The present study investigated the induction of immunological, hormonal and histological changes in the freshwater fish, Pseudetroplus maculatus after sublethal exposure of chlorpyrifos. Fish were exposed to chlorpyrifos at one-tenth (0.661 mu g/L) and one-fifth (1.32 mu g/L) of LC50 value, for 15 and 30 d, along with the respective control group. Innate and adaptive immune responses of the fish against the toxicant exposure were measured using lysozyme, complement (ACH(50)) levels, phagocytic, nitroblue tetrazolium (NBT), myeloperoxidase (MPO), anti-protease and hemagglutination activities, and IgM concentration. The results revealed that sublethal exposure of chlorpyrifos caused significant (p < 0.05) reduction in lysozyme, ACH(50), phagocytic, and anti-protease activities whereas there was significant (p < 0.05) increase in NBT, MPO and hemagglutination levels along with serum IgM concentration. Chlorpyrifos treatment showed significant (p < 0.05) decline in the serum levels of cortisol, thyroid, testosterone and estradiol hormones in duration- and concentration-dependent manner. The major histological lesions noted in liver includes necrosis, vacuolization, hepatocytic and cytoplasmic degeneration, while kidneys showed vacoules, necrosis and rupture in renal tubules and glomerulus, whereas spleen were found with melanomacrophage aggregation and necrosis. Similarly, testis showed remarkable changes like reduction in the number of spermatozoa and disintegrated seminiferous tubules while ovarian lesions include degenerated and empty follicles, few atretic oocytes, reduced size of follicles, and broken theca granulosa. The current findings revealed that the use of chlorpyrifos in domestic and agricultural purposes even at sublethal concentration could affect the non-target organisms including fish, and thereby alter the health status of aquatic ecosystems.