Comparative Morpho-Physiological, Biochemical, and Gene Expressional Analyses Uncover Mechanisms of Waterlogging Tolerance in Two Soybean Introgression Lines.

Waterlogging is one of the key abiotic factors that severely impedes the growth and productivity of soybeans on a global scale. To develop soybean cultivars that are tolerant to waterlogging, it is a prerequisite to unravel the mechanisms governing soybean responses to waterlogging. Hence, we explored the morphological, physiological, biochemical, and transcriptional changes in two contrasting soybean introgression lines, A192 (waterlogging tolerant, WT) and A186 (waterlogging sensitive, WS), under waterlogging. In comparison to the WT line, waterlogging drastically decreased the root length (RL), shoot length (ShL), root fresh weight (RFW), shoot fresh weight (ShFW), root dry weight (RDW), and shoot dry weight (ShDW) of the WS line. Similarly, waterlogging inhibited soybean plant growth by suppressing the plant's photosynthetic capacity, enhancing oxidative damage from reactive oxygen species, and decreasing the chlorophyll content in the WS line but not in the WT line. To counteract the oxidative damage and lipid peroxidation, the WT line exhibited increased activity of antioxidant enzymes such as peroxidase (POD), superoxide dismutase (SOD), and catalase (CAT), as well as higher levels of proline content than the WS line. In addition, the expression of antioxidant enzyme genes (POD1, POD2, FeSOD, Cu/ZnSOD, CAT1, and CAT2) and ethylene-related genes (such as ACO1, ACO2, ACS1, and ACS2) were found to be up-regulated in WT line under waterlogging stress conditions. In contrast, these genes showed a down-regulation in their expression levels in the stressed WS line. The integration of morpho-physiological, biochemical, and gene expression analyses provide a comprehensive understanding of the responses of WT and WS lines to waterlogging conditions. These findings would be beneficial for the future development of soybean cultivars that can withstand waterlogging.

T2T reference genome assembly and genome-wide association study reveal the genetic basis of Chinese bayberry fruit quality.

Chinese bayberry (Myrica rubra or Morella rubra; 2n = 16) produces fruit with a distinctive flavor, high nutritional, and economic value. However, previous versions of the bayberry genome lack sequence continuity. Moreover, to date, no large-scale germplasm resource association analysis has examined the allelic and genetic variations determining fruit quality traits. Therefore, in this study, we assembled a telomere-to-telomere (T2T) gap-free reference genome for the cultivar 'Zaojia' using PacBio HiFi long reads. The resulting 292.60 Mb T2T genome, revealed 8 centromeric regions, 15 telomeres, and 28 345 genes. This represents a substantial improvement in the genome continuity and integrity of Chinese bayberry. Subsequently, we re-sequenced 173 accessions, identifying 6 649 674 single nucleotide polymorphisms (SNPs). Further, the phenotypic analyses of 29 fruit quality-related traits enabled a genome-wide association study (GWAS), which identified 1937 SNPs and 1039 genes significantly associated with 28 traits. An SNP cluster pertinent to fruit color was identified on Chr6: 3407532 to 5 153 151 bp region, harboring two MYB genes (MrChr6G07650 and MrChr6G07660), exhibiting differential expression in extreme phenotype transcriptomes, linked to anthocyanin synthesis. An adjacent, closely linked gene, MrChr6G07670 (MLP-like protein), harbored an exonic missense variant and was shown to increase anthocyanin production in tobacco leaves tenfold. This SNP cluster, potentially a quantitative trait locus (QTL), collectively regulates bayberry fruit color. In conclusion, our study presented a complete reference genome, uncovered a suite of allelic variations related to fruit-quality traits, and identified functional genes that could be harnessed to enhance fruit quality and breeding efficiency of bayberries.

Toxicologic effect and transcriptome analysis for sub-chronic exposure to carbendazim, prochloraz, and their combination on the liver of mice.

Carbendazim (CBZ) and prochloraz (PCZ) are broad-spectrum fungicides used in agricultural peat control. Both fungicides leave large amounts of residues in fruits and are toxic to non-target organisms. However, the combined toxicity of the fungicides to non-target organisms is still unknown. Therefore, we characterized the toxic effects of dietary supplementation with CBZ, PCZ, and their combination for 90 days in 6-week-old male Institute of Cancer Research (ICR) mice. CBZ-H (100 mg/kg day), PCZ-H (10 mg/kg day), and their combination treatments increased the relative liver weights and caused liver injury. The serum total cholesterol (TC), triglyceride (TG), glucose (Glu), pyruvate (PYR), low-density lipoprotein cholesterol (LDL-C), and high-density lipoprotein cholesterol (HDL-C) levels were reduced, and synergistic toxicity was observed. Hepatic transcriptome revealed that 326 differentially expressed genes (DEGs) of liver were observed in the CBZ treatment group, 149 DEGs in the PCZ treatment group, and 272 DEGs in the combination treatment group. According to KEGG enrichment analysis, the fungicides and their combination affected lipid metabolism, amino acid metabolism, and ferroptosis. In addition, the relative mRNA levels of key genes involved in lipid metabolism were also examined. Compared with individual exposure, combined exposure to CBZ and PCZ caused a more obvious decrease in the expression of some genes related to glycolipid metabolism. Furthermore, the relative mRNA levels of some key genes in the combination treatment group were lower than those in the CBZ and PCZ treated groups. In summary, CBZ, PCZ, and their combination generally caused hepatotoxicity and glycolipid metabolism disorders, which could provide new insights for investigating the combined toxicity of multiple fungicides to animals.

