Abstract Trichlorfon is an organophosphorus (OP) insecticide that is used as an agriculture pesticide to destroy insects, a human medicine to combat internal parasites, and an ectoparasiticide in the livestock and aquaculture industries, but which has caused aquatic toxicity in the prawn industry. The aim of this study was to investigate the effects of trichlorfon on molecular and enzymatic processes of the immunological response of the giant freshwater prawn, Macrobrachium rosenbergii, at 0, 0.2, and 0.4mgL−1 with 0, 3, 6, 12, and 24h of exposure. The total hemocyte count (THC), respiratory bursts (RBs), phenoloxidase (PO) activity, and superoxide dismutase (SOD) activity were examined to evaluate immunological responses and oxidative stress. Results showed that THCs of the prawn exposed to trichlorfon at both concentrations (0.2 and 0.4mgL−1) had increased after 12 and 24h; SOD and PO activities had significantly increased at 3h, whereas production of RBs had dramatically increased as oxidative stress at each sampling time after exposure to trichlorfon compared to the control. A potential biomarker of OPs, acetylcholinesterase (AChE) revealed a significant decrease after exposure for 6h, and showed a time-dependent tendency. Immune gene expressions, including prophenoloxidase (proPO), the lipopolysaccharide- and β-1,3-glucan-binding protein (LGBP), peroxinectin (PE), α2-macroglubulin (α2M), transglutaminase (TG), and copper, zinc (Cu,Zn)-SOD, of prawns exposed to trichlorfon at 0, 0.2, and 0.4mgL−1 for 0, 6, and 24h were further evaluated. Expressions of all of the immune genes significantly decreased when prawns were exposed to 0.4mgL−1 trichlorfon for 24h, and among them, an increase in SOD expression was seen after exposure to 0.4mgL−1 for 6h. Prawns exposed to trichlorfon within 24h exhibited the decrease of circulating hemocytes, and also the induction of oxidative stress, which caused subsequent damage to DNA formation of immune genes. From these results, we concluded that immunological responses and immune gene expressions of prawn exposed to trichlorfon at 0.4mgL−1 for 24h were perturbed, thus causing a deficiency in immunity and subsequent increased susceptibility to pathogen infections.