CaLAP1 and CaLAP2 orchestrate anthocyanin biosynthesis in the seed coat of Cicer arietinum.

MAIN CONCLUSION: Our findings shed light on the regulation of anthocyanin and proanthocyanidin biosynthesis in chickpea seed coats. Expression of R2R3-MYB transcription factors CaLAP1 and CaLAP2 enhanced the anthocyanins and proanthocyanidins content in chickpea. The seed coat color is a major economic trait in leguminous crop chickpea (Cicer arietinum). Anthocyanins and proanthocyanidins (PAs) are two classes of flavonoids that mainly contribute to the flower, seed coat and color of Desi chickpea cultivars. Throughout the land plant lineage, the accumulation of anthocyanins and PAs is regulated by MYB and bHLH transcription factors (TFs), which form an MBW (MYB, bHLH, and WD40) complex. Here, we report two R2R3-MYB TFs in chickpea belonging to the anthocyanin-specific subgroup-6, CaLAP1 (Legume Anthocyanin Production 1), and CaLAP2 (Legume Anthocyanin Production 2), which are mainly expressed in the flowers and developmental stages of the seeds. CaLAP1 and CaLAP2 interact with TT8-like CabHLH1 and WD40, forming the MBW complex, and bind to the promoter sequences of anthocyanin- and PA biosynthetic genes CaCHS6, CaDFR2, CaANS, and CaANR, leading to anthocyanins and PA accumulation in the seed coat of chickpea. Moreover, these CaLAPs partially complement the anthocyanin-deficient phenotype in the Arabidopsis thaliana sextuple mutant seedlings. Overexpression of CaLAPs in chickpea resulted in significantly higher expression of anthocyanin and PA biosynthetic genes leading to a darker seed coat color with higher accumulation of anthocyanin and PA. Our findings show that CaLAPs positively modulate anthocyanin and PA content in seed coats, which might influence plant development and resistance to various biotic and abiotic stresses.

Using RNA sequencing to unravel molecular changes underlying the defense response in chickpea induced by Phytophthora medicaginis.

Phytophthora root rot (PRR), caused by Phytophthora medicaginis, is a major soil-borne disease of chickpea in Australia. Breeding for PRR resistance is an effective approach to avoid significant yield loss. Genetic resistance has been identified in cultivated chickpea (Cicer arietinum) and in the wild relative C. echinospermum, with previous studies identifying independent genetic loci associated with each of these sources. However, the molecular mechanisms associated with PRR resistance are not known. RNA sequencing analysis employed in this study identified changes in gene expression in roots of three chickpea genotypes grown hydroponically, early post-infection with P. medicaginis zoospores. Analyses of differentially expressed genes (DEG) identified the activation of a higher number of non-specific R-genes in a PRR-susceptible variety than in the resistant genotypes, suggesting a whole plant resistance response occurring in chickpea against the pathogen. Contrasting molecular changes in signaling profiles, proteolysis and transcription factor pathways were observed in the cultivated and wild Cicer-derived resistant genotypes. DEG patterns supported a hypothesis that increased root elongation and reduced adventitious root formation limit the pathogen entry points in the genotype containing the wild Cicer source of PRR resistance. Candidate resistance genes, including an aquaporin and a maltose transporter in the wild Cicer source and GDSL esterases/lipases in the cultivated source of resistance, were oppositely regulated. Increased knowledge of these genes and pathways will improve our understanding of molecular mechanisms controlling PRR resistance in chickpea, and support the development of elite chickpea varieties through molecular breeding approaches.

Inheritance of Resistance to Chickpea Fusarium Wilt Disease (Fusarium oxysporum f. sp. ciceris Race 2) in a Wide-Cross Cicer arietinum × Cicer reticulatum Mapping Family.

