Lipids are key nutrients that affect the development of the ovary in aquatic animals. This study aimed to investigate the effects of different lipid sources on the ovarian development and health of Macrobrachium rosenbergii broodstock. The M. rosenbergii broodstock, with an initial body weight of 8.07±0.74 g, were fed diets containing 5 % fish oil (FO), soya bean oil (SO), rapeseed oil (RO), coconut oil (CO), and a 1:1 mixture of fish oil and soybean oil (MO) as lipid sources for a period of 56 days. The results indicated that there were no significant differences in survival rate (SR) and hepatopancreatic index (HSI) among the different groups (P > 0.05). However, the specific growth rate (SGR) and gonadosomatic index (GSI) in the MO group were significantly higher than other groups (P < 0.05). Additionally, fatty acid levels in diets directly influenced the fatty acid composition of prawn hepatopancreas, showing a similar trend to that observed in the diet itself. Furthermore, compared to the other trial groups, haemolymph levels of 17-β estradiol (E2) and progesterone (PROG) were significantly increased in the MO group (P < 0.05). The ovarian section also exhibited better histomorphology and larger oocyte diameter in this group. Moreover, total antioxidant capacity was significantly increased in the MO group (P < 0.05). Saturated fatty acids in RO and CO groups could promote the expression of lipid synthesis-related genes fatty acid synthase (fas) and acetyl-CoA carboxylase (acc), concurrently suppressing the expression of lipid catabolism-related genes acyl-CoA oxidase-1 (acox-1) and AMP-activated protein kinase alpha (ampkα) (P < 0.05). Conversely, polyunsaturated fatty acids in the MO group significantly up-regulated the expression levels of fatty acid binding protein-3 (fabp-3) and peroxisome proliferator-activated receptor gamma (pparγ) (P < 0.05). Based on the findings of hepatopancreatic tissue structure, it was determined that the use of a single vegetable oil resulted in some damage to the hepatopancreatic tissue structure of M. rosenbergii broodstock, leading to increased oxidative stress. In conclusion, it is recommended that a 1:1 mixture of fish oil and soybean oil be used as a dietary lipid source to effectively meet the nutritional requirements during the ovarian development period of M. rosenbergii broodstock. This combination also enhances antioxidant properties, promotes the synthesis of haemolymph steroid hormones and lipid metabolism in female prawns, thereby facilitating ovarian development and overall health of M. rosenbergii broodstock.