Assessing the effects of nickel on, e.g., Medicago sativa L. nodules using multidisciplinary approach

Industrial wastes and fertilizers can introduce excessive levels of nickel (Ni) into the environment, potentially causing threats to plants, animals, as well as human beings. However, the number of studies on the effects of Ni toxicity on nodules is fairly limited. To address this issue, the effects of increasing Ni concentration on alfalfa nodules were assessed at chemical, biochemical, and transcriptomic levels. For this purpose, plants were grown in soils supplied with Ni (control, 0 mg/kg; C1, 50 mg/kg; C2, 150 mg/kg; C3, 250 mg/kg; and C4, 500 mg/kg) for 90 days. Ni loads in leaves, roots, and nodules were monitored after the exposure period. A set of biochemical biomarkers of oxidative stress was determined in nodules including antioxidants and metal homeostasis as well as lipid peroxidation. Gene expression levels of the main targets involved in oxidative stress and metal homeostasis were assessed. Our data indicated a high concentration of Ni in leaves, roots, and nodules where values reached 25.64 ± 3.04 mg/kg, 83.23 ± 5.16 mg/kg, and 125.71 ± 4.53 mg/kg in dry weight, respectively. Moreover, a significant increase in nodule biomass was observed in plants exposed to C4 in comparison to control treatment and percentage increased by 63%. Then, lipid peroxidation increased with a rate of 95% in nodules exposed to C4. Enzymatic activities were enhanced remarkably, suggesting the occurrence of oxidative stress, with increased superoxide dismutase (SOD), glutathione reductase (GR), and ascorbate peroxidase (APX). Our results showed also a significant upregulation of SOD, GR and APX genes in nodules. Nodule homoglutathione (HGSH) levels increased with the different Ni concentrations, with a remarkable decrease of glutathione S-transferase (GST) activity and glutathione (GSH) content for the highest Ni concentration with 43% and 52% reduction, respectively. The phytochelatin (PC) and metallothionein (MT) concentrations increased in nodules, which implied the triggering of a cellular protection mechanism for coping with Ni toxicity. The results suggested that Ni promotes a drastic oxidative stress in alfalfa nodules, yet the expression of MT and PC to reduce Ni toxicity could be used as Ni stress bioindicators. Our findings provide new insights into the central role of alfalfa nodules in limiting the harmful effects of soil pollution. Therefore, nodules co-expressing antioxidant enzymes may have high phytoremediation potential.