Pearl millet (PM) is a nutrient-rich, climate-resilient cereal with potential for low-glycemic food applications. This study examined molecular and microstructural determinants of in vitro starch digestibility in 11 PM genotypes. Variation in starch composition (amylose: 18.27-27.68%; amylopectin: 30.62-51.12%) influenced the distribution of digestible fractions. Rapidly digestible starch ranged from 26.32% (HHB-67 Improved) to 44.74% (PC-701), while slowly digestible starch varied from 14.65% (Pusa-1803) to 27.44% (Chanana Bajri) and was negatively correlated with predicted glycemic index (pGI: 54.63-58.87; r =-0.633). Rheological analysis showed an inverse relationship between peak viscosity and pGI (r =-0.828), suggesting restricted starch gelatinization reduces enzymatic accessibility. X-ray diffraction revealed that higher crystallinity (18.44-26.97%) was also associated with lower pGI (r =-0.788). The Principal Component Analysis grouped Chanana Bajri, PC-701 and 86M94 as structurally compact, slowly digestible types, while Dhodsar Local and Pusa-1803 aligned with higher RDS and pGI, confirming microstructure-governed digestibility variation.

Modern genomic tools can accelerate molecular breeding of underutilized crops such as teff (Eragrostis tef), a gluten-free cereal that is a staple in the Horn of Africa. Here, we report the development of a pangenome comprising chromosome-scale assemblies of 26 teff accessions, including both landraces and improved cultivars, and representing the species’ gene pool. The teff pangenome reveals a highly conserved genome architecture characterized by structural stability across different accessions, with a content of repetitive elements increased by 8% with respect to previous annotations. We describe the features of the pangenome and use it to advance teff biology, targeting seed color, a priority trait for breeding. By defining k-mer-based haplotypes at genomic loci previously known to be associated with the trait, we build a prediction model capable of accurately predicting the color of the seed. Our supervised classification analysis shows that seed color determination cannot be explained by less than four haplotype combinations at loci on chromosomes 4B and 6A. Our results represent a key resource to advance teff breeding and demonstrate that a pangenomic dimension can expand association studies to fully dissect the determination of complex traits.

Tef ( Eragrostis tef ) is an indigenous African cereal that is gaining global attention as a gluten-free “superfood” with high protein, mineral, and fibre contents. However, tef yields are limited by lodging and by losses during harvest owing to its small grain size (150× lighter than wheat). Breeders must also consider a strong cultural preference for white-grained over brown-grained varieties. Tef is relatively understudied with limited “omics” resources. Here, we resequence 220 tef accessions from an Ethiopian diversity collection and also perform multi-locational phenotyping for 25 agronomic and grain traits. Grain metabolome profiling reveals differential accumulation of fatty acids and flavonoids between white and brown grains. k -mer and SNP-based genome-wide association uncover important marker-trait associations, including a significant 70 kb peak for panicle morphology containing the tef orthologue of rice qSH1 —a transcription factor regulating inflorescence morphology in cereals. We also observe a previously unknown relationship between grain size, colour, and fatty acids. These traits are highly associated with retrotransposon insertions in homoeologues of TRANSPARENT TESTA 2 , a known regulator of grain colour. Our study provides valuable resources for tef research and breeding, facilitating the development of improved cultivars with desirable agronomic and nutritional properties.

Lipid-induced digestive resistance could be an affordable management strategy to lower the glycemic amplitude of dietary starch. This study evaluated the influence of fatty acid (FA) composition, chain length, and saturation of five cooking fats-ghee (GH), coconut oil (CO), sunflower oil (SO), mustard oil (MO), and til oil (TO)-on the inherent glycemic potential (IGP) of starches from pearl millet (PM) and rice. Starch-lipid (S-L) complexes were analyzed using in vitro starch hydrolysis kinetics. The inclusion of cooking fats in starches resulted in higher resistant starch (RS) content, which was attributed to the formation of stable S-L structures. Fourier transform infrared spectroscopy and x-ray diffraction revealed that GH and MO-induced complexes exhibited longer and shorter starch molecule assemblies in PM and rice, which ultimately limited the IGP to 57.13% and 58.87%, respectively. Subsequently, the in vitro glucose diffusion assay validated the lesser glucose bioavailability from MO-induced starch complexes in the system, revealing the correlation among chain length and degree of saturation of cooking fats in the context of IGP of dietary starches. Henceforth, by understanding these S-L interactions, newer food prototypes could be designed in the near future.

