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Since its discovery in 2015, the mobile colistin resistance gene mcr-1 has been reported in bacteria from > 50 countries. Although aquaculture-associated bacteria may act as a significant reservoir for colistin resistance, systematic investigations of mcr-1 in the aquaculture supply chain are scarce. We investigated the presence of colistin resistance determinants in the aquaculture supply chain in south China and determined their characteristics and relationships. A total of 250 samples were collected from a duck-fish integrated fishery, slaughter house, and market in Guangdong Province, China, in July 2017. Colistin-resistant bacteria were isolated on colistin-supplemented CHROMagar Orientation plates, and the species were identified by matrix-assisted laser desorption/ionization time-of-flight assay. The presence of mcr genes was confirmed by polymerase chain reaction analysis. We examined the minimum inhibitory concentrations (MICs) of 16 antimicrobial agents against the isolates using agar diffusion and broth microdilution methods. Whole-genome sequencing (WGS) was used to explore the molecular characteristics and relationships of mcr-1 -positive Escherichia coli (MCRPEC). Overall, 143 (57.2%) colistin-resistant bacteria were isolated, of which, 56 (22.4%, including 54 Escherichia coli and two Klebsiella pneumoniae) and four Aeromonas species were positive for mcr-1 and mcr-3 , respectively. The animal-derived MCRPEC were significantly more prevalent in integrated fishery samples (40.0%) than those in market (4.8%, P < 0.01) samples but not in slaughter house (28.0%, P = 0.164). All MCRPEC were highly resistant to ampicillin, tetracycline, and compound sulfamethoxazole (>90%) but were susceptible to carbapenems and tigecycline. WGS analysis suggested that mcr-1 was mainly contained on plasmids, including IncHI2 (29.6%), IncI2 (27.8%), IncX4 (14.8%), and IncP (11.1%). Genomic analysis suggested mcr-1 transmission via the aquatic food chain. MCRPEC were highly prevalent in the aquaculture supply chain, with the isolates showing resistance to most antibiotics. The data suggested mcr-1 could be transferred to humans via the aquatic food chain. Taking the "One Health" perspective, aquaculture should be incorporated into systematic surveillance programs with animal, human, and environmental monitoring. • We reported a high prevalence (21.6%) of MCRPEC in aquaculture supply chain. • All MCRPEC isolates were resistant to most antimicrobial agents, in particular, those officially used in aquaculture. • We provide the evidence that mcr-1 could be transferred from animals to humans via the aquaculture supply chain. [ABSTRACT FROM AUTHOR]