An electrochemical biosensor was constructed for quantitatively detecting Skeletonema costatum (S. costatum) by combining the electrode modification material (NC-Au) with a strand displacement reaction (SDR). The SDR process addresses the issues of steric hindrance and electrostatic repulsion resulting from the large size of genomic DNA. It enhances the efficiency of the interfacial hybridization reaction and endows the biosensor with remarkable sensitivity. The limit of detection (LOD) and limit of quantification (LOQ) were 33.43 fg/μL (831 cells/L) and 87.21 fg/μL (2112 cells/L), respectively. Additionally, the biosensor has demonstrated excellent accuracy compared to methods such as microscopy and ddPCR (P >
0.05). Subsequent assessment of the adjacent waters of the Beibu Gulf using biosensors indicated a low risk of S. costatum red tide outbreaks in the region, which is consistent with the findings of the ecological survey. Therefore, we believe that this biosensor can provide a completely new idea for the dynamic monitoring and early warning of S. costatum red tide.