Contemporary Enterovirus D68 strains show enhanced replication and translation at 37°C

Abstract

Enterovirus D68 (EV-D68) emerged in 2014 as an important pathogen linked to severe lower respiratory disease and acute flaccid myelitis outbreaks. Historically associated with mild common-cold-like symptoms, clusters of severe disease attributed to EV-D68 appeared during a series of outbreaks in 2014, 2016, and 2018. Previous studies of historic EV-D68 strains demonstrated attenuated replication at temperatures of the lower respiratory tract (37{degrees}C), when compared to the upper respiratory tract (32{degrees}C). By testing a panel of historic and contemporary EV-D68 strains at 32{degrees}C and 37{degrees}C, we demonstrate that contemporary strains of EV-D68 undergo little to no attenuation at increased temperatures. Contemporary strains produced higher levels of viral proteins at 32{degrees}C and 37{degrees}C than historic strains, although both strains infected similar numbers of cells and had comparable amounts of replication complexes. IRES activity assays with dual-luciferase reporter plasmids demonstrated enhanced translation in recent EV-D68 strains mapped to regions of variability in the 5 UTR found only in contemporary strains. Using an infectious clone system, we demonstrate that the translation advantage dictated by the 5 UTR does not solely mediate temperature sensitivity. The strain-dependent effects of temperature on the EV-D68 life cycle gives insight into the susceptibility of the lower respiratory system to contemporary strains. IMPORTANCEEnterovirus-D68 (EV-D68) emerged in 2014 as a causative agent of biannual severe pediatric respiratory disease and acute flaccid myelitis (AFM). We show that recent EV-D68 viruses have gained the ability to replicate at 37{degrees}C. Enhanced virus protein translation seemed to correlate with enhanced virus replication at 37{degrees}C but other genetic factors are also contributing to this phenotype. An enhanced ability to replicate at core body temperature may have allowed EV-D68 to penetrate both lower in the respiratory tract and into the central nervous system, explaining the recent surge in severe disease associated with virus infection.