Genomic Analyses of Acute Flaccid Myelitis Cases Among a Potential Cluster in Arizona Provide Further Evidence of Enterovirus D68 Role. human metagenome

Enteroviruses are a common cause of respiratory and gastrointestinal illness, and multiple subtypes, including poliovirus, can cause neurologic disease. In recent years, enterovirus D68 (EV-D68) has been associated with serious neurologic illnesses, including acute flaccid myelitis (AFM), frequently preceded by respiratory disease. A cluster of eleven suspect cases of pediatric AFM was identified in September of 2016 in Phoenix AZ. To determine if these cases were associated with EV-D68, we performed multiple genomic analyses of nasopharyngeal swabs and CSF material from the patients, including real-time PCR and amplicon sequencing targeting the EV-D68 VP1 gene, and unbiased microbiome sequencing (i.e., 16S rRNA gene and DNA and RNA shotgun metagenomic sequencing). Four of the 11 patients were classified as confirmed cases of AFM, and an additional case was classified as probable AFM. Real-time PCR and amplicon sequencing detected EV-D68 virus RNA in the three AFM patients from which NP swabs were collected, as well as in a fourth patient diagnosed with acute disseminated encephalomyelitis, a disease that commonly follows bacterial or viral infections, including enterovirus. No other obvious etiological causes for AFM were found by 16S or RNA/DNA metagenomic sequencing in these cases, strengthening the likelihood that EV-D68 is a cause. Herpes simplex viral DNA was identified in the CSF of the fourth case of AFM and in one additional suspect case from the cluster. The rapid molecular assays developed for this study could be used to diagnose patients with suspected EV-D68 respiratory illness, to aid in AFM diagnosis, and for future EV-D68 surveillance and epidemiology.