Influence of salt stress on biochemical processes in chickpea,Cicer arietinum L.

Summary Soybean plants were grown in pots with or without vesicular-arbuscular myocorrhizal (VAM) fungi in three soils of low plant-available P content, different texture and different water-holding capacities. Mineral nutrients, except P, were provided in a complete nutrient solution. The biomass of non-VAM plants was positively and fungal colonization negatively correlated with increasingly coarse soil texture. There was no correlation of soil P with host or endophyte growth. Plant growth enhancement was positively correlated with soil water content at −1.5 MPa. These observations suggest soil water status and the mycorrhizal condition interact in influencing plant growth.     

Isolation,characterization, and effect of phosphate-zinc-solubilizing bacterial strains on chickpea (Cicer arietinum L.) growth

ObjectivePhosphate (P) and zinc (Zn) are essential plant nutrients required for nodulation, nitrogen-fixation, plant growth and yield. Mostly applied P and Zn nutrients in the soil are converted into unavailable form. A small number of soil microbes have the ability to transform unsolvable forms of P and Zn to an available form. P-Zn-solubilizing rhizobacteria are potential alternates for P and Zn supplement. In the present study, the effect of two P-Zn-solubilizing bacterial strains (Bacillus sp. strain AZ17 and Pseudomonas sp. strain AZ5) was evaluated on the growth of chickpea plant.MethodologyBoth strains were purified from the rhizospheric soil of chickpea plant grown-up in sandy soil and rain-fed area (Thal desert). In vitro, both strains solubilize P and Zn as well both strain produce IAA and organic acids. In the field experiments, conducted in the rain-fed area, the positive influence of inoculation with both bacterial isolates AZ5 and AZ17 on chickpea growth was observed.ResultsThe application of inoculum (strains AZ5 and AZ17) resulted in up to 17.47% and 17.34% increase in grain yield of both types of chickpea grown in fertilized and non-fertilized soil, respectively over non-inoculated control. Strain AZ5 was the most effective inoculum, increasing up to 17.47%, 16.04%, 26.32%, 22.53%, 26.12% and 22.59% in grain yield, straw weight, nodules number, dry weight of nodules, Zn uptake and P uptake respectively, over control.ConclusionThese results indicated that Pseudomonas sp. strain AZ5 and Bacillus sp. strain AZ17 can serve as effective microbial inocula for chickpea, particularly in the rain-fed area.     

The effect of soil pH on chickpea (Cicer arietinum) genotype sensitivity to isoxaflutole

A glasshouse experiment was conducted to investigate the effect of soil pH on chickpea (Cicer arietinum) tolerance to isoxaflutole applied pre-emergence at 0, 75 (recommended rate) and 300 g a.i. ha⁻¹. For this study, the variables examined were two desi chickpea genotypes (97039-1275 as a tolerant line and 91025-3021 as a sensitive line) and four pH levels (5.1, 6.9, 8.1, and 8.9). The results demonstrated differential tolerances among chickpea genotypes to isoxaflutole at different rates and soil pH levels. Isoxaflutole applied pre-emergence resulted in increased phytotoxicity with increases in soil pH and herbicide rate. Even the most tolerant chickpea genotype was damaged when exposed to higher pH and herbicide rates, as indicated by increased leaf chlorosis and significant reductions in plant height, and shoot and root dry weight. The effects were more severe with the sensitive genotype. The susceptibility of chickpea to this herbicide depends on genotype and soil pH which should be taken into account in breeding new lines, and in the agronomy of chickpea production.     

The effects of salinity and sodicity upon nodulation and nitrogen fixation in chickpea (Cicer arietinum)

Production of grain legumes is severely reduced in salt-affected soils because their ability to form and maintain nitrogen-fixing nodules is impaired by both salinity and sodicity (alkalinity). Genotypes of chickpea, Cicer arietinum, with high nodulation capacity under stress were identified by field screening in a sodic soil in India and subsequently evaluated quantitatively for nitrogen fixation in a glasshouse study in a saline but neutral soil in the UK. In the field, pH 8.9 was the critical upper limit for most genotypes studied but genotypes with high nodulation outperformed all others at pH 9.0-9.2. The threshold limit of soil salinity for shoot growth was at ECe 3 dS m(-1), except for the high-nodulation selection for which it was ECe 6. Nodulation was reduced in all genotypes at salinities above 3 dS m(-1) but to a lesser extent in the high-nodulation selection, which proved inherently superior under both non-saline and stress conditions. Nitrogen fixation was also much more tolerant of salinity in this selection than in the other genotypes studied. The results show that chickpea genotypes tolerant of salt-affected soil have better nodulation and support higher rates of symbiotic nitrogen fixation than sensitive genotypes.     