Chickpea (Cicer arietinum) is a major food legume providing high quality nutrition, especially in developing regions. Chickpea wilt (Fusarium oxysporum f. sp. ciceris) causes significant annual losses. Integrated disease management of Fusarium wilt is supported by resistant varieties. Relatively few resistance genes are known so there is value in exploring genetic resources in chickpea wild relatives. This study investigates the inheritance of Fusarium wilt resistance (race 2) in recombinant inbred lines (RILs) from a cross between a cultivated susceptible chickpea variety (Gokce) and a wild resistant Cicer reticulatum line (Kayat-077). RILs, parents, resistant and susceptible tester lines were twice grown in the greenhouse with inoculation and disease symptoms scored. DNA was extracted from dried leaves and individuals were single nucleotide polymorphism (SNP) genotyped. SNPs were placed on the reference chickpea genome and quantitative trait locus (QTL) mapping was performed. Significant QTL regions were examined using PulseDB to identify candidate genes. The results showed the segregation of Fusarium wilt resistance conforming to a single gene inheritance. One significant QTL was found at the start of chromosome 8, containing 138 genes, three of which were disease-resistance candidates for chickpea breeding.

Implementation of theoretical non-photochemical quenching (NPQ(T)) to investigate NPQ of chickpea under drought stress with High-throughput Phenotyping.

Non-photochemical quenching (NPQ) is a protective mechanism for dissipating excess energy generated during photosynthesis in the form of heat. The accelerated relaxation of the NPQ in fluctuating light can lead to an increase in the yield and dry matter productivity of crops. Since the measurement of NPQ is time-consuming and requires specific light conditions, theoretical NPQ (NPQ(T)) was introduced for rapid estimation, which could be suitable for High-throughput Phenotyping. We investigated the potential of NPQ(T) to be used for testing plant genetic resources of chickpea under drought stress with non-invasive High-throughput Phenotyping complemented with yield traits. Besides a high correlation between the hundred-seed-weight and the Estimated Biovolume, significant differences were observed between the two types of chickpea desi and kabuli for Estimated Biovolume and NPQ(T). Desi was able to maintain the Estimated Biovolume significantly better under drought stress. One reason could be the effective dissipation of excess excitation energy in photosystem II, which can be efficiently measured as NPQ(T). Screening of plant genetic resources for photosynthetic performance could take pre-breeding to a higher level and can be implemented in a variety of studies, such as here with drought stress or under fluctuating light in a High-throughput Phenotyping manner using NPQ(T).

Development and evaluation of Fusarium wilt-resistant and high-yielding chickpea advanced breeding line, KCD 11.

Fusarium wilt (FW) is the most severe soil-borne disease of chickpea that causes yield losses up to 100%. To improve FW resistance in JG 11, a high-yielding variety that became susceptible to FW, we used WR 315 as the donor parent and followed the pedigree breeding method. Based on disease resistance and yield performance, four lines were evaluated in station trials during 2017-2018 and 2018-2019 at Kalaburagi, India. Further, two lines, namely, Kalaburagi chickpea desi 5 (KCD 5) and KCD 11, which possesses the resistance allele for a specific single-nucleotide polymorphism marker linked with FW resistance, were evaluated across six different locations (Bidar, Kalaburagi, Raichur, Siruguppa, Bhimarayanagudi and Hagari) over a span of 3 years (2020-2021, 2021-2022 and 2022-2023). KCD 11 exhibited notable performance, showcasing yield advantages of 8.67%, 11.26% and 23.88% over JG 11, and the regional checks Super Annigeri 1 (SA 1) and Annigeri 1, respectively, with enhanced FW resistance in wilt sick plot. Further, KCD 11 outperformed JG 11, SA 1 and Annigeri 1 in multi-location trials conducted across three seasons in the North Eastern Transition Zone, North Eastern Dry Zone, and North Dry Zones of Karnataka. KCD 11 was also tested in trials conducted by All India Coordinated Research Project on chickpea and was also nominated for state varietal trials for its release as a FW-resistant and high-yielding variety. The selected line is anticipated to cater the needs of chickpea growers with the dual advantage of yield increment and disease resistance.

Chickpea (Cicer arietinum) PHO1 family members function redundantly in Pi transport and root nodulation.

