Maintaining a healthy intestine is essential for growth and host defense mechanisms of farmed fish, particularly those densely stocked. In this context, maintaining a healthy intestinal microbiota for proper digestion, absorption, and integrity of the mucosa is of utmost importance. Nano-form (SeNPs)-enriched Pediastrum boryanum (Turpin) exhibits antimicrobial, antioxidant, and antitumor properties. Therefore, SeNPs are valuable nutrition supplements for farmed aquatic animals. Algogenic Se nanoparticles (SeNPs, 79.26nm) were synthesized from the green microalga Pediastrum boryanum. A Nile tilapia (Oreochromis niloticus) basal diet was blended with 0.75 or 1.5mg/kg dry feed of the SeNPs and fed for 8 weeks. Comprehensive arrays of intestinal biomarker-related parameters were investigated. Supplementation with SeNPs, particularly at a 1.5 mg/kg diet, improved growth performance, amylase and lipase activity, serotonin metabolite level, and intestinal morphometry. In addition, SeNPs supplementation suppressed serotonin levels. SeNPs at a 1.5 mg/kg diet were observed to be the optimal dose, as they showed better results concerning the increase in beneficial microbes and decrease in pathogenic microbes, demonstrating the potential role of SeNPs in growth improvement and enhanced intestinal functions. The sum of our data underscores the benefits of supplementing Nile tilapia feed with SeNPs to promote the growth and profitability of aquaculture.