The multifaceted roles of Arbuscular Mycorrhizal Fungi in peanut responses to salt, drought, and cold stress

Abstract

Background: Arbuscular Mycorrhizal Fungi (AMF) are beneficial microorganisms in soil-plant interactions; however, the underlying mechanisms regarding their roles in legumes environmental stress remain elusive. Present trials were undertaken to study the effect of AMF on the ameliorating of salt, drought, and cold stress in peanut (Arachis hypogaea L.) plants. A new product of AMF combined with Rhizophagus irregularis SA, Rhizophagus clarus BEG142, Glomus lamellosum ON393, and Funneliformis mosseae BEG95 (1: 1: 1: 1, w/w/w/w) was inoculated with peanut and the physiological and metabolomic responses of the AMF-inoculated and non-inoculated peanut plants to salt, drought, and cold stress were comprehensively characterized, respectively.

Results: AMF-inoculated plants exhibited higher plant growth, leaf relative water content (RWC), net photosynthetic rate, maximal photochemical efficiency of photosystem II (PSII) (Fv/Fm), activities of antioxidant enzymes, and K⁺: Na⁺ ratio while lower leaf relative electrolyte conductivity (REC), concentration of malondialdehyde (MDA), and the accumulation of reactive oxygen species (ROS) under stressful conditions. Moreover, the structures of chloroplast thylakoids and mitochondria in AMF-inoculated plants were less damaged by these stresses. Non-targeted metabolomics indicated that AMF altered numerous pathways associated with organic acids and amino acid metabolisms in peanut roots under both normal-growth and stressful conditions, which were further improved by the osmolytes accumulation data.

Conclusion: This study provides a promising AMF product and demonstrates that this AMF combination could enhance peanut salt, drought, and cold stress tolerance through improving plant growth, protecting photosystem, enhancing antioxidant system, and regulating osmotic adjustment.

Keywords: AMF; Environmental stress; Legumes; Metabolic pathway; Plant physiology.

Fig. 1

Effect of AMF on (a) plant growth, (b) plant height, (c) shoot dry weight, (d) relative water content (RWC), and (e) relative electrolyte conductivity (REC) in peanut plants under stressful conditions. The peanut seeds were inoculated with/without AMF before exposing to normal/salt/drought/cold growth conditions. The measurements were conducted on the 8^th day after the onset of stress treatments. Bars represent the mean values of three biological replicates with standard deviation; asterisks indicate a significant difference in comparison to non-AMF according to Tukey's test (P < 0.05)

Fig. 2

Effect of AMF on gas exchange in the third fully expanded leaves of peanut plants under stressful conditions. (a) Net photosynthetic rate (Pn), (b) stomatal conductance (Gs), (c) intercellular CO₂ concentration (Ci), and (d) transpiration rate (Tr). The peanut seeds were inoculated with/without AMF before exposing to normal/salt/drought/cold growth conditions. The gas exchange parameters were measured on the 8^th day after the onset of stress treatments. Bars represent the mean values of three biological replicates with standard deviation; asterisks indicate a significant difference in comparison to non-AMF according to Tukey's test (P < 0.05)

Fig. 3

Effect of AMF on chlorophyll fluorescence and chlorophyll content in the third fully expanded leaves of peanut plants under stressful conditions. The peanut seeds were inoculated with/without AMF before exposing to normal/salt/drought/cold growth conditions. On 8^th day after the onset of stress treatments, the images showing (a) the maximal photochemical efficiency of photosystem II (PSII) (Fv/Fm) were taken. The false color code depicted at the bottom of the image ranges from 0 (black) to 1 (purple). Vertical bar = 1 cm. Meanwhile, the value of (b) the total chlorophyll content, (c) Fv/Fm, (d) photochemical quenching coefficient (qP), and (e) the quantum yield of PSII photochemistry (ΦPSII) were determined. Bars represent the mean values of three biological replicates with standard deviation; asterisks indicate a significant difference in comparison to non-AMF according to Tukey's test (P < 0.05)

Fig. 4

Effect of AMF on the integrity of thylakoids and mitochondria in the third fully expanded leaves of peanut plants under stressful conditions. The peanut seeds were inoculated with/without AMF before exposing to normal/salt/drought/cold growth conditions. On the 8^th day after the onset of stress treatments, the leaves were collected for cytochemical staining and observed with a transmission electron microscopy

Fig. 5

Effect of AMF on the accumulations of H₂O₂, O₂^-., and concentration of MDA in the third fully expanded leaves of peanut plants under stressful conditions. The peanut seeds were inoculated with/without AMF before exposing to normal/salt/drought/cold growth conditions. On the 8^th day after the onset of stress treatments, the leaves were excised, and the histochemical staining of (a) H₂O₂ (DAB staining) and O₂^-. (NBT staining) were performed. Vertical bar = 1 cm. Meanwhile, the leaves were collected and the (b) concentration of H₂O₂, (c) O₂^-. production rate, and (d) concentration of MDA were measured. Bars represent the mean values of three biological replicates with standard deviation; asterisks indicate a significant difference in comparison to non-AMF according to Tukey's test (P < 0.05)

Fig. 6

Effect of AMF on antioxidant enzymes in the third fully expanded leaves of peanut plants under stressful conditions. The peanut seeds were inoculated with/without AMF before exposing to normal/salt/drought/cold growth conditions. The activities of (a) superoxide dismutase (SOD), (b) guaiacol peroxidase (G-POD), (c) catalase (CAT), and (d) ascorbate peroxidase (APX) were measured on the 8^th day after the onset of stress treatments. Bars represent the mean values of three biological replicates with standard deviation; asterisks indicate a significant difference in comparison to non-AMF according to Tukey's test (P < 0.05)

Fig. 7

(a) OPLS-DA models, (b) volcano plots, and (c) heat map showing the differential metabolites with MS2 based on the non-target metabolomics in the peanut root samples of “AMF vs Control”

Fig. 8

(a) KEGG classification and (b) biosynthetic pathway analysis based on the non-target metabolomics in the peanut root samples of “AMF vs Control”

Fig. 9

The working model illustrating the mechanisms of AMF in alleviating peanut salt, drought, and cold stress