Diversity of pollinators and their role in the pollination biology of chickpea,Cicer arietinum L. (Fabaceae)

Chickpea (Cicer arietinum L.) is an important source of food for people worldwide. In the current study, we studied its pollination biology with special reference to floral visitors along with their visitation rate, frequency and pollen load during 2012 and 2013. We also explored the effect of floral visitors on the capsule weight, seed weight, seed numbers and seed germination. Results revealed three bees, two wasps, five flies, one moth and three butterfly species on the flowers of chickpea. Apis dorsata, A. florea, Amegilla sp. and Eristalinus aeneus were the major species with 434–474, 223–311, 69–74 and 81–136 individuals, respectively in both years. Floral visitors differed significantly in term of visitation frequency with A. florea as the most frequent visitor (9.13–9.86 visits/flower/5 min) followed by E. aeneus (5.43–5.58 visits/flower/5 min) and A. dorsata (1.72–2.31 visits/flower/5 min) in both years. Similarly, A. florea had statistically highest visitation rate (16.85–19.99 flowers visited/min) followed by E. aeneus (9.73–10.68 flowers visited/min). A. dorsata had significantly higher pollen load on its body (84629–85,104 pollen grains) followed by A. florea (64940–65,135 pollen grains) and Amegilla sp. (64020–65,120 pollen grains). The open-pollinated flowers had significantly higher capsule weight (0.27 ± 0.01 g), seed weight (0.18 ± 0.01 g), seed numbers (1.67 ± 0.07 seeds) and seed germination (95 ± 1.38%) as compared to flowers deprived of pollinators in cages. The results suggested A. florea, A. dorsata and E. aeneus could be effective pollinators of chickpea. Hence these three species can be properly utilized on commercial scale to increase crop yield.     

Biochemical and molecular biological studies on infection (Ascochyta rabiei)-induced thaumatin-like proteins from chickpea plants (Cicer arietinum L.)

A pathogenesis-related protein induced by infection with Ascochyta rabiei was purified from intercellular washing fluid of chickpea (Cicer arietinum L.) leaves. Amino-terminal sequencing identified the protein, named PR-5a, as a thaumatin-like protein. The isoelectric point was determined with 6.5 and the molecular mass is 16 kDa. Therefore, chickpea PR-5a is the first dicot member of a TLP subgroup containing small TLPs with a molecular weight between 15 and 18 kDa. PR-5a shows no antifungal activity towards A. rabiei. Screening of a chickpea cDNA library led to the isolation of a cDNA clone (p5a-241) for this protein. A second cDNA clone (ELR112) encoding a TLP was isolated using differential hybridisation of cDNA libraries obtained from elicited and water treated cell suspension cultures of chickpea. The deduced protein (PR-5b) has a molecular mass of 22 kDa. PR-5b is postulated to be located in the vacuole due to the presence of a respective N-terminal signal peptide and a carboxy-terminal extension. Southern blot analyses showed that ELR112 and p5a-241 represent single copy genes. During fungal infection of chickpea plants expression of both genes proceeds much faster in an A. rabiei resistant cultivar than in a susceptible one.     

Partial purification and some biochemical properties of acid phosphatase in germinating chickpea (Cicer arietinum) seeds

An acid phosphatase (EC 3.1.3.2.) from the embryonic axes of chickpea seeds ( Cicer arietinum L. cv. Castellana) was purified by ammonium sulphate precipitation, chromatography on Sephacryl S-200 and polyacrylamide gel electrophoresis. The preparation has an apparent molecular weight of 39 kDa, pH optimum for p -nitrophenylphosphate hydrolysis of 5.25, and K _m of 0.57 m M . The enzyme hydrolyzed all the mono- and di-phosphorylated sugars tested, but had no effect on ATP, ADP, AMP and phosphoenolpyruvate. Phosphate was a competitive inhibitor. Mg²⁺. Ca²⁺, Hg²⁺, Fe³⁺, arsenate, K⁺ and Zn²⁺ were inhibitory. Mn²⁺, dithiothreitol and EDTA had no effect, and polyamines were activators.     

