The pear (Pyrus spp.), a perennial fruit tree, is subjected to genetic alterations over decades or even centuries to adapt to complex climatic and cultivation conditions. Genome-wide studies of deleterious mutations remain limited in perennial fruit trees, particularly regarding the effects of domestication on deleterious mutations. In this study, 232 pear accessions were resequenced, and 9 909 773 single-nucleotide polymorphisms (SNPs), and 139 335 deleterious mutation sites, were identified genome wide. A higher proportion of deleterious mutations in coding regions (1.4%) were observed in the pear genome than annual crops. During domestication, a reduction in deleterious mutations in Pyrus pyrifolia/P. bretschneideri was found to be associated with their decreases in selective sweep regions. Conversely, an increase in the number of deleterious mutations in P. communis was observed, which may be related to a higher occurrence within selective sweep regions. In P. ussuriensis, an overall increasing trend in deleterious mutations was identified, which was determined to be unrelated to domestication or gene introgression but instead linked to its relatively high heterozygosity. Differential deleterious mutation genes were identified during the domestication process. Among these, the PyMYC2 gene, associated with stone cell synthesis, was identified through GWAS, overexpression of PyMYC2 in pear callus significantly promoter lignin biosynthesis, PyMYC2 contains three nonsynonymous deleterious mutations that were selected during the domestication of Asian pears. This research provides new insights into developing future breeding strategies aimed at improving agronomic traits and offers a framework for studying deleterious mutation patterns in the domestication of perennial fruit trees.

To produce mature seed, flowering plants must undergo successful male and female gametogenesis and pollination followed by fruit set, growth, and ripening. This sequential process involves complex genetic programming and less understood epigenetic reprogramming. Here we report a previously unidentified CHROMOMETHYLASE3-directed epi-control in pollen mother cell (PMC)-to-microspore transition that determines male fertility to affect seed formation. We generated and characterized hairpin RNA-mediated RNAi and CRISPR/Cas9 transgenic tomato lines in which CHROMOMETHYLASE3 (CMT3) was either knockdown (KD) or knockout (KO). CHROMOMETHYLASE3 has pleiotropic effects on vegetative and reproductive growth, including leaf, flower, and seed development, besides its influence on tomato ripening and fruit size. However, CMT3 KD plants exhibited stronger effects than KO plants in terms of these vegetative and reproductive processes. Real-time quantitative PCR analysis suggested that genetic compensation might contribute to the less impact of KO plants on pollen and seed development. Integrated RNA-seq and whole-genome bisulfite sequencing reveal that CMT3 functions as an epi-switch via a self-feedback mechanism to modulate gene expression and governs early development of microspores from PMCs prior to the tetrad stage during microsporogenesis to microgametogenesis, possibly through the pectin catabolic process, to establish pollen fertility that affects seed production in tomato.

Monoterpenoids are vital compounds that impart a distinctive floral flavor. They exist in both glycosidic and free forms in grapes. The breeding of improved monoterpenoid varieties has consistently been a topic of interest, yet only a limited number of molecular markers have been documented. This study employed a genome-wide association study (GWAS) on an F₁ population crossed between a typical muscat variety (‘Muscat of Alexandria’) and a non-aromatic variety (‘Christmas Rose’), conducted over two consecutive years. A total of 4089 significant single nucleotide polymorphism sites (sigSNPs) and 892 candidate genes associated with monoterpenoids were identified. The sigSNPs corresponding to the glycosidic and total (glycosidic plus free) concentrations of various monoterpenoid compounds exhibited a high similarity. The majority of sigSNPs were located on chromosome 5, indicating the existence of a monoterpenoid-related marker cluster. Sixty-one lead SNPs located within the gene region and stably appearing in 2 years were selected and verified using a germplasm population. The alleles of the 25 lead SNPs were confirmed to be highly associated with monoterpenoid levels. The genes containing these lead SNPs were mainly glycoside hydrolase, ABC transporter, as well as the previously reported 1-deoxy-D-xylulose-5-phosphate synthase (VvDXS1) and geranylgeranyl pyrophosphate synthase large subunit (VvGGPPS-LSU). The function of VvGGPPS-LSU in regulating monoterpenoid levels was elucidated through in vivo overexpression, demonstrating the reliability of the marker cluster. The present study proposes a molecular marker set for the breeding with the objective of improving aroma, and a candidate gene network for the regulation of monoterpenoid synthesis in grapevine.

