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Most infections are associated with flu-like symptoms, but paralysis may develop in young children. It has been suggested that infection only with recent viral isolates can cause paralysis. To address the hypothesis that EV-D68 has recently acquired neurotropism, murine organotypic brain slice cultures, induced human motor neurons and astrocytes, and mice lacking the alpha\/beta interferon receptor were infected with multiple virus isolates. All EV-D68 isolates, from 1962 to the present, can infect neural cells, astrocytes, and neurons. Furthermore, our results show that sialic acid binding does not play a role in EV-D68 neuropathogenesis. 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