In the present study, decapsulated cysts from eleven Tunisian Artemia salina populations were analysed for their fatty acid profile. Results showed that palmitic (16:0), palmitoleic (16:1n-7), stearic (18:0), cis-vaccenic (18:1n-7), oleic (18:1n-9), linoleic (18:2n-6), linolenic (18:3n-3) and eicosapentaenoic (20:5n-3) were the major fatty acids. The ratio of C16:0/C16:1 fatty acids fluctuated between 0.8 and 3.8. Docosahexaenoic acid (22:6n-3) was absent or found in trace (<0.2%) and arachidonic acids (20:4n-6) was found in higher quantity in all marine-type cysts than in freshwater-type cysts samples. Furthermore, based on the fatty acid profile of the studied Artemia salina populations, we can concluded that Sijoumi, Sahline, Bekalta, Mcheguig and El Adhibet strains could be ascribed to marine-type Artemia, whereas the population from Moknine, Sidi El Hani, Sfax, El Melah, Zarzis and Mhabeul could be categorized as freshwater-type. Principal components analysis showed that palmitoleic acid, linolenic acid, eicosapentaenoic acid, arachidonic acid and C16:0/C16:1 ratio are the most important fatty acids variable between cysts samples, with a total contribution of 68.73% relatively to the first component, whereas, for the second component, palmitic acid, cis-vaccenic acid and oleic acid, are the most important variables, with a total contribution of 56.25%. Moreover, palmitoleic acid, linolenic acid, eicosapentaenoic acid, arachidonic acid and C16:0/C16:1 ratio are the most important fatty acids which contribute to the discrimination between freshwater and marine-type Artemia; while palmitic acid, cis-vaccenic acid and oleic acids represent the major fatty acids permitting differentiation between strains from the same Artemia type, especially for freshwater-type Artemia.