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Background Gamma ray induced mutation breeding has emerged as an excellent method for expedited development of improved varieties of rice, a staple food for more than half the world's population. However, the assessment of radiation induced variations are primarily phenotypic in nature. In this direction, evaluation of the metabolic signature of bio-active ingredients, which confer beneficial properties to rice, could be employed as a tool to select varieties which not only retain the health benefits of the parent variety but also exhibit better agronomic traits. The present study was, therefore, aimed at evaluating the metabolomic changes in the mutants of Gathuwan, an indigenous Indian rice with immunomodulatory properties. The mutant varieties were developed through gamma irradiation, and liquid chromatography-tandem mass spectrometry (LC–MS/MS)-based metabolic profiling was performed. Results A total of 274 differentially expressed compounds were identified among Gathuwan and four of its mutants (mutant 6, mutant 7, mutant 8 and mutant 12), indicating that gamma irradiation induced stable metabolic alterations. Significant differences were observed in the phytochemical composition of mutants relative to the parent, emphasizing the importance of metabolic screening in functional rice breeding. Cluster analysis and phytochemical profiling revealed that mutant 6 was metabolically closest to the parent variety. Additionally, distinct phytohormonal variations among the mutants were observed which may account for the phenotypic differences in growth and development. Conclusions Our study demonstrates that radiation-induced improvement in agronomic traits are accompanied by distinct alterations in phytochemical and phytohormonal profiles in stable rice mutants. These metabolic changes support the functional potential of the mutants and provide insights into the biological mechanisms underlying their traits. Among the mutants, mutant 6 emerges as a promising candidate due to its similarity to the parent in metabolite composition. Therefore, inclusion of metabolomic profiling as a selection criterion offers a powerful tool to identify robust and functionally superior rice varieties.

Rice production needs to be sustained in the coming decades, as the changeable climatic conditions are becoming more conducive to disease outbreaks. The majority of rice diseases cause enormous economic damage and yield instability. Among them, rice blast caused by Magnaportheoryzae is a serious fungal disease and is considered one of the major threats to world rice production. This pathogen can infect the above-ground tissues of rice plants at any growth stage and causes complete crop failure under favorable conditions. Therefore, management of blast disease is essentially required to sustain global food production. When looking at the drawback of chemical management strategy, the development of durable, resistant varieties is one of the most sustainable, economic, and environment-friendly approaches to counter the outbreaks of rice blasts. Interestingly, several blast-resistant rice cultivars have been developed with the help of breeding and biotechnological methods. In addition, 146 R genes have been identified, and 37 among them have been molecularly characterized to date. Further, more than 500 loci have been identified for blast resistance which enhances the resources for developing blast resistance through marker-assisted selection (MAS), marker-assisted backcross breeding (MABB), and genome editing tools. Apart from these, a better understanding of rice blast pathogens, the infection process of the pathogen, and the genetics of the immune response of the host plant are very important for the effective management of the blast disease. Further, high throughput phenotyping and disease screening protocols have played significant roles in easy comprehension of the mechanism of disease spread. The present review critically emphasizes the pathogenesis, pathogenomics, screening techniques, traditional and molecular breeding approaches, and transgenic and genome editing tools to develop a broad spectrum and durable resistance against blast disease in rice. The updated and comprehensive information presented in this review would be definitely helpful for the researchers, breeders, and students in the planning and execution of a resistance breeding program in rice against this pathogen.

