Polychaete are diverse species of the soft-bottom community, and are often used as indicators in environment monitoring programs. However, the effects of anthropogenic activities and natural environmental variation on polychaete assemblage are rarely addressed. The goals of this study are to identify the effects of natural environmental variation and anthropogenic stress on polychaete assemblage, and to explore the relationship between the polychaete assemblage structure and anthropogenic stress without considering the natural environmental variation. Based on the data collected from the surveys conducted in the tidal flat of Jiaozhou Bay, the relationship between polychaete assemblage structure and environmental variables was determined using multivariate statistical methods including hierarchical cluster analysis, multidimensional scaling(MDS) and canonical correspondence analysis(CCA). The results showed that the polychaete assemblage was dominated by two species, Amphictene japonica and Heteromastus filiformis, and could be divided into two subgroups characterized by high and low species abundance. CCA illustrated that the natural environmental variables including water temperature and the distance from coast had primary effects on the polychaete assemblage structure; while stress of contaminants, such as As and Hg, had the secondary influences; and stress from the aquacultured species, mainly Ruditapes philippinarum, had a limited effect. Colinearity between the natural environmental variables and anthropogenic stress variables caused a critical divergence in the interpretation of CCA results, which highlighted the risk of a lack of information in environment assessment. Glycinde gurjanovae, Sternaspis scutata and Eulalia bilineata may serve as the ‘contamination indicators’, which need to be confirmed in future studies. Polychaete are diverse species of the soft-bottom community, and are often used as indicators in environment monitoring programs. However, the effects of anthropogenic activities and natural environmental variation on polychaete assemblage are rarely addressed. The goals of this study are to identify the effects of natural environmental variation and anthropogenic stress on polychaete assemblage, and to explore the relationship between the polychaete assemblage structure and anthropogenic stress without considering the natural environmental variation. Based on the data collected from the surveys conducted in the tidal flat of Jiaozhou Bay, the relationship between polychaete assemblage structure and environmental variables was determined using multivariate statistical methods including hierarchical cluster analysis, multidimensional scaling (MDS) and canonical correspondence analysis (CCA). The results showed that the polychaete assemblage was dominated by two species, Amphictene japonic a and Heteromastus filiformis, and could be divided into two subgroups characterized by high and low species abundance. CCA illustrated that the natural environmental variables including water temperature and the distance from coast had primary effects on the polychaete assemblage structure; while stress of contaminants, such as As and Hg, had the secondary influences; and stress from the aquacultured species, mainly Ruditapes philippinarum, had a limited effect. Colinearity between the natural environmental variables and anthropogenic stress variables caused a critical divergence in the interpretation of CCA results, which highlighted the risk of a lack of information in environment assessment. Glycinde gurjanovae, Sternaspis scutata and Eulalia bilineata may serve as the ’contamination indicators’, which need to be confirmed in future studies.