The ubiquitin-proteasome system (UPS) is important for protein post-translational modification in plants. E2 (ubiquitin-conjugating enzyme) and E3 (ubiquitin ligases enzyme), key enzymes of UPS, play crucial roles in all aspects of plant development, growth, and environmental stresses. Despite extensive knowledge of UPS roles in crop growth and development, E2-E3 pair functions in potato tuber development and stress responses remain understudied. Here, we describe the role of StUBC18 (a potato E2) in drought stress tolerance. It is determined that StUBC18 (E2)-StPUB40 (E3) pair plays important roles in drought stress tolerance and potato tuber yield. StUBC18 and StPUB40 expression was downregulated under various stresses (drought, salt, polyethylene glycol, and H₂O₂). Overexpression of StUBC18 and StPUB40 in potatoes decreased drought stress tolerance, while interfering with the expression of StUBC18 and StPUB40 increased drought stress tolerance, respectively. The protein interaction test demonstrated that StUBC18 interacts with StPUB40 in the plant cell. Co-overexpression of StUBC18-StPUB40 in potato enhanced reactive oxygen species (ROS) accumulation and induced pleiotropic changes, reducing drought tolerance. Our findings revealed how the StUBC18-StPUB40 pair regulates potato drought stress tolerance by altering leaf anatomy (palisade and spongy tissue thickness) and influences tuber yield.

Grapevine powdery mildew (GPM), caused by Erysiphe necator, poses a significant threat to all green grapevine tissues, leading to substantial economic losses in viticulture. Traditional grapevine cultivars derived from Vitis vinifera are highly susceptible to GPM, whereas the wild Chinese accession Baishui-40 (BS-40) of V. piasezkii var. pagnucii exhibits robust resistance. To illuminate the genetic basis of resistance, we sequenced and assembled the chromosome-level genome of ‘BS-40’, achieving a total mapped length of 578.6 Mb distributed across nineteen chromosomes. A comprehensive annotation identified 897 nucleotide-binding leucine-rich repeat (NLR) genes in the ‘BS-40’ genome, which exhibited high sequence similarity across Vitis genomes. 284 of these NLR genes were differentially expressed upon GPM infection. A hybrid population of ‘BS-40’ and V. vinifera was constructed and 195 progenies were whole-genome re-sequenced. A new GPM-resistant locus, designated Ren17, located within the 0.74-1.23 Mb region on chromosome 1 was identified using genome-wide association study, population selection, and QTL analysis. Recombinant events indicated that an NLR gene cluster between 1 045 489 and 1 089 719 bp on chromosome 1 is possibly the key contributor to GPM resistance in ‘BS-40’. Based on an SNP within this region, a dCAPS marker was developed that can predict the GPM resistance in ‘BS-40’-derived materials with 99.4% accuracy in the progenies of ‘BS-40’ and V. vinifera. This chromosome-level genome assembly of V. piasezkii var. pagnucii provides a valuable resource not only for grapevine evolution, genetic analysis, and pan-genome studies but also a new locus Ren17 as a promising target for GPM-resistant breeding in grapevine.

Self-incompatibility (SI) is a complex molecular mechanism in flowering plants that prevents self-fertilization and promotes outcrossing. We conducted metabolome analysis of ornamental kale (Brassica oleracea var. acephala) pistils following SI and compatible pollination (CP), revealing significant alterations in lipid metabolism, particularly the accumulation of free fatty acid (FFA) metabolites during CP. Treatment of stigmas with acetyl-CoA and malonyl-CoA, key precursors in fatty acid (FA) synthesis, broke down SI and enhanced CP. Conversely, inhibiting acetyl-CoA carboxylase (ACCase), the rate-limiting enzyme in de novo FA synthesis, significantly reduced compatible pollen attachment and tube growth, highlighting the critical role of FA metabolism in mediating pollination success. We identified a novel interaction between the FERONIA (BoFER) receptor kinase and the biotin carboxyl carrier protein 1 (BoBCCP1), a subunit of the ACCase complex. Suppressing the expression of BoBCCP1 in the stigma reduced CP response, suggesting that the FER-BCCP1 module may play a crucial role in regulating FA biosynthesis and determining the outcome of pollen-stigma interactions. Our findings provide new insights into the identification of key metabolic pathways and signaling modules controlling pollen-stigma interactions, and offer a valuable resource for the targeted improvement of Brassica crop breeding.

