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Globally, drought stress poses a significant threat to crop productivity. Improving the drought tolerance of crops with microbial biostimulants is a sustainable strategy to meet a growing population’s demands. This research aimed to elucidate microbial biostimulants’ (Plant Growth Promoting Rhizobacteria) role in alleviating drought stress in oil-seed crops. In total, 15 bacterial isolates were selected for drought tolerance and screened for plant growth-promoting (PGP) attributes like phosphate solubilization and production of indole-3-acetic acid, siderophore, hydrogen cyanide, ammonia, and exopolysaccharide. This research describes two PGPR strains: Acinetobacter calcoaceticus AC06 and Bacillus amyloliquefaciens BA01. The present study demonstrated that these strains (AC06 and BA01) produced abundant osmolytes under osmotic stress, including proline (2.21 and 1.75 µg ml − 1 ), salicylic acid (18.59 and 14.21 µg ml − 1 ), trehalose (28.35 and 22.74 µg mg − 1 FW) and glycine betaine (11.35 and 7.74 mg g − 1 ) respectively. AC06 and BA01 strains were further evaluated for their multifunctional performance by inoculating in Arachis hypogaea L. (Groundnut) under mild and severe drought regimes (60 and 40% Field Capacity). Inoculation with microbial biostimulants displayed distinct osmotic-adjustment abilities of the groundnut, such as growth parameters, plant biomass, photosynthetic pigments, relative water content, proline, and soluble sugar in respective to control during drought. On the other hand, plant sensitivity indexes such as electrolyte leakage and malondialdehyde (MDA) contents were decreased as well as cooperatively conferred plant drought tolerance by induced alterations in stress indicators such as catalase (CAT), ascorbate peroxidase (APX), and superoxide dismutase (SOD). Thus, Acinetobacter sp. AC06 and Bacillus sp. BA01 can be considered as osmolyte producing microbial biostimulants to simultaneously induce osmotic tolerance and metabolic changes in groundnuts under drought stress.

• Background and Aims The genus Arachis contains 80 described species. Section Arachis is of particular interest because it includes cultivated peanut, an allotetraploid, and closely related wild species, most of which are diploids. This study aimed to analyse the genetic relationships of multiple accessions of section Arachis species using two complementary methods. Microsatellites allowed the analysis of inter- and intraspecific variability. Intron sequences from single-copy genes allowed phylogenetic analysis including the separation of the allotetraploid genome components. • Methods Intron sequences and microsatellite markers were used to reconstruct phylogenetic relationships in section Arachis through maximum parsimony and genetic distance analyses. • Key Results Although high intraspecific variability was evident, there was good support for most species.However, some problems were revealed, notably a probable polyphyletic origin for A. kuhlmannii. The validity of the genome groups was well supported. The F, K and D genomes grouped close to the A genome group. The 2n = 18 species grouped closer to the B genome group. The phylogenetic tree based on the intron data strongly indicated that A. duranensis and A. ipaënsis are the ancestors of A. hypogaea and A. montícola. Intron nucleotide substitutions allowed the ages of divergences of the main genome groups to be estimated at a relatively recent 2.3-2.9 million years ago. This age and the number of species described indicate a much higher speciation rate for section Arachis than for legumes in general. • Conclusions The analyses revealed relationships between the species and genome groups and showed a generally high level of intraspecific genetic diversity. The improved knowledge of species relationships should facilitate the utilization of wild species for peanut improvement. The estimates of speciation rates in section Arachis are high, but not unprecedented. We suggest these high rates may be linked to the peculiar reproductive biology of Arachis.