Effect of chloride and sulphate types of salinity on characteristics of chlorophyll content,photosynthesis and respiration of chickpea (Cicer arietinum L.)

Various regimes of predominantly chloride and sulphate salinity reduced chlorophyll (Chl) (a +b) content and net photosynthetic rate (Pn) in two cultivars ofCicer arietinum L. However, the rate of respiration (Rd) was stimulated up to 6 dS m^-1 of salinity and thereafter it declined with increase in salinity levels. Chloride salinity was more detrimental than the sulphate one as far as Chl (a +b) and Pn were concerned, but R_D was reduced more under sulphate salinity in cv. H-75-35 especially in 110 d-old plants. The cultivar L-144 was relatively more salt sensitive than the cv. H-75-35.     

Synergistic effect of Pseudomonas putida and Bacillus amyloliquefaciens ameliorates drought stress in chickpea (Cicer arietinum L.)

Two plant growth promoting rhizobacteria (PGPR) Pseudomonas putida NBRIRA and Bacillus amyloliquefaciens NBRISN13 with ability to tolerate abiotic stress along with multiple PGP traits like ACC deaminase activity, minerals solubilisation, hormones production, biofilm formation, siderophore activity were evaluated for their synergistic effect to ameliorate drought stress in chickpea. Earlier we have reported both the strains individually for their PGP attributes and stress amelioration in host plants. The present study explains in detail the possibilities and benefits of utilizing these 2 PGPR in consortium for improving the chickpea growth under control and drought stressed condition. In vitro results clearly demonstrate that both the PGPR strains are compatible to each other and their synergistic growth enhances the PGP attributes. Greenhouse experiments were conducted to evaluate the effect of inoculation of both strains individually and consortia in drought tolerant and sensitive cultivars (BG362 and P1003). The growth parameters were observed significantly higher in consortium as compared to individual PGPR. Colonization of both PGPR in chickpea rhizosphere has been visualized by using gfp labeling. Apart from growth parameters, defense enzymes, soil enzymes and microbial diversity were significantly modulated in individually PGPR and in consortia inoculated plants. Negative effects of drought stress has been ameliorated and apparently seen by higher biomass and reversal of stress indicators in chickpea cultivars treated with PGPR individually or in consortia. Findings from the present study demonstrate that synergistic application has better potential to improve plant growth promotion under drought stress conditions.     

Active extrusion of protons and exudation of carboxylic acids in response to iron deficiency by roots of chickpea (Cicer arietinum L.)

A chickpea cultivar, K-850, acidified the nutrient solution in response to iron deficiency, with subsequent re-greening of chlorotic leaves. No recovery of chlorosis was observed when the nutrient solution was buffered at a pH 6.3. During the period of acidification induced by iron deficiency, the roots of K-850 exuded more carboxylic acids than when supplied with sufficient iron. However, the rate of extrusion of protons was much higher than the rate of exudation of carboxylic acids during the acidification period. The extrusion of protons was inhibited by the addition of vanadate at the beginning of the decrease in pH. It appeared that acidification of the solution in response to iron deficiency was mediated by a proton-pumping ATPase, located at the plasma membrane. The presence of cations in the solution was essential for the extrusion of protons under iron deficiency, but the species of cation made no significant difference to the rate of extrusion of protons from roots. Therefore, we concluded that non-specific H+/cation antiport was involved in the acidification process.     

Response of field-grown chickpea (Cicer arietinum L.) to phosphorus fertilization for yield and nitrogen fixation

Application of phosphorus at 40, 60, 80 and 100 kg P₂O₅ ha^–1 in the presence of a uniform dressing of nitrogen (N) and potash (K₂O) each applied at 20 and 24 kg ha^–1 to chickpea (CM-88) grown in sandy loam soil in a replicated field experiment improved the nodulation response of the crop, increased its grain yield (ka ha^–1) by 18, 59, 40 and 14 percent, biomass yield (ka ha^–1) by 32, 32, 54 and 14 percent, biomass N (kg ha^–1) by 31, 48, 49, 19 percent, and biomass P (kg ha^–1) by 26, 40, 41 and 11 percent, respectively. The effect of phosphorus on the nitrogenase activity of the excised roots of chickpea was, however, inconsistent.     

Abundance and polymorphism of di-, tri-and tetra-nucleotide tandem repeats in chickpea (Cicer arietinum L.)

