Chlorpyrifos ethyl is a popular insecticide widely used in agriculture within the Vietnamese Mekong delta, including for rice farming. Here, local farmers often apply pesticides at very high rates which leads to contamination of the surrounding environment. Silver barb (Barbonymus gonionotus) is a fish species indigenous to the delta, which resides in a variety of water bodies and is also commonly cultivated in rice–fish systems. As a result, this species is at high risk of exposure to chlorpyrifos ethyl. This study aims to determine the lethal concentration (LC50) of chlorpyrifos ethyl, as well as the effects of sub-lethal concentrations on the activity of cholinesterase and growth of Silver barb. Lethal concentration testing was conducted in a static non-renewed system. Three concentrations of chlorpyrifos ethyl (1%, 10% and 20% LC50-96 h) were conducted in triplicate to assess the effects of chlorpyrifos ethyl on the brain cholinesterase (ChE) of fingerling fish for 15 days, and on their growth for 60 days. Results showed that chlorpyrifos ethyl was highly toxic to fingerling Silver barb with a LC50-96 h of 0.119 ppm. The lowest observed effect concentration (LOEC) was 1%LC50-96 h for ChE and 10%LC50-96 h for growth. No observed effect concentration (NOEC) of chlorpyrifos ethyl for growth was 1%LC50-96 h. The result from this study suggests that ChE activity is significantly inhibited at environmentally realistic concentrations in the Vietnamese Mekong delta and can be used as a biomarker of pesticide exposure. Further study in the rice fields as well as in the canals or rivers is required.