Neonatal Escherichia coli (E. coli) sepsis is increasing. There is limited data on the factors contributing to increased mortality and severity of illness in neonatal E. coli sepsis. A retrospective review of neonates (<30 days) admitted to a Level IV NICU in the United States from 2008 to 2022 diagnosed with E. coli bloodstream or cerebrospinal fluid infection was conducted. Primary outcome was defined as mortality from or severe illness during E. coli infection (defined as a need for inotropic support or metabolic acidosis). E. coli neonatal sepsis rate increased from 2008 to 2022 (average of 1.12 per 1000 live births). The primary outcome, which occurred in 57.4% of cases, was independently associated with prematurity, neutropenia, and thrombocytopenia. Ampicillin resistance was not associated with the primary outcome. GA, neutropenia, and thrombocytopenia but not ampicillin resistance, are associated with mortality or severe illness from E. coli sepsis.