Oats (Avena sativa L) is a temperate cereal and an important healthy cereal cultivated for food and feed. Therefore, understanding drought responses in oats could significantly impact oat production under harsh climatic conditions. In particular, drought during anthesis (flowering) affects grain filling, quality and yield. Here, we characterised metabolite responses of two Mediterranean oat (Avena sativa L.) cultivars, Flega and Patones, during drought stress at anthesis. In the more drought-tolerant Patones, the developing grains from the top (older) and bottom (younger) spikelets of primary panicle were found to be larger in size in response to drought, suggesting accelerated grain development. Flega showed a more rapid transition to flowering and grain development under drought. The metabolomes of source (sheath, flag leaf, rachis) and sink (developing grains) tissues from Patones showed differential accumulation in fatty acids levels, including α-linolenic acid, sugars and amino acids with drought. Flega showed enhanced energy metabolism in both source and sink tissues. Lower levels of glutathione in source tissues and the accumulation of ophthalmic acid in the grains of Flega were indicators of oxidative stress. Our study revealed two distinct metabolite regulatory patterns in these cultivars during drought at anthesis. In Patones, α-linolenic acid-associated processes may accelerate grain-filling, while in Flega oxidative stress appears to influence traits such as flowering time. Overall, this work provides a first insight into the metabolite regulation in oat's source and sink tissues during anthesis under drought stress.

Arabidopsis (Arabidopsis thaliana) seeds expressing the feedback-insensitive form of cystathionine & gamma;-synthase (AtD-CGS), the key gene of methionine (Met) synthesis, under the control of a seed-specific phaseolin promoter (SSE plants) show a significant increase in Met content. This elevation is accompanied by increased levels of other amino acids (AAs), sugars, total protein, and starch, which are important from a nutritional aspect. Here, we investigated the mechanism behind this phenomenon. Gas chromatography-mass spectrometry (GC-MS) analysis of SSE leaves, siliques, and seeds collected at 3 different developmental stages showed high levels of Met, AAs, and sugars compared to the control plants. A feeding experiment with isotope-labeled AAs showed an increased flux of AAs from nonseed tissues toward the developing seeds of SSE. Transcriptome analysis of leaves and seeds displayed changes in the status of methylation-related genes in SSE plants that were further validated by methylation-sensitive enzymes and colorimetric assay. These results suggest that SSE leaves have higher DNA methylation rates than control plants. This occurrence apparently led to accelerated senescence, together with enhanced monomer synthesis, which further resulted in increased transport of monomers from the leaves toward the seeds. The developing seeds of SSE plants, however, show reduced Met levels and methylation rates. The results provide insights into the role of Met in DNA methylation and gene expression and how Met affects the metabolic profile of the plant. Increasing the level of Met in seeds leads to changing the methylation state in leaves and to higher contents of soluble metabolites in leaves and seeds.

Salinity induced metabolite responses resulted in differential accumulation of flavonoids and antioxidant metabolites in shoots and roots suggesting improved antioxidant capacity in providing salt-adaptive phenotype of tef seedling. Tef [(Eragrostis tef) (Zucc.) Trotter] is an important ‘cash crop’ of Ethiopia grown mainly for human food, and development of elite tef cultivars with better performance is vital to Ethiopian farmers and breeders. Soil salinity is one of the key constraints that affects tef yield in the Ethiopian lowlands and Rift valley where cultivation of tef is limited. Being a minor crop, the responses of tef towards salinity is unknown. Salinity involves physiological and metabolite reprogramming that can have major impact on germination and seedling establishment. Here we evaluate the in vitro effect of NaCl on tef germination and associate this with metabolomic approaches to suggest salt tolerance mechanisms. In this study, 19 tef varieties were screened for NaCl tolerance and were investigated using untargeted metabolomics. Screened tef varieties showed differential germination rates with NaCl treatment varying from < 20 to 100

