CONCLUSION

Dietary supplementation with fermented L. leucocephala enhanced several innate immune responses in _M. _rosenbergii challenged with _A. _hydrophila. Fermentation improved the nutritional quality of L. leucocephala by increasing essential amino acid content and reducing mimosine concentration, while supplementation of experimental diets increased TPC, TFC, antioxidant activity, and reducing power. Prawns receiving fermented L. leucocephala, particularly at the 20% inclusion level, exhibited enhanced hemocyte proliferation, increased hematopoietic activity, elevated PO activity, and upregulated expression of key immune- and stress-related genes, including CHF, proPO, IMD, Relish, HSP70, Cu/Zn-SOD, and ALF. These responses were accompanied by improved resistance to _A. _hydrophila infection and the maintenance of normal intestinal morphology, indicating that the evaluated dietary inclusion levels were well tolerated.

From a practical perspective, the findings suggest that fermented L. leucocephala has potential as a functional feed ingredient and partial fishmeal substitute for freshwater prawn culture, providing both nutritional and immunological benefits. The study further demonstrates that fermentation with L. plantarum may reduce ANFs while enhancing the bioactive and antioxidant properties of plant-derived feed ingredients.

A major strength of this study is the comprehensive evaluation of immune responses through hematological, histological, molecular, and antioxidant-related assessments following bacterial challenge.

Future studies should employ isonitrogenous and amino acid-balanced diets, evaluate long-term growth and production performance, characterize fermentation-derived metabolites and microbial dynamics, quantify pathogen clearance, and investigate gut microbiota responses. Such studies will help clarify the mechanisms underlying immune modulation and establish the optimal inclusion level of fermented L. leucocephala for commercial aquaculture applications.

Overall, the present study provides evidence that fermented L. leucocephala can serve as a promising functional feed ingredient that enhances innate immunity, antioxidant status, and disease resistance in _M. _rosenbergii, supporting its potential application in sustainable prawn aquaculture.