The identification and development of a new plant-based feed ingredient as an alternative protein source to FM have gained the interest of the aquafeed industrial players. Therefore, this study aimed to investigate the physical, biochemical, and bacteriological properties of dietary FWM and the impacts on the growth and reproductive performances of farmed female stinging catfish, H. fossilis broodstock. Five experimental diets were formulated with different FWM inclusion (0, 25, 50, 75, and 100%). Fatty acid profiles such as 4:0, 10:0, 20:0, 21:0, 22:0, 24:0, 20:1n9, 18:3n6, 20:3n6, 20:4n6, and 22:6n3 were found in higher levels in FWM compared to the water spinach meal (WM). Meanwhile, there were no significant differences in the physical properties of the FWM experimental diets (p > 0.05). Furthermore, the experimental feed with 0, 25, 50, and 75% FWM were more palatable to the broodstock than 100% FWM. The number of total bacteria (TB) and lactic acid bacteria (LAB) in catfish diets exhibited a rising trend with an increase in FWM, while 50% of FWM-fed fish intestines had a significantly (p < 0.05) higher TB and LAB than other treatment groups. The growth, feed utilization, and reproductive variables of H. fossilis were significantly (p < 0.05) influenced by FWM inclusion at various levels. Moreover, the significantly (p < 0.05) highest oocytes weight, fertilization, egg ripeness, and ovipositor diameter were observed in the treatment of 50% FWM diet treatment group. In addition, the spawning response was 100% in all treatments except for the control group (66.67%). Significant differences (p < 0.05) were found in the hematological and serum biochemical indices in most treatment groups. In addition, the histological analysis of H. fossilis midintestinal tissue indicated that the fish fed with a 50% FWM diet had an unbroken epithelial barrier with more goblet cell arrangements and a well-organized villi structure and tunica muscularis compared to other treatment groups. These outcomes suggested that FWM at 50% inclusion is an adequate protein supplement for fish feed, resulting in better growth, reproductive performance, and health of H. fossilis broodstock development.