Enterovirus D68 (EV-D68) is an emerging virus known to cause sporadic disease and occasional epidemics of severe lower respiratory tract infection. However, the true prevalence of infection with EV-D68 is unknown, due in part to the lack of a rapid and specific nucleic acid amplification test as well as the infrequency with which respiratory samples are analyzed by enterovirus surveillance programs. During the 2014 EV-D68 epidemic in the United States, we noted an increased frequency of “low-positive” results for human rhinovirus (HRV) detected in respiratory tract samples using the GenMark Diagnostics eSensor respiratory viral panel, a multiplex PCR assay able to detect 14 known respiratory viruses but not enteroviruses. We simultaneously noted markedly increased admissions to our Pediatric Intensive Care Unit for severe lower respiratory tract infections in patients both with and without a history of reactive airway disease. Accordingly, we hypothesized that these “low-positive” RVP results were due to EV-D68 rather than rhinovirus infection. Sequencing of the picornavirus 5’ untranslated region (5’-UTR) of 49 samples positive for HRV by the GenMark RVP revealed that 33 (67.3%) were in fact EV-D68. Notably, the mean intensity of the HRV RVP result was significantly lower in the sequence-identified EV-D68 samples (20.3 nA) compared to HRV (129.7 nA). Using a cut-off of 40 nA for the differentiation of EV-D68 from HRV resulted in 94% sensitivity and 88% specificity. The robust diagnostic characteristics of our data suggest that the cross-reactivity of EV-D68 and HRV on the GenMark Diagnostics eSensor RVP platform may be an important factor to consider in making accurate molecular diagnosis of EV-D68 at institutions utilizing this system or other molecular respiratory platforms that may also cross-react.

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