Nitrogen-Fixing Bacteria Promote the Growth of P. Y. Li by Regulating Physiological and Biochemical Reactions and Protecting Enzyme System-Related Gene Expression.

The widespread application of chemical fertilizers and pesticides has resulted in environmental pollution. With the growing emphasis on ecological agriculture in traditional Chinese medicine, microbial fertilizers are increasingly recognized for their potential. The aim of this study is to investigate the effect of inoculating nitrogen-fixing bacteria on the soil (yellow loam, river sand, and organic fertilizer in a 2:1:1 ratio) of , with a focus on the leaf changes in terms of physiological parameters, antioxidant enzyme activity, and corresponding gene expression levels. The experiment involved three nitrogen-fixing bacteria, namely , , and , with a total of eight treatment groups. The objective was to assess how these bacterial treatments influenced physiological parameters, photosynthetic characteristics, pigment content, and both antioxidant enzyme activities and gene expression in the leaves of . The experimental results demonstrated statistically significant reductions ( < 0.05) in malondialdehyde (MDA) content and stomatal limitation value (LS) in leaves under treatment conditions relative to the control group (CK). The most substantial decreases were observed dual-inoculation with and (N5), showing reductions of 38.24% and 20.94% in MDA and LS compared to CK values. Additionally, leaf area, leaf thickness, stem thickness, plant height, photosynthetic parameters, pigment content, soluble sugars, soluble proteins, proline levels, and the activities of superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) exhibited varying degrees of increase. Compared to the CK group, the SOD, POD, and CAT activities of the N5 group increased by 141.06%, 160.59%, and 106.23%, respectively. The relative gene expression patterns of SOD, POD, and CAT corresponded with the trends observed in their respective antioxidant enzyme activities. Pearson correlation analysis further demonstrated that leaf area and net photosynthetic rate (Pn) were significantly correlated with respect to SOD, POD, and CAT activities, as well as their corresponding gene expression levels. In conclusion, inoculation with nitrogen-fixing bacteria improved the growth and stress tolerance of , with the combined application of and yielding the most effective results. This study establishes that different rhizosphere nitrogen-fixing bacteria, either individually or in combination, influence the photosynthetic characteristics, physiological and biochemical parameters, and protective enzyme systems of . These findings provide a theoretical foundation for the selection of nitrogen-fixing bacteria as biofertilizers in the artificial cultivation of and highlight their potential application in the cultivation of traditional Chinese medicinal materials.