Investigating the mechanisms underlying phytoprotection by plant growth‐promoting rhizobacteria in Spartina densiflora under metal stress

Abstract

Pollution of coasts by toxic metals and metalloids is a worldwide problem for which phytoremediation using halophytes and associated microbiomes is becoming relevant. Metal(loid) excess is a constraint for plant establishment and development, and plant growth promoting rhizobacteria (PGPR) mitigate plant stress under these conditions. However, mechanisms underlying this effect remain elusive. The effect of toxic metal(loid)s on activity and gene expression of ROS-scavenging enzymes in roots of the halophyte Spartina densiflora grown on real polluted sediments in a greenhouse experiment was investigated. Sediments of the metal-polluted joint estuary of Tinto and Odiel rivers and control, unpollutred samples from the Piedras estuary were collected and submitted to ICP-OES. Seeds of S. densiflora were collected from the polluted Odiel marshes and grown in polluted and unpolluted sediments. Rhizophere biofilm-forming bacteria were selected based on metal tolerance and inoculated to S. densiflora and grown for 4 months. Fresh or frozen harvested plants were used for enzyme assays and gene expression studies, respectively. Metal excess induced SOD (five-fold increase), whereas CAT and ascorbate peroxidase displayed minor induction (twofold). A twofold increase of TBARs indicated membrane damage. Our results showed that metal-resistant PGPR (P. agglomerans RSO6 and RSO7 and B. aryabhattai RSO25) contributed to alleviate metal stress, as deduced from lower levels of all antioxidant enzymes to levels below those of non-exposed plants. The oxidative stress index (OSI) decreased between 50 and 75% upon inoculation. The results also evidenced the important role of PAL, involved in secondary metabolism and/or lignin synthesis, as a pathway for metal stress management in this halophyte upon inoculation with appropriate PGPR, since the different inoculation treatments enhanced PAL expression between 3.75- and five-fold. Our data confirm, at the molecular level, the role of PGPR in alleviating metal stress in S. densiflora and evidence the difficulty of working with halophytes for which little genetic information is available.

Safe Cultivation of Medicago sativa in Metal-Polluted Soils from Semi-Arid Regions Assisted by Heat- and Metallo-Resistant PGPR

“…To estimate the overall oxidative stress, the OSI was proposed [34,45]. The high value of OSI was recorded in non-inoculated plants grown at HMC, for which the OSI was almost twice than the control.…”

“…In order to understand the molecular mechanisms underlying plants root protection by bacteria, expression levels of several stress related genes were investigated. Based on their involvement in metal stress management, nine genes were selected which were representative of the different pathways for stress alleviation [34] and their involvement in bacteria-plant symbiosis [39] (Table 1), as follows: Histone 1 ( His1 ) was used as the housekeeping gene; NRAMP1 (natural resistance-associated macrophage protein) is a Mn transporter which is known to be overexpressed under As and Cd stress in rice [48,49]; glutathione reductase ( GR ) is involved in the maintenance of the redox state [36]; and phytochelatin synthase ( PCS ) is involved in the synthesis of phytochelatins for metal complexation [50]. The gene CP450 coding for cytochrome P450 is involved in stress management under xenobiotics and metal exposure [51,52].…”

“…Oxidative stress index or OSI is a parameter recently introduced [34,45] to express the extent of the oxidative stress and it is calculated as the average of the prorated values of the different antioxidant enzymes in reference to their values in the control situation, as follows: OSI=([SOD]/[SOD]0 + [CAT]/[CAT]0 + [APX]/[APX]0 + [GPX]/[GPX]0)/4 where [SOD], [CAT], [APX], and [GPX] are the values of these enzymes in the presence of heavy metals (with or without bacteria). [SOD]0, [CAT]0, [APX]0, and [GPX]0 were the control values, i.e., in the absence of both metals and bacteria.…”

“…[31,32]. Once inside the cell, heavy metals induce oxidative, osmotic, and ionic stresses in plants [33,34], which are accompanied by the induction of antioxidant enzymes such as superoxide dismutase (SOD), ascorbate peroxidase (APX), glutathione peroxidase (GR), catalase (CAT), and low molecular weight deactivating molecules such as ascorbic acid and thiols such as glutathione and proline. Moreover, the intracellular presence of organic ligands, namely metallothionins, phytochelatins, organic acids, and amino acids, ensures complexation and therefore the detoxification of metal ions [35,36,37].…”

“…From the point of view of omics, adaptation to metals implies the overexpression of multiple genes belonging to different pathways such as cysteine synthase, glutathione transferase, and glutathione reductase for the metabolism of glutathione and phytochelatins, genes of secondary metabolism such as phenylalanine ammonia lyase, and genes involved in drought stress such as the proline synthesis pathway, metal transporters, etc. [34,38,39]. Therefore, studies are needed to understand the plant–bacteria relationship and to identify genes and pathways that participate in the management of metal stress.…”