ObjectiveTo determine the extent of vitamin D3 deficiency and levels in pregnant First Nations and non-First Nations women in SK. Also, to determine the distribution of vitamin D3 values in the general population in SK. MethodsVitamin D3 levels were measured by LC–MS/MS from 19,181 consecutive patient blood/serum samples received at the Saskatchewan Disease Control Laboratory, and from 743 First Nations, and 301 non-First Nations pregnant women in SK. ResultsThe ages of the 19,181 patient samples ranged from day1 (0years) to 102years. Of the total, 14,658 were female, and 4523 were males. 30.8% had relative vitamin D3 insufficiency (50–75nmol/L), and 22.5% were in the deficient range (<50nmol/L). In summer, a larger percentage of SK patients are in the optimum range, whereas in winter, the number of patients in the vitamin D3 deficiency range increased to 33.0% from 14.1%. Samples from pregnant women were collected during the first trimester of pregnancy. Whereas non-First Nations pregnant women had similar vitamin D3 levels to non-pregnant women in SK, vitamin D3 levels were significantly lower than the optimum of 75nmol/L in pregnant First Nations women than in non-First Nations women. 29.7% of First Nations pregnant women were in the relative insufficiency range, and 45.6% were vitamin D3 deficient. ConclusionsFirst Nations pregnant women have lower vitamin D3 levels than non-First Nations pregnant women. This puts them and their unborn babies at high risk of a diverse range of disorders associated with vitamin D3 deficiency or insufficiency.