The mode of action of honey bee pollen (HBP) was investigated in Nile tilapia (Oreochromis niloticus) challenged with Aeromonas hydrophila. Thus, fish with an average weight of 29 ± 3 g were divided into four groups, and fed with HBP-free diet (control), and 1%, 2.5% and 4% (w/v) HBP incorporated into basal diet for 10, 20 and 30 days. Immunological, hematological, biochemical and growth parameters were measured, and sub-groups of fish were challenged with A. hydrophila via intraperitoneal injection. HBP significantly increased the growth performance parameters [body weight, length, average daily gain (ADG), specific growth rate (SGR), and feed efficiency ratio (FER)] and immunological (phagocytic activity, serum bactericidal activity and nitroblue tetrazolium assay (NBT)), hematological (hematocrit (Hct), leucocrit (Lct), the numbers of neutrophils, monocytes and lymphocytes) and biochemical parameters (serum total protein, albumin and globulin ratios). Furthermore, all treated fish exhibited significant protection against challenge with A. hydrophila, with the highest protection (93%) observed in the group fed with 2.5% (w/v) HBP for 20 and 30 days.