Enterovirus D68 (EV-D68) has caused recurring respiratory disease outbreaks in the United States since 2014. The dominant circulating EV-D68 strain has evolved from clade B1 to the more recent B2 and B3 clades. As recurrent outbreaks and continued virus evolution are expected for EV-D68, a robust real-time PCR assay that detects known strains as well as potential emerging strains is critical for national surveillance and clinical diagnostics. We describe a type-specific EV-D68 real-time RT-PCR (rRT-PCR) assay termed CDC2022, which targets sequences encoding conserved amino acid regions of all extant EV-D68 strains. We targeted three motifs conserved among all strains in the last 60 years. The assay achieved 100% (270/270) sensitivity and 100% (344/344) specificity when tested with a collection of 613 respiratory specimens, compared to the gold-standard EV semi-nested VP1 PCR and sequencing assay (snPCR/Seq). CDC2022 gave negative results with 289/289 non-target viruses, including 104 EV A-D isolates, 165 rhinovirus (RV) isolates or clinical specimens, and 14 other common respiratory viruses. The assay can detect as few as 0.28 CCID<inf>50</inf> per reaction. An in silico "phylo-primer-mismatch" analysis was performed to visualize primer/probe mismatches and to compare CDC2022 with other EV-D68 rRT-PCR assays, including the previous CDC assay (CDC2015) developed in 2014 for clade B1 strains. It showed that CDC2022 has the fewest primer/probe mismatches among all assays analyzed and is suitable for all clades. We additionally tested 11 EV-D68-positive clinical specimens from 2022 that were confirmed by snPCR/Seq, and all were detected. CDC2022 assay could provide a critical tool for molecular surveillance of EV-D68. Copyright The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. This article is a US Government work. It is not subject to copyright under 17 USC 105 and is also made available for use under a CC0 license.

We report the whole-genome sequences of new enterovirus D94 and D111 strains, isolated from cultures from stool specimens collected from acute flaccid paralysis (AFP) cases for poliovirus surveillance in Angola during 2010.

BACKGROUND: In 2014, a nationwide outbreak of severe respiratory illness occurred in the United States, primarily associated with enterovirus D-68 (EV-D68). A proportion of illness was associated with rhinoviruses and other enteroviruses, which we aimed to characterize further. METHODS: Respiratory specimens from pediatric and adult patients with respiratory illness were submitted to CDC during August - November 2014. While initial laboratory testing focused on identification of EV-D68, the negative specimens were typed by molecular sequencing to identify additional enterovirus (EV) and rhinovirus (RV) types. Testing for other pathogens was not conducted. We compared available clinical and epidemiologic characteristics among patients with EV-D68 and RV species A-C identified. RESULTS: Among 2629 typed specimens, 1012 were EV-D68 (39%) and 81 (3.1%) represented 24 other EV types; 968 were RVs (37%) covering 114 types and grouped into three human RV species (RV-A=446, RV-B=133, RV-C=389); and 568 (22%) had no RV or EV detected. EV-D68 was more frequently identified in patients presenting earlier in the investigation period. Among patients with EV-D68, RV-A, RV-B, or RV-C identified, the age distributions markedly differed. Clinical syndromes and ICU admissions by age were largely similar among those with EV-D68 and RV species. CONCLUSIONS: RVs were commonly associated with severe respiratory illness, alongside EV-D68, during the nationwide outbreak. Clinical characteristics were largely similar among those with EV-D68, RV-A, RV-B, or RV-C. A better understanding of the epidemiology of RVs and EVs is needed to help inform development and use of diagnostic tests, therapeutics, and preventive measures.