Phosphorus (P), a macronutrient, plays key roles in plant growth, development, and yield. Phosphate (Pi) transporters (PHTs) and PHOSPHATE1 (PHO1) are central to Pi acquisition and distribution. Potentially, PHO1 is also involved in signal transduction under low P. The current study was designed to identify and functionally characterize the PHO1 gene family in chickpea (CaPHO1s). Five CaPHO1 genes were identified through a comprehensive genome-wide search. Phylogenetically, CaPHO1s formed two clades, and protein sequence analyses confirmed the presence of conserved domains. CaPHO1s are expressed in different plant organs including root nodules and are induced by Pi-limiting conditions. Functional complementation of atpho1 mutant with three CaPHO1 members, CaPHO1, CaPHO1;like, and CaPHO1;H1, independently demonstrated their role in root to shoot Pi transport, and their redundant functions. To further validate this, we raised independent RNA-interference (RNAi) lines of CaPHO1, CaPHO1;like, and CaPHO1;H1 along with triple mutant line in chickpea. While single gene RNAi lines behaved just like WT, triple knock-down RNAi lines (capho1/like/h1) showed reduced shoot growth and shoot Pi content. Lastly, we showed that CaPHO1s are involved in root nodule development and Pi content. Our findings suggest that CaPHO1 members function redundantly in root to shoot Pi export and root nodule development in chickpea.

Cicer super-pangenome provides insights into species evolution and agronomic trait loci for crop improvement in chickpea.

Chickpea (Cicer arietinum L.)-an important legume crop cultivated in arid and semiarid regions-has limited genetic diversity. Efforts are being undertaken to broaden its diversity by utilizing its wild relatives, which remain largely unexplored. Here, we present the Cicer super-pangenome based on the de novo genome assemblies of eight annual Cicer wild species. We identified 24,827 gene families, including 14,748 core, 2,958 softcore, 6,212 dispensable and 909 species-specific gene families. The dispensable genome was enriched for genes related to key agronomic traits. Structural variations between cultivated and wild genomes were used to construct a graph-based genome, revealing variations in genes affecting traits such as flowering time, vernalization and disease resistance. These variations will facilitate the transfer of valuable traits from wild Cicer species into elite chickpea varieties through marker-assisted selection or gene-editing. This study offers valuable insights into the genetic diversity and potential avenues for crop improvement in chickpea.

Uncovering DNA methylation landscapes to decipher evolutionary footprints of phenotypic diversity in chickpea.

Genetic diversity and environmental factors are long believed to be the dominant contributors to phenotypic diversity in crop plants. However, it has been recently established that, besides genetic variation, epigenetic variation, especially variation in DNA methylation, plays a significant role in determining phenotypic diversity in crop plants. Therefore, assessing DNA methylation diversity in crop plants becomes vital, especially in the case of crops like chickpea, which has a narrow genetic base. Thus, in the present study, we employed whole-genome bisulfite sequencing to assess DNA methylation diversity in wild and cultivated (desi and kabuli) chickpea. This revealed extensive DNA methylation diversity in both wild and cultivated chickpea. Interestingly, the methylation diversity was found to be significantly higher than genetic diversity, suggesting its potential role in providing vital phenotypic diversity for the evolution and domestication of the Cicer gene pool. The phylogeny based on DNA methylation variation also indicates a potential complementary role of DNA methylation variation in addition to DNA sequence variation in shaping chickpea evolution. Besides, the study also identified diverse epi-alleles of many previously known genes of agronomic importance. The Cicer MethVarMap database developed in this study enables researchers to readily visualize methylation variation within the genes and genomic regions of their interest (http://223.31.159.7/cicer/public/). Therefore, epigenetic variation like DNA methylation variation can potentially explain the paradox of high phenotypic diversity despite the narrow genetic base in chickpea and can potentially be employed for crop improvement.

Meta QTL analysis for dissecting abiotic stress tolerance in chickpea.

