Eastern equine encephalitis virus rapidly infects and disseminates in the brain and spinal cord of cynomolgus macaques following aerosol challenge

Abstract

Eastern equine encephalitis virus (EEEV) is mosquito-borne virus that produces fatal encephalitis in humans. We recently conducted a first of its kind study to investigate EEEV clinical disease course following aerosol challenge in a cynomolgus macaque model utilizing the state-of-the-art telemetry to measure critical physiological parameters. Here, we report the results of a comprehensive pathology study of NHP tissues collected at euthanasia to gain insights into EEEV pathogenesis. Viral RNA and proteins as well as microscopic lesions were absent in the visceral organs. In contrast, viral RNA and proteins were readily detected throughout the brain including autonomic nervous system (ANS) control centers and spinal cord. However, despite presence of viral RNA and proteins, majority of the brain and spinal cord tissues exhibited minimal or no microscopic lesions. The virus tropism was restricted primarily to neurons, and virus particles (~61–68 nm) were present within axons of neurons and throughout the extracellular spaces. However, active virus replication was absent or minimal in majority of the brain and was limited to regions proximal to the olfactory tract. These data suggest that EEEV initially replicates in/near the olfactory bulb following aerosol challenge and is rapidly transported to distal regions of the brain by exploiting the neuronal axonal transport system to facilitate neuron-to-neuron spread. Once within the brain, the virus gains access to the ANS control centers likely leading to disruption and/or dysregulation of critical physiological parameters to produce severe disease. Moreover, the absence of microscopic lesions strongly suggests that the underlying mechanism of EEEV pathogenesis is due to neuronal dysfunction rather than neuronal death. This study is the first comprehensive investigation into EEEV pathology in a NHP model and will provide significant insights into the evaluation of countermeasure.

Sequelae and Animal Modeling of Encephalitic Alphavirus Infections

“…At least half or as high as 90% of neurologic cases develop lasting sequelae [ 14 , 24 , 27 ], including seizures, convulsions, altered mental status, somnolence, and intellectual disability [ 14 , 26 , 28 ]. This results in a very high cost of typically institutionalized care, often for life [ 29 ].…”

“…Animal modeling of EEE in mice, guinea pigs, and macaques suggests that viral entry into the brain, inducing the associated neurologic disease and sequelae, may occur through the olfactory bulb in the case of aerosol exposure [ 23 , 27 , 32 ]. However, the related VEE and WEE viruses can enter murine brains through the circumventricular organs before spreading to other regions after peripheral infection; this non-olfactory entry route cannot be ruled out for EEEV following peripheral infection [ 33 ].…”

“…However, the related VEE and WEE viruses can enter murine brains through the circumventricular organs before spreading to other regions after peripheral infection; this non-olfactory entry route cannot be ruled out for EEEV following peripheral infection [ 33 ]. Moreover, the cause of neurologic signs may be from neurologic dysfunction as opposed to neuronal death, as suggested by the lack of histopathological lesions within the brain [ 27 ]. As EEEV has previously been studied as a potential bioweapon, understanding all potential mechanisms of neurologic infiltration and disease following respiratory infection is crucial for vaccine and therapeutic development.…”

“…The development of therapies to minimize sequelae and promote recovery relies on animal modeling. Current lethal models resulting in neurologic signs include mice, golden hamsters, guinea pigs, and non-human primates including cynomolgus macaques and the common marmoset (see Table 1 ) [ 23 , 27 , 32 , 34 , 35 , 36 , 37 , 38 , 39 , 40 , 41 , 42 ]. The majority of modeling is focused on intranasal or aerosol exposure, which may be relevant from a biodefense perspective [ 32 , 34 , 35 , 37 , 38 , 40 , 41 ].…”

Entry receptors — the gateway to alphavirus infection

“…In 1995, an epidemic with 75,000 patients occurred in Columbia with 300 deaths and 3,000 patients with neurological complications (16). Both VEEV and EEEV were weaponized as aerosol agents by the United States and the Soviet Union during the Cold War (17)(18)(19). WEEV has a case fatality rate of 3% to 7% (7).…”