Grain filling in cereals is complex process that determines the final grain yield and quality. Abiotic stresses can have major impact on grain filling. Oats ( Avena sativa L.) is sensitive to drought which adversely affect yield and productivity. In this study, we characterised the grain filling responses of two Mediterranean oat cultivars Flega and Patones under severe drought. Grains from the top (older) and bottom (younger) spikelets of primary panicle were larger in size in response to drought, particularly in Patones, suggesting accelerated grain development. The metabolomes of source (sheath, flag leaf) and sink (developing grains) tissues were profiled to describe source-sink partitioning. In Patones, the developing grains showed increased sugars and amino acids which indicate accelerated grain filling. These were associated with elevated α-linolenic acid levels in source tissues but decreased in developing grains under drought. There was also a significant decrease in C18 fatty acids (FA) and jasmonates (JA) derivatives in the developing grains which suggested a role for JA signalling in Patones with drought. Flega showed a different response, with accelerated flowering and enhanced energy metabolism in both source and sink organs. The accumulation of ophthalmic acid in grains of Flega and lower levels of reduced glutathione in source tissues suggested greater oxidative stress than Patones under drought may be driving the grain filling phenotype. This study suggests that oats cultivars can use α-linolenic acid-linked signalling or oxidative events influences accelerated grain filling with drought. These could be important traits in developing oat cultivars that maintain yield in drought-prone environments. Highlight The impact on drought in one tolerant and one susceptible oat cultivar was assessed at the grain filling stage. The drought tolerant cultivar, Patones, showed accelerated grain development which could be a strategy to escape drought. Metabolite mapping of flag leaves, sheath and grains of Flega suggested that alpha linolenic acid could be regulating the altered sink-source relationships. The drought susceptible cultivar, Metabolomics shifts in Flega suggested that oxidative stress accelerated flowering.

The emergence of communicable and non-communicable diseases has posed a health challenge for millions of people worldwide and is a major threat to the economic and social development in the coming century. The occurrence of the recent pandemic, SARS-CoV-2, caused by lethal severe acute respiratory syndrome coronavirus 2, is one such example. Rapid research and development of drugs for the treatment and management of these diseases have become an incredibly challenging task for the pharmaceutical industry. Although, substantial attention has been paid to the discovery of therapeutic compounds from natural sources having significant medicinal potential, their synthesis has made a slow progress. Hence, the discovery of new targets by the application of the latest biotechnological and synthetic biology approaches is very much the need of the hour. Polyketides (PKs) and non-ribosomal peptides (NRPs) found in bacteria, fungi and plants are a diverse family of natural products synthesized by two classes of enzymes: polyketide synthases (PKS) and non-ribosomal peptide synthetases (NRPS). These enzymes possess immense biomedical potential due to their simple architecture, catalytic capacity, as well as diversity. With the advent of the latest in-silico and in-vitro strategies, these enzymes and their related metabolic pathways, if targeted, can contribute highly towards the biosynthesis of an array of potentially natural drug leads that have antagonist effects on biopolymers associated with various human diseases. In the face of the rising threat from multidrug-resistant pathogens, this will further open new avenues for the discovery of novel and improved drugs by combining natural and synthetic approaches. This review discusses the relevance of polyketides and non-ribosomal peptides and the improvement strategies for the development of their derivatives and scaffolds, and how they will be beneficial for future bioprospecting and drug discovery.

Tef (Eragrostis tef), is a gluten-free orphan cereal, crop of nutritional and economical significance. Here we used untargeted metabolomics to survey metabolite variation in 14 diverse tef accessions at 15-days post germination. Tef genotypes were classified into four metabolomic groups where variation was linked to flavones and flavonols. Further analysis on white seeded accessions shows variation related to sucrose and important vitamins, nicotinamides (vitamin B3) riboflavin (vitamin B2) and folate (vitamin B9). Coloured seeded accessions showed variation in metabolism related to amino acid and sugars. This study highlights the potential of metabolomics in exploring the nutritional traits in tef.

