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MBio. 2018 Oct 16;9(5). pii: e01954-18. doi: 10.1128/mBio.01954-18.

Contemporary Circulating Enterovirus D68 Strains Have Acquired the Capacity for Viral Entry and Replication in Human Neuronal Cells.

Author information

1
J. Craig Venter Institute, Rockville, Maryland, USA.
2
Neuroscience Program and Medical Scientist Training Program, University of Colorado School of Medicine, Aurora, Colorado, USA.
3
J. Craig Venter Institute, La Jolla, California, USA.
4
Department of Neurology, University of Colorado School of Medicine, Aurora, Colorado, USA.
5
Denver VA Medical Center, Denver, Colorado, USA.
6
Department of Immunology and Microbiology, University of Colorado School of Medicine, Aurora, Colorado, USA.
7
Department of Medicine, University of Colorado School of Medicine, Aurora, Colorado, USA.
8
J. Craig Venter Institute, La Jolla, California, USA RScheuermann@jcvi.org.
9
Department of Pathology, University of California, La Jolla, California, USA.

Abstract

Enterovirus D68 (EV-D68) has historically been associated with respiratory illnesses. However, in the summers of 2014 and 2016, EV-D68 outbreaks coincided with a spike in polio-like acute flaccid myelitis/paralysis (AFM/AFP) cases. This raised concerns that EV-D68 could be the causative agent of AFM during these recent outbreaks. To assess the potential neurotropism of EV-D68, we utilized the neuroblastoma-derived neuronal cell line SH-SY5Y as a cell culture model to determine if differential infection is observed for different EV-D68 strains. In contrast to HeLa and A549 cells, which support viral infection of all EV-D68 strains tested, SH-SY5Y cells only supported infection by a subset of contemporary EV-D68 strains, including isolates from the 2014 outbreak. Viral replication and infectivity in SH-SY5Y were assessed using multiple assays: virus production, cytopathic effects, cellular ATP release, and VP1 capsid protein production. Similar differential neurotropism was also observed in differentiated SH-SY5Y cells, primary human neuron cultures, and a mouse paralysis model. Using the SH-SY5Y cell culture model, we determined that barriers to viral binding and entry were at least partly responsible for the differential infectivity phenotype. Transfection of genomic RNA into SH-SY5Y generated virions for all EV-D68 isolates, but only a single round of replication was observed from strains that could not directly infect SH-SY5Y. In addition to supporting virus replication and other functional studies, this cell culture model may help identify the signatures of virulence to confirm epidemiological associations between EV-D68 strains and AFM and allow for the rapid identification and characterization of emerging neurotropic strains.IMPORTANCE Since the EV-D68 outbreak during the summer of 2014, evidence of a causal link to a type of limb paralysis (AFM) has been mounting. In this article, we describe a neuronal cell culture model (SH-SY5Y cells) in which a subset of contemporary 2014 outbreak strains of EV-D68 show infectivity in neuronal cells, or neurotropism. We confirmed the difference in neurotropism in vitro using primary human neuron cell cultures and in vivo with a mouse paralysis model. Using the SH-SY5Y cell model, we determined that a barrier to viral entry is at least partly responsible for neurotropism. SH-SY5Y cells may be useful in determining if specific EV-D68 genetic determinants are associated with neuropathogenesis, and replication in this cell line could be used as rapid screening tool for identification of neurotropic EV-D68 strains. This may assist with better understanding of pathogenesis and epidemiology and with the development of potential therapies.

KEYWORDS:

EV-D68; SH-SY5Y; enterovirus; myelitis; neurotropism; neurovirology

Comment in

PMID:
30327438
PMCID:
PMC6191546
DOI:
10.1128/mBio.01954-18
[Indexed for MEDLINE]
Free PMC Article

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