Abstract: Drought is one of the major abiotic stresses affecting plant growth and development, which severely restricts crop productivity. This study aimed to screen plant growth-promoting rhizobacteria (PGPB) with excellent growth-promoting and drought-resistant traits from the rhizosphere soil of Onobrychis viciifolia (sainfoin) in different drought habitats, and to evaluate their regulatory effects on the growth, development and physiological responses of Medicago sativa L. seedlings under drought stress. A total of 903 culturable bacterial strains belonging to 82 species were isolated via multi-medium cultivation combined with 16S rRNA gene sequencing identification. These strains were predominantly affiliated to the phyla Proteobacteria, Actinobacteria and Bacteroidetes, with the genera Serratia and Pseudomonas as the dominant ones. Analysis of growth-promoting traits revealed that the 82 rhizobacterial strains exhibited varying degrees of growth-promoting properties. Specifically, strains OvR165 (Enterobacter sp.), OvR548 (Acinetobacter sp.) and OvR687 (Pseudomonas sp.) possessed the capabilities of phosphate solubilization, nitrogen fixation, siderophore production, indole-3-acetic acid (IAA) secretion and extracellular polysaccharide (EPS) synthesis. Among them, OvR548 and OvR687 also exhibited strong biofilm formation abilities. Pot experiments demonstrated that inoculation with these three strains significantly alleviated the inhibitory effects of drought on Medicago sativa seedlings: it promoted the accumulation of plant height, root length, leaf area and biomass, increased chlorophyll content and relative water content, reduced cell membrane damage and reactive oxygen species (ROS) accumulation, and effectively mitigated drought stress-induced damage to plants. Additionally, these strains showed a strong potential for improving plant tolerance to drought, further enhancing the ecological adaptability of crops under water scarcity conditions. In conclusion, the rhizosphere of Onobrychis viciifolia in different arid regions of Qinghai Province harbors abundant and functionally diverse resources of growth-promoting and drought-tolerant bacteria, which have great ecological value and application prospects. This study provides excellent strain resources and theoretical support for sustainable grassland management and microbial agent development in arid regions, helping to improve the productivity and resilience of crops in drought-prone areas.