Tef is a highly nutritious gluten-free Ethiopian cereal with food-feed potential. However, its productivity is affected by lodging, weed infestation, terminal drought, small seed size, and shattering. Following the recent availability of tef genome sequences, we highlight the need to harness the benefits that this underutilised crop offers to improve food security.

Phytophthora capsici is one of the most destructive pathogens causing quick wilt (foot rot) disease in black pepper (Piper nigrum L.) to which no effective resistance has been defined. To better understand the P. nigrum–P. capsici pathosystem, we employed metabolomic approaches based on flow-infusion electrospray–high-resolution mass spectrometry. Changes in the leaf metabolome were assessed in infected and systemic tissues at 24 and 48 hpi. Principal Component Analysis of the derived data indicated that the infected leaves showed a rapid metabolic response by 24 hpi whereas the systemic leaves took 48 hpi to respond to the infection. The major sources of variations between infected leaf and systemic leaf were identified, and enrichment pathway analysis indicated, major shifts in amino acid, tricarboxylic acid cycle, nucleotide and vitamin B6 metabolism upon infection. Moreover, the individual metabolites involved in defensive phytohormone signalling were identified. RT-qPCR analysis of key salicylate and jasmonate biosynthetic genes indicated a transient reduction of expression at 24 hpi but this increased subsequently. Exogenous application of jasmonate and salicylate reduced P. capsici disease symptoms, but this effect was suppressed with the co-application of abscisic acid. The results are consistent with abscisic acid reprogramming, salicylate and jasmonate defences in infected leaves to facilitate the formation of disease. The augmentation of salicylate and jasmonate defences could represent an approach through which quick wilt disease could be controlled in black pepper.

Drought is a major abiotic stress that limits crop productivity and is driving the need to introduce new tolerant crops with better economic yield. Tef (Eragrostis tef) is a neglected (orphan) Ethiopian warm-season annual gluten-free cereal with high nutritional and health benefits. Further, tef is resilient to environmental challenges such as drought, but the adaptive mechanisms remain poorly understood. In this study, metabolic changes associated with drought response in 11 tef accessions were identified using phenomic and metabolomic approaches under controlled conditions. Computerized image analysis of droughted plants indicated reductions in leaf area and green pigments compared with controls. Metabolite profiling based on flow-infusion electrospray-high-resolution mass spectroscopy (FIE-HRMS) showed drought associated changes in flavonoid, phenylpropanoid biosynthesis, sugar metabolism, valine, leucine and isoleucine biosynthesis, and pentose phosphate pathways. Flavonoid associated metabolites and TCA intermediates were lower in the drought group, whereas most of the stress-responsive amino acids and sugars were elevated. Interestingly, after drought treatment, one accession Enatite (Ent) exhibited a significantly higher plant area than the others, and greater accumulation of flavonoids, amino acids (serine and glycine), sugars (ribose, myo-inositol), and fatty acids. The increased accumulation of these metabolites could explain the increased tolerance to drought in Ent compared with other accessions. This is the first time a non-targeted metabolomics approach has been applied in tef, and our results provide a framework for a better understanding of the tef metabolome during drought stress that will help to identify traits to improve this understudied potential crop.

The low level of methionine, an essential sulfur-containing amino acid, limits the nutritional quality of seeds. Two main factors can control the level of protein-bound methionine: the level of free methionine that limits protein accumulation and the methionine residues inside the storage proteins. To reveal the main limiting factor, we generated transgenic Arabidopsis thaliana seed-specific plants expressing the methionine-rich sunflower seed storage (SSA) protein (A1/A2). The contents of protein-bound methionine in the water-soluble protein fraction that includes the SSA in A1/A2 were 5.3- and 10.5-fold, respectively, compared to control, an empty vector (EV). This suggests that free methionine can support this accumulation. To elucidate if the level of free methionine could be increased further in the protein-bound methionine, these lines were crossed with previously characterized plants having higher levels of free methionine in seeds (called SSE). The progenies of the crosses (A1S, A2S) exhibited the highest level of protein-bound methionine, but this level did not differ significantly from A2, suggesting that all the methionine residues of A2 were filled with methionine. It also suggests that the content of methionine residues in the storage proteins is the main limiting factor. The results also proposed that the storage proteins can change their content in response to high levels of free methionine or SSA. This was assumed since the water-soluble protein fraction was highest in A1S/A2S as well as in SSE compared to EV and A1/A2. By using these seeds, we also aimed at gaining more knowledge about the link between high free methionine and the levels of metabolites that usually accumulate during abiotic stresses. This putative connection was derived from a previous analysis of SSE. The results of metabolic profiling showed that the levels of 29 and 20 out of the 56 metabolites were significantly higher in SSE and A1, respectively, that had higher level of free methionine, compared A1S/A2S, which had lower free methionine levels. This suggests a strong link between high free methionine and the accumulation of stress-associated metabolites.

