Eight bifidobacterial strains isolated from human breast milk have been tested for their abilities to convert linoleic acid (LA) andα-linolenic acid (LNA) to conjugated linoleic acid (CLA) and conjugatedα-linolenic acid (CLNA), respectively. These bioactive lipids display important properties that may contribute to the maintenance and improvement human health. Three selectedBifidobacterium brevestrains produced CLA from LA and CLNA from LNA in MRS (160–170 and 210–230 μg mL⁻¹, resp.) and, also, in reconstituted skim milk (75–95 and 210–244 μg mL⁻¹, resp.). These bifidobacterial strains were also able to simultaneously produce both CLA (90–105 μg mL⁻¹) and CLNA (290–320 μg mL⁻¹) in reconstituted skim milk. Globally, our findings suggest that these bifidobacterial strains are potential candidates for the design of new fermented dairy products naturally containing very high concentrations of these bioactive lipids. To our knowledge, this is the first study describing CLNA production and coproduction of CLA and CLNA byBifidobacterium brevestrains isolated from human milk in reconstituted skim milk.