Association Analysis of the Genomic and Functional Characteristics of Halotolerant Glutamicibacter endophyticus J2-5-19 from the Rhizosphere of Suaeda salsa

Abstract

Halotolerant plant growth-promoting bacteria (HT-PGPB) have attracted considerable attention for their significant potential in mitigating salt stress in crops. However, the current exploration and development of HT-PGPB remain insufficient to meet the increasing demands of agriculture. In this study, an HT-PGPB isolated from coastal saline-alkali soil in the Yellow River Delta was identified as Glutamicibacter endophyticus J2-5-19. The strain was capable of growing in media with up to 13% NaCl and producing proteases, siderophores, and the plant hormone IAA. Under 4‱ salt stress, inoculation with strain J2-5-19 significantly increased the wheat seed germination rate from 37.5% to 95%, enhanced the dry weight of maize seedlings by 41.92%, and notably improved the development of maize root systems. Moreover, this work presented the first whole-genome of Glutamicibacter endophyticus, revealing that G. endophyticus J2-5-19 resisted salt stress by expelling sodium ions and taking up potassium ions through Na⁺/H⁺ antiporters and potassium uptake proteins, while also accumulating compatible solutes such as betaine, proline, and trehalose. Additionally, the genome contained multiple key plant growth-promoting genes, including those involved in IAA biosynthesis, siderophore production, and GABA synthesis. The findings provide a theoretical foundation and microbial resources for the development of specialized microbial inoculants for saline-alkali soils.

Keywords: Genomic analysis; Glutamicibacter endophyticus; Halotolerant plant growth-promoting bacteria; saline-alkali soil.

Figure 1

(A) Colony morphology of strain J2-5-19 grown on an LB agar plate. (B) Gram-staining of strain J2-5-19 observed at 1000× magnification; scale bar = 10 μm. (C) Cellular morphology of strain J2-5-19 observed by SEM at 25,000× magnification; scale bar = 300 nm.

Figure 2

Neighbor-joining phylogenetic tree based on 16S rRNA gene sequences of strain J2-5-19 and representative Glutamicibacter species. Bootstrap values at each branch node indicate the percentage of support based on 1000 resampling replicates. Strain J2-5-19 clusters with G. endophyticus EGI 6500322T within the same clade. The scale bar represents 0.002 nucleotide substitutions per site.

Figure 3

Genomic circle map of G. endophyticus J2-5-19. From inside to outside, the first circle represents the scale, the second circle represents GC skew, the third circle represents GC content, the fourth and seventh circles represent the COG to which each coding sequence (CDS) belongs, and the fifth and sixth circles represent the location of CDS, transfer RNA, and ribosomal RNA on the genome.

Figure 4

GBDP tree inferred with FastME 2.1.6.1 from GBDP distances calculated from genome sequences. The branch lengths are scaled in terms of GBDP distance formula d5. The numbers above branches are GBDP pseudo-bootstrap support values >60% from 100 replications, with an average branch support of 92.1%.

Figure 5

Growth curves of strain J2-5-19 under different NaCl concentrations. Data points represent the mean of three biological replicates (n = 3), with error bars indicating the standard deviation (SD) of the mean.

Figure 6

(A) Structure of the mrp operon (B) The predicted structure of the Mrp protein complex.

Figure 7

The biosynthetic mechanisms of several compatible solutes (A) Biosynthetic pathway of glycine betaine; (B) Biosynthetic pathways of glutamate, glutamine, and proline; (C) Biosynthetic pathway of trehalose; (D) Biosynthetic pathway of inositol; (E) Biosynthetic pathway of sorbitol.

Figure 8

(A) Growth of strain J2-5-19 on casein agar plates; (B) Growth of strain J2-5-19 on CAS agar plates; (C) IAA production by strain J2-5-19 in LB medium and LB supplemented with tryptophan (5 mM). Data are presented as mean ± SD (n = 5 for each treatment). Statistical significance was analyzed using independent sample t-tests. Error bars represent the standard deviation (SD) of the mean. Asterisks indicate statistical significance (****, p < 0.0001).

Figure 9

(A) Seed germination rates after treatment with different concentrations of bacterial suspensions under salt stress. Data are presented as median with interquartile range (n = 3 for each treatment). Statistical significance was determined using the Kruskal–Wallis H test with Bonferroni correction. An asterisk indicates statistical significance (*, p < 0.05). (B) Wheat seedlings treated with G. endophyticus J2-5-19 show improved growth compared to the control.

Figure 10

(A) Overall plant condition 30 days after treatment (CK: control); (B) Effects of strain J2-5-19 on the agronomic traits of maize. Statistical significance was analyzed using independent sample t-tests. Error bars represent the SD of the mean (n = 8 for each treatment). Asterisks indicate statistical significance (*, p < 0.05; **, p < 0.01; ***, p < 0.001).

Figure 11

Predicted siderophore biosynthetic gene cluster (BGC) in strain J2-5-19 (upper panel) compared with the desferrioxamine E biosynthetic gene cluster (BGC0001572) from Pantoea agglomerans (lower panel). Lines indicate homologous regions between the two clusters.

Figure 12

(A) IAA biosynthesis pathway involving tryptophan and its intermediates; (B) GABA biosynthesis from L-ornithine through putrescine and related intermediates; (C) Conversion of acetoin to (R,R)-butane-2,3-diol.