Habitat loss, human persecution, and infectious diseases all threaten declining reptile populations. Lake Erie watersnakes (Nerodia sipedon insularum, LEWS), once classified as an endangered species in part due to human persecution, have recovered to stable population levels but have been observed with a high prevalence of ophidiomycosis. Strategies are needed to mitigate the current disease threat, including assessing overall wellness. Hematologic analysis provides information about the presence of inflammation and infection and thus informs health-based conservation efforts, but has not been previously performed in LEWS. The objective of this study was to evaluate hematologic parameters in LEWS and identify differences based on ophidiomycosis status. Blood was drawn from wild-caught snakes at nine sites in 2018 and 2019 and complete blood counts were performed in 180 individuals. For apparently healthy snakes, packed cell volume was significantly higher in males (median = 32.5%) compared to females (median = 26.5%; P = 0.03). Animals classified as having possible or apparent ophidiomycosis, or those with skin lesions, had a relative azurophilia and lymphopenia compared to individuals classified as negative or Ophidiomyces present, or those without skin lesions (P < 0.01). This is the first study to investigate hematology in a free-ranging population of LEWS and will serve as a baseline for future investigations that aim to improve conservation efforts through population health monitoring.