Chromosome-scale pearl millet genomes reveal CLAMT1b as key determinant of strigolactone pattern and Striga susceptibility.

The yield of pearl millet, a resilient cereal crop crucial for African food security, is severely impacted by the root parasitic weed Striga hermonthica, which requires host-released hormones, called strigolactones (SLs), for seed germination. Herein, we identify four SLs present in the Striga-susceptible line SOSAT-C88-P10 (P10) but absent in the resistant 29Aw (Aw). We generate chromosome-scale genome assemblies, including four gapless chromosomes for each line. The Striga-resistant Aw lacks a 0.7 Mb genome segment containing two putative CARLACTONOIC ACID METHYLTRANSFERASE1 (CLAMT1) genes, which may contribute to SL biosynthesis. Functional assays show that P10CLAMT1b produces the SL-biosynthesis intermediate methyl carlactonoate (MeCLA) and that MeCLA is the precursor of P10-specific SLs. Screening a diverse pearl millet panel confirms the pivotal role of the CLAMT1 section for SL diversity and Striga susceptibility. Our results reveal a reason for Striga susceptibility in pearl millet and pave the way for generating resistant lines through marker-assisted breeding or direct genetic modification.

Genome-wide SNPs and candidate genes underlying the genetic variations for protein and amino acids in pearl millet (Pennisetum glaucum) germplasm.

MAIN CONCLUSION: A total of 544 significant marker-trait associations and 286 candidate genes associated with total protein and 18 amino acids were identified. Thirty-three candidate genes were found near the strong marker trait associations (- log10P ≥ 5.5). Pearl millet (Pennisetum glaucum) is largely grown as a subsistence crop in South Asia and sub-Saharan Africa. It serves as a major source of daily protein intake in these regions. Despite its importance, no systematic effort has been made to study the genetic variations of protein and amino acid content in pearl millet germplasm. The present study was undertaken to dissect the global genetic variations of total protein and 18 essential and non-essential amino acids in pearl millet, using a set of 435 K Single Nucleotide Polymorphisms (SNPs) and 161 genotypes of the Pearl Millet Inbred Germplasm Association Panel (PMiGAP). A total of 544 significant marker-trait associations (at P < 0.0001; - log10P ≥ 4) were detected and 23 strong marker-trait associations were identified using Bonferroni's correction method. Forty-eight pleiotropic loci were found in the genome for the studied traits. In total, 286 candidate genes associated with total protein and 18 amino acids were identified. Thirty-three candidate genes were found near strongly associated SNPs. The associated markers and the candidate genes provide an insight into the genetic architecture of the traits studied and are going to be useful in breeding improved pearl millet varieties in the future. Availabilities of improved pearl millet varieties possessing higher protein and amino acid compositions will help combat the rising malnutrition problem via diet.

Consensus genetic linkage map and QTL mapping allow to capture the genomic regions associated with agronomic traits in pearl millet.

MAIN CONCLUSION: A genetic linkage map representing the pearl millet genome was constructed with SNP markers. Major and stable QTL associated with flowering, number of productive tillers, ear head length, and test weight were mapped on chromosomes 1 and 3. Pearl millet (Pennisetum glaucum) is a major cereal and fodder crop in arid and semi-arid regions of Asia and Africa. Agronomic traits are important traits in pearl millet breeding and genetic and environmental factors highly influence them. In the present study, an F9 recombinant inbred line (RIL) population derived from a cross between PT6029 and PT6129 was evaluated for agronomic traits in three environments. Utilizing a genotyping by sequencing approach, a dense genetic map with 993 single nucleotide polymorphism markers covering a total genetic distance of 1035.4 cM was constructed. The average interval between the markers was 1.04 cM, and the seven chromosomes varied from 115.39 to 206.72 cM. Quantitative trait loci (QTL) mapping revealed 35 QTL for seven agronomic traits, and they were distributed on all pearl millet chromosomes. These QTL individually explained 11.35 to 26.71% of the phenotypic variation, with LOD values ranging from 2.74 to 5.80. Notably, four QTL (qDFF1.1, qNPT3.1, qEHL3.1, and qTW1.1) associated with days to fifty percent flowering, the number of productive tillers, ear head length, and test weight were found to be major and stable QTL located on chromosomes 1 and 3. Collectively, our results provide an important base for understanding the genetic architecture of agronomic traits in pearl millet, which is useful for accelerating the genetic gain toward crop improvement.

Transpiration efficiency variations in the pearl millet reference collection PMiGAP.

