Antibiotic resistance genes (ARGs) are emerging contaminants that pose a potential risk to human health worldwide. In this study, a metagenomic analysis was performed to investigate the distribution of ARGs in paddy field ecosystems, crayfish monoculture pond ecosystems, and rice-crayfish cultivation field ecosystems. The results showed that MacB and BcrA are two dominant ARGs, and macrolide is the dominant antibiotic not only in the water, but also in the sediment and gut of crayfish, in both the crayfish monoculture and the rice-crayfish cultivation ecosystems. Meanwhile, some lower-abundance ARGs in the water and sediment of crayfish cultivation ecosystems were significantly different in their abundance than those in rice paddy fields, and the ARGs in crayfish culture system and rice paddy fields showed even higher dissimilarities of diversity. Comprehensive analysis with redundancy analysis(RDA)and the distribution of dominant ARGs showed that the dissimilarity was related to the higher concentrations of total nitrogen (TN), total phosphorus (TP), chlorophyll a (Chla), permanganate index (CODMn), and nitrate in the water of rice paddy fields, and was related to the higher contents of N, P, K, and organic matter (OM) in the sediment of rice paddy fields. The source trackers of ARGs within the crayfish cultivation ponds and the rice-crayfish cultivation fields showed that the sediment in crayfish cultivation ponds mainly played the role of the ‘sink’ for ARGs, and the water and gut of crayfish mutually contributed to the high rates of ARGs. The ARG contribution rates of crayfish gut and sediment decreased and increased, respectively, in rice-crayfish cultivation fields and in crayfish monoculture ponds, which might be related to the lower crayfish biomass and the lower water depth in rice-crayfish cultivation fields.