Fig. 5: Evaluation of the roles of Ammoniphilus sp. in the promotion of root apoplastic Fe reutilization.

a Neighbor-joining tree showed the phylogenetic relationship among different Ammoniphilus isolates obtained in this study. The numbers in parentheses indicated the sequence similarity of each isolate with OTU5. Accession numbers of isolates obtained from the NCBI database were shown in regular letters (in bold). Five isolates including AS12, AS15, AS27, AS63, and AS72 that were highlighted with red arrows were selected for the gnotobiotic and hydroponic experiments. b Living or heat-killed bacterial strains (10⁹ CFU mL⁻¹) were streaked onto tryptic soy agar (TSA) plates and co-incubated with 2-old-day seedlings at 28 °C in dark for 24 h. These seedlings were then transferred to gnotobiotic systems and treated with bacterial suspensions at a final concentration of 5 × 107 CFU g⁻¹ soil. After 10 days of culture, c plant phenotypes, d total chlorophyll content, e growth rescue activity, f shoot and g root soluble Fe, and h root apoplastic Fe content were measured in these plants (n = 15 plants per group). i Two-week-old tomato plants were transferred to hydroponic systems and exposed to -Fe (0 μM Fe) treatment with the presence of living or heat-killed bacterial strains at a final concentration of 5 × 107 CFU mL⁻¹. j Representative images of plant phenotype, k total chlorophyll content, and l shoot soluble content were determined after 12 days of treatments. The data were analyzed by one-way ANOVA with Tukey’s post hoc test at p < 0.05 (n = 15 plants per group; *p < 0.05, **p < 0.01). Different lowercase letters indicated significant differences in the growth rescue activity among different treatments using Duncan’s multiple range tests at p < 0.05 (n = 15). Error bars indicate standard deviation of the mean.