Transpiration efficiency (TE), the biomass produced per unit of water transpired, is a key trait for crop performance under limited water. As water becomes scarce, increasing TE would contribute to increase crop drought tolerance. This study is a first step to explore pearl millet genotypic variability for TE on a large and representative diversity panel. We analyzed TE on 537 pearl millet genotypes, including inbred lines, test-cross hybrids, and hybrids bred for different agroecological zones. Three lysimeter trials were conducted in 2012, 2013 and 2015, to assess TE both under well-watered and terminal-water stress conditions. We recorded grain yield to assess its relationship with TE. Up to two-fold variation for TE was observed over the accessions used. Mean TE varied between inbred and testcross hybrids, across years and was slightly higher under water stress. TE also differed among hybrids developed for three agroecological zones, being higher in hybrids bred for the wetter zone, underlining the importance of selecting germplasm according to the target area. Environmental conditions triggered large Genotype x Environment (GxE) interactions, although TE showed some high heritability. Transpiration efficiency was the second contributor to grain yield after harvest index, highlighting the importance of integrating it into pearl millet breeding programs. Future research on TE in pearl millet should focus (i) on investigating the causes of its plasticity i.e. the GxE interaction (ii) on studying its genetic basis and its association with other important physiological traits.

Transcriptome analysis of Pennisetum americanum × Pennisetum purpureum and Pennisetum americanum leaves in response to high-phosphorus stress.

Excessive phosphorus (P) levels can disrupt nutrient balance in plants, adversely affecting growth. The molecular responses of Pennisetum species to high phosphorus stress remain poorly understood. This study examined two Pennisetum species, Pennisetum americanum × Pennisetum purpureum and Pennisetum americanum, under varying P concentrations (200, 600 and 1000 µmol·L- 1 KH2PO4) to elucidate transcriptomic alterations under high-P conditions. Our findings revealed that P. americanum exhibited stronger adaption to high-P stress compared to P. americanum× P. purpureum. Both species showed an increase in plant height and leaf P content under elevated P levels, with P. americanum demonstrating greater height and higher P content than P. americanum× P. purpureum. Transcriptomic analysis identified significant up- and down-regulation of key genes (e.g. SAUR, GH3, AHP, PIF4, PYL, GST, GPX, GSR, CAT, SOD1, CHS, ANR, P5CS and PsbO) involved in plant hormone signal transduction, glutathione metabolism, peroxisomes, flavonoid biosynthesis, amino acid biosynthesis and photosynthesis pathways. Compared with P. americanum× P. purpureum, P. americanum has more key genes in the KEGG pathway, and some genes have higher expression levels. These results contribute valuable insights into the molecular mechanisms governing high-P stress in Pennisetum species and offer implications for broader plant stress research.

DNAJ protein gene expansion mechanism in Panicoideae and PgDNAJ functional identification in pearl millet.

KEY MESSAGE: Analyze the evolutionary pattern of DNAJ protein genes in the Panicoideae, including pearl millet, to identify and characterize the biological function of PgDNAJ genes in pearl millet. Global warming has become a major factor threatening food security and human development. It is urgent to analyze the heat-tolerant mechanism of plants and cultivate crops that are adapted to high temperature conditions. The Panicoideae are the second largest subfamily of the Poaceae, widely distributed in warm temperate and tropical regions. Many of these species have been reported to have strong adaptability to high temperature stress, such as pearl millet, foxtail millet and sorghum. The evolutionary differences in DNAJ protein genes among 12 Panicoideae species and 10 other species were identified and analyzed. Among them, 79% of Panicoideae DNAJ protein genes were associated with retrotransposon insertion. Analysis of the DNAJ protein pan-gene family in six pearl millet accessions revealed that the non-core genes contained significantly more TEs than the core genes. By identifying and analyzing the distribution and types of TEs near the DNAJ protein genes, it was found that the insertion of Copia and Gypsy retrotransposons provided the source of expansion for the DNAJ protein genes in the Panicoideae. Based on the analysis of the evolutionary pattern of DNAJ protein genes in Panicoideae, the PgDNAJ was obtained from pearl millet through identification. PgDNAJ reduces the accumulation of reactive oxygen species caused by high temperature by activating ascorbate peroxidase (APX), thereby improving the heat resistance of plants. In summary, these data provide new ideas for mining potential heat-tolerant genes in Panicoideae, and help to improve the heat tolerance of other crops.