BACKGROUND: Chickpea is prone to many abiotic stresses such as heat, drought, salinity, etc. which cause severe loss in yield. Tolerance towards these stresses is quantitative in nature and many studies have been done to map the loci influencing these traits in different populations using different markers. This study is an attempt to meta-analyse those reported loci projected over a high-density consensus map to provide a more accurate information on the regions influencing heat, drought, cold and salinity tolerance in chickpea. RESULTS: A meta-analysis of QTL reported to be responsible for tolerance to drought, heat, cold and salinity stress tolerance in chickpeas was done. A total of 1512 QTL responsible for the concerned abiotic stress tolerance were collected from literature, of which 1189 were projected on a chickpea consensus genetic map. The QTL meta-analysis predicted 59 MQTL spread over all 8 chromosomes, responsible for these 4 kinds of abiotic stress tolerance in chickpea. The physical locations of 23 MQTL were validated by various marker-trait associations and genome-wide association studies. Out of these reported MQTL, CaMQAST1.1, CaMQAST4.1, CaMQAST4.4, CaMQAST7.8, and CaMQAST8.2 were suggested to be useful for different breeding approaches as they were responsible for high per cent variance explained (PVE), had small intervals and encompassed a large number of originally reported QTL. Many putative candidate genes that might be responsible for directly or indirectly conferring abiotic stress tolerance were identified in the region covered by 4 major MQTL- CaMQAST1.1, CaMQAST4.4, CaMQAST7.7, and CaMQAST6.4, such as heat shock proteins, auxin and gibberellin response factors, etc. CONCLUSION: The results of this study should be useful for the breeders and researchers to develop new chickpea varieties which are tolerant to drought, heat, cold, and salinity stresses.

Novel insights into drought-induced regulation of ribosomal genes through DNA methylation in chickpea.

Modifications within the epigenome of an organism in response to external environmental conditions allow it to withstand the hostile stress factors. Drought in chickpea is a severely limiting abiotic stress factor which is known to cause huge yield loss. To analyse the methylome of chickpea in response to drought stress conditions and how it affects gene expression, we performed whole-genome bisulfite sequencing (WGBS) and RNA-seq of two chickpea genotypes which contrast for drought tolerance. It was observed that the mCHH was most variable under drought stress and the drought tolerant (DT) genotype exhibited substantial genome-wide hypomethylation as compared to the drought sensitive (DS) genotype. Specifically, there was substantial difference in gene expression and methylation for the ribosomal genes for the tolerant and sensitive genotypes. The differential expression of these genes was in complete agreement with earlier reported transcriptomes in chickpea. Many of these genes were hypomethylated (q < 0.01) and downregulated under drought stress (p < 0.01) in the sensitive genotype. The gene RPS6 (ribosomal protein small subunit) was found to be downregulated and hypomethylated in the drought sensitive genotype which could possibly lead to reduced ribosomal biosynthesis. This study provides novel insights into regulation of drought-responsive genes in chickpea.

MicroRNA164e suppresses NAC100 transcription factor-mediated synthesis of seed storage proteins in chickpea.

Development of protein-enriched chickpea varieties necessitates an understanding of specific genes and key regulatory circuits that govern the synthesis of seed storage proteins (SSPs). Here, we demonstrated the novel involvement of Ca-miR164e-CaNAC100 in regulating SSP synthesis in chickpea. Ca-miRNA164e was significantly decreased during seed maturation, especially in high-protein accessions. The miRNA was found to directly target the transactivation conferring C-terminal region of a nuclear-localized transcription factor, CaNAC100 as revealed using RNA ligase-mediated-rapid amplification of cDNA ends and target mimic assays. The functional role of CaNAC100 was demonstrated through seed-specific overexpression (NACOE) resulting in significantly augmented seed protein content (SPC) consequential to increased SSP transcription. Further, NACOE lines displayed conspicuously enhanced seed weight but reduced numbers and yield. Conversely, a downregulation of CaNAC100 and SSP transcripts was evident in seed-specific overexpression lines of Ca-miR164e that culminated in significantly lowered SPC. CaNAC100 was additionally demonstrated to transactivate the SSP-encoding genes by directly binding to their promoters as demonstrated using electrophoretic mobility shift and dual-luciferase reporter assays. Taken together, our study for the first time established a distinct role of CaNAC100 in positively influencing SSP synthesis and its critical regulation by CamiR164e, thereby serving as an understanding that can be utilized for developing SPC-rich chickpea varieties.

