Abstract:[Background] Plant endophytes represent the treasure for investigating microbial diversity and adaptive evolution. Cistanche is a genus of medicinal plants belonging to Orobanchaceae and honored as “ginseng of the deserts”. However, few studies have been carried out on the endophytes of Cistanche. [Objective] To investigate the diversity of endophytic bacteria in different Cistanche species and explore the endophytic bacterial resources. [Methods] The plants of Cistanche tubulosa (Schrenk) R. Wight (GH) and C. deserticola Y.C. Ma (HM) were sampled and subjected to high-throughput sequencing. The conventional microbial isolation method was used to isolate endophytic bacteria, and the isolates were identified by 16S rRNA gene sequence alignments. Subsequently, the stress tolerance, plant growth-promoting effects, and antimicrobial activities of the strains were evaluated. [Results] The high-throughput sequencing revealed a total of 1 086 OTUs of endophytic bacteria in GH and HM samples, among which 46 (4.24%) common OTUs were shared by the two species. The OTUs belonged to 226 genera, 104 orders of 19 phyla and some uncategorized bacteria. Proteobacteria was the dominant phylum in both samples, accounting for 69.46%. Significant differences were observed in the community structure of endophytic bacteria between the two species. In addition, 65 strains of endophytic bacteria were isolated from the plants, belonging to 34 species, 20 genera of 3 phyla (Bacillota, Proteobacteria, and Actinomycetota), and Bacillota had the highest relative abundance. Furthermore, the stress tolerance, plant growth-promoting effects, and antimicrobial activities of the isolates were evaluated. [Conclusion] The community structure of endophytic bacteria presented significant differences between Cistanche species. This study isolated several endophytic bacterial strains, which provided a basis for subsequent applications.