[Background] Phage lysin has good safety and antibacterial properties, and is expected to replace antibiotics as a feed additive to improve the intestinal flora of livestock and poultry and prevent animal diseases. [Objective] White feather broilers were fed with complex phage lyase and antibiotic preparations to evaluate the effect of phage lysin on the intestinal flora of livestock and poultry and its application potential as an alternative feed additive. [Methods] Three-day-old healthy white feather broilers were fed, and the basic diet was supplemented with (200 mg/kg) complex phage lysin prepared from Thermus bacteriophage lysin (TSPphg) and Subthermus bacteriophage lysin (MMPphg) the lysin preparation and the 50 μg/g chlortetracycline preparation were respectively compared for feeding management. The high-throughput sequencing on the intestinal flora of white feather broilers at 28 days using the third-generation high-throughput sequencing technology, With the help of bioinformatics analysis of intestinal flora diversity and the relative abundance of dominant flora (top ten relative abundance), reveal the differences between and within each group, and further compare the liver oxidase activities. [Results] Complex phage lysin and chlortetracycline preparation had a certain effect on the intestinal flora of white feather broilers, both of which decreased the abundance and diversity of intestinal flora, and the Shannon index was significantly different compared with the blank group (P<0.05). In the dominant intestinal bacteria, Bacteroidetes significantly increased and Firmicutes significantly decreased in the complex phage lysin group compared with chlortetracycline group and blank group (P<0.05). At the same time, at the genus level, the relative abundance of Faecalibacterium increased significantly, while the relative abundance of Helicobacter decreased significantly (P<0.05), this trend was similar to that of most alternative antibiotic products after feeding poultry intestinal flora, which was more conducive to the health of poultry intestines. In addition, compared with chlortetracycline group and blank group, the activities of catalase (CAT) and superoxide dismutase (SOD) in complex phage lysin group were significantly increased go up (P<0.05). [Conclusion] Phage lysin can effectively improve the intestinal flora of livestock and poultry, promote the micro-ecological health of the intestinal livestock and poultry and improve the immunity of the body. It has a good prospect to replace antibiotics with phage lysin in livestock and poultry breeding and is worth further study.