Summary Peanut stem is a vital organ to provide mechanical support and energy for aerial tissue development. However, the transcriptional regulatory mechanisms underlying stem development at a single‐cell resolution remain unclear. Herein, single‐nuclei isolation coupled with fluorescent‐activated cell sorting was employed to construct a cell atlas of peanut seedling stems using microdroplets‐based single‐nuclei RNA‐sequencing. This approach yielded 29 308 cells with 53 349 expressed genes underlying the identification of five cell types characterized by known marker genes. Additionally, 2053 differentially expressed genes (DEGs) were identified across different cell types. Furthermore, 3306 core‐DEGs involved in cell development trajectories were used to construct a transcription factor (TF) interaction network, providing insights into specific biological pathways and transcriptional regulation dynamics underlying cell‐type differentiation. Additionally, 1446 DEGs associated with different cell‐cycle profile were identified, revealing that peanut stem elongation and cell expansion are closely linked to auxin‐responsive pathway. This was supported by the examination of endogenous phytohormones and the identification of 10 hormone‐responsive DEGs. Moreover, AhWRKY70 was localized in the nucleus and is highly enriched in stem cortex and xylem cells and exhibits a tissue‐specific expression pattern that regulates stem growth. Overexpression of AhWRKY70 in Arabidopsis led to accelerated stem growth by modulating the phytohormone signalling pathway, influencing the expression of sixteen auxin and ethylene‐responsive genes as demonstrated by transcriptome sequencing. In conclusion, the single‐cell atlas provides a foundational dataset for understanding gene expression heterogeneity in peanut seedling stems. The elucidation of AhWRKY70 function expands our understanding of the roles of WRKY family members in peanut.

Background Intercropping, a diversified planting pattern, increases land use efficiency and farmland ecological diversity. We explored the changes in soil physicochemical properties, nutrient uptake and utilization, and microbial community composition in wide-strip intercropping of maize and peanut. Results The results from three treatments, sole maize, sole peanut and intercropping of maize and peanut, showed that intercropped maize had a marginal advantage and that the nutrient content of roots, stems and grains in side-row maize was better than that in the middle row of intercropped maize and sole maize. The yield of intercropped maize was higher than that of sole cropping. The interaction between crops significantly increased soil peroxidase activity, and significantly decreased protease and dehydrogenase activities in intercropped maize and intercropped peanut. The diversity and richness of bacteria and fungi decreased in intercropped maize rhizosphere soil, whereas the richness of fungi increased intercropped peanut. RB41 , Candidatus-udaeobacter , Stropharia , Fusarium and Penicillium were positively correlated with soil peroxidase activity, and negatively correlated with soil protease and dehydrogenase activities. In addition, intercropping enriched the functional diversity of the bacterial community and reduced pathogenic fungi. Conclusion Intercropping changed the composition and diversity of the bacterial and fungal communities in rhizosphere soil, enriched beneficial microbes, increased the nitrogen content of intercropped maize and provided a scientific basis for promoting intercropping in northeastern China.

Background Drought stress significantly affects peanut ( Arachis hypogaea L.) growth and yield, necessitating strategies to enhance crop resilience. This study evaluates the impact of foliar-applied Metformin, gibberellic acid (GA₃), and indole-3-acetic acid (IAA) at concentrations of 5.0, 7.5, and 10.0 mg L⁻ 1 under different irrigation regimes (100%, 80%, and 60% of the recommended irrigation rate). Methods A two-year field experiment was conducted under arid conditions to assess the effects of these treatments on plant growth, yield, photosynthetic pigments, nutrient uptake, and water use efficiency (WUE). Peanut plants were exposed to three irrigation levels (100%, 80%, and 60%), and foliar treatments were applied at specific growth stages. Photosynthetic parameters, including chlorophyll and carotenoid content, were measured alongside growth and yield attributes to determine treatment efficacy. Results The application of Metformin at 7.5 mg L⁻ 1 under 80% irrigation significantly improved plant height (76.9 cm), branch number (17.7 per plant), fresh weight (2928.5 kg acre⁻ 1 ), dry biomass (329.1 kg acre⁻ 1 ), and total seed yield (1593.9 kg acre⁻ 1 ) compared to other treatments. Additionally, water use efficiency (WUE) increased by 50.8% in plants treated with Metformin at 7.5 mg L⁻ 1 under 80% irrigation compared to untreated plants. The highest chlorophyll content (1.27 mg g⁻ 1 FW) and carotenoid levels (2.87 mg g⁻ 1 FW) were observed with Metformin at 7.5 mg L⁻ 1 under 100% irrigation, indicating improved photosynthetic performance. Conclusion Foliar application of Metformin at 7.5 mg L⁻ 1 under 80% irrigation effectively enhances peanut growth, yield, and WUE, providing a sustainable strategy to mitigate drought stress effects. This treatment balances crop productivity and water conservation, making it a viable approach for peanut cultivation in water-limited environments. Highlights - Foliar application of Metformin at 7.5 mg L⁻¹ significantly improves peanut growth, yield, and water use efficiency under drought stress. - Peanut plants treated with Metformin exhibit increased plant height, branch number, fresh and dry biomass, and seed yield. - The combination of Metformin at 7.5 mg L⁻¹ with 80% irrigation optimally enhances yield while conserving water resources. - Metformin application increases chlorophyll and carotenoid content, supporting improved photosynthesis and plant vigor. - The study demonstrates that Metformin can serve as an effective, eco-friendly bio-stimulant for peanut cultivation in water-scarce regions.

