This study aimed to investigate the effects of dietary supplementation with Machilus thunbergii (MT), a natural coastal plant resource, on the growth performance, feed utilization, innate immune response, antioxidant status, and resistance to Vibrio parahaemolyticus causing acute hepatopancreatic necrosis disease (AHPND) of Penaeus vannamei. Experimental diets containing 0.0%, 0.5%, and 1.0% MT extract powder (designated as MT0.0, MT0.5, and MT1.0, respectively) were formulated. After the 5-week feeding trial, shrimp fed the MT0.5 diet showed significantly higher final body weight, weight gain, and protein efficiency ratio, along with a lower feed conversion ratio, than shrimp fed the MT0.0 diet. The activities of lysozyme, superoxide dismutase, and glutathione peroxidase, along with the expression of crustin and penaeidin-3a, were significantly higher in shrimp fed the MT0.5 diet, suggesting modulation of innate immune and antioxidant responses. During the AHPND challenge test, no significant differences in final survival were observed among the dietary treatments; however, shrimp fed the MT1.0 diet showed a tendency toward higher survival. Histopathological analysis also indicated partial restoration of hepatopancreatic tissue structure in the MT0.5 and MT1.0 groups at 150 h post-infection. To our knowledge, limited information is available regarding the dietary application of MT in shrimp aquaculture. The present findings suggest that MT supplementation may influence growth performance, immune responses, and antioxidant status of P. vannamei, suggesting its potential application as a phytogenic feed additive.

Abstract
Introduction
Materials and Methods
    1. Experimental diets and design
    2. Experimental shrimp and rearing conditions
    3. Sample collection
    4. Innate immune parameters
    5. Quantitative PCR analysis for immune-related genes
    6. Challenge trial
    7. Statistical analysis
Results
    1. Growth performance and feed utilization
    2. Innate immune and antioxidant responses
    3. Immune-related gene expression
    4. Responses to AHPND challenge
Acknowledgement
References

• Chorong Lee(Research Professor, Institute of Smart Space Agriculture (ISSA), Gyeongsang National University, Jinju 52828, Korea)
• Seong Kyoon Choi(Principal Researcher, Core Protein Resources Center, DGIST, Daegu 42988, Korea)
• Hye Jin Jeon(Postdoctoral Researcher, Institute for Veterinary Biomedical Science, Kyungpook National University, Daegu 41566, Korea)
• Jee Eun Han(Associate Professor, Institute for Veterinary Biomedical Science, Kyungpook National University, Daegu 41566, Korea)
• Song Park(Assistant Professor, Division of Animal Science, College of Agriculture and Life Science, Gyeongsang National University, Jinju 52828, Korea) Corresponding author