OVATE family proteins (OFPs) constitute a class of transcription factors regulating various developmental processes in plants. Nevertheless, their precise regulatory functions in melon (Cucumis melo L.) fruit development remain elusive. In this study, we identified expression profiling of melon OFP genes and revealed the molecular function of CmOFP6-19b gene mediating fruit size variation. Quantitative analysis revealed predominant CmOFP expression in reproductive organs (female/male flowers and ovaries), with distinct differential expression patterns observed among paralogs. Through melon genetic transformation, we revealed that CmOFP6-19b gene functions as a negative regulator in fruit enlargement. Overexpression of the CmOFP6-19b gene resulted in reduced fruit size, while its downregulation led to increased fruit size. Bimolecular fluorescence complementation and yeast two-hybrid assays confirmed nuclear-localized physical interaction between CmOFP6-19b and CmKNOX16. Overexpression of CmKNOX16 in melon produced smaller fruits, phenocopying the CmOFP6-19b-Oe lines. Quantitative real-time PCR (RT-qPCR) analysis showed negative correlation between CmOFP6-19b/CmKNOX16 expression level and fruit size, with peak expression levels observed in a cultivar displaying minimal longitudinal diameter. The results of histological section and expression analysis suggest that CmOFP6-19b and CmKNOX16 may affect melon fruit size by regulating genes related to cell division and cell expansion. In conclusion, our findings systematically characterized the phylogenetic architecture and expression divergence of CmOFP genes, and elucidated the function and molecular mechanism of CmOFP6-19b-CmKNOX16 regulatory module in mediating melon fruit development, providing a theoretical foundation for melon breeding.

Flavonol glycosides have many prominent benefits to human health and significant contributions to the growth and development of tea plant as well as the color and taste of tea infusion. In this study, a gene isolated from tea plant was found to encode a 52.2-kDa protein located on the plasma membrane and in the cytoplasm with activity of flavonol glycosyltransferase (CsFGT). The prokaryotically expressed recombinant CsFGT (rCsFGT) exhibited its main glucosyl transfer activity towards rutin to produce quercetin 3-O-β-d-glucopyranosyl-(1→3)-α-l-rhamnopyranosyl-(1→6)-β-d-glucopyranoside (Q-g-r-g), and showed a minor galactosyl transfer activity towards delphinidin to produce delphinidin 3-O galactoside. The maximum activity of rCsFGT was observed at 30°C and pH 8.0. The main function of rCsFGT seems to be catalysis of the biosynthesis of Q-g-r-g rather than delphinidin 3-O galactoside since its affinity and catalytic efficiency are much higher towards rutin than towards delphinidin. Molecular docking and site-directed mutation reveal that amino acid residues G290, E292, R319, and Q352 play important roles in the catalytic specificity of CsFGT. The Q-g-r-g content in leaves of different tea cultivars was significantly correlated with the CsFGT expression level. Injection of antisense oligodeoxyribonucleotides remarkably downregulated endogenous CsFGT expression and consequently reduced the Q-g-r-g content significantly. These findings will help elucidate the differential accumulation mechanism of flavonol glycosides in different tea germplasms.

Horticultural crops have important economic value in the world. Biotic stress has serious impacts on horticultural crops’ growth and development as well as yield. Melatonin, a multifunctional signaling molecule, has been increasingly documented to play a pivotal role in mediating plant defense responses against diverse biotic stressors, including bacterial, fungal, and viral pathogens in horticultural crop species. Previous studies showed that exogenous melatonin treatment significantly improved horticultural crops growth and increased their tolerance to biotic stress. Although there are numerous studies to show that exogenous melatonin treatment can markedly improve the tolerance for horticultural crops in response to biotic stress, the role of melatonin in biotic stress responses remains unclear and requires clarification. In the review, we summarize the effects of melatonin on horticultural crops’ disease resistance. Moreover, we assess future perspectives in melatonin research and its applications to improve horticultural crop production and tolerance for biotic stress. This review explores future research directions and potential applications to enhance the productivity and biotic stress tolerance of horticultural crops, and also provides a theoretical basis for enhancing the scientific understanding of the role of melatonin in response to biotic stress in horticultural crops.