Greater ecophysiological stress tolerance in the core environment than in extreme environments of wild chickpea (Cicer reticulatum).

Global climate change and land use change underlie a need to develop new crop breeding strategies, and crop wild relatives (CWR) have become an important potential source of new genetic material to improve breeding efforts. Many recent approaches assume adaptive trait variation increases towards the relative environmental extremes of a species range, potentially missing valuable trait variation in more moderate or typical climates. Here, we leveraged distinct genotypes of wild chickpea (Cicer reticulatum) that differ in their relative climates from moderate to more extreme and perform targeted assessments of drought and heat tolerance. We found significance variation in ecophysiological function and stress tolerance between genotypes but contrary to expectations and current paradigms, it was individuals from more moderate climates that exhibited greater capacity for stress tolerance than individuals from warmer and drier climates. These results indicate that wild germplasm collection efforts to identify adaptive variation should include the full range of environmental conditions and habitats instead of only environmental extremes, and that doing so may significantly enhance the success of breeding programs broadly.

Evaluation of different Kabuli chickpea genotypes against Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) in relation to biotic and abiotic factors.

Background: The chickpea pod borer Helicoverpa armigera (Hübner) is a significant insect pest of chickpea crops, causing substantial global losses. Methods: Field experiments were conducted in Central Punjab, Pakistan, to investigate the impact of biotic and abiotic factors on pod borer population dynamics and infestation in nine kabuli chickpea genotypes during two cropping seasons (2020-2021 and 2021-2022). The crops were sown in November in both years, with row-to-row and plant-to-plant distances of 30 and 15 cm, respectively, following a randomized complete block design (RCBD). Results: Results showed a significant difference among the tested genotypes in trichome density, pod wall thickness, and leaf chlorophyll contents. Significantly lower larval population (0.85 and 1.10 larvae per plant) and percent damage (10.65% and 14.25%) were observed in genotype Noor-2019 during 2020-2021 and 2021-2022, respectively. Pod trichome density, pod wall thickness, and chlorophyll content of leaves also showed significant variation among the tested genotypes. Pod trichome density and pod wall thickness correlated negatively with larval infestation, while chlorophyll content in leaves showed a positive correlation. Additionally, the larval population positively correlated with minimum and maximum temperatures, while relative humidity negatively correlated with the larval population. Study results explore natural enemies as potential biological control agents and reduce reliance on chemical pesticides.

Effect of drought stress on the expression pattern of genes involved in ABA biosynthesis in Desi-type chickpea (Cicer arietinum L.).

BACKGROUND: The behavior of Abscisic acid (ABA) as a stress phytohormone may be involved in mechanisms leading to tolerance and survival in adverse environmental conditions such as drought stress. METHODS: Here, we evaluated ABA-mediated responses at physio-biochemical and molecular levels in drought-stressed seedlings of two different Desi-type chickpea genotypes (10 as a tolerant genotype and 247 as a sensitive one). RESULTS: Under drought stress, two chickpea genotypes showed a decrease in their relative water content (RWC), and the intense decrease was related to the sensitive genotype (73.9%) in severe stress. Hydrogen peroxide (H2O2) and malondialdehyde (MDA) concomitant with the severity of stress increased in genotypes and the higher increase was in the sensitive genotype (5.8-fold and 3.43-fold, respectively). In the tolerant genotype, the enhanced accumulation of total phenolic content (1.75-fold) and radical scavenging action, based on 1,1-diphenyl-2-picrylhydrazyl test (DPPH), (1.69-fold) were simultaneous with ABA accumulation (1.53-fold). In the tolerant genotype, transcriptional analysis presented upregulation of Zeaxanthin epoxidase (ZEP) (1.35-fold), 9-cis-epoxycarotenoid dioxygenase (NCED) (5.16-fold), and Abscisic aldehyde oxidase (AAO) (1.52-fold compared to control conditions) genes in severe stress in comparison with mild stress. The sensitive genotype had a declining trend in total chlorophyll (up to 70%) and carotenoid contents (36%). The main conclusion to be drawn from this investigation is that ABA with its regulatory effects can affect drought tolerance mechanisms to alleviate adverse effects of unsatisfactory environmental conditions. CONCLUSIONS: In this paper, we tried to indicate that drought stress induces overexpression of genes triggering ABA-mediated drought responses simultaneously in two genotypes while more increment expression was related to the tolerant genotype. At first thought, it seems that the tolerant genotype compared to the sensitive genotype has a genetically inherent ability to cope with and drop adverse effects of drought stress through over-accumulation of ABA as drought.

