Microbial inoculants derived from plant growth-promoting rhizobacteria (PGPR) offer eco-friendly alternatives to traditional chemical fertilizers, maintaining microbiota balance in agricultural systems. However, limited research has explored the combined effects of microbial inoculants and chemical fertilizers on crop growth and soil properties. In this study, we investigated seven fertilizer combinations, ranging from no fertilizer to various proportions of chemical fertilizers with microbial inoculants, on timothy (Phleum pratense L.) growth, chlorophyll content, soil properties, enzyme activities, and soil microbial communities. A randomized block design was employed to analyze these effects. The results indicate that the combination of 85% chemical fertilizer with microbial inoculants significantly increased timothy yield and chlorophyll content. In addition, a reduction to 55% chemical fertilizer in conjunction with microbial inoculants resulted in comparable yield to that of 100% fertilizer with no inoculants. The microbial inoculants treatments notably elevated soil catalase, urease, acid phosphatase, and invertase activities, along with soil fast-acting nutrient content. The sequencing results show that the abundance of beneficial bacteria increased, while that of fungi decreased in the soil rhizosphere after the application of microbial inoculants. This study underscored the potential of microbial inoculants combined with reductions in chemical fertilizers to enhance soil microbiology, nutrient content, and beneficial microbial abundance while suppressing pathogenic fungi, thereby promoting timothy growth and yield. These findings provide a theoretical basis for the use of microbial inoculants in sustainable agricultural practices, providing valuable insights for optimizing microbial inoculants and chemical fertilizer formulations to mitigate the sustainability challenges posed by conventional fertilizers.