Transcriptomic and physiological analysis provide new insight into seed shattering mechanism in Pennisetum alopecuroides 'Liqiu'.

KEY MESSAGE: Through the histological, physiological, and transcriptome-level identification of the abscission zone of Pennisetum alopecuroides 'Liqiu', we explored the structure and the genes related to seed shattering, ultimately revealing the regulatory network of seed shattering in P. alopecuroides. Pennisetum alopecuroides is one of the most representative ornamental grass species of Pennisetum genus. It has unique inflorescence, elegant appearance, and strong stress tolerance. However, the shattering of seeds not only reduces the ornamental effect, but also hinders the seed production. In order to understand the potential mechanisms of seed shattering in P. alopecuroides, we conducted morphological, histological, physiological, and transcriptomic analyses on P. alopecuroides cv. 'Liqiu'. According to histological findings, the seed shattering of 'Liqiu' was determined by the abscission zone at the base of the pedicel. Correlation analysis showed that seed shattering was significantly correlated with cellulase, lignin, auxin, gibberellin, cytokinin and jasmonic acid. Through a combination of histological and physiological analyses, we observed the accumulation of cellulase and lignin during 'Liqiu' seed abscission. We used PacBio full-length transcriptome sequencing (SMRT) combined with next-generation sequencing (NGS) transcriptome technology to improve the transcriptome data of 'Liqiu'. Transcriptomics further identified many differential genes involved in cellulase, lignin and plant hormone-related pathways. This study will provide new insights into the research on the shattering mechanism of P. alopecuroides.

Pearl millet a promising fodder crop for changing climate: a review.

The agricultural sector faces colossal challenges amid environmental changes and a burgeoning human population. In this context, crops must adapt to evolving climatic conditions while meeting increasing production demands. The dairy industry is anticipated to hold the highest value in the agriculture sector in future. The rise in the livestock population is expected to result in an increased demand for fodder feed. Consequently, it is crucial to seek alternative options, as crops demand fewer resources and are resilient to climate change. Pearl millet offers an apposite key to these bottlenecks, as it is a promising climate resilience crop with significantly low energy, water and carbon footprints compared to other crops. Numerous studies have explored its potential as a fodder crop, revealing promising performance. Despite its capabilities, pearl millet has often been overlooked. To date, few efforts have been made to document molecular aspects of fodder-related traits. However, several QTLs and candidate genes related to forage quality have been identified in other fodder crops, which can be harnessed to enhance the forage quality of pearl millet. Lately, excellent genomic resources have been developed in pearl millet allowing deployment of cutting-edge genomics-assisted breeding for achieving a higher rate of genetic gains. This review would facilitate a deeper understanding of various aspects of fodder pearl millet in retrospect along with the future challenges and their solution. This knowledge may pave the way for designing efficient breeding strategies in pearl millet thereby supporting sustainable agriculture and livestock production in a changing world.

Exploration of the pearl millet phospholipase gene family to identify potential candidates for grain quality traits.

BACKGROUND: Phospholipases constitute a diverse category of enzymes responsible for the breakdown of phospholipids. Their involvement in signal transduction with a pivotal role in plant development and stress responses is well documented. RESULTS: In the present investigation, a thorough genome-wide analysis revealed that the pearl millet genome contains at least 44 phospholipase genes distributed across its 7 chromosomes, with chromosome one harbouring the highest number of these genes. The synteny analysis suggested a close genetic relationship of pearl millet phospholipases with that of foxtail millet and sorghum. All identified genes were examined to unravel their gene structures, protein attributes, cis-regulatory elements, and expression patterns in two pearl millet genotypes contrasting for rancidity. All the phospholipases have a high alpha-helix content and distorted regions within the predicted secondary structures. Moreover, many of these enzymes possess binding sites for both metal and non-metal ligands. Additionally, the putative promoter regions associated with these genes exhibit multiple copies of cis-elements specifically responsive to biotic and abiotic stress factors and signaling molecules. The transcriptional profiling of 44 phospholipase genes in two genotypes contrasting for rancidity across six key tissues during pearl millet growth revealed a predominant expression in grains, followed by seed coat and endosperm. Specifically, the genes PgPLD-alpha1-1, PgPLD-alpha1-5, PgPLD-delta1-7a, PgPLA1-II-1a, and PgPLD-delta1-2a exhibited notable expression in grains of both the genotypes while showing negligible expression in the other five tissues. The sequence alignment of putative promoters revealed several variations including SNPs and InDels. These variations resulted in modifications to the corresponding cis-acting elements, forming distinct transcription factor binding sites suggesting the transcriptional-level regulation for these five genes in pearl millet. CONCLUSIONS: The current study utilized a genome-wide computational analysis to characterize the phospholipase gene family in pearl millet. A comprehensive expression profile of 44 phospholipases led to the identification of five grain-specific candidates. This underscores a potential role for at least these five genes in grain quality traits including the regulation of rancidity in pearl millet. Therefore, this study marks the first exploration highlighting the possible impact of phospholipases towards enhancing agronomic traits in pearl millet.

