Currently, the riverine ecosystem is one of the most threatened habitats and become a research hotspot due to the declining diversity. This loss is mainly attributed to human influence, such as overexploitation, habitat destruction, invasion of exotic species and aquatic pollution, as well as the dam’s construction, which have had drastic impact on the migration route of diadromous fish (Hilsha, salmon, freshwater prawn). Recently, the analysis of environmental DNA (eDNA) has been revolutionizing biodiversity monitoring and can used for the detection of hidden organismic diversity, and alien species at a large scale without a priori knowledge of the composition of species assemblages in riverine environment. eDNA has proven to be a valuable tool for characterizing fish species due to its cost-effectiveness and non-invasiveness, making it eco-friendly approach. eDNA is defined as total DNA obtained from environmental samples such as water, sediment, soil, or air, subsuming DNA from various sources such as unicellular or small multicellular taxa or tissue particles and gametes of multicellular organisms. In this article, we highlight an innovative eDNA approach and its diverse applications. Top of FormAdditionally, we provide basic guidelines for using eDNA to prevent contamination.