulla oblongata, and cerebellum [11][12][13][14]. In contrast, EEEV could not be detected in the heart, liver, lung, spleen, and kidney of guinea pigs and marmosets [11,14].…”

“…The pathology is comprised of severe pulmonary edema and congestion, multifocal hemorrhage, splenic atrophy, myocarditis, and necrosis [21,23,26,28,48,52,57]. The lack of similar pathology in visceral organs and tissues in animals infected via an aerosol route demonstrates that the route of infection substantially alters the virus dissemination [11][12][13][14]. Limited studies have examined EEEV pathogenesis in the brain utilizing TEM [16,29,36,67].…”

Erratum to: Characterization and pathogenesis of aerosolized eastern equine encephalitis in the common marmoset (Callithrix jacchus)

“…Upon publication of the original article [ 1 ], it was noticed that the author's correction requests for Table 2 had not been incorporated, due to an error in the systems. These corrections are listed below and have also been corrected in the original article:…”