Robinia pseudoacacia is widely planted on the Loess Plateau as a strong drought-tolerant and salt-tolerant species for vegetation restoration. However, this mode of pure plantation has triggered great concern over the soil ecosystem. The aim of this study was to explore the effects of the plantation on soil physiochemical properties, soil microorganisms, and the relationship between them in Robinia pseudoacacia plantations of different ages. Four different ages of Robinia pseudoacacia stands, including 10-year-old, 15-year-old, 25-year-old, and 40-year-old (abbreviated as Y10, Y15, Y25, and Y40, respectively) were selected, and 20 soil physicochemical and biological indicators were determined. The variation in soil microbial biomass was influenced by sampling depth, and consistent with the variations in TN (soil total nitrogen) and SOC (soil organic carbon) during 25 years’ artificial forestation. Soil moisture increased significantly at Y15 and then decreased at Y40 but other soil properties remained relatively stable. The contents of phosphor lipid fatty acid (PLFA) of different microbial groups followed the order of B (Bacteria) > G⁻ (Gram-negative) > G⁺ (Gram-positive) > A (Actinomycetes) > F (Fungi). The ratios of F/B (Fungi to Bacteria) and Sat/Mono (Saturated PLFAs to Monosaturated PLFAs) of different ages of plantations showed a similar trend, i.e., declined first, then rose, and declined again. The ratios of Cy/Pre (Cyclopropyl PLFAs to Precursor PLFAs) and G⁺/G⁻ (Gram-positive to Gram-negative) of the soil of all ages of plantations showed a trend of slow growth and a trend of rapid growth, respectively. Redundancy analysis showed that the contents of individual PLFAs and total PLFA were positively correlated with SOC and TN, but variations of soil PLFA ratios mostly depended on other soil properties. After artificial forestation, the ratios of F/B and Sat/Mono were lower than before forestation, while the ratio of Cy/Pre varied with different soil layers. The ratio of G⁺/G⁻ increased with the increase in afforestation time, peaking at the 25th year. The contents of individual PLFAs and total PLFA may be sensitive indicators of SOC and TN within 25 years’ plantation. Lower ratio of F/B and higher G⁺/G⁻ suggest that the sustainability of the ecosystem is weaker and the fertility of the soil is lower after plantation of Robinia pseudoacacia.

Phospholipid fatty acids (PLFAs) can be used as biomarkers for qualitative and quantitative analyses of soil microbial community diversity. In this study, we collected soil samples at 10-cm intervals to a depth of 1 m from Robinia pseudoacacia plantations of four different ages (10, 15, 25 and 40 years) in a loess area and analysed the soil microbial community structure by PLFA analysis. A total of 97 PLFAs were detected in soils of R. pseudoacacia plantations of different ages. The individual PLFA contents gradually decreased in the 0- to 40-cm soil layers, with little variation in the 40- to 100-cm soil layers. The individual PLFAs were similarly distributed in the soils of R. pseudoacacia plantations of different ages, and there was a clear variation with stand age and soil depth. The individual PLFA contents in the 0- to 20-cm soil layers were highest for the 25-year-old plantation, while those in the 20- to 40-cm soil layers were relatively high for the 25- and 40-year-old plantations; the 16:0 content was the highest among individual PLFAs. The total PLFA content and the PLFA contents of different microbial groups [bacteria, fungi, Gram-positive bacteria (G⁺), Gram-negative bacteria (G^-) and actinomycetes] initially increased before decreasing in the soils of R. pseudoacacia plantations with increasing stand age, whereas these contents gradually decreased with increasing soil depth; the highest PLFA contents was found in the 25-year-old plantation. The total PLFA content and the contents of fungal, G^- and actinomycete PLFAs in the soils of R. pseudoacacia plantations differed significantly among stands of different ages in the 0- to 10-cm, 10- to 20-cm and 30- to 40-cm soil layers, while no significant differences were found in the 20- to 30-cm soil layers; the G⁺ and bacterial PLFAs contents in each of the 0- to 40-cm soil layers were significantly different. The PLFA ratios between different microbial groups differed among the stands of different ages. The fungi/bacteria ratio showed a “decrease-increase-decrease” trend with stand age, while the G⁺/G^- ratio showed an “increase-decrease” trend. The saturated/monounsaturated PLFA ratio initially decreased before plateauing, while the opposite trend was observed for the cyclopropyl/precursor ratio. The PLFA contents of different microbial groups were ranked as follows: bacteria > G^- > G⁺ > actinomycetes > fungi. In the principle component analysis, 18:1ω9c, 10Me18:0, i17:0, a17:0, 18:1ω7c, 18:1ω5c and 18:0 made the greatest contribution to principal component 1, and a14:0, i14:0 3OH, i14:0, i14:1ω7c and 14:0 made the greatest contribution to principal component 2. In conclusion, soil nutrient status and other soil eco-environmental stress factors should be considered in 10- to 25-year-old (particularly ~15-year-old) plots for the management of R. pseudoacacia plantations to prevent forest soil degradation and improve forest stand quality, thereby achieving better soil and water conservation and environmental improvement in R. pseudoacacia plantations.