Table 1.
Viability of P. agglomerans ISIB55 and B. caribensis ISIB40 during the immobilization process in nanofibers produced by the electrospinning technique.
Fig 1.
Beneficial properties of P. agglomerans ISIB55 (ISIB55) and B. caribensis ISIB40 (ISIB40).
(A) Indole-3-acetic acid (IAA) production from nanofiber-immobilized cultures (nanofiber) and fresh cultures (culture) of ISIB55 and ISIB40. The data express the mean values ± standard error of IAA concentration expressed as μg·ml-1. (B) Phosphate solubilization from nanofiber and culture of ISIB55 and ISIB40. The data express the mean values ± standard error of phosphate solubilization index (PSI). The images below the bars show clear halos around the ISIB55 and ISIB40 colonies, indicating solubilizing capacity of phosphate in the different conditions evaluated. (C) Siderosphore synthesis from nanofiber and culture of ISIB55 and ISIB40. The data express the mean values ± standard error of halo diameters. The images below the bars show the color change from blue to orange that appeared around the colonies producing siderophores in the CAS-blue agar medium. Different letters in the graphs indicate statistically significant differences (P < 0.05) in IAA production, phosphate solubilization or siderosphore synthesis between the different conditions assayed for each bacterial strain, in accordance with Tukey's test. (D) Nitrogen fixation from nanofiber and culture of ISIB55 and ISIB40. The imagen shows the color change in the NFb semisolid medium when the bacteria fixed the nitrogen. A. brasilense Az39 and E. coli BW25113 were used as positive and negative controls, respectively.
Fig 2.
Viability of P. agglomerans ISIB 55 (ISIB55) and B. caribensis ISIB40 (ISIB40) in soybean seeds (TEC 5936 IPRO and RSF 7166 IPRO varieties).
(A) Seeds treated with (■) nanofiber-immobilized ISBI55 or (●) ISBI55 culture; and (B) seeds treated with (■) nanofiber-immobilized ISBI40 or (●) ISBI40 culture. After treatment, the seeds were stored for 30 days at room temperature. Data are plotted as the mean values of viable cell numbers (log10 CFU·seed-1). Different letters indicate statistically significant differences (P < 0.05) in the number of viable cells between seed treatment, seed variety and storage time, in accordance with Tukey's test.
Fig 3.
Macro and micro-appearance of soybean seeds (TEC 5936 IPRO variety).
(A) Photographs of soybean seeds not treated (Untreated-seeds), inoculated with rhizobacteria cultures (Inoculated-seeds), and coated with nanofiber-immobilized rhizobacteria (Coated-seeds). (B) Scanning electron microscopy micrographs of untreated seeds (Untreated), inoculated with P. agglomerans ISIB55 (ISIB55) or B. caribensis ISIB40 (ISIB40) cultures and coated with nanofiber-immobilized Yeast Mannitol Broth medium (YMB-PVA), nanofiber-immobilized ISIB55 (ISIB55-PVA) or nanofiber-immobilized ISIB40 (ISIB40-PVA). The micrographs were observed at 5000x magnification. The results are representative of two independent experiments. Similar results were observed in the seed variety RSF 7166 IPRO.
Table 2.
Effects of seed coating with nanofiber-immobilized beneficial rhizobacteria on germination, seedling growth and vigor index of soybean.
Fig 4.
Scanning electron microscopy micrographs of soybean roots from seeds (TEC 5936 IPRO variety).
Seeds (A) untreated, (B) inoculated with P. agglomerans ISIB55 (ISIB55) or (C) B. caribensis ISIB40 (ISIB40) cultures and (D) coated with nanofiber-immobilized Yeast Mannitol Broth medium (YMB-PVA), (E) nanofiber-immobilized ISIB55 (ISIB55-PVA) or (F) nanofiber-immobilized ISIB40 (ISIB40-PVA). Images observed at 5000× magnification show a visibly greater colonization of rhizobacteria on the roots from the coated seeds compared to the inoculated seeds. Similar results were observed in the seed variety RSF 7166 IPRO. The results are representative of two independent experiments.
Fig 5.
Effect of seed coating with nanofiber-immobilized rhizobacteria on growth promotion in soybean plants.
(A) Leaf number, (B) length of shoot and main root and (C) dry weight of shoot and root determined in plants from seeds (TEC 5936 IPRO and RSF 7166 IPRO varieties) inoculated with Yeast Mannitol Broth medium (YMB), coated with nanofiber-immobilized YMB (YMB-PVA), nanofiber-immobilized P. agglomerans ISIB55 (ISIB55-PVA) or nanofiber-immobilized B. caribensis ISIB40 (ISIB40-PVA) and inoculated with P. agglomerans ISIB55 (ISIB55) or B. caribensis ISIB40 (ISIB40) cultures. Data are plotted as mean values ± standard error. Different letters indicate statistically significant differences between seed treatments in the same variety (P < 0.05). (D) Photographs of soybean plants (TEC 5936 IPRO and RSF 7166 IPRO varieties) obtained after 25 days of seed seeding treated, as described above. Results are representative of two independent experiments.