Background: Enterovirus D68 (EV-D68) caused a widespread outbreak of respiratory illness in the United States in 2014, predominantly affecting children. We describe EV-D68 rates, spectrum of illness and risk factors from prospective, population-based acute respiratory illness (ARI) surveillance at a large U.S. pediatric hospital. Methods: Children <13 years of age with ARI and residence in Hamilton County, Ohio were enrolled from the inpatient and emergency department (ED) settings at a children's hospital in Cincinnati, Ohio, from July 1-October 31, 2014. For each participant, we interviewed parents, reviewed medical records, and tested nasal and throat swabs for EV-D68 using real-time RT-PCR assay. Results: EV-D68 infection was detected in 51/207 (25%) inpatients and 58/505 (11%) ED patients. Rates of EV-D68 hospitalization and ED visit were 1.3 (95% CI: 1.0-1.6) and 8.4 per 1,000 children <13 years of age, respectively. Pre-existing asthma was associated with EV-D68 infection (aOR 3.2; 95 %CI 2.0-5.1). Compared with other ARI, children with EV-D68 were more likely to be admitted from the ED (p=<0.001), receive supplemental oxygen (p=0.001), and require ICU admission (p=0.04); however, mechanical ventilation was uncommon (2/51 inpatients, p=0.64), and no deaths occurred. Conclusions: During the 2014 EV-D68 epidemic, high rates of pediatric hospitalizations and ED visits were observed. Children with asthma were at increased risk for medically attended EV-D68 illness. Preparedness planning for a high-activity EV-D68 season in the U.S. should take into account increased healthcare utilization, particularly among children with asthma, during the late summer and early fall.

The genomic sequences of three 2016 enterovirus D68 (EV-D68) strains were obtained from respiratory samples of patients from Florida, Texas, and New York. These EV-D68 sequences share highest nucleotide identities with strains that circulated in North America, Europe, and Asia in 2014-2015.

BACKGROUND: Sporadic cases of parotitis are generally assumed to be mumps, which often requires a resource-intensive public health response. This project surveyed the frequency of viruses detected among such cases. METHODS: During 2009-2011, 8 jurisdictions throughout the United States investigated sporadic cases of parotitis. Epidemiologic information, serum, and buccal and oropharyngeal swabs were collected. Polymerase chain reaction methods were used to detect a panel of viruses. Anti-mumps virus immunoglobulin M (IgM) antibodies were detected using a variety of methods. RESULTS: Of 101 specimens, 38 were positive for a single virus: Epstein-Barr virus (23), human herpesvirus (HHV)-6B (10), human parainfluenza virus (HPIV)-2 (3), HPIV-3 (1), and human bocavirus (1). Mumps virus, enteroviruses (including human parechovirus), HHV-6A, HPIV-1, and adenoviruses were not detected. Early specimen collection did not improve viral detection rate. Mumps IgM was detected in 17% of available specimens. Patients in whom a virus was detected were younger, but no difference was seen by sex or vaccination profile. No seasonal patterns were identified. CONCLUSIONS: Considering the timing of specimen collection, serology results, patient vaccination status, and time of year may be helpful in assessing the likelihood that a sporadic case of parotitis without laboratory confirmation is mumps.

The National Research Council has recommended that at least one, preferably two, polio antiviral drugs be developed as a supplement to the tools currently available for control of polio outbreaks post-eradication. The primary application of such drugs is expected to be the resolution of chronic poliovirus excretion in persons with primary immunodeficiency disorders. We have assessed the in vitro activity of AG-7404 (also known as "compound 1"), an inhibitor of picornaviral 3C protease, against a large panel of programmatically important poliovirus strains and its activity in combination with two poliovirus capsid inhibitors, V-073 and BTA798. AG-7404 was active against all viruses in this panel, with EC50 values ranging from 0.080 to 0.674muM. Similarly, BTA798 was active against all viruses in this panel, with EC50 values ranging from 0.003 to 0.591muM. By comparison, values for V-073 were 0.003-0.126muM. BTA798 was active against V-073-resistant variants with an alanine to valine change in VP3 at position 24. However, BTA798 was inactive against the V-073-resistant strains with amino acid substitutions at VP1 amino acids 194 (equivalent to 192 in type 3) and 236. As expected from its different mechanism of action, AG-7404 was fully active against all V-073-resistant variants, with EC50 values ranging from 0.218 to 0.819muM, compared to values of 0.202-0.407muM for the V-073-susceptible parental strains. In vitro drug combination experiments demonstrated synergy between AG-7404 and either V-073 or BTA798, whereas the combination of the two capsid inhibitors acted additively.

Human enterovirus 68 (EV68) infections are rarely reported. We describe a respiratory outbreak associated with EV68 among 18 children admitted to a remote Indian Health Service facility during August 11-September 14, 2010. Clinical illness was characterized by pneumonia and wheezing. EV68 should be considered as an etiology in outbreaks of lower respiratory tract illness.