[Objectives] Crested Ibis Nipponia nippon is protected at the national level in China and is also one of the most critically endangered bird species globally. Unfortunately, the population of wild Crested Ibises has experienced a drastic decline due to environmental degradation. Research indicates that the gut microorganisms of Crested Ibises play vital roles in nutrient absorption, immune regulation and healthy growth. The objective of this study was to culturally analyze the intestinal microbiota of Crested Ibis nestlings for beneficial microorganisms. In this study, the gut microbial diversity and extracellular enzyme activity of Crested Ibises were examined to further understand their healthy development. [Methods] The intestinal microorganisms in Crested Ibis nestlings were obtained through pure culture methodology. The identification of the bacteria involved a combination of Gram staining, physiological and biochemical analysis, and 16S rRNA gene amplification and sequence analysis. A phylogenetic tree was subsequently constructed using 16S rRNA gene sequences of intestinal bacteria derived from Crested Ibis nestlings, including representative isolates and related species. Strains that produced amylase, protease, cellulase, or lipase were screened through hydrolytic cycle degradation. [Results] A total of 254 strains of bacteria were isolated from the fresh excreta of artificially-reared Crested Ibis nestlings, comprising 2 phyla and 9 genera (Table 2). Thirty-three gram- positive strains and 221 strains of gram-negative bacteria were identified (Fig. 1). Among these strains, 211 were identified as Proteobacteria, accounting for 83.07% of the total isolates. Among them, 179 were Escherichia sp., representing 70.47% of the total number of isolates, additionally, 18 strains of Citrobacter sp. (7.09%), 14 strains of Enterobacter sp. (5.51%), and 14 strains of Klebsiella sp. (5.51%) were also isolated. Of the total number of isolates, 7.09% were identified as follows:fourteen strains of Enterobacter sp. (5.51%), 4 strains of Klebsiella sp. (1.57%), 4 strains of Shigella sp. (1.57%), two strains of Aeromonas sp. (0.79%), and another 2 strains of Aeromonas sp. (0.79%). Forty-three strains, accounting for 16.93% of all the isolates, belonged to Firmicutes, in which 27 strains (10.63%) belonged to Enterococcus sp., 4 strains belonged to Lysinibacillus sp. (1.57%), and Bacillus sp. had 4 strains (1.57%). Physiological and biochemical identification revealed that the traits of each strain were in accordance with their respective 16S rRNA genes (Table 1). The differences between enzyme-producing strains were evaluated (Fig. 4), and the types and efficacy of extracellular enzyme production in these cultivable bacteria were analyzed (Fig. 5). The results revealed that 244 strains produced protease, 19 strains produced lipase, 10 strains produced amylase, and 4 strains produced cellulose (Fig. 3). [Conclusion] The Crested Ibis conservation station aims to research intestinal microorganisms to prevent deaths resulting from intestinal diseases during the summer and fall seasons each year. We found that the bacteria isolated from the intestines of Crested Ibis nestlings were classified into nine genera belonging to two phyla. Escherichia, a member of the phylum Proteobacteria, was the most abundant genus, accounting for 70.47% of the isolates, and 86.22% of the strains produced proteases. The findings can offer technical insights into the breeding of Crested Ibises.