Rapeseed (Brassica napus L.) with determinate inflorescence (DTI) exhibits desirable traits, including reduced plant height, enhanced lodging resistance, and consistent maturity, making them valuable breeding resources. DTI is modulated by BnaA10.TFL1 and BnaC09.TFL1 (BnaA10/C09.TFL1), which encode the TERMINAL FLOWER 1 protein, a key regulator of flowering time and meristem identity. However, the underlying functional and regulatory mechanisms remain unclear. In this study, we demonstrated that variations in the promoter region of BnaA10/C09.TFL1, rather than the coding region, contributed to the transition from indeterminate inflorescence (IDTI) to DTI in B. napus. Specifically, BnaA10.SEP inhibited BnaA10/C09.TFL1 expression by binding to the GT1-motif in the promoter region of BnaA10/C09.TFL1^DTI, contributing to the IDTI phenotype under short-day conditions. Meanwhile, two novel DTI mutants were successfully generated through the simultaneous knockout of BnaA10/C09.TFL1 using the CRISPR/Cas9 system. Furthermore, BnaA10/C09.TFL1 and its homolog BnaA02.FT interacted with BnaA07.14-3-3 instead of directly binding to BnaA08.FD to regulate the development of different inflorescence architectures. Overall, the BnaA10.SEP-BnaA10/C09.TFL1-BnaA07.14-3-3-BnaA08.FD module revealed a new mechanism for DTI formation and a promising strategy for modifying inflorescence architecture traits in B. napus.

Roses (Rosa hybrida) are the most popular cut flower plants worldwide, accounting for over a third of the global cut flower industry. Gray mold, caused by Botrytis cinerea, is often referred to as the postharvest "cancer" of cut roses and represents the most significant disease impacting the postharvest preservation of these flowers in China. Currently, research progress in this area has been limited. Our study utilized single-cell RNA sequencing technology to elucidate the mechanisms underlying B. cinerea resistance in R. hybrida “Jumilia.” We identified seven distinct cell groups within rose petals. The rose epidermis acts as the physical barrier of defense against B. cinerea, while the infection rate may be accelerated through vascular tissues. Furthermore, we identified several key genes, including pectin methylesterases, pathogenesis-related proteins, glutathione S-transferase, and endochitinase EP3, which may play crucial roles in the stress response. The biosynthesis of secondary metabolites temporarily mitigates the infection process, and pathogenesis-related proteins have been recognized as key regulatory genes. This preliminary study elucidates the cellular changes and molecular mechanisms involved in B. cinerea infection in rose petals at the single-cell level. Our findings provide new insights into the defense mechanisms of roses against fungal diseases.

Citrus production is threatened by biotic and abiotic stresses, particularly Huanglongbing (HLB), creating an urgent need for efficient engineering of citrus for disease resistance. Gene editing, especially transgene-free approaches, offers a promising alternative to traditional breeding, which is slow and constrained by citrus’ long juvenile phase. However, producing transgene-free, genome-edited citrus remains challenging. Here, we present a novel method to significantly enhance the efficiency of transgene-free gene editing in citrus using Agrobacterium-mediated transient expression of Cas9 and gRNAs. By treating Agrobacterium cells and citrus explants and applying a 3-day transient kanamycin selection, we achieved a 17-fold increase in transgene-free editing efficiency. The transient kanamycin-mediated suppression of shoot regeneration from non-Agrobacterium-infected cells not only improved the efficiency of identifying edited plants but also enhanced shoot regeneration efficiency from Agrobacterium-infected cells, regardless of whether these cells had stably incorporated T-DNA or not. This enhancement was likely due to reduced competition for space and nutrients from shoots regenerated from noninfected cells. In experiments targeting the phytoene desaturase (PDS) gene, transgene-free mutant shoot recovery increased from 0.017% to 0.291% of the total shoots produced. With an efficient screening method for gene-edited plants, the development of transgene-free gene-edited plants becomes relatively easy and practicable. These results suggest that this optimized protocol could be applicable to other perennial crops, offering a valuable tool for improving citrus varieties and other economically important plants.