Delineation of genes for a major QTL governing heat stress tolerance in chickpea.

Chickpea (Cicer arietinum) is a cool season grain legume experiencing severe yield loss during heat stress due to the intensifying climate changes and its associated gradual increase of mean temperature. Hence, understanding the genetic architecture regulating heat stress tolerance has emerged as an important trait to be addressed for enhancing yield and productivity of chickpea under heat stress. The present study is intended to identify the major genomic region(s) governing heat stress tolerance in chickpea. For this, an integrated genomics-assisted breeding strategy involving NGS-based high-resolution QTL-seq assay, QTL region-specific association analysis and molecular haplotyping was deployed in a population of 206 mapping individuals and a diversity panel of 217 germplasm accessions of chickpea. This combinatorial strategy delineated a major 156.8 kb QTL genomic region, which was subsequently narrowed-down to a functional candidate gene CaHSFA5 and its natural alleles associated strongly with heat stress tolerance in chickpea. Superior natural alleles and haplotypes delineated from the CaHSFA5 gene have functional significance in regulating heat stress tolerance in chickpea. Histochemical staining, interaction studies along with differential expression profiling of CaHSFA5 and ROS scavenging genes suggest a cross talk between CaHSFA5 with ROS homeostasis pertaining to heat stress tolerance in chickpea. Heterologous gene expression followed by heat stress screening further validated the functional significance of CaHSFA5 for heat stress tolerance. The salient outcomes obtained here can have potential to accelerate multiple translational genomic analysis including marker-assisted breeding and gene editing in order to develop high-yielding heat stress tolerant chickpea varieties.

Genome-wide identification and analysis of SPL gene family in chickpea (Cicer arietinum L.).

A transcription factor in plants encodes SQUAMOSA promoter binding protein-like (SPL) serves a broad spectrum of important roles for the plant, like, growth, flowering, and signal transduction. A gene family that encodes SPL proteins is documented in various model plant species, including Arabidopsis thaliana and Oryza sativa. Chickpea (Cicer arietinum), a leguminous crop, has not been thoroughly explored with regard to the SPL protein-encoding gene family. Chickpea SPL family genes were located and characterized computationally using a genomic database. Gene data of chickpea were obtained from the phytozome repository and was examined using bioinformatics methods. For investigating the possible roles of SPLs in chickpea, genome-wide characterization, expression, as well as structural analyses of this SPL gene family were performed. Cicer arietinum genome had 19 SPL genes, whereas, according to phylogenetic analysis, the SPLs in chickpea are segregated among four categories: Group-I has 2 introns, Group-II and IV have 1-2 introns (except CaSPL13 and CaSPL15 having 3 introns), and Group-III has 9 introns (except CaSPL1 and CaSPL11 with 1 and 8 introns, respectively). The SBP domain revealed that SPL proteins featured two zinc-binding sites, i.e., C3H and C2HC and one nuclear localization signal. All CaSPL proteins are found to contain highly conserved motifs, i.e., Motifs 1, 2, and 4, except CaSPL10 in which Motifs 1 and 4 were absent. Following analysis, it was found that Motifs 1 and 2 of the chickpea SBP domain are Zinc finger motifs, and Motif 4 includes a nuclear localization signal. All pairs of CaSPL paralogs developed by purifying selection. The CaSPL promoter investigation discovered cis-elements that are responsive to stress, light, and phytohormones. Examination of their expression patterns highlighted major CaSPLs to be evinced primarily among younger pods and flowers. Indicating their involvement in the plant's growth as well as development, along with their capacity to react as per different situations by handling the regulation of target gene's expression, several CaSPL genes are also expressed under certain stress conditions, namely, cold, salt, and drought. The majority of the CaSPL genes are widely expressed and play crucial roles in terms of the plant's growth, development, and responses to the environmental-stress conditions. Our work provides extensive insight into the gene family CaSPL, which might facilitate further studies related to the evolution and functions of the SPL genes for chickpea and other plant species.