Quinolone synthase from Aegle marmelos (AmQNS) is a Rutacean-specific plant type III polyketide synthase that synthesizes quinolone, acridone, and benzalacetone with therapeutic potential. Simple architecture and broad substrate affinity of AmQNS make it as one of the target enzymes to produce novel structural scaffolds. Another unique feature of AmQNS despite its high similarity to acridone forming type III polyketide synthase from Citrus microcarpa is the variation in the product formation. Hence, to explore the characteristic features of AmQNS, an in-depth sequence and structure-based bioinformatics analyses were performed. Our studies indicated that AmQNS and its nearest homologs have evolved by a series of gene duplication events and strong purifying selection pressure constrains them in the evolutionary process. Additionally, some amino acid alterations were identified in the functionally important region(s), which can contribute to the functional divergence of the enzyme. Prediction of favorable amino acid substitutions will be advantageous in the metabolic engineering of AmQNS for the production of novel compounds. Furthermore, comparative modeling and docking studies were utilized to investigate the structural behavior and small molecule interaction pattern of AmQNS. The observations and results reported here are crucial for advancing our understanding of AmQNS's phylogenetic position, selection pressure, evolvability, interaction pattern and thus providing the foundation for further studies on the structural and reaction mechanism.

Nitric oxide (NO) and elicitors have been found to enhance the defense responses in plants. In this study, the role of NO in the regulation of chalcone synthase (CHS) gene during jasmonic acid (JA)-mediated defense response under wound stress in ginger were investigated. It was observed that NO is responsible for the up-regulation of CHS during wound stress, which further stimulates the JA-mediated stress responses. Confocal microscopy and qRT-PCR were used to study the accumulation of NO and CHS (ZoCHS) transcript levels, respectively, at the wound site. Relative expression of ZoCHS was found to increase by double fold in wounded plants upon treatment with NO and JA. Confocal studies confirmed the accumulation of NO at the wound sites which further triggered the JA-mediated stress responses. Overall results in this work implicate the role of NO in positive modulation of JA defense signaling responses and expression of CHS in ginger during wounding. These findings will help in the development of plants with improved metabolic and defense traits.

Zingiberaceae or ‘ginger family’ is the largest family in the order ‘Zingiberales’ with more than 1300 species in 52 genera, which are mostly distributed throughout Asia, tropical Africa and the native regions of America with their maximum diversity in Southeast Asia. Many of the members are important spice, medicinal or ornamental plants including ginger, turmeric, cardamom and kaempferia. These plants are distinguished for the highly valuable metabolic products, which are synthesised through phenylpropanoid pathway, where type III polyketide synthase is the key enzyme. In our present study, we used sequence, structural and evolutionary approaches to scrutinise the type III polyketide synthase (PKS) repertoire encoded in the Zingiberaceae family. Highly conserved amino acid residues in the sequence alignment and phylogram suggested strong relationships between the type III PKS members of Zingiberaceae. Sequence and structural level investigation of type III PKSs showed a small number of variations in the substrate binding pocket, leading to functional divergence among these PKS members. Molecular evolutionary studies indicate that type III PKSs within Zingiberaceae evolved under strong purifying selection pressure, and positive selections were rarely detected in the family. Structural modelling and protein-small molecule interaction studies on Zingiber officinale PKS ‘a representative from Zingiberaceae’ suggested that the protein is comparatively stable without much disorder and exhibited wide substrate acceptance.