Here we describe a case of facultative parthenogenesis in a diamondback water snake (Nerodia rhombifer). An adult female N. rhombifer kept in captivity produced four unfertilized ova, one stillborn and one live neonate in July 2016. The neonates had characteristic abnormalities in morphology and were determined as males, suggestive of a parthenogenetic event. Stillborn and live neonates along with the mother and four unrelated N. rhombifer were genetically screened at twelve microsatellite loci. Results confirmed that the reproductive event represented a case of facultative parthenogenesis. In light of previous reports of facultative parthenogenesis in the genera Nerodia and Thamnophis, these results suggest that this reproductive mode may be a widespread phenomenon in the Natricinae as a whole. © 2020 Folium Publishing Company.