The recent advancements in forward genetics have expanded the applications of mutation techniques in advanced genetics and genomics, ahead of direct use in breeding programs. The advent of next-generation sequencing (NGS) has enabled easy identification and mapping of causal mutations within a short period and at relatively low cost. Identifying the genetic mutations and genes that underlie phenotypic changes is essential for understanding a wide variety of biological functions. To accelerate the mutation mapping for crop improvement, several high-throughput and novel NGS based forward genetic approaches have been developed and applied in various crops. These techniques are highly efficient in crop plants, as it is relatively easy to grow and screen thousands of individuals. These approaches have improved the resolution in quantitative trait loci (QTL) position/point mutations and assisted in determining the functional causative variations in genes. To be successful in the interpretation of NGS data, bioinformatics computational methods are critical elements in delivering accurate assembly, alignment, and variant detection. Numerous bioinformatics tools/pipelines have been developed for such analysis. This article intends to review the recent advances in NGS based forward genetic approaches to identify and map the causal mutations in the crop genomes. The article also highlights the available bioinformatics tools/pipelines for reducing the complexity of NGS data and delivering the concluding outcomes.

Nitrogen inputs for sustainable crop production for a growing population require the enhancement of biological nitrogen fixation. Efforts to increase biological nitrogen fixation include bioprospecting for more effective nitrogen-fixing bacteria. As bacterial nitrogenases are extremely sensitive to oxygen, most primary isolation methods rely on the use of semisolid agar or broth to limit oxygen exposure. Without physical separation, only the most competitive strains are obtained. The distance between strains provided by plating on solid media in reduced oxygen environments has been found to increase the diversity of culturable potential diazotrophic bacteria. To obtain diverse nitrogen-fixing isolates from natural grasslands, we plated soil suspensions from 27 samples onto solid nitrogen-free agar and incubated them under atmospheric and oxygen-reducing conditions. Putative nitrogen fixers were confirmed by subculturing in liquid nitrogen-free media and PCR amplification of the nifH genes. Streaking of the 432 isolates on nitrogen-rich R2A revealed many cocultures. In most cases, only one community member then grew on NFA, indicating the coexistence of nonfixers in coculture with fixers when growing under nitrogen-limited conditions. To exclude isolates able to scavenge residual nitrogen, such as that from vitamins, we used a stringent nitrogen-free medium containing only 6.42 μmol/L total nitrogen and recultured them in a nitrogen-depleted atmosphere. Surprisingly, PCR amplification of nifH using various primer pairs yielded amplicons from only 17% of the 442 isolates. The majority of the nifH PCR-negative isolates were Bacillus and Streptomyces. It is unclear whether these isolates have highly effective uptake systems or nitrogen reduction systems that are not closely aligned with known nitrogenase families. We advise caution in determining the nitrogen fixation ability of plants from growth on nitrogen-free media, even where the total nitrogen is very limited.

Mutation breeding offers a simple, fast and efficient way to rectify major defects without altering their original identity. The present study deployed radiation (gamma rays @ 300Gy)-induced mutation breeding for the improvement and revival of three traditional rice landraces, viz., Samundchini, Vishnubhog and Jhilli. Among the various putative mutants identified in the M2 generation, only three, ten and five rice mutants of Samundchini, Vishnubhog and Jhilli, respectively, were advanced to the M4, M5 and M6 generations, along with their parents and three checks for evaluations based on 13 agro-morphological and 16 grain quality traits. Interestingly, all the mutants of the three landraces showed a reduction in days to 50% flowering and plant height as compared to their parents in all the three generations. The reduction in days to 50% flowering ranges from 4.94% (Vishnubhog Mutant V-67) to 21.40% (Jhilli Mutant J-2-13), whereas the reduction in plant height varies from 11.28% (Vishnubhog Mutant V-45-2, Vishnubhog Mutant V-67) to 37.65% (Jhilli Mutant J-15-1). Furthermore, two, six and three mutants of Samundchini, Vishnubhog and Jhilli have increased their yield potential over their corresponding parents, respectively. Interestingly, Samundchini Mutant S-18-1 (22.45%), Vishnubhog Mutant V-74-6 (36.87%) and Jhilli Mutant J-13-5 (25.96%) showed the highest yield advantages over their parents. Further, a pooled analysis of variance based on a randomized complete block design revealed ample variations among the genotypes for the studied traits. In addition, all the traits consistently showed high to moderate PCV and GCV and a slight difference between them in all three generations indicated the negligible effect of the environment. Moreover, in the association analysis, the traits, viz., fertile spikelets/panicle, panicle length, total tillers/plant, spikelet fertility percent and 100-seed weight showed the usual grain yield/plant, whereas the traits hulling (%) and milling (%) with HRR (%) consistently showed high direct effects and significant positive correlations. The SSR marker-based genome similarity in rice mutants and corresponding parents ranged from 95.60% to 71.70% (Vishnubhog); 95.62% to 89.10% (Samundchini) and 95.62% to 80.40% (Jhilli), indicating the trueness of the mutants. Moreover, the UPGMA algorithm and Gower distance-based dendrogram, neighbour joining tree and PCA scatter diagram assured that mutants were grouped with their respective parents and fell into separate clusters showing high similarity between mutants and parents and dissimilarity among the 24 genotypes. Overall, the information and materials generated from the current study will be very useful and informative for students, researchers and plant breeders. Additionally, our results also showed that irradiation could generate a considerable amount of genetic variability and provide new avenues for crop improvement and diversification.

