Carbon Metabolism Characteristics of Rhizosphere Soil Microbial Communities in Different-Aged Alfalfa (Medicago sativa L.) and Their Covarying Soil Factors in the Semi-Arid Loess Plateau

The carbon metabolism activity of rhizosphere soil microbial communities is an essential indicator for assessing soil ecosystem health, as it directly affects soil nutrient cycling and the stability of organic matter. However, there is a limited understanding of the carbon metabolism characteristics of rhizosphere soil microorganisms in alfalfa (Medicago sativa L.) of different ages and their relationships with soil physicochemical properties. This study used Biolog EcoPlates to evaluate the carbon metabolism activity, functional diversity, and carbon-source utilization preferences of rhizosphere soil microbial communities in 5-, 7-, and 9-year-old alfalfa grasslands on the semi-arid Loess Plateau of western China. We analyzed the relationships between soil physicochemical properties and microbial carbon metabolism characteristics, considering their potential covariation. The results showed that, with the extension of alfalfa planting years, the rhizosphere soil water content decreased significantly, pH decreased slightly, but soil organic carbon, total nitrogen, and total phosphorus contents increased significantly. The rhizosphere soil microbial community of 9-year-old alfalfa exhibited the highest carbon metabolism activity, Shannon diversity index, and carbon-source utilization. Rhizosphere soil microorganisms from different-aged alfalfa showed significantly different preferences for carbon-source utilization, with microorganisms from 9-year-old alfalfa preferentially utilizing carbon sources such as N-acetyl-D-glucosamine, D-mannitol, and D-cellobiose. Redundancy analysis revealed that soil water content was among the most important factors influencing the carbon metabolism activity of rhizosphere soil microbial communities while acknowledging that the relative contributions of soil water content, organic carbon, and nitrogen require careful interpretation, owing to their potential collinearity. This study demonstrates that, under rain-fed conditions in the semi-arid Loess Plateau, the continuous cultivation of alfalfa for nine years led to a significant decrease in soil water content but enhanced the rhizosphere soil nutrient status and microbial carbon metabolism activity, with no apparent signs of microbial functional degradation, although soil water depletion was observed. These findings highlight the complex interactions among multiple soil factors in influencing microbial carbon metabolism, providing valuable microbiological insights for understanding the sustainability of alfalfa grasslands and a theoretical basis for the scientific management of alfalfa grasslands in the semi-arid Loess Plateau region. Future research should consider longer planting periods to determine the critical age of alfalfa grassland degradation under semi-arid conditions and its associated microbial mechanisms.