ACC Deaminase Producing Methylobacterium oryzae CBMB20 and Exogenous Trehalose Application Alleviate Salinity Stress in Arabidopsis

Abstract

Farming communities are very concerned about salt stress because of its negative impact on crop productivity. This study evaluated the ability of the Methylobacterium oryzae CBMB20 and exogenous trehalose treatments on Arabidopsis thaliana growth under salt stress conditions. A. thaliana growth was enhanced using M. oryzae CBMB20 as a bioinoculant in both saline and non-saline environments. In addition to better photosynthetic efficiency and endogenous trehalose content, the inoculation of M. oryzae CBMB20 produced improved growth parameters, such as increased rosette fresh weight and shoot length. Reduced levels of proline and malondialdehyde (MDA) under salt stress (150 mM NaCl) further indicated that the inoculated plants had enhanced tolerance to salinity. GFP-tagged M. oryzae CBMB20 was also used in spatial distribution experiments, which showed that the bacteria colonized A. thaliana's root, shoot, and hypocotyl. By increasing shoot length and total fresh weight, the exogenous application of trehalose also markedly enhanced plant growth. Proline and MDA contents were decreased by exogenous trehalose during salt stress, while the endogenous trehalose concentration in A. thaliana remained unaffected. The use of trehalose and M. oryzae CBMB20 can both have a good impact on plant development and stress tolerance in saline environments.

Keywords: Arabidopsis thaliana; Methylobacterium oryzae; PGPR; osmolytes; salt stress; trehalose.

Fig. 1. The growth of Arabidopsis thaliana plants after inoculation with M. oryzae CBMB20 under salt stress and non-stress conditions.

Bacteria+ (treated with M. oryzae CBMB20); Bacteria- (not treated with M. oryzae CBMB20).

Fig. 2. Effects of M. oryzae CBMB20 on plant growth under salt stress and non-stress conditions.

Arabidopsis development phenotypes were analyzed in response to salt stress, inoculated with (B+) and without (B-) M. oryzae CBMB20 under 0 mM NaCl and 150 mM NaCl. Figures show (A) Rosette Fresh Weight, (B) Shoot Length, (C) Root Length, and (D) Whole Fresh Weight. Different letters indicate the significant difference among treatments (p < 0.05); according to one way ANOVA test; Data are the mean ± SD, n = 21.

Fig. 3. Effects of M. oryzae CBMB20 on photosynthetic pigments of Arabidopsis plants grown under salt stress and non-stress conditions.

Figures show (A) Chlorophyll a, (B) Chlorophyll b, and (C) Carotenoid. The data represents the mean ± SD of three replicates and the different letters indicate the significant difference among treatments (p < 0.05); according to one way ANOVA test. Treatments details are the same as in Fig. 2.

Fig. 4. Effects of M. oryzae CBMB20 on (A) Proline content, (B) MDA content and (C) Cellular trehalose concentration of leaves in Arabidopsis plants grown under salt stress and non-stress conditions.

The data represents the mean ± SD of three replicates and the different letters indicate the significant difference among treatments (p < 0.05); according to one way ANOVA test. Treatments details are the same as in Fig. 2.

Fig. 5. Confocal microscopic images of the persistence of GFP-tagged M. oryzae CBMB20 in roots, shoot and hypocotyl of Arabidopsis thaliana.

(A) In root under non-stress condition, (B) in shoot under non-stressed condition, (C) in hypocotyl under non-stress condition, (D) in root under salt stress condition, (E) in shoot under salt stress condition, and (F) in hypocotyl under salt stress condition.

Fig. 6. The growth of Arabidopsis thaliana plants after exogenous trehalose application under salt stress and non- stress conditions.

Fig. 7. Effect of exogenous applied trehalose on plant growth under salt stress and non-stress conditions.

Arabidopsis development phenotypes were analyzed in response to salt stress, application (Tre⁺) or without (Tre^-) of 10 mM Trehalose under 0 mM NaCl and 150 mM NaCl concentration. (A) Whole Fresh Weight, (B) Shoot Length, (C) Rosette Fresh Weight, and (D) Root Length. Different letters indicate the significant difference among treatments (p < 0.05); according to one way ANOVA test; Data are the mean ± SD, n = 21.

Fig. 8. Effects of exogenous trehalose application on (A) Proline content, (B) MDA content, and (C) Cellular trehalose concentration of leaves in Arabidopsis plants grown under salt stress and non-stress conditions.

The data represents the mean ± SD of three replicates and the different letters indicate the significant difference among treatments (p < 0.05); according to one way ANOVA test. Treatments details are the same as in Fig. 7.