Background Rapid alkalinization factors (RALFs) are small peptides hormones that regulate plant growth and stress responses. Although RALFs have been identified in a broad range of land plant species, their roles in peanuts ( Arachis hypogaea L.) remain largely unexplored. Result A total of 24 AhRALF genes we identified in the peanut genome and classified them into three clades through phylogenetic analysis. Whole genome duplication (WGD) or segmental duplication primarily drives the expansion of AhRALFs . Gene transcription analysis revealed that two genes from clade II ( AhRALF1 and AhRALF12 ) and three from clade III ( AhRALF8 , AhRALF10 , and AhRALF21 ) are highly expressed across 18 different tissues. Notably, AhRALF11 and AhRALF24 , paralogous genes from clade II, are specifically expressed in immature buds and flowers. Additionally, AhRALF1 , AhRALF12 , AhRALF8 , and AhRALF21 exhibited elevated expression under aluminum (Al) stress. Functional analysis of AhRALF1 confirmed its secretory function and inhibitory effect on root growth in Arabidopsis . Moreover, AhRALF1 -silenced plants displayed reduced tolerance to Al stress, with altered antioxidant enzyme activities and increased oxidative damage. Conclusion This study provides a comprehensive analysis of the AhRALF gene family in peanut, highlighting their roles in growth regulation and stress responses. The function of AhRALF1 in enhancing peanut tolerance to Al stress was preliminary revealed. Our findings provide valuable insights into the roles of AhRALFs in peanuts and lay the groundwork for future functional studies and breeding programs.

PREMISE OF THE STUDY: Several species of Arachis have been cultivated for their edible seeds, historically and to the present day. The diploid species that have a history of cultivation show relatively small signatures of domestication. In contrast, the tetraploid species A. hypogaea evolved into highly domesticated forms and became a major world crop, the cultivated peanut. It seems likely that allotetraploidization (hybridity and/or tetraploidization) in some way enhanced attractiveness for cultivation. Here we investigate this using six different hybridization and tetraploidization events, from distinct Arachis diploid species, including one event derived from the same wild species that originated peanut. METHODS: Twenty‐six anatomical, morphological, and physiological traits were examined in the induced allotetraploid plants and compared with their wild diploid parents. KEY RESULTS: Nineteen traits were transgressive (showed strong response to hybridization and chromosome duplication): allotetraploids had larger leaves, stomata and epidermal cells than did their diploid parents. In addition, allotetraploids produced more photosynthetic pigments. These traits have the same trend across the different hybrid combinations, suggesting that the changes are more likely due to ploidy rather than hybridity. In contrast, seed dimensions and seed mass did not significantly change in response to hybridization or tetraploidization. CONCLUSIONS: We suggest that the original allotetraploid that gave rise to cultivated peanut may have been attractive because of an increase in plant size, different transpiration characteristics, higher photosynthetic capacity, or other characteristics, but contrary to accepted knowledge, increased seed size was unlikely to have been important in the initial domestication.

