BACKGROUND: Folate is essential for the synthesis and integrity of DNA, normal cell formation, and body growth. Folate deficiency among women of reproductive age (WRA) increases the risk of poor birth outcomes including neural tube defect (NTD)-affected pregnancies. Folate status is largely dependent on dietary intakes. OBJECTIVES: We aimed to explore the spatial distribution of biomarkers of folate status and their association with farming systems among nonpregnant WRA in Ethiopia. METHODS: Serum and RBC folate concentration data were derived from the Ethiopia National Micronutrient Survey of 2015. The spatial dependencies of folate concentration of WRA were investigated and its relation with the dominant local farming system was explored. RESULTS: The median serum folate and RBC folate concentrations were 12.3 nmol/L and 567.3 nmol/L, respectively. The national prevalence of folate deficiency using homocysteine concentration as a metabolic indicator based on serum and RBC folate concentration was 11.6% and 5.7%, respectively. The majority of women (77.9%) had low RBC folate concentrations consistent with increased risk of NTD-affected pregnancies. Folate nutrition was spatially dependent at distances of ≤ 300 km. A marked variability in folate concentration was observed between farming systems: greater RBC folate concentration (median: 1036 nmol/L) was found among women from the Lake Tana fish-based system, whereas the lowest RBC folate concentration (median: 386.7 nmol/L) was observed in the highland sorghum chat mixed system. CONCLUSIONS: The majority (78%) of WRA in Ethiopia had low folate status potentially increasing the risk of NTD-affected pregnancies. These findings may help national and subnational nutrition intervention strategies to target the most affected areas in the country.