Protective Effects of Andrographis paniculata Extract on Nilem ( Osteochilus vittatus ) Spermatozoa During Low-Temperature Preservation: Physiological and Ultrastructural Evidence

Introduction: Nilem ( Osteochilus vittatus ) is a freshwater fish with considerable potential for aquaculture. However, information regarding the preservation of its spermatozoa remains limited. Objectives: This study aimed to evaluate the effects of Andrographis paniculata extract on sperm motility, viability, morphological abnormalities, and ultrastructure of nilem ( O. vittatus ) spermatozoa during cold storage. Methods: Sperm samples were diluted in fish Ringer’s solution supplemented with A. paniculata extract at concentrations of 0% (P0), 8% (P1), 9% (P2), and 10% (P3). The samples were stored at 4°C for 48 hours. Sperm quality was evaluated based on motility, viability, and morphological abnormalities, while ultrastructural changes were examined using cryogenic scanning electron microscopy (cryo-SEM). Results: The concentration of A. paniculata extract significantly affected sperm quality. Treatment P2 produced the highest motility (53.55% ± 4.87%) and viability (93.87% ± 1.23%) and the lowest percentage of morphological abnormalities (50.31% ± 8.86%). In contrast, the control group exhibited the lowest motility and the highest percentage of abnormalities. Cryo-SEM observations showed that spermatozoa in P2 retained relatively intact head, midpiece, and flagellar structures. In comparison, membrane disruption, head deformation, and flagellar abnormalities were observed more prominently in P0 and P3. Conclusion: Supplementation with 9% A. paniculata extract effectively maintained the physiological quality and ultrastructural integrity of nilem spermatozoa during storage at 4°C for 48 hours. This protective effect may be related to the extract’s antioxidant properties, although direct measurement of oxidative stress biomarkers is needed to confirm this mechanism.