Abstract: Amphibians play a pivotal role in the food chain and ecosystem. Effective assessment and monitoring of amphibian diversity are crucial steps in biodiversity conservation. In May 2023, 10 investigation sites were selected in each of the four habitats around Chaohu Lake: ponds, rivers, croplands and wetlands. This study used environmental DNA technology to explore the diversity of amphibians and analyze the impact of environmental factors on the distribution pattern and the habitat preferences of amphibians. The results revealed that: (1) A total of 7 amphibians belonging to 4 families and 5 genera were detected at 40 sites in Chaohu Lake. Fejervarya multistriata was the dominant species in terms of dominance index and relative abundance. (2) Significant differences were observed in community structure and dominant species in different habitats. Ponds and wetlands showed greater dissimilarity in community structure compared to rivers and cropland, and the species richness in river habitats was lower. (3) Fejervarya multistriata was dominant in ponds, farmlands, and wetland habitats, while Rana zhenhaiensis preferred pond habitats. (4) Generalized Additive Models (GAMs) showed that species richness was significantly negatively correlated with altitude and water depth. Redundancy analysis (RDA) showed that water width, temperature, and pH also significantly influenced the distribution patterns of amphibians. The study showed that there were differences in community structure and dominant species among ponds, rivers, croplands and wetlands habitats in Chaohu Lake, and that the diversity and distribution pattern of amphibians were affected by a variety of environmental factors, proving that environmental DNA can be an effective method for investigating amphibians in Chaohu Lake.