Optimization of compound bacterial inoculant enhances soil nutrients and bacterial community richness in alpine grassland.

Grassland degradation results in nutrient loss and ecosystem function impairment, making it crucial to explore green and effective restoration technologies. In this study, three strains (Acinetobacter calcoaceticus J1, Pseudomonas piscium J5, Bacillus subtilis Y3) isolated from alpine grassland, and mixed in equal volume (VJ1:VJ5:VY3 = 1:1:1) to prepare the compound bacteria. The fermentation medium and conditions of the compound bacteria were optimized, then pot and field experiments were conducted to validate the application of the optimized compound bacterial inoculant. The optimal conditions were: soluble starch 2.62 g L-1, yeast extract 20.26 g L-1, KCl 5.01 g L-1, pH 6.0, incubated at 25 °C for 48 h, with 20 % liquid volume, 3 % inoculation ratio, and 200 r·min-1. Under these conditions, the number of viable bacteria was 128.47 times higher than before optimization. The obtained compound bacterial inoculant resulted in a significant increase of 34.24 %, 52.61 %, 57.74 %, 62.50 %, and 53.92 % in the height, total root length, root surface area, volume, and branch number of Elymus nutans, with better effects than the single bacterial. The field experiment revealed that the compound bacterial inoculant could significantly increase above-ground biomass (10.63 %), organic matter (16.99 %), available nitrogen (12.58 %), total nitrogen, and phosphorus (15.79 % and 11.54 %). This study has developed an environmentally friendly compound bacterial inoculant (G1), that accelerates soil nutrient cycling by reducing soil pH and increasing the richness of the bacterial community, thus restoring degraded grassland. This result realized the principle of "near-natural restoration" of degraded alpine grassland and provided a new biological restoration technology for the degraded alpine grassland.