Arbuscular mycorrhizae reduced arsenic induced oxidative stress by coordinating nutrient uptake and proline-glutathione levels in Cicer arietinum L. (chickpea).

Accumulation of Arsenic (As) generates oxidative stress by reducing nutrients availability in plants. Arbuscular mycorrhizal (AM) symbiosis can impart metalloid tolerance in plants by enhancing the synthesis of sulfur (S)-rich peptides (glutathione- GSH) and low-molecular-weight nitrogenous (N) osmolytes (proline- Pro). The present study, therefore investigated the efficiency of 3 AM fungal species (Rhizoglomus intraradices-Ri, Funneliformis mosseae -Fm and Claroideoglomus claroideum- Cc) in imparting As (arsenate-AsV -40 at 60 mg kg-1 and arsenite- AsIII at 5 and 10 mg kg-1) tolerance in two Cicer arietinum (chickpea) genotypes (HC 3 and C 235). As induced significantly higher negative impacts in roots than shoots, which was in accordance with proportionately higher reactive oxygen species (ROS) in the former, with AsIII more toxic than AsV. Mycorrhizal symbiosis overcame oxidative stress by providing the host plants with necessary nutrients (P, N, and S) through enhanced microbial enzyme activities (MEAs) in soil, which increased the synthesis of Pro and GSH and established a redox balance in the two genotypes. This coordination between nutrient status, Pro-GSH levels, and antioxidant defense was stronger in HC 3 than C 235 due to its higher responsiveness to the three AM species. However, Ri was most beneficial in inducing redox homeostasis, followed by Fm and Cc, since the Cicer arietinum-Ri combination displayed the maximum ability to boost antioxidant defense mechanisms and establish a coordination with Pro synthesis. Thus, the results highlighted the importance of selecting specific chickpea genotypes having an ability to establish effective mycorrhizal symbiosis for imparting As stress tolerance.

Novel Alleles from Cicer reticulatum L. for Genetic Improvement of Cultivated Chickpeas Identified through Genome Wide Association Analysis.

The availability of wild chickpea (Cicer reticulatum L.) accessions has the potential to be used for the improvement of important traits in cultivated chickpeas. The main objectives of this study were to evaluate the phenotypic and genetic variations of chickpea progeny derived from interspecific crosses between C. arietinum and C. reticulatum, and to establish the association between single nucleotide polymorphism (SNP) markers and a series of important agronomic traits in chickpea. A total of 486 lines derived from interspecific crosses between C. arietinum (CDC Leader) and 20 accessions of C. reticulatum were evaluated at different locations in Saskatchewan, Canada in 2017 and 2018. Significant variations were observed for seed weight per plant, number of seeds per plant, thousand seed weight, and plant biomass. Path coefficient analysis showed significant positive direct effects of the number of seeds per plant, thousand seed weight, and biomass on the total seed weight. Cluster analysis based on the agronomic traits generated six groups that allowed the identification of potential heterotic groups within the interspecific lines for yield improvement and resistance to ascochyta blight disease. Genotyping of the 381 interspecific lines using a modified genotyping by sequencing (tGBS) generated a total of 14,591 SNPs. Neighbour-joining cluster analysis using the SNP data grouped the lines into 20 clusters. The genome wide association analysis identified 51 SNPs that had significant associations with different traits. Several candidate genes associated with early flowering and yield components were identified. The candidate genes and the significant SNP markers associated with different traits have a potential to aid the trait introgression in the breeding program.

