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ABSTRACT The widespread prevalence of saline environments poses a significant global environmental challenge. Salt stress, induced by saline soils, disrupts soil microecology and affects the plant-microbe-soil cycling process. Utilizing microbial fungicides stands as a primary strategy to mitigate salt stress-induced damage to plants and soils. This study investigated the influence of Bacillus subtilis (Bs) inoculation on the microbial community, assembly processes, and functional changes in bacteria and fungi in Glycyrrhiza uralensis Fisch. (licorice) seedlings under varying salt stress levels, primarily employing microbiomics techniques. Soil enzyme activities displayed a declining trend with increasing salt stress, which was mitigated by Bs inoculation. Microbiome analysis revealed a significant increase in bacterial and fungal operational taxonomic units, particularly in Ascomycetes and Nitrogen-fixing Bacteria, thereby enhancing soil denitrification. The abundance of Proteobacteria, Actinobacteriota, Bacteroidota, and Firmicutes in bacteria, as well as Ascomycota in fungi, increased with higher salt stress levels, a process facilitated by Bs inoculation. However, functional predictions indicated a reduction in the relative abundance of Dung Saprotrophs with Bs inoculation. Salt stress disrupted soil assembly processes, showcasing a continuous decline in diffusion limitation with increased salt concentration, where Bs inoculation reached a peak under moderate stress. In summary, this research elucidates the communication mechanism of Bs in enhancing salt tolerance in licorice from a microbiome perspective, contributing to a comprehensive understanding of abiotic and biotic factors.IMPORTANCELicorice is a herb that grows in deserts or saline soils. Enhancing the salt tolerance of licorice is necessary to maintain the quality of cultivated licorice and to ensure the supply of medicinal herbs. In the past, we have demonstrated the effectiveness of inoculation with Bacillus subtilis (Bs) to enhance the salt tolerance of licorice and revealed the key metabolic pathways for the development of salt tolerance through multi-omics. In this study, we used the microbiomics approach to reveal the plant-microbe-soil interactions at the level of inoculation of Bs affecting the dynamics of soil microbial communities from bacterial and fungal perspectives, thus bridging the interactions between biotic and abiotic factors.

WRKY is one of the largest transcription factor families in higher plants, playing a crucial role in plant growth, development, and responses to environmental stresses by binding to the W-box motif in the target gene promoters. Glycyrrhiza uralensis Fish. (G. uralensis), known for its extensive medicinal value and adaptation to saline-alkali environments, has garnered attention. However, information regarding the members and functions of the G. uralensis WRKY gene family is relatively limited. In this study, we identified 52 GuWRKY genes from the G. uralensis genome and comprehensively investigated their characteristics through phylogenetic analysis, examination of gene duplication events, analysis of gene structures, and conservation of protein motifs. The GuWRKY family was classified into seven groups based on phylogenetic analysis, and collinearity analysis revealed that many GuWRKY genes originated from segmental duplications, demonstrating a highly conserved domain. Promoter cis-element analysis indicated the presence of elements related to hormones, stress, and growth and development in GuWRKYs. Transcriptomic analysis revealed distinct expression patterns of GuWRKYs under salt stress, with GuWRKY31 identified as a potential key regulator in salt stress responses. Subcellular localization experiments validated some of the predicted results. The provided biological information lays a crucial foundation for further exploration of the functions of GuWRKY genes.

Introduction: Research on Glycyrrhiza uralensis, a nonhalophyte that thrives in saline–alkaline soil and a traditional Chinese medicinal component, is focused on improving its ability to tolerate salt stress to increase its productivity and preserve its “Dao-di” characteristics. Furthermore, the inoculation of bioagents such as Bacillus subtilis to increase plant responses to abiotic stressors is currently a mainstream strategy. Mitogen-activated protein kinase (MAPK), a highly conserved protein kinase, plays a significant role in plant responses to various abiotic stress pathways.Methods: This investigation involved the identification of 21 members of the GuMAPK family from the genome of G. uralensis, with an analysis of their protein conserved domains, gene structures, evolutionary relationships, and phosphorylation sites using bioinformatics tools.Results: Systematic evolutionary analysis of the 21 GuMAPKs classified them into four distinct subgroups, revealing significant differences in gene structure and exon numbers. Collinearity analysis highlighted the crucial role of segmental duplication in expanding the GuMAPK gene family, which is particularly evident in G. uralensis and shows a close phylogenetic relationship with Arabidopsis thaliana, tomato, and cucumber. Additionally, the identification of phosphorylation sites suggests a strong correlation between GuMAPK and various physiological processes, including hormonal responses, stress resistance, and growth and development. Protein interaction analysis further supported the role of GuMAPK proteins in regulating essential downstream genes. Through examination of transcriptome expression patterns, GuMAPK16-2 emerged as a prospective pivotal regulatory factor in the context of salt stress and B. subtilis inoculation, a finding supported by its subcellular localization within the nucleus.Discussion: These discoveries offer compelling evidence for the involvement of GuMAPK in the salt stress response and for the exploration of the mechanisms underlying B. subtilis’ enhancement of salt tolerance in G. uralensis.