SummaryDespite progress in understanding the role of nitric oxide (NO) in the pathogenesis of helminth infections, the role in strongyloidosis is unknown. Firstly, we studied the production of NO in mice infected with Strongyloides venezuelensis as well as in macrophage cultures stimulated with parasite antigens. Somatic larvae 3 (L3) and excretory–secretory female antigens stimulate specific NO production measured by Griess reaction and expression of inducible NO synthase by RT‐PCR and quantitative PCR. Moreover, mice infected with S. venezuelensis produce NO in migration stages. Secondly, we analysed the effect of NO production on L3 and females of S. venezuelensis using NO donors such as diethylenetriamine and 3,3‐bis(aminoethyl)‐1‐hydroxy‐2‐oxo‐1‐triazene. Parasites died after NO donor treatment in a dose‐dependent manner. Finally, apoptotic mechanisms are involved in the death of S. venezuelensis larvae.