Omics-driven utilization of wild relatives for empowering pre-breeding in pearl millet.

MAIN CONCLUSION: Pearl millet wild relatives harbour novel alleles which could be utilized to broaden genetic base of cultivated species. Genomics-informed pre-breeding is needed to speed up introgression from wild to cultivated gene pool in pearl millet. Rising episodes of intense biotic and abiotic stresses challenge pearl millet production globally. Wild relatives provide a wide spectrum of novel alleles which could address challenges posed by climate change. Pre-breeding holds potential to introgress novel diversity in genetically narrow cultivated Pennisetum glaucum from diverse gene pool. Practical utilization of gene pool diversity remained elusive due to genetic intricacies. Harnessing promising traits from wild pennisetum is limited by lack of information on underlying candidate genes/QTLs. Next-Generation Omics provide vast scope to speed up pre-breeding in pearl millet. Genomic resources generated out of draft genome sequence and improved genome assemblies can be employed to utilize gene bank accessions effectively. The article highlights genetic richness in pearl millet and its utilization with a focus on harnessing next-generation Omics to empower pre-breeding.

Introgression of Δ1-pyrroline-5-carboxylate synthetase (PgP5CS) confers enhanced resistance to abiotic stresses in transgenic tobacco.

Δ1-pyrroline-5-carboxylate synthetase (P5CS) is one of the key regulatory enzymes involved in the proline biosynthetic pathway. Proline acts as an osmoprotectant, molecular chaperone, antioxidant, and regulator of redox homeostasis. The accumulation of proline during stress is believed to confer tolerance in plants. In this study, we cloned the complete CDS of the P5CS from pearl millet (Pennisetum glaucum (L.) R.Br. and transformed into tobacco. Three transgenic tobacco plants with single-copy insertion were analyzed for drought and heat stress tolerance. No difference was observed between transgenic and wild-type (WT) plants when both were grown in normal conditions. However, under heat and drought, transgenic plants have been found to have higher chlorophyll, relative water, and proline content, and lower malondialdehyde (MDA) levels than WT plants. The photosynthetic parameters (stomatal conductance, intracellular CO2 concentration, and transpiration rate) were also observed to be high in transgenic plants under abiotic stress conditions. qRT-PCR analysis revealed that the expression of the transgene in drought and heat conditions was 2-10 and 2-7.5 fold higher than in normal conditions, respectively. Surprisingly, only P5CS was increased under heat stress conditions, indicating the possibility of feedback inhibition. Our results demonstrate the positive role of PgP5CS in enhancing abiotic stress tolerance in tobacco, suggesting its possible use to increase abiotic stress-tolerance in crops for sustained yield under adverse climatic conditions.

Genome-wide characterization of the NBLRR gene family provides evolutionary and functional insights into blast resistance in pearl millet (Cenchrus americanus (L.) Morrone).

