BACKGROUND: Enterovirus infections are very common and typically cause mild illness, although neonates are at higher risk for severe illness. In 2007, the Centers for Disease Control and Prevention (CDC) received multiple reports of severe neonatal illness and death associated with coxsackievirus B1 (CVB1), a less common enterovirus serotype not previously associated with death in surveillance reports to the CDC. METHODS: This report includes clinical, epidemiologic, and virologic data from cases of severe neonatal illness associated with CVB1 reported during the period from 2007 through 2008 to the National Enterovirus Surveillance System (NESS), a voluntary, passive surveillance system. Also included are data on additional cases reported to the CDC outside of the NESS. Virus isolates or original specimens obtained from patients from 25 states were referred to the CDC picornavirus laboratory for molecular typing or characterization. RESULTS: During 2007-2008, the NESS received 1079 reports of enterovirus infection. CVB1 accounted for 176 (23%) of 775 reported cases with known serotype, making it the most commonly reported serotype for the first time ever in the NESS. Six neonatal deaths due to CVB1 infection were also reported to the CDC during that time. Phylogenetic analysis of the 2007 and 2008 CVB1 strains indicated that the increase in cases resulted from widespread circulation of a single genetic lineage that had been present in the United States since at least 2001. CONCLUSIONS: Healthcare providers and public health departments should be vigilant to the possibility of continuing CVB1-associated neonatal illness, and testing and continued reporting of enterovirus infections should be encouraged.