Abstract: Objective To isolate Escherichia coli-specific bacteriophages from hospital waste water, analyze their biological characteristics and detect their cleavage effcts on bacterial biofilms.Methods The phages were isolated and purified by double-layer agar plate method and their morphology was observed by transmission electron microscopy. Their one-step growth curve, in vitro lysis curve, tem- perature and acid-base stability were determined. Whole genome sequencing was used to analyze the genome composition and characteristics of bacteriophages. The crystal violet staining was used to evaluate the scavenging effects of phage on host biofilms. Results A novel Escherichia coli-specific bacteriophage named vB_ EcoP-R1 was isolated from hospital sewage. Transmission electron microscopy revealed that the morphology of vB_ EcoP-R1 was similar to that of Podoviridae, a member of Caudovirales, and it could survive stably in the temperature range of -20 C to50 9C and pH 5.0 to 11.0. The total length of the vB_ EcoP-R1 genome was 40 875 bp and the GC content was 49.94%. There were no virulence genes and drug resistance genes in the phage genome. vB_ EcoP-R1 was capable of lysing clinically isolated Escherichia coli and showed high species and genus specificity. After the phage acted on the host bacteria preformned biofilm for 4 h, the clearance rate of the biofilm reached 80. 16%. Conclusion vB_ EcoP-R1 should be a novel Escherichia coli-specif-ic bacteriophage with strong lysis and high specificity for species and genus. The bacteriophage could lyse the biofilm formed by the host bacteria, and is expected to be used as an alternative treatment for Escherichia coli infection.