Fittonia albivenis, commonly known as the nerve plant, is an ornamental species native to the Peruvian rainforest, valued for its vibrant and diverse leaf coloration. Understanding the genetic mechanisms underlying this coloration is crucial for enhancing its ornamental value and adaptation to environmental stressors. Here, by leveraging advanced sequencing technologies such as PacBio HiFi, Oxford Nanopore, and Hi-C, we achieved a nearly complete haplotype-phased genome assembly for F. albivenis, revealing a 2.08-Gb genome composed of 18 chromosome pairs and containing 66 telomeres. This assembly enabled the identification of subgenome-specific repetitive sequences, elucidating their impact on gene expression and structural variations. Through RNA sequencing, metabolomic profiling, and resequencing, we dissected the regulatory networks influencing chlorophyll and anthocyanin biosynthesis, identifying key genes and transcription factors driving leaf color variation. Our findings highlight the roles of gene duplication and specific transcription factors in pigment synthesis pathways, providing a foundation for future genetic studies and breeding programs aimed at enhancing ornamental and adaptive traits in F. albivenis and related species.

Volatile aroma compounds make significant contributions to human perception of flowers. Osmanthus fragrans is a famous aroma plant, and linalool is proved to be the dominant aroma active compound. Although some terpene synthases have been characterized, a comprehensive study of the hub metabolic gene and its transcriptional regulation remain to be revealed. Here, we selected a specific cultivar Boyeyingui with the highest content of linalool among 20-wide-cultivated cultivars for genome and transcriptome sequencings. Among the 25 new putative OfTPSs, only OfTPS6, OfTPS7 could exclusively produce linalool in planta. Biochemical analysis demonstrated that OfTPS6, OfTPS7 were able to catalyze geranyl diphosphate into linalool and a small proportion of other monoterpenes in vitro. Spatial and temporal correlation analysis further confirmed the expression level of OfTPS7 was strongly correlated with linalool content in a panel of 20 cultivars, suggesting OfTPS7 was the essential linalool synthase gene. Combined with yeast one-hybrid screen and weighted correlation network analysis, a nucleus-localized transcriptional factor OfWRKY33 was identified as a prospective modulator. Y1H, LUC, and EMSA demonstrated that OfWRKY33 directly bound to the W-box of OfTPS7 promoter to stimulate its transcription. OfWRKY33 could coordinately induce the expressions of OfTPS7 and 1-deoxy-d-xylulose 1, thereby promoting the linalool formation. The results first identified the key linalool synthase gene OfTPS7 and a novel transcription factor playing a role in the complex regulatory network of linalool biosynthesis in O. fragrans flowers.

Quercus fabri is a common timber oak tree species widely distributed in subtropical areas of China. In this study, we presented a chromosome-scale reference genome assembly of Q. fabri achieved by integrating PacBio Sequel II, DNBseq™, and Hi-C sequencing platforms, and the results indicated the Q. fabri genome has a size of 836.74 Mb. Through the analysis of significantly expanded gene families, we identified that many of the top-ranked KEGG pathways are associated with amino acid metabolism. Subsequently, we performed an amino acid metabolic profile analysis on Q. fabri and related species, including Quercus aliena, Quercus acutissima, and Quercus variabilis. The findings revealed that the content of amino acids in Q. fabri was significantly higher than that in the other three oak species. Additionally, we found a significantly higher content of flavor amino acids, such as glutamic acid (Glu), aspartic acid (Asp), and glycine (Gly), in Q. fabri. Considering these results, we designed experiments to assess the nutrient content in mushrooms cultivated from the four oak trees. The results indicated that the total amino acid and protein content of mushrooms cultivated using Q. fabri as a substrate was significantly greater than that of mushrooms grown on the other three oak species. This characteristic may explain why Q. fabri wood is particularly effective as a substrate for cultivating more flavorful mushrooms. This study presents the complete genome and evolutionary information of Q. fabri, and integrates metabolic profiling to explore the underlying reasons for the enhanced flavor of mushrooms cultivated from it.