Genetic Analysis of Partially Resistant and Susceptible Chickpea Cultivars in Response to Ascochyta rabiei Infection.

The molecular mechanism involved in chickpea (Cicer arietinum L.) resistance to the necrotrophic fungal pathogen Ascochyta rabiei is not well documented. A. rabiei infection can cause severe damage in chickpea, resulting in significant economic losses. Understanding the resistance mechanism against ascochyta blight can help to define strategies to develop resistant cultivars. In this study, differentially expressed genes from two partially resistant cultivars (CDC Corinne and CDC Luna) and a susceptible cultivar (ICCV 96029) to ascochyta blight were identified in the early stages (24, 48 and 72 h) of A. rabiei infection using RNA-seq. Altogether, 3073 genes were differentially expressed in response to A. rabiei infection across different time points and cultivars. A larger number of differentially expressed genes (DEGs) were found in CDC Corinne and CDC Luna than in ICCV 96029. Various transcription factors including ERF, WRKY, bHLH and MYB were differentially expressed in response to A. rabiei infection. Genes involved in pathogen detection and immune signalings such as receptor-like kinases (RLKs), Leucine-Rich Repeat (LRR)-RLKs, and genes associated with the post-infection defence response were differentially expressed among the cultivars. GO functional enrichment and pathway analysis of the DEGs suggested that the biological processes such as metabolic process, response to stimulus and catalytic activity were overrepresented in both resistant and susceptible chickpea cultivars. The expression patterns of eight randomly selected genes revealed by RNA-seq were confirmed by quantitative PCR (qPCR) analysis. The results provide insights into the complex molecular mechanism of the chickpea defence in response to the A. rabiei infection.

Evaluation of biocontrol efficacy of rhizosphere dwelling bacteria for management of Fusarium wilt and Botrytis gray mold of chickpea.

BACKGROUND: Chickpea (Cicer arietinum L.) production is affected by many biotic factors, among them Fusarium wilt caused by Fusarium oxysporum f. sp. ciceri and Botrytis gray mold caused by Botrytis cinerea led to severe losses. As fungicide application is not advisable, biological management is the best alternative for plant protection. The rhizosphere-dwelling antagonistic bacteria are one of the important successful alternative strategy to manage these diseases of chickpea. Rhizosphere dwelling bacteria serve as biocontrol agents by different mechanisms like producing antibiotics, different enzymes, siderophores against pathogens and thereby reducing the growth of pathogens. RESULTS: The present study aimed to isolate rhizospheric bacteria from the soils of different chickpea fields to evaluate biocontrol efficacy of the isolated bacteria to manage Fusarium wilt and Botrytis gray mold in chickpea. A total of 67 bacteria were isolated from chickpea rhizosphere from Bundelkhand region of India. Study revealed the isolated bacteria could reduce the Fusarium oxysporum f. sp. ciceris and Botrytis cinerea infection in chickpea between 17.29 and 75.29%. After screening of all the bacteria for their biocontrol efficacy, 13 most promising bacterial isolates were considered for further study out of which, three bacterial isolates (15d, 9c and 14a) have shown the maximum in vitro antagonistic effects against Fusarium oxysporum f. sp. ciceri and Botrytis cinerea comparable to in vivo effects. However, Isolate (15d) showed highest 87.5% and 82.69% reduction in disease against Fusarium wilt and Botrytis gray mold respectively, under pot condition. Three most potential isolates were characterized at molecular level using 16S rRNA gene and found to be Priestia megaterium (9c and 14a) and Serratia marcescens (15d). CONCLUSION: This study identified two native biocontrol agents Priestia megaterium and Serratia marcescens from the rhizospheric soils of Bundelkhand region of India for control of Fusarium wilt, Botrytis gray mold. In future, efforts should be made to further validate the biocontrol agents in conjugation with nanomaterials for enhancing the synergistic effects in managing the fungal diseases in chickpea. This study will definitely enhance our understanding of these bioagents, and to increase their performance by developing effective formulations, application methods, and integrated strategies.