A Seed Endophytic Bacterium Cronobacter dublinensis BC-14 Enhances the Growth and Drought Tolerance of Echinochloa crus-galli

Abstract

Successful seed germination and plant seedling growth often require association with endophytic bacteria. Barnyard grass (Echinochloa crus-galli (L.) P. Beauv.) is a main weed during rice cultivation and has frequently been found in drought-prone fields such as cornfields in recent years. To determine whether endophytic bacteria enhance the survival chances of barnyard grass in dryland conditions, endophytic bacteria were collected from barnyard grass seeds. An endophytic bacterial strain, BC-14, was selected and confirmed as Cronobacter dublinensis based on its morphology, physiology, biochemistry, and genomic information. Moreover, C. dublinensis BC-14 secreted IAA in the Luria-Bertani broth up to 28.44 mg/L after 5 days; it could colonize the roots of barnyard grass. After the inoculation with seeds or the well-mixed planting soil, the bacterium can significantly increase the root length and plant height of barnyard grass under drought conditions. When comparing with the control group on the 28th day, it can be seen that the bacterium can significantly increase the contents of chlorophyll b (up to 7.58 times) and proline (37.21%); improve the activities of superoxide dismutase, catalase, and peroxidase (36.90%, 51.51%, and 12.09%, respectively); and reduce the content of malondialdehyde around 25.92%, which are correlated to the drought tolerance. The bacterial genomic annotation revealed that it contains growth-promoting and drought-resistant functional genes. In a word, C. dublinensis BC-14 can help barnyard grass suppress drought stress, promote plant growth, and enhance biomass accumulation, which is helpful to interpret the mechanism of weed adaptability in dry environments.

Keywords: barnyard grass; drought stress tolerance; plant growth promotion; plant–microbe interaction; seed endophytic bacteria.

Figure 1

The effects on barnyard grass ((A) root length; (B) leaf length; (C) fresh weight; (D) dry weight; and (E) seed germination rate) applied with four endophytic bacteria. There are significant differences between data groups represented by different lowercase letters (p < 0.05).

Figure 2

Identification of Cronobacter dublinensis BC-14. (A) Cultural characteristics. (B) Gram staining. (C) Heatmap of ANI analysis based on the genome sequences of 10 Cronobacter strains. (D) The phylogenetic tree based on single-copy ortholog gene sequences of genomes using the maximum likelihood (ML) method. Bootstrap values from 1000 replicates are shown at nodes. Cronobacter condimenti LMG 26250 was selected as the outgroup. ^T: type strain. Scale bar: B = 25 μm.

Figure 3

GFP-tagged Cronobacter dublinensis BC-14 localization in barnyard grass roots using confocal laser microscopic images ((A) white light; (B) fluorescence mode; and (C) overlay image). The white arrows represented bacteria colonizing the roots.

Figure 4

The growth appearance (A1,B1), root length (A2,B2), and plant height (A3,B3) of barnyard grass treated with Cronobacter dublinensis BC-14 without (A: 0% PEG) and under (B: 20% PEG) drought stress conditions at 7, 14, 21, and 28 days. C−: The seeds were soaked in sterile water and then sown in aseptic soil; C+: The seeds were soaked in sterile water and then sown in bacterial (BC-14) mixed soil; S−: The seeds were soaked in the bacterial inoculum (BC-14, 10⁸ CFU/mL) and then sown in aseptic soil. ***: significant difference compared with C− and C+, p < 0.05; **: significant difference compared with C− and S−, p < 0.05; *: significant difference compared with C, p < 0.05. The same figure below.

Figure 5

The variations of chlorophyll (A1,A2), chlorophyll a (B1,B2), chlorophyll b (C1,C2), proline (D1,D2), superoxide dismutase (E1,E2), malondialdehyde (F1,F2), catalase (G1,G2), and peroxidase (H1,H2) in barnyard grass at 7, 14, 21, and 28 days after the treatment with Cronobacter dublinensis BC-14 without ((A1–H1): 0% PEG) and under ((A2–H2): 20% PEG) drought stress conditions.

Figure 6

The whole genome sequence of Cronobacter dublinensis BC-14.