The photoperiod is essential to flower induction, and the exact timing of the process can be precisely regulated based on the relative duration of light and darkness. However, the mechanisms linking photoperiod and flower induction in woody plants remain largely unexplored. Using RNA-seq, we identified a photoperiod response factor PmNAC32, which is predominantly expressed in early-flowering varieties. Overexpression of PmNAC32 in Arabidopsis thaliana, tobacco, and Prunus mume calli resulted in accelerated flowering. Binding and activation analyses revealed that PmNAC32 can be directly suppressed by REVEILLE 1 (RVE1) and REVEILLE 3 (RVE3), implying that PmNAC32 plays a role in the photoperiodic signaling pathway. Further studies established that PmNAC32 functions as a positive regulator of CONSTANS-LIKE 5 (COL5) and a negative regulator of CONSTANS-LIKE 4 (COL4). Interestingly, we identified two homologs of PmNAC32, namely PmNAC29 and PmNAC47. These three proteins can interact with each other and enhance the regulation of PmCOL4 and PmCOL5. Although PmNAC29 and PmNAC47 can promote flower induction respectively, neither of them responded to the photoperiod. Thus, our results reveal a novel mechanism by which PmNAC32 regulates flower induction in Prunus mume.

Drought is a major abiotic stress. WRKYs are one of the largest families of transcription factors (TFs) in plants. The effects of most WRKYs on developmental regulation and drought adaptation in Citrus remain largely unclear. Citrus reticulata cv. Sanhu hongju (Sanhu) is a drought-tolerant variety from Jiangxi Province, China. Here, we report a differentially expressed CrWRKY57 gene in drought-treated Sanhu leaves through transcriptome analysis. Its transcriptional expression could be induced by abscisic acid (ABA) treatment and water deficit. Overexpression of CrWRKY57 in lemon (Citrus limon) and tobacco (Nicotiana tabacum) confers enhanced drought tolerance, while RNA interference (RNAi)-mediated silencing in Sanhu increases dehydration susceptibility and reduces root volume. Moreover, virus-induced gene silencing-mediated knockdown of CrWRKY57 in Sanhu reduces primary root length and lateral root number by nearly 50% compared to the control. The results of yeast two-hybrid, co-immunoprecipitation assays and bimolecular fluorescence complementation demonstrate that CrWRKY57 interacts with CrABF3, a key TF in ABA signaling. Silencing ClABF3, its homolog in lemon, also increases drought sensitivity and disrupts root system development. Together, CrWRKY57 and CrABF3 directly activate the promoter of the cell cycle gene CrCYCD6;1 by binding to W-box and ABRE elements, respectively. Furthermore, silencing CrCYCD6;1 in Sanhu also severely reduces primary root length and lateral root number. Collectively, our findings provide a new perspective of CrWRKY57 as a positive player in drought response and highlight the role of the CrWRKY57-CrABF-CrCYCD6;1 module in enhancing drought tolerance by modulating root development.

Apple (Malus × domestica) is one of the most popular fruits grown and consumed worldwide, contributing to human health with significant amounts of polyphenols and other bioactive compounds, and providing positive impacts to the economy and society. Understanding the diversity and inheritance of health-active compounds in apple can provide novel selection criteria for future breeding and cultivar development, as consumers increasingly prioritize the health benefits of their food choices. We therefore conducted an untargeted metabolomic analysis using ultra-high-performance liquid chromatography-mass spectrometry (UPLC-MS) to investigate thousands of semipolar chemicals, mainly phenolic compounds, in 439 diverse apple accessions, and quantified 2066 features in positive ion mode. To identify key areas of genetic control for apple metabolite abundance, we performed a metabolomic genome-wide association study (mGWAS) on the quantified mass features using ~280 000 single nucleotide polymorphisms (SNPs). The mGWAS revealed >630 significant loci with hotspots for various groups of known and unknown phenolic compounds including flavonols on Chromosome 1, dihydrochalcones on Chromosome 5, and flavanols on Chromosomes 15 and 16. The most significant hotspot on Chromosome 16 included bHLH and C2H2 transcription factors that may play a role in controlling the abundance and complexity of phenolic compounds through regulation of the flavonoid biosynthesis pathway. Our analysis links the apple metabolome with candidate genes and biosynthetic mechanisms and establishes a foundation for marker-assisted breeding and gene editing to improve and modify phenolic compounds in apple for marketability and the benefit of human health.