Salt stress is one of the most severe environmental stresses limiting the productivity of crops, including rice. However, there is a lack of information on how salt-stress sensitivity varies across different developmental stages in rice. In view of this, a comparative evaluation of contrasting rice varieties CSR36 (salt tolerant) and Jaya (salt sensitive) was conducted, wherein NaCl stress (50 mM) was independently given either at seedling (S-stage), tillering (T-stage), flowering (F-stage), seed-setting (SS-stage) or throughout plant growth, from seedling till maturity. Except for S-stage, CSR36 exhibited improved NaCl stress tolerance than Jaya, at all other tested stages. Principal component analysis (PCA) revealed that the improved NaCl stress tolerance in CSR36 coincided with enhanced activities/levels of enzymatic/non-enzymatic antioxidants (root ascorbate peroxidase for T- (2.74-fold) and S+T- (2.12-fold) stages and root catalase for F- (5.22-fold), S+T- (2.10-fold) and S+T+F- (2.61-fold) stages) and higher accumulation of osmolytes (shoot proline for F-stage (5.82-fold) and S+T+F- (2.31-fold) stage), indicating better antioxidant capacitance and osmotic adjustment, respectively. In contrast, higher shoot accumulation of Na+ (14.25-fold) and consequent increase in Na+/K+ (14.56-fold), Na+/Mg+2 (13.09-fold) and Na+/Ca+2 (8.38-fold) ratio in shoot, were identified as major variables associated with S-stage salinity in Jaya. Higher root Na+ and their associated ratio were major deriving force for other stage specific and combined stage salinity in Jaya. In addition, CSR36 exhibited higher levels of Fe3+, Mn2+ and Co3+ and lower Cl− and SO42−, suggesting its potential to discriminate essential and non-essential nutrients, which might contribute to NaCl stress tolerance. Taken together, the findings provided the framework for stage-specific salinity responses in rice, which will facilitate crop-improvement programs for specific ecological niches, including coastal regions.