Background The cultivated peanut is an important oil and cash crop grown worldwide. To meet the growing demand for peanut production each year, genetic studies and enhanced selection efficiency are essential, including linkage mapping, genome-wide association study, bulked-segregant analysis and marker-assisted selection. Specific locus amplified fragment sequencing (SLAF-seq) is a powerful tool for high density genetic map (HDGM) construction and quantitative trait loci (QTLs) mapping. In this study, a HDGM was constructed using SLAF-seq leading to identification of QTL for seed weight and size in peanut. Results A recombinant inbred line (RIL) population was advanced from a cross between a cultivar ‘Huayu36’ and a germplasm line ‘6–13’ with contrasting seed weight, size and shape. Based on the cultivated peanut genome, a HDGM was constructed with 3866 loci consisting of SLAF-seq and simple sequence repeat (SSR) markers distributed on 20 linkage groups (LGs) covering a total map distance of 1266.87 cM. Phenotypic data of four seed related traits were obtained in four environments, which mostly displayed normal distribution with varied levels of correlation. A total of 27 QTLs for 100 seed weight (100SW), seed length (SL), seed width (SW) and length to width ratio (L/W) were identified on 8 chromosomes, with LOD values of 3.16–31.55 and explaining phenotypic variance (PVE) from 0.74 to 83.23%. Two stable QTL regions were identified on chromosomes 2 and 16, and gene content within these regions provided valuable information for further functional analysis of yield component traits. Conclusions This study represents a new HDGM based on the cultivated peanut genome using SLAF-seq and SSRs. QTL mapping of four seed related traits revealed two stable QTL regions on chromosomes 2 and 16, which not only facilitate fine mapping and cloning these genes, but also provide opportunity for molecular breeding of new peanut cultivars with improved seed weight and size.

Abiotic stress is a limiting factor in peanut production. Peanut is an important oil crop and cash crop in China. Peanut yield is vulnerable to abiotic stress due to its seeds grown underground. Jasmonic acid (JA) is essential for plant growth and defense against adversity stresses. However, the regulation and mechanism of the jasmonic acid biosynthesis pathway on peanut defense against abiotic stresses are still limitedly understood. In this study, a total of 64 genes encoding key enzymes of JA biosynthesis were identified and classified into lipoxygenases (AhLOXs), alleno oxide synthases (AhAOSs), allene oxide cyclases (AhAOCs), and 12-oxo-phytodienoic acid reductases (AhOPRs) according to gene structure, conserved motif, and phylogenetic feature. A cis-regulatory element analysis indicated that some of the genes contained stress responsive and hormone responsive elements. In addition to proteins involved in JA biosynthesis and signaling, they also interacted with proteins involved in lipid biosynthesis and stress response. Sixteen putative Ah-miRNAs were identified from four families targeting 35 key genes of JA biosynthesis. A tissue expression pattern analysis revealed that AhLOX2 was the highest expressed in leaf tissues, and AhLOX32 was the highest expressed in shoot, root, and nodule tissues. AhLOX16, AhOPR1, and AhOPR3 were up-regulated under drought stress. AhLOX16, AhAOS3, AhOPR1, and AhAOC4 had elevated transcript levels in response to cold stress. AhLOX5, AhLOX16, AhAOC3, AhOPR1, and AhOPR3 were up-regulated for expression under salt stress. Our study could provide a reference for the study of the abiotic stress resistance mechanism in peanut.

Key messageCo-localized intervals and candidate genes were identified for major and stable QTLs controlling pod weight and size on chromosomes A07 and A05 in an RIL population across four environments.Cultivated peanut (Arachis hypogaea L.) is an important legume crops grown in > 100 countries. Hundred-pod weight (HPW) is an important yield trait in peanut, but its underlying genetic mechanism was not well studied. In this study, a mapping population (Xuhua 13 × Zhonghua 6) with 187 recombinant inbred lines (RILs) was developed to map quantitative trait loci (QTLs) for HPW together with pod length (PL) and pod width (PW) by both unconditional and conditional QTL analyses. A genetic map covering 1756.48 cM was constructed with 817 markers. Additive effects, epistatic interactions, and genotype-by-environment interactions were analyzed using the phenotyping data generated across four environments. Twelve additive QTLs were identified on chromosomes A05, A07, and A08 by unconditional analysis, and five of them (qPLA07, qPLA05.1, qPWA07, qHPWA07.1, and qHPWA05.2) showed major and stable expressions in all environments. Conditional QTL mapping found that PL had stronger influences on HPW than PW. Notably, qHPWA07.1, qPLA07, and qPWA07 that explained 17.93–43.63% of the phenotypic variations of the three traits were co-localized in a 5 cM interval (1.48 Mb in physical map) on chromosome A07 with 147 candidate genes related to catalytic activity and metabolic process. In addition, qHPWA05.2 and qPLA05.1 were co-localized with minor QTL qPWA05.2 to a 1.3 cM genetic interval (280 kb in physical map) on chromosome A05 with 12 candidate genes. This study provides a comprehensive characterization of the genetic components controlling pod weight and size as well as candidate QTLs and genes for improving pod yield in future peanut breeding.