MAIN CONCLUSION: The investigation is the first report on genome-wide identification and characterization of NBLRR genes in pearl millet. We have shown the role of gene loss and purifying selection in the divergence of NBLRRs in Poaceae lineage and candidate CaNBLRR genes for resistance to Magnaporthe grisea infection. Plants have evolved multiple integral mechanisms to counteract the pathogens' infection, among which plant immunity through NBLRR (nucleotide-binding site, leucine-rich repeat) genes is at the forefront. The genome-wide mining in pearl millet (Cenchrus americanus (L.) Morrone) revealed 146 CaNBLRRs. The variation in the branch length of NBLRRs showed the dynamic nature of NBLRRs in response to evolving pathogen races. The orthology of NBLRRs showed a predominance of many-to-one orthologs, indicating the divergence of NBLRRs in the pearl millet lineage mainly through gene loss events followed by gene gain through single-copy duplications. Further, the purifying selection (Ka/Ks < 1) shaped the expansion of NBLRRs within the lineage of pear millet and other members of Poaceae. Presence of cis-acting elements, viz. TCA element, G-box, MYB, SARE, ABRE and conserved motifs annotated with P-loop, kinase 2, RNBS-A, RNBS-D, GLPL, MHD, Rx-CC and LRR suggests their putative role in disease resistance and stress regulation. The qRT-PCR analysis in pearl millet lines showing contrasting responses to Magnaporthe grisea infection identified CaNBLRR20, CaNBLRR33, CaNBLRR46 CaNBLRR51, CaNBLRR78 and CaNBLRR146 as putative candidates. Molecular docking showed the involvement of three and two amino acid residues of LRR domains forming hydrogen bonds (histidine, arginine and threonine) and salt bridges (arginine and lysine) with effectors. Whereas 14 and 20 amino acid residues of CaNBLRR78 and CaNBLRR20 showed hydrophobic interactions with 11 and 9 amino acid residues of effectors, Mg.00g064570.m01 and Mg.00g006570.m01, respectively. The present investigation gives a comprehensive overview of CaNBLRRs and paves the foundation for their utility in pearl millet resistance breeding through understanding of host-pathogen interactions.

Abiotic stress tolerance in pearl millet: Unraveling molecular mechanisms via transcriptomics.

Pearl millet (Pennisetum glaucum (L.)) is a vital cereal crop renowned for its ability to thrive in challenging environmental conditions; however, the molecular mechanisms governing its salt stress tolerance remain poorly understood. To address this gap, next-generation RNA sequencing was conducted to compare gene expression patterns in pearl millet seedlings exposed to salt stress with those grown under normal conditions. Our RNA sequencing analysis focused on shoots from 13-day-old pearl millet plants subjected to either salinity stress (150 mmol of NaCl for 3 days) or thermal stress (50°C for 60 s). Of 36,041 genes examined, 17,271 genes with fold changes ranging from 2.2 to 19.6 were successfully identified. Specifically, 2388 genes were differentially upregulated in response to heat stress, whereas 4327 genes were downregulated. Under salt stress conditions, 2013 genes were upregulated and 4221 genes were downregulated. Transcriptomic analysis revealed four common abiotic KEGG pathways that play crucial roles in the response of pearl millet to salt and heat stress: phenylpropanoid biosynthesis, photosynthesis-antenna proteins, photosynthesis, and plant hormone signal transduction. These metabolic pathways are necessary for pearl millet to withstand and adapt to abiotic stresses caused by salt and heat. Moreover, the pearl millet shoot heat stress group showed specific transcriptomics related to KEEG metabolic pathways such as cytochrome P450, cutin, suberine, and wax biosynthesis, zeatin biosynthesis, crocin biosynthesis, ginsenoside biosynthesis, saponin biosynthesis, and biosynthesis of various plant secondary metabolites. In contrast, pearl millet shoots exposed to salinity stress exhibited transcriptomic changes associated with KEEG metabolic pathways related to carbon fixation in photosynthetic organisms, mismatch repair, and nitrogen metabolism. Our findings underscore the remarkable cross-tolerance of pearl millet to simultaneous salt and heat stress, elucidated through the activation of shared abiotic KEGG pathways. This study emphasizes the pivotal role of transcriptomics analysis in unraveling the molecular responses of pearl millet under abiotic stress conditions.

Comprehensive genomic analysis of Bacillus subtilis and Bacillus paralicheniformis associated with the pearl millet panicle reveals their antimicrobial potential against important plant pathogens.