Essential genes are crucial for bacterial growth and the pathogenicity. However, how gene essentiality varies under distinct growth conditions and its role in virulence remains poorly understood. In this study, we constructed high-saturation Tn5 transposon mutant libraries for Xanthomonas citri subsp. citri (Xac), the causative agent of citrus canker disease, which results in substantial economic losses to the citrus industry. These libraries were generated under two growth conditions: nutrient-rich medium (NB) and plant-mimicking medium (XVM2). The libraries achieved an average insertion density of 6.85 base pairs, ensuring high-quality coverage. Using these libraries, we identified 568 genes in both bacterial populations as essential genes of Xac, while the 61 (NB) and 54 (XVM2) as conditional essential genes. To validate the findings, two allele-replacement mutants, ∆ublA and ∆pgl, were generated based on variations in insertion density and gap ratios between two growth conditions. These mutants exhibited significant growth defects in XVM2 and compromised virulence in citrus plants, as evidenced by weaker canker symptoms and reduced bacterial populations in planta. Additionally, we performed a comparative analysis of essential gene conservation across the Xanthomonas genus and other species within the class Gammaproteobacteria. A pan-essentialome dataset was assembled from four Xanthomonas strains, revealing a broad pan-essentialome and a precise core-essentialome. To facilitate further research, we developed XanthoBrowser (http://xacdb.lumoxuan.cn/), an online resource that provides user-friendly interface for searching and comparison of essential genes across four Xanthomonas strains. In conclusion, this study provides comprehensive insights into bacterial adaptation mechanisms and highlight potential targets for bactericide development.

Early vigour (EV) and precocity are important traits for orchard establishment and profitability in macadamia. EV determines tree growth and adaptation, while precocity facilitates early yield, offering economic benefits. Although, a positive relationship between these traits has been observed in other tree crops, their association in macadamia remains unclear. This study aimed to identify genetic links between EV and precocity by assessing genetic variability, heritability, and correlations in a 5-year-old macadamia breeding population. The population comprised 904 progenies planted across six sites in Queensland, Australia. Genome-wide association studies (GWAS) were conducted on a subset of 220 accessions genotyped with 7401 SNP markers. A linear mixed model incorporating a kinship matrix and principal components to account for population structure was used to perform association analysis in TASSEL. Phenotypic analyses in ASReml-R revealed that precocity had higher broad- (H² = 0.25-0.84) and narrow-sense (h² = 0.10-0.77) heritability compared to EV (H² = 0-0.61, h² = 0-0.49). EV and precocity showed positive phenotypic (0.25-0.42) and genetic (0.21-0.31) correlations. GWAS identified 11 significant markers (false discovery rate < 0.05), including two pleiotropic markers (Mint10079 and Mint4004) associated with both EV and precocity. Putative genes linked to these markers were involved in cell wall modelling, pathogen defence, abiotic stress tolerance, flowering, overall growth, and development in other tree species. These significant markers, postvalidation, hold substantial promise for utilization in marker-assisted selection (MAS). Integrating putative pleiotropic markers into MAS can enhance genetic gain by reducing the selection time for and enabling simultaneous selection for EV and precocity.

The approximately 20-year breeding cycle has severely restricted the progress of genetic improvement in poplar. Genomic selection (GS) breeding has been demonstrated as an effective approach to accelerate this process. However, its application in forest tree species remains at an early stage. To advance the genetic improvement of target traits in Populus deltoides, the primary species of poplar plantations in China, we systematically implemented GS breeding using 765 hybrid progenies from 32 multi-generational full-sib families. Firstly, we assembled a high-quality genome of one core parent P. deltoides ‘Danhong’, with a genome size of 419.4 Mb and scaffold N50 of 22.0 Mb, which is also the first telomere-to-telomere (T2T) level genome of P. deltoides. Through comparative genomic analysis, we identified 1395 specific structural variants closely associated with growth and development. Subsequently, through genome-wide association studies (GWAS), we identified 135 quantitative trait nucleotides (QTNs) associated with growth and wood quality traits. By systematically evaluating reference genomes, statistical models, and various marker selection strategies, we developed optimal genomic prediction (GP) models for six traits, with the highest prediction accuracy (PA) reaching 0.730 for DBH. Compared with using all markers, the PA was improved by an average of 136.34%. Furthermore, by integrating GP, GWAS, and RNA-seq results, we identified core breeding parents and elite clones for P. deltoides genetic improvement and discovered important candidate genes. Our results provide a promising strategy for accelerating breeding cycles and genetic improvement, offering valuable breeding and genetic resources for forest tree improvement.

