Vitamin D deficiency can be regarded as one of the overgrowing health problem in all of the world. Evidence from a clinical trial suggested a role for probiotic bacteria in increasing vitamin D. However, probiotic's effect is strain specific and this effect should be confirmed about different strains. The objective was to determine if yogurt fortification with probiotic bacteria, Lactobillus acidophilus La‐B5, Bifidobacterium lactis Bb‐12 either alone or in combination with vitamin D can be a complementary treatment for vitamin D deficiency. The end‐points were vitamin D, cardio metabolic lipid profile, anthropometric indices (weight, height, waist, hip, fat mass, lean body mass) and dietary intake. A 10‐week parallel‐group, double‐blind, randomized and controlled trial was conducted on 140 obese men and women. The participants were randomly allocated to receive 100 grams either 1) plain low‐fat yogurt or 2) probiotic yogurt or 3) vitamin D‐fortified yogurt or 4) probiotic and vitamin D cofortified yogurt. All groups received low‐calorie diet. Vitamin D increased significantly in group 4 (p = .008), group 3 (p = .001) and group 1 (p = .012 with no difference between groups. Vitamin D‐fortified yogurt had the most effect size and showed a significant difference versus plain (p = .018) and probiotic yogurt (p = .002). Regarding lipid profile, there were no significant differences between groups. Data from this study does not support the hypothesis that yogurt fortified with probiotic bacteria, Lactobillus acidophilus La‐5 and Bifidobacterium lactis Bb‐12 either alone or in combination with vitamin D might impose any increasing effect on serum level of vitamin D in comparison with vitamin D‐fortified yogurt.