The abundance and polymorphism of 38 different simple-sequence repeat motifs was studied in four accessions of cultivated chickpea (Cicer arietinum L.) by in-gel hybridization of synthetic oligonucleotides to genomic DNA digested with 14 different restriction enzymes. Among 38 probes tested, 35 yielded detectable hybridization signals. The abundance and level of polymorphism of the target sequences varied considerably. The probes fell into three broad categories: (1) probes yielding distinct, polymorphic banding patterns; (2) probes yielding distinct, monomorphic banding patterns, and (3) probes yielding blurred patterns, or diffused bands superimposed on a high in lane background. No obvious correlation existed between abundance, fingerprint quality, and the sequence characteristics of a particular motif. Digestion with methyl-sensitive enzymes revealed that simple-sequence motifs are enriched in highly methylated genomic regions. The high level of intraspecific polymorphism detected by oligonucleotide fingerprinting suggests the suitability of simple-sequence repeat probes as molecular markers for genome mapping.     

Effect of phosphorus and zinc on nodulation and nitrogen fixation in chickpea (Cicer arietinum L.)

A green house study was conducted on the effect of P and Zn on nodulation and N fixation in chickpea (Cicer arietinum L.) in a loamy sand (Typic Torripsamments) using treatment combinations of five levels of P (0, 25, 50, 100 and 250 ppm), and six levels of Zn (0, 5, 10, 20, 40 and 100 ppm). The number, dry matter and leghaemoglobin content of nodules, and amount of N fixed generally increased with Zn alone upto 19 ppm and P alone upto 50 ppm, and decreased with their higher levels. Application of 25 to 50 ppm P and 5 to 10 ppm Zn counteracted to a greater extent the adverse effect of 40 and 100 ppm Zn, and 250 ppm P, resp. Maximum nodulation and N fixation (91 to 145% over zero P and Zn, at maturity) was recorded with 25 to 50 ppm P applied along with 5 to 10 ppm Zn. At 64 days, depletion in soil-N was noted, particularly when P was applied, whereas at maturity there was a gain in soil-N, ranging from 10.5 to 44.5 kg/2×10⁶ kg soil depending upon P and Zn treatments. The increase in nodulation and N fixation with balanced P and Zn nutrition might be attributed to an increase in leghaemoglobin, and K and Fe concentration in nodules, and increased plant growth, resulting into enhanced activity of N fixing organisms. The results showed that balanced P and Zn nutrition is essential not only for plant growth but also for maximum activity of Rhizobium for N fixation. Work done at Harvana Agricultural University, Hissar, India.     

Isolation and characterisation of endophytic actinobacteria and their effect on the growth and nodulation of chickpea (Cicer arietinum)

Plant and Soil - Phyllostachys praecox is one of the bamboo species that cultivated extensively in southern China. An organic material mulching technique is commonly adopted in winter to increase...     

Effects of supplemental nitrate and thermal regime on the nitrogen nutrition of chickpea (Cicer arietinum L.)

Summary Nodulated chickpea plants were grown in pots in a glasshouse programmed to simulate either hot (32.5°C day/18°C night) or warm (25°/18°C) thermal regimes characteristic of those experienced by crops grown in different seasons or locations in the semi-arid tropics. The plants were irrigated with nutrient solution either devoid of inorganic nitrogen or containing 0.71, 1.43 or 2.86 mM nitrate. Increasing concentrations of supplemental nitrate stimulated the rate of dry matter production by vegetative plants in both thermal regimes. Differences between vegetative dry weight of plants given nitrate and those relying exclusively on symbiotic dinitrogen fixation were greatest in the hot regime where the durations of vegetative growth were shorter. However, symbiotically-dependent plants and those given 0.71 mM nitrate continued to produce branches throughout the reproductive period, particularly in the warm regime. As they matured, these plants became more comparable in vegetative stature to those which had received greater concentrations of nitrate and had established final branch numbers earlier (i.e prior to main pod-fill). Potential seed yields were determined primarily by the number of potential reproductive sites (nodes) available (i.e. by the extent of branching) which largely determined the number of seeds harvested. Since final branch numbers in all nitrate treatments were greatest in the warm regime, yields were also larger than those at 32.5°C. The implications of these data for the nitrogen economy of chickpea crops is discussed.One of a series of papers resulting from a collaborative project with the International Crops Research Institute for the Semi-Arid Tropics, India; sponsored by the UK Overseas Development Administration.     

Genetic characterization of the acetohydroxyacid synthase (AHAS) gene responsible for resistance to imidazolinone in chickpea (Cicer arietinum L.)

