The work described in this thesis was conducted to evaluate if groundwater containing naturally high levels of iodide, consumed by lactating cows, contributes to increased levels of iodine in bulk tank milk. The study included 80 dairy herds in eastern (n=58) and southwestern (n=22) Ontario with consistently low, elevated, or high bulk milk iodine (BMI) levels. The first study objective was to evaluate the variation in BMI levels, and identify potential explanatory variables that may contribute to increased milk iodine levels on farms. Results of this analysis indicate that higher iodide content in groundwater is significantly associated with BMI concentration. In addition, BMI content was associated with the coverage goal of post-milking disinfectants on teats, and iodine content of feed. The second objective was to evaluate the isotopic composition of iodine in bulk tank milk (BTM) produced and the water consumed by the lactating herds, and to compare the respective isotopic 129I/127I ratios to determine if groundwater is a major contributing source of iodine in BTM. The isotopic fingerprint of raw milk and groundwater consumed by lactating cattle contained similar radiogenic and geogenic identities, further indicating that BMI levels can be altered by iodine inputs from groundwater consumed by the milking herd. Overall, the current field-based study provides strong evidence that groundwater is a significant contributing source of iodine to BTM in dairy herds located in eastern and southwestern Ontario.