BACKGROUND: Plant microbiome confers versatile functional roles to enhance survival fitness as well as productivity. In the present study two pearl millet panicle microbiome member species Bacillus subtilis PBs 12 and Bacillus paralicheniformis PBl 36 found to have beneficial traits including plant growth promotion and broad-spectrum antifungal activity towards taxonomically diverse plant pathogens. Understanding the genomes will assist in devising a bioformulation for crop protection while exploiting their beneficial functional roles. RESULTS: Two potential firmicute species were isolated from pearl millet panicles. Morphological, biochemical, and molecular characterization revealed their identities as Bacillus subtilis PBs 12 and Bacillus paralicheniformis PBl 36. The seed priming assays revealed the ability of both species to enhance plant growth promotion and seedling vigour index. Invitro assays with PBs 12 and PBl 36 showed the antibiosis effect against taxonomically diverse plant pathogens (Magnaporthe grisea; Sclerotium rolfsii; Fusarium solani; Alternaria alternata; Ganoderma sp.) of crops and multipurpose tree species. The whole genome sequence analysis was performed to unveil the genetic potential of these bacteria for plant protection. The complete genomes of PBs 12 and PBl 36 consist of a single circular chromosome with a size of 4.02 and 4.33 Mb and 4,171 and 4,606 genes, with a G + C content of 43.68 and 45.83%, respectively. Comparative Average Nucleotide Identity (ANI) analysis revealed a close similarity of PBs 12 and PBl 36 with other beneficial strains of B. subtilis and B. paralicheniformis and found distant from B. altitudinis, B. amyloliquefaciens, and B. thuringiensis. Functional annotation revealed a majority of pathway classes of PBs 12 (30) and PBl 36 (29) involved in the biosynthesis of secondary metabolites, polyketides, and non-ribosomal peptides, followed by xenobiotic biodegradation and metabolism (21). Furthermore, 14 genomic regions of PBs 12 and 15 of PBl 36 associated with the synthesis of RiPP (Ribosomally synthesized and post-translationally modified peptides), terpenes, cyclic dipeptides (CDPs), type III polyketide synthases (T3PKSs), sactipeptides, lanthipeptides, siderophores, NRPS (Non-Ribosomal Peptide Synthetase), NRP-metallophone, etc. It was discovered that these areas contain between 25,458 and 33,000 secondary metabolite-coding MiBiG clusters which code for a wide range of products, such as antibiotics. The PCR-based screening for the presence of antimicrobial peptide (cyclic lipopeptide) genes in PBs 12 and 36 confirmed their broad-spectrum antifungal potential with the presence of spoVG, bacA, and srfAA AMP genes, which encode antimicrobial compounds such as subtilin, bacylisin, and surfactin. CONCLUSION: The combined in vitro studies and genome analysis highlighted the antifungal potential of pearl millet panicle-associated Bacillus subtilis PBs12 and Bacillus paralicheniformis PBl36. The genetic ability to synthesize several antimicrobial compounds indicated the industrial value of PBs 12 and PBl 36, which shed light on further studies to establish their action as a biostimulant for crop protection.

Understanding genetic diversity in drought-adaptive hybrid parental lines in pearl millet.

Information on genetic diversity and population structure is helpful to strategize enhancing the genetic base of hybrid parental lines in breeding programs. The present study determined the population structure and genetic diversity of 109 pearl millet hybrid parental lines, known for their better adaptation and performance in drought-prone environments, using 16,472 single nucleotide polymorphic (SNP) markers generated from GBS (genotyping-by-sequencing) platforms. The SNPs were distributed uniformly across the pearl millet genome and showed considerable genetic diversity (0.337), expected heterozygosity (0.334), and observed heterozygosity (0.031). Most of the pairs of lines (78.36%) had Identity-by-State (IBS) based genetic distances of more than 0.3, indicating a significant amount of genetic diversity among the parental lines. Bayesian model-based population stratification, neighbor-joining phylogenetic analysis, and principal coordinate analysis (PCoA) differentiated all hybrid parental lines into two clear-cut major groups, one each for seed parents (B-lines) and pollinators (R-lines). Majority of parental lines sharing common parentages were found grouped in the same cluster. Analysis of molecular variance (AMOVA) revealed 7% of the variation among subpopulations, and 93% of the variation was attributable to within sub-populations. Chromosome 3 had the highest number of LD regions. Genomic LD decay distance was 0.69 Mb and varied across the different chromosomes. Genetic diversity based on 11 agro-morphological and grain quality traits also suggested that the majority of the B- and R-lines were grouped into two major clusters with few overlaps. In addition, the combined analysis of phenotypic and genotypic data showed similarities in the population grouping patterns. The present study revealed the uniqueness of most of the inbred lines, which can be a valuable source of new alleles and help breeders to utilize these inbred lines for the development of hybrids in drought-prone environments.

Glutaredoxin regulation of primary root growth is associated with early drought stress tolerance in pearl millet.

