Comparative Physiological and Metabolic Analysis Reveals a Complex Mechanism Involved in Drought Tolerance in Chickpea (Cicer arietinum L.) Induced by PGPR and PGRs.

The plant growth promoting rhizobacteria (PGPR) and plant growth regulators (PGRs) can be applied to improve the growth and productivity of plants, with potential to be used for genetic improvement of drought tolerance. However, for genetic improvement to be achieved, a solid understanding of the physiological and biochemical changes in plants induced by PGPR and PGR is required. The present study was carried out to investigate the role of PGPR and PGRs on the physiology and biochemical changes in chickpea grown under drought stress conditions and their association with drought tolerance. The PGPR, isolated from the rhizosphere of chickpea, were characterized on the basis of colony morphology and biochemical characters. They were also screened for the production of indole-3-acetic acid (IAA), hydrogen cyanide (HCN), ammonia (NH₃), and exopolysaccharides (EPS) production. The isolated PGPR strains, named P1, P2, and P3, were identified by 16S-rRNA gene sequencing as Bacillus subtilis, Bacillus thuringiensis, and Bacillus megaterium, respectively. The seeds of two chickpea varieties, Punjab Noor-2009 (drought sensitive) and 93127 (drought tolerant) were soaked for 2-3 h prior to sowing in 24 h old cultures of isolates. The salicylic acid (SA) and putrescine (Put) were sprayed (150 mg/L) on 25 day old chickpea seedlings. The results showed that chickpea plants treated with a consortium of PGPR and PGRs significantly enhanced the chlorophyll, protein, and sugar contents compared to irrigated and drought conditions. Leaf proline content, lipid peroxidation, and activities of antioxidant enzymes (CAT, APOX, POD, and SOD) all increased in response to drought stress but decreased due to the PGPR and PGRs treatment. An ultrahigh performance liquid chromatography-high resolution mass spectrometry (UPLC-HRMS) analysis was carried out for metabolic profiling of chickpea leaves planted under controlled (well-irrigated), drought, and consortium (drought plus PGPR and PGRs) conditions. Proline, L-arginine, L-histidine, L-isoleucine, and tryptophan were accumulated in the leaves of chickpea exposed to drought stress. Consortium of PGPR and PGRs induced significant accumulation of riboflavin, L-asparagine, aspartate, glycerol, nicotinamide, and 3-hydroxy-3-methyglutarate in the leaves of chickpea. The drought sensitive chickpea variety showed significant accumulation of nicotinamide and 4-hydroxy-methylglycine in PGPR and PGR treated plants at both time points (44 and 60 days) as compared to non-inoculated drought plants. Additionally, arginine accumulation was also enhanced in the leaves of the sensitive variety under drought conditions. Metabolic changes as a result of drought and consortium conditions highlighted pools of metabolites that affect the metabolic and physiological adjustments in chickpea that reduce drought impacts.