This study evaluated the optimal concentration of the dietary incorporation of β-1,3-1,6-glucan (Glu) from Schizophyllum commune for enhancing the resistance to Photobacterium damselae subsp. piscicida and Streptococcus iniae infections in cobia (Rachycentron canadum). In experiment 1, fish were fed diets containing graded levels (0%, 0.1%, 0.2%, 0.4%, 0.8%, and 1.6%) of supplemental Glu for 10, 20, 30, and 40 days. Following these dietary treatments, the fish were challenged by an intraperitoneal injection of a P. damselae subsp. piscicida solution. The results showed that cobia fed the diet containing 0.4%, 0.8%, and 1.6% Glu showed a significantly (p < 0.01) enhanced resistance against P. damselae subsp. piscicida. In experiment 2, the weight gain and survival of cobia did not significantly differ (p > 0.05) after being fed the diets containing 0%, 0.5%, 1.0%, and 2.0% Glu for 15 weeks. The challenge tests showed that fish fed the diet containing 0.5% Glu for 10, 20, and 30 days showed significantly (p < 0.01) enhanced resistance against P. damselae subsp. piscicida. On the other hand, cobia fed the diet containing 0.5% Glu for 10 days showed significantly (p < 0.001) enhanced resistance against S. iniae. Results obtained from the two experiments strongly demonstrated that oral administration of 0.5% Glu for 10 days enhanced the resistance of cobia against P. damselae subsp. piscicida and S. iniae infections.