Seedling root traits impact plant establishment under challenging environments. Pearl millet is one of the most heat and drought tolerant cereal crops that provides a vital food source across the sub-Saharan Sahel region. Pearl millet's early root system features a single fast-growing primary root which we hypothesize is an adaptation to the Sahelian climate. Using crop modeling, we demonstrate that early drought stress is an important constraint in agrosystems in the Sahel where pearl millet was domesticated. Furthermore, we show that increased pearl millet primary root growth is correlated with increased early water stress tolerance in field conditions. Genetics including genome-wide association study and quantitative trait loci (QTL) approaches identify genomic regions controlling this key root trait. Combining gene expression data, re-sequencing and re-annotation of one of these genomic regions identified a glutaredoxin-encoding gene PgGRXC9 as the candidate stress resilience root growth regulator. Functional characterization of its closest Arabidopsis homolog AtROXY19 revealed a novel role for this glutaredoxin (GRX) gene clade in regulating cell elongation. In summary, our study suggests a conserved function for GRX genes in conferring root cell elongation and enhancing resilience of pearl millet to its Sahelian environment.

Pearl millet is a staple food for over 90 million people living in regions of Africa and India that typically experience high temperatures and little rainfall. It was domesticated about 4,500 years ago in the Sahel region of West Africa and is one of the most heat and drought tolerant cereal crops worldwide. In most plants, organs known as roots absorb water and essential nutrients from the soil. Young pearl millet plants develop a fast-growing primary root, but it is unclear how this unique feature helps the crop to grow in hot and dry conditions. Using weather data collected from the Sahel over a 20-year period, Fuente, Grondin et al. predicted by modelling that early drought stress is the major factor limiting pearl millet growth and yield in this region. Field experiments found that plants with primary roots that grow faster within soil were better at tolerating early drought than those with slower growing roots. Further work using genetic approaches revealed that a gene known as PgGRXC9 promotes the growth of the primary root. To better understand how this gene works, the team examined a very similar gene in a well-studied model plant known as Arabidopsis. This suggested that PgGRXC9 helps the primary root to grow by stimulating cell elongation within the root. Since it is well adapted to dry conditions, pearl millet is expected to play an important role in helping agriculture adjust to climate change. The findings of Fuente, Grondin et al. may be used by plant breeders to create more resilient and productive varieties of pearl millet.

Unraveling the interplay between root exudates, microbiota, and rhizosheath formation in pearl millet.

BACKGROUND: The rhizosheath, a cohesive soil layer firmly adhering to plant roots, plays a vital role in facilitating water and mineral uptake. In pearl millet, rhizosheath formation is genetically controlled and influenced by root exudates. Here, we investigated the impact of root exudates on the microbiota composition, interactions, and assembly processes, and rhizosheath structure in pearl millet using four distinct lines with contrasting soil aggregation abilities. RESULTS: Utilizing 16S rRNA gene and ITS metabarcoding for microbiota profiling, coupled with FTICR-MS metabonomic analysis of metabolite composition in distinct plant compartments and root exudates, we revealed substantial disparities in microbial diversity and interaction networks. The ß-NTI analysis highlighted bacterial rhizosphere turnover driven primarily by deterministic processes, showcasing prevalent homogeneous selection in root tissue (RT) and root-adhering soil (RAS). Conversely, fungal communities were more influenced by stochastic processes. In bulk soil assembly, a combination of deterministic and stochastic mechanisms shapes composition, with deterministic factors exerting a more pronounced role. Metabolic profiles across shoots, RT, and RAS in different pearl millet lines mirrored their soil aggregation levels, emphasizing the impact of inherent plant traits on microbiota composition and unique metabolic profiles in RT and exudates. Notably, exclusive presence of antimicrobial compounds, including DIMBOA and H-DIMBOA, emerged in root exudates and RT of low aggregation lines. CONCLUSIONS: This research underscores the pivotal influence of root exudates in shaping the root-associated microbiota composition across pearl millet lines, entwined with their soil aggregation capacities. These findings underscore the interconnectedness of root exudates and microbiota, which jointly shape rhizosheath structure, deepening insights into soil-plant-microbe interactions and ecological processes shaping rhizosphere microbial communities. Deciphering plant-microbe interactions and their contribution to soil aggregation and microbiota dynamics holds promise for the advancement of sustainable agricultural strategies. Video Abstract.

Iron and zinc biofortification of rice by synergistic expression of OsNAS2 gene with monocot (Pennisetum glaucum) and dicot (Phaseolus vulgaris) ferritins.