Most genomic studies start by mapping sequencing data to a reference genome. The quality of reference genome assembly, genetic relatedness to the studied population, and the mapping method employed directly impact variant calling accuracy and subsequent genomic analyses, introducing reference bias and resulting in erroneous conclusions. However, the impacts of reference bias have gained limited attention. This study compared population genomic analyses using four different reference genomes of mango (Mangifera indica), including the two haploid assemblies of haplotype-resolved telomere-to-telomere (T2T) genome assembly, a pangenome, and an older version of the reference genome available on NCBI. The choice of reference genome dramatically impacted the mapping efficiency and resulted in notable differences in calling the genetic variants, particularly structural variations (SVs). Phylogenetic analysis was more sensitive to the reference genome compared to genetic differentiation. Population genomic analyses of artificial selection in domestication and SV hotspot regions varied across reference genomes. Notably, the gene enrichment analyses showed significant differences in the top enriched biological processes depending on the reference genome used. Overall, the mango pangenome outperformed the other reference genomes across various metrics, followed by T2T reference genomes, as they captured greater diversity and effectively reduced reference bias. Our findings highlight the role of the mango pangenome in reducing reference bias and underscore the critical role of reference genome selection, suggesting that it is one of the most important factors in population genomic studies.

The MEGGIC (Magic EGGplant InCanum) population here presented is the first highly inbred eggplant (Solanum melongena) multiparent advanced generation intercross (MAGIC) population developed so far, derived from seven cultivated accessions and one wild Solanum incanum from arid regions. The final 325 S5 lines were high-throughput genotyped using low-coverage whole-genome sequencing (lcWGS) at 3X, yielding 293 783 high-quality SNPs after stringent filtering. Principal component analysis (PCA) and neighbor-joining clustering revealed extensive genetic diversity driven by the unique genetic profile of the wild founder, and lack of genetic structure, suggesting a well-mixed population with a high degree of recombination. The eight founders and a core subset of 212 lines were phenotyped for above-and belowground traits, revealing wide phenotypic diversity. Root morphology traits displayed moderate heritability values, and strong correlation were found between root and aerial traits, suggesting that a well-developed root system supports greater aboveground growth. Genome-wide association studies (GWAS) identified a genomic region on chromosome 6 associated with root biomass (RB), total root length (RL), and root surface area (SA). Within this region, SmLBD13, an LOB-domain protein involved in lateral root development, was identified as a candidate gene. The S. incanum haplotype in this region was linked to reduced lateral root branching density, a trait that may enhance deeper soil exploration and resource uptake. These findings provide key insights into root genetics in eggplant, demonstrating MEGGIC potential for high-resolution trait mapping. Furthermore, they highlight the role of exotic wild germplasm in breeding more resilient cultivars and rootstocks with improved root architecture and enhanced nutrient uptake efficiency.

Pear propagation is primarily achieved through asexual reproduction via grafting. During the graft union healing process, there is metabolic exchange between the rootstock and the scion. However, a multi-omics systematic study on the role of sugar in the graft union healing process has not been reported. In this study, using micrografting techniques, we comparatively analyzed the metabolic changes during the healing process in homograft and heterograft of pear through metabolomics and transcriptomics. We found significant differences in sugar metabolism pathways after grafting. In the fructose and mannose metabolic pathways, sorbitol exhibited opposite trends in homograft and heterograft. Subsequent transcriptomics analysis confirmed that these metabolite changes were caused by differential expression of related synthetic and converting enzyme genes. Furthermore, spatial metabolomics identified sorbitol accumulation in the scion after homologous grafting. To further verify the role of sorbitol, exogenous sorbitol treatment was applied, revealing that it enhanced tissue adhesion, shortened the time required for callus growth, promoted high expression of xylem formation genes and cambium differentiation genes, and facilitated the reconnection of xylem and phloem, thereby playing a positive role in graft union healing. This study systematically analyzed changes in sugar metabolism during the grafting process and confirmed that sorbitol can promote graft union healing.