Controlled-release Nitrogen Fertilizers (CRNFs) are an effective fertilization technique by minimizing nutrient loss and making Nitrogen (N) available to plants as they grow. Biochar-based CRNF (BCRNF) technologies have been demonstrated very promising in increase of corn yield. Despite the beneficial effects of BCRNFs, their impacts on prokaryotic and fungal soil communities are not well evaluated. Different formulations of BCRNF were developed to investigate their effects on corn productivity. We analyzed the soil microbes and their functional potential under different BCRNF regimes using amplified V3–V4 region of 16s rRNA for determining prokaryotic, and ITS genes for fungal communities. The soil prokaryotic diversity was similar across the treatments, with differences in prokaryotic genera with relative abundance of 0.1% or less in the soil (p < 0.05). In contrast, the fungal community diversity was different only for unfertilized soil. It had a high relative abundance for Aspergillus. Genus level comparison showed that Pseudofabraea was higher in Bioasphalt-based BCRNF compared to other treatments. Moreover, the N-fixing communities in soil were also similar across the treatments. At genus level, Microvirga, Azospirillum, and Methyloprofundus were highest in no-fertilizer control. The functional potential predictions using PICRUSt2 portrayed a consistent N-cycling functions across the treatments. However, the predicted gene functions related to nitrous-oxide reductase (nosZ) and hydroxylamine reductase (hcp) were significantly lower in soil receiving BCRNF containing biosolid. Overall, BCRNF treatments previously identified to increase corn yield displayed minimal shifts in the soil microbial communities. Thus, such novel fertilization would enable increased crop yield without affecting soil communities leading to sustainable crop production.

The quantity of grass-root exudates varies by season, suggesting temporal shifts in soil microbial community composition and activity across a growing season. We hypothesized that bacterial community and nitrogen cycle-associated prokaryotic gene expressions shift across three phases of the growing season. To test this hypothesis, we quantified gene and transcript copy number of nitrogen fixation (nifH), ammonia oxidation (amoA, hao, nxrB), denitrification (narG, napA, nirK, nirS, norB, nosZ), dissimilatory nitrate reduction to ammonia (nrfA), and anaerobic ammonium oxidation (hzs, hdh) using the pre-optimized Nitrogen Cycle Evaluation (NiCE) chip. Bacterial community composition was characterized using V3-V4 of the 16S rRNA gene, and PICRUSt2 was used to draw out functional inferences. Surprisingly, the nitrogen cycle genes and transcript quantities were largely stable and unresponsive to seasonal changes. We found that genes and transcripts related to ammonia oxidation and denitrification were different for only one or two time points across the seasons (p < 0.05). However, overall, the nitrogen cycling genes did not show drastic variations. Similarly, the bacterial community also did not vary across the seasons. In contrast, the predicted functional potential was slightly low for May and remained constant for other months. Moreover, soil chemical properties showed a seasonal pattern only for nitrate and ammonium concentrations, while ammonia oxidation and denitrification transcripts were strongly correlated with each other. Hence, the results refuted our assumptions, showing stability in N cycling and bacterial community across growing seasons in a natural grassland.

BACKGROUND : Bradyrhizobium fixes nitrogen symbiotically with soybean and is an agriculturally significant bacterium. Much is known about the Bradyrhizobium species that nodulate soybeans. Conversely, prevalence of Bradyrhizobium in soil and the rhizosphere is known only to the genus level as culture independent approaches have provided only partial 16S rRNA gene sequences, so that nodulating and non-nodulating species could not be distinguished. METHODS : To track which species in bulk soil proliferate in the rhizosphere, and then nodulate, we sought to study population dynamics of Bradyrhizobium in soybean fields and rhizosphere at the species level. Recent advances in Oxford Nanopore Technologies provided us with higher fidelity and increased number of reads which enabled us to track Bradyrhizobium populations at the species level. RESULTS : We found evidence for 74 species of Bradyrhizobium within a community of 10,855 bacterial species in bulk soil and rhizosphere from three different soybean fields in South Dakota. The most predominant species in bulk soil and rhizosphere included B. liaoningense, B. americanum, and B. diversitatus, however none of these were isolated from nodules. Isolates from nodules included B. japonicum, B. elkanii and B. diazoefficiens. These nodulators also maintained populations in bulk soil and rhizosphere, although they were not the most prevalent Bradyrhizobium. CONCLUSIONS : Our findings reveal the rich diversity and community dynamics of Bradyrhizobium species in soybean field soil as well as in the rhizosphere. Our results showed that many species of the genus maintain populations in soybean field soil, even in the long-term absence of potential nodulating partners.