Iron and zinc deficiencies are the most prevalent cause of global hidden hunger. Rice, being one of the most consumed crops worldwide, is suitable to target for Fe and Zn biofortification. In present study, we generated rice transgenic lines to meet the recommended dietary requirement of iron and zinc through endosperm specific expression of dicot (kidney bean) and monocot (pearl millet) Ferritins along with constitutive expression of rice nicotianamine synthase 2 (OsNAS2) gene. Visualization through perls' prussian staining and quantification by ICP-MS showed significant improvement in grain iron content in all the transgenic lines. The transgenic lines expressing any of the three selected gene combinations (PvFerrtin-OsNAS2, feedPgFerrtin-OsNAS2 and foodPgFerritin-OsNAS2), showed the potential to surpass the 30% of the estimated average requirement (13 µg/g Fe and 28 µg/g Zn) proposed for rice in HarvestPlus breeding program. Though the expression of PvFerritin along with OsNAS2 gene in IET10364 (indica) variety showed the best result, providing up to 4.2- and 3.5-fold increase in iron (30.56 µg/g) and zinc (60.1 µg/g) content, respectively; in polished grains compared to non-transgenic control. Thus, the lines developed in our study can be used for further breeding purpose to enhance the iron and zinc content in commercial rice varieties.

Genome-wide association analyses of agronomic traits and Striga hermonthica resistance in pearl millet.

Pearl millet (Pennisetum glaucum [L.] R. Br.) is a nutrient-dense, relatively drought-tolerant cereal crop cultivated in dry regions worldwide. The crop is under-researched, and its grain yield is low (< 0.8 tons ha-1) and stagnant in the major production regions, including Burkina Faso. The low productivity of pearl millet is mainly attributable to a lack of improved varieties, Striga hermonthica [Sh] infestation, downy mildew infection, and recurrent heat and drought stress. Developing high-yielding and Striga-resistant pearl millet varieties that satisfy the farmers' and market needs requires the identification of yield-promoting genes linked to economic traits to facilitate marker-assisted selection and gene pyramiding. The objective of this study was to undertake genome-wide association analyses of agronomic traits and Sh resistance among 150 pearl millet genotypes to identify genetic markers for marker-assisted breeding and trait introgression. The pearl millet genotypes were phenotyped in Sh hotspot fields and screen house conditions. Twenty-nine million single nucleotide polymorphisms (SNPs) initially generated from 345 pearl millet genotypes were filtered, and 256 K SNPs were selected and used in the present study. Phenotypic data were collected on days to flowering, plant height, number of tillers, panicle length, panicle weight, thousand-grain weight, grain weight, number of emerged Striga and area under the Striga number progress curve (ASNPC). Agronomic and Sh parameters were subjected to combined analysis of variance, while genome-wide association analysis was performed on phenotypic and SNPs data. Significant differences (P < 0.001) were detected among the assessed pearl millet genotypes for Sh parameters and agronomic traits. Further, there were significant genotype by Sh interaction for the number of Sh and ASNPC. Twenty-eight SNPs were significantly associated with a low number of emerged Sh located on chromosomes 1, 2, 3, 4, 6, and 7. Four SNPs were associated with days-to-50%-flowering on chromosomes 3, 5, 6, and 7, while five were associated with panicle length on chromosomes 2, 3, and 4. Seven SNPs were linked to thousand-grain weight on chromosomes 2, 3, and 6. The putative SNP markers associated with a low number of emerged Sh and agronomic traits in the assessed genotypes are valuable genomic resources for accelerated breeding and variety deployment of pearl millet with Sh resistance and farmer- and market-preferred agronomic traits.

Improved pearl millet genomes representing the global heterotic pool offer a framework for molecular breeding applications.

High-quality reference genome assemblies, representative of global heterotic patterns, offer an ideal platform to accurately characterize and utilize genetic variation in the primary gene pool of hybrid crops. Here we report three platinum grade de-novo, near gap-free, chromosome-level reference genome assemblies from the active breeding germplasm in pearl millet with a high degree of contiguity, completeness, and accuracy. An improved Tift genome (Tift23D2B1-P1-P5) assembly has a contig N50 ~ 7,000-fold (126 Mb) compared to the previous version and better alignment in centromeric regions. Comparative genome analyses of these three lines clearly demonstrate a high level of collinearity and multiple structural variations, including inversions greater than 1 Mb. Differential genes in improved Tift genome are enriched for serine O-acetyltransferase and glycerol-3-phosphate metabolic process which play an important role in improving the nutritional quality of seed protein and disease resistance in plants, respectively. Multiple marker-trait associations are identified for a range of agronomic traits, including grain yield through genome-wide association study. Improved genome assemblies and marker resources developed in this study provide a comprehensive framework/platform for future applications such as marker-assisted selection of mono/oligogenic traits as well as whole-genome prediction and haplotype-based breeding of complex traits.