Key message

A point mutation in the AHAS1 gene leading to resistance to imidazolinone in chickpea was identified. The resistance is inherited as a single gene. A KASP marker targeting the mutation was developed.

Abstract

Weed control in chickpea (Cicer arietinum L.) is challenging due to poor crop competition ability and limited herbicide options. A chickpea genotype with resistance to imidazolinone (IMI) herbicides has been identified, but the genetic inheritance and the mechanism were unknown. In many plant species, resistance to IMI is caused by point mutation(s) in the acetohydroxyacid synthase (AHAS) gene resulting in an amino acid substitution preventing herbicide attachment to the molecule. The main objective of this research was to characterize the resistance to IMI herbicides in chickpea. Two homologous AHAS genes namely AHAS1 and AHAS2 sharing 80 % amino acid sequence similarity were identified in the chickpea genome. Cluster analysis indicated independent grouping of AHAS1 and AHAS2 across legume species. A point mutation in the AHAS1 gene at C675 to T675 resulting in an amino acid substitution from Ala205 to Val205 confers the resistance to IMI in chickpea. A KASP marker targeting the point mutation was developed and effectively predicted the response to IMI herbicides in a recombinant inbred (RI) population of chickpea. The RI population was used in molecular mapping where the major locus for the reaction to IMI herbicide was mapped to chromosome 5. Segregation analysis across an F₂ population and RI population demonstrated that the resistance is inherited as a single gene in a semi-dominant fashion. The simple genetic inheritance and the availability of KASP marker generated in this study would speed up development of chickpea varieties with resistance to IMI herbicides.     

Appearance of different phosphatase forms and phosphorus partitioning in nodules of chickpea (Cicer arietinum L.) during development

Acid and alkaline phosphatase and phytase activities were determined in the bacteroid free fractions of chickpea (Cicer arietinum L.) nodules at 15 days intervals, from 40 days after sowing (DAS) to 85 DAS. In general, the activities and specific activity of both the acid and alkaline phosphatases declined at 55 DAS. Out of the various substrates studied, ATP was the best substrate for both phosphatases. Activities of phosphatases with glucose-6-phosphate and fructose-6-phosphate were low in comparison to these with fructose 1,6 bisphosphate. The efficiency of acid phosphatase for utilizing fructose 1,6 bis phosphate as a substrate increased with nodule development. A fructose 1,6 bis phosphate specific acid phosphatase with elution volume to void volume (Ve/Vo) ratio of around 2.0 was observed in mature nodules (80 DAS). Acid phosphatase at 40 DAS was resolved into two peaks which were eluted at Ve/Vo of about 1.5 and 1.8. However, at 60 DAS the peak with Ve/Vo of 1.5 could not be detected. With ATP as substrate, a high (Ve/Vo of 1.2) and low MM form (Ve/Vo of 2.1) alkaline phosphatases were observed at 40 DAS however at 60 DAS stage only one peak with Ve/Vo of 1.7 was detected. Although, a low activity of acid phytase was observed in nodules at all stages of development but neither alkaline phytase nor phytic acid could be detected. It appears that the nodules acquire inorganic phosphate from the roots. The higher content of water soluble organic phosphorus in mature nodules could be due to the low activities of phosphatases at maturity.     

Isolation and characterization of cDNA clones encoding ADP-glucose pyrophosphorylase (AGPase) large and small subunits from chickpea (Cicer arietinum L.)

Four cDNA clones encoding two large subunits and two small subunits of the starch regulatory enzyme ADP-glucose pyrophosphorylase (AGPase) were isolated from a chickpea (Cicer arietinum L.) stem cDNA library. DNA sequence and Southern blot analyses of these clones, designated CagpL1, CagpL2 (large subunits) and CagpS1 and CagpS2 (small subunits), revealed that these isoforms represented different AGPase large and small subunits. RNA expression analysis indicated that CagpL1 was expressed strongly in leaves with reduced expression in the stem. No detectable expression was observed in seeds and roots. CagpL2 was expressed moderately in seeds followed by weak expression in leaves, stems and roots. Similar analysis showed that CagpS1 and CagpS2 displayed a spatial expression pattern similar to that observed for CagpL2 with the exception that CagpS1 showed a much higher expression in seeds than CagpS2. The spatial expression patterns of these different AGPase subunit sequences indicate that different AGPase isoforms are used to control starch biosynthesis in different organs during chickpea development.