Drought Tolerant Enterobacter sp./ Leclercia adecarboxylata Secretes Indole-3-acetic Acid and Other Biomolecules and Enhances the Biological Attributes of Vigna radiata (L.) R. Wilczek in Water Deficit Conditions

Abstract

Drought or water stress is a limiting factor that hampers the growth and yield of edible crops. Drought-tolerant plant growth-promoting rhizobacteria (PGPR) can mitigate water stress in crops by synthesizing multiple bioactive molecules. Here, strain PAB19 recovered from rhizospheric soil was biochemically and molecularly characterized, and identified as Enterobacter sp./Leclercia adecarboxylata (MT672579.1). Strain PAB19 tolerated an exceptionally high level of drought (18% PEG-6000) and produced indole-3-acetic acid (176.2 ± 5.6 µg mL^-1), ACC deaminase (56.6 ± 5.0 µg mL^-1), salicylic acid (42.5 ± 3.0 µg mL^-1), 2,3-dihydroxy benzoic acid (DHBA) (44.3 ± 2.3 µg mL^-1), exopolysaccharide (204 ± 14.7 µg mL^-1), alginate (82.3 ± 6.5 µg mL^-1), and solubilized tricalcium phosphate (98.3 ± 3.5 µg mL^-1), in the presence of 15% polyethylene glycol. Furthermore, strain PAB19 alleviated water stress and significantly (p ≤ 0.05) improved the overall growth and biochemical attributes of Vigna radiata (L.) R. Wilczek. For instance, at 2% PEG stress, PAB19 inoculation maximally increased germination, root dry biomass, leaf carotenoid content, nodule biomass, leghaemoglobin (LHb) content, leaf water potential (ΨL), membrane stability index (MSI), and pod yield by 10%, 7%, 14%, 38%, 9%, 17%, 11%, and 11%, respectively, over un-inoculated plants. Additionally, PAB19 inoculation reduced two stressor metabolites, proline and malondialdehyde, and antioxidant enzymes (POD, SOD, CAT, and GR) levels in V. radiata foliage in water stress conditions. Following inoculation of strain PAB19 with 15% PEG in soil, stomatal conductance, intercellular CO₂ concentration, transpiration rate, water vapor deficit, intrinsic water use efficiency, and photosynthetic rate were significantly improved by 12%, 8%, 42%, 10%, 9% and 16%, respectively. Rhizospheric CFU counts of PAB19 were 2.33 and 2.11 log CFU g^-1 after treatment with 15% PEG solution and 8.46 and 6.67 log CFU g^-1 for untreated controls at 40 and 80 DAS, respectively. Conclusively, this study suggests the potential of Enterobacter sp./L. adecarboxylata PAB19 to alleviate water stress by improving the biological and biochemical features and of V. radiata under water-deficit conditions.

Keywords: Enterobacter sp./L. adecarboxylata; Vigna radiata (L.) R. Wilczek; drought stress; gas exchange parameters; growth improvement; growth regulating substances.

Figure 1

Neighbor-joined phylogenetic tree of Enterobacter sp./Leclercia adecarboxylata PAB19. The tree was constructed based on 16S rRNA partial gene sequences (taken from NCBI BLAST search tool) from some closely related phylogenetic species (type cultures). Sequences were aligned using the Clustal W sequence alignment tool in MEGA 7.0 software. Bootstrap percentage values as obtained from 1000 replications of the data set are given at nodes. The GenBank accession numbers of isolates and other genera or species are presented in parenthesis. The scale bar corresponds to the mean number of nucleotide substitutions per site.

Figure 2

Effects of different levels of water stress (2%, 5%, 10%, and 15% PEG solution) on biofilm development (A) and their associated traits; EPS production (B), alginate production (C), and cell surface hydrophobicity (D) of Enterobacter sp./L. adecarboxylata PAB 19. In these figures, bar diagrams represent the mean values of three replicates. Mean values followed by different letters were significantly different (p ≤ 0.05) as determined by the Duncan’s Multiple Range Test (DMRT). Bar and scatter plots represent means ± SDs.

Figure 3

Germination efficiency (A), seed vigor indices (B), leaf water potential (C), and membrane stability indices (D) of V. radiata (L.) cultivated under different levels of water stress (2%, 5%, 10% and 15% PEG-6000) and inoculated with drought-tolerant Enterobacter sp./L. adecarboxylata PAB19 under green-house conditions. Bar diagrams and curves represent the mean values of experiments done in triplicate. Mean values followed by different letters are significantly different at p ≤ 0.05 as determined by the DMRT. Bar and scatter plots represent means ± SDs.

Figure 4

Impact of the bio-inoculation of Enterobacter sp./L. adecarboxylata PAB19 on biological features; plant length at 50 DAS (A) and 80 DAS (B), fresh weight at 50 DAS (C) and 80 DAS (D), dry biomass at 50 DAS (E) and 80 DAS (F) of V. radiata (L.) cultivated under different levels of water stress (2%, 5%, 10% and 15% PEG-6000) under greenhouse conditions. Bar diagrams represent the mean values of three replicates. Mean values followed by different letters are significantly different (p ≤ 0.05) as determined by the DMRT. Bar and scatter plots represent means ± SDs.

Figure 5

Impacts of the bio-inoculation of Enterobacter sp./L. adecarboxylata PAB19 on leaf pigments (A), shoot P (B), root P (C), shoot N (D), and root N (E) of V. radiata (L.) cultivated under different levels of drought stress (2%, 5%, 10% and 15% PEG-6000) under greenhouse conditions. Mean values followed by different letters are significantly different (p ≤ 0.05) as determined by the DMRT test. Bar and scatter plots represent means ± SDs. Bar and scatter plots represent means ± SDs.

Figure 6

Impact of Enterobacter sp./L. adecarboxylata PAB19 on symbiotic features such as nodule number (A), nodule biomass (B), and LHb content (C) at 50 DAS of V. radiata (L.) in the presence of different levels of water stress (2%, 5%, 10% and 15% PEG-6000) under greenhouse conditions. Bar diagrams represent the mean values of three replicates. Mean values followed by different letters are significantly different (p ≤ 0.05) as determined by the DMRT. Bar and scatter plots represent means± SDs.

Figure 7

Impact of drought-tolerant Enterobacter sp./L. adecarboxylata PAB19 on stress-related parameters; proline (A) and MDA (B) contents and antioxidant enzyme activities; CAT (C), SOD (D), POD (E), and GR (F) of V. radiata cultivated in the presence of different levels of water stress (2%, 5%, 10% and 15% PEG-6000) raised under greenhouse conditions. Bar diagrams represent the mean values of three replicates. Mean values followed by different letters are significantly different (p ≤ 0.05) as determined by the DMRT. Bar and scatter plots represent means ± SDs.

Figure 8

Impact of Enterobacter sp./L. adecarboxylata PAB19 on gas exchange parameters; stomatal conductance (A), intercellular CO₂ concentration (B), transpiration rate (C), vapor pressure deficit (D), intrinsic water use efficiency (E), and photosynthetic rate (F) of V. radiata cultivated in the presence of different levels of water stress under green-house conditions. Bar and scatter plots represent means± SDs. Mean values followed by different letters are significantly different (p ≤ 0.05) as determined by the DMRT.