The Effects of Accompanying Ryegrass on Bayberry Trees by Change of Soil Property, Rhizosphere Microbial Community Structure, and Metabolites.

As a subtropical and tropical tree, bayberry (Myrica rubra) is an important fruit tree grown commercially in southern China. Interestingly, our studies found that the fruit quality of bayberry with accompanying ryegrass was significantly improved, but its mechanism remains unclear. The aim of this study was to explore the mechanism of accompanying ryegrass on the beneficial effect of the fruit quality of bayberry by measuring the vegetative growth parameters, fruit parameters with economic impact, physical and chemical properties of rhizosphere soil, microbial community structure, and metabolites of the bayberry with/without ryegrass. Notably, the results revealed a significant difference between bayberry trees with and without accompanying ryegrass in fruit quality parameters, soil physical and chemical properties, microbial community structure, and metabolites. Compared with the control without accompanying ryegrass, the planting of ryegrass increased the titratable sugar, vitamin C, and titratable flavonoid contents of bayberry fruits by 2.26%, 28.45%, and 25.00%, respectively, and decreased the titratable acid contents by 9.04%. Furthermore, based on 16S and ITS amplicon sequencing of soil microflora, the accompanying ryegrass caused a 12.47% increment in Acidobacteriota while a 30.04% reduction in Actinobacteria was recorded, respectively, when compared with the bayberry trees without ryegrass. Redundancy discriminant analysis of microbial communities and soil properties indicated that the main variables of the bacterial community included available nitrogen, available phosphorus, exchangeable aluminum, and available kalium, while the main variables of the fungal community included exchangeable aluminum, available phosphorus, available kalium, and pH. In addition, the change in microbial community structure was justified by the high correlation analysis between microorganisms and secondary metabolites. Indeed, GC-MS metabolomics analysis showed that planting ryegrass caused a 3.83%-144.36% increase in 19 metabolites such as 1,3-Dipentyl-heptabarbital and carbonic acid 1, respectively, and a 23.78%-51.79% reduction of 5 metabolites compared to the bayberry trees without the accompanying ryegrass. Overall, the results revealed the significant change caused by the planting of ryegrass in the physical and chemical properties, microbiota, and secondary metabolites of the bayberry rhizosphere soils, which provides a new insight for the ecological improvement of bayberry.

The completed genome sequence of Pestalotiopsis versicolor, a pathogenic ascomycete fungus with implications for bayberry production.

The pathogenic fungus Pestalotiopsis versicolor is a major etiological agent of fungal twig blight disease affecting bayberry trees. However, the lack of complete genome sequence information for this crucial pathogenic fungus hinders the molecular and genetic investigation of its pathogenic mechanism. To address this knowledge gap, we have generated the complete genome sequence of P. versicolor strain XJ27, employing a combination of Illumina, PacBio, and Hi-C sequencing technologies. This comprehensive genome sequence, comprising 7 chromosomes with an N50 contig size of 7,275,017 bp, a GC content ratio of 50.16%, and a total size of 50.80 Mb, encompasses 13,971 predicted coding genes. By performing comparative genomic analysis between P. versicolor and the genomes of eleven plant-pathogenic fungi, as well as three closely related fungi within the same group, we have gained initial insights into its evolutionary trajectory, particularly through gene family analysis. These findings shed light on the distinctive characteristics and evolutionary history of P. versicolor. Importantly, the availability of this high-quality genetic resource will serve as a foundational tool for investigating the biology, molecular pathogenesis, and virulence of P. versicolor. Furthermore, it will facilitate the development of more potent antifungal medications by uncovering potential vulnerabilities in its genetic makeup.

The MAP Kinase PvMK1 Regulates Hyphal Development, Autophagy, and Pathogenesis in the Bayberry Twig Blight Fungus Pestalotiopsis versicolor.

Bayberry twig blight caused by the ascomycete fungus Pestalotiopsis versicolor is a devastating disease threatening worldwide bayberry production. However, the molecular basis underlying the pathogenesis of P. versicolor is largely unknown. Here, we identified and functionally characterized the MAP kinase PvMk1 in P. versicolor through genetic and cellular biochemical approaches. Our analysis reveals a central role of PvMk1 in regulating P. versicolor virulence on bayberry. We demonstrate that PvMk1 is involved in hyphal development, conidiation, melanin biosynthesis, and cell wall stress responses. Notably, PvMk1 regulates P. versicolor autophagy and is essential for hyphal growth under nitrogen-depleting conditions. These findings suggest the multifaceted role of PvMk1 in regulating P. versicolor development and virulence. More remarkably, this evidence of virulence-involved cellular processes regulated by PvMk1 has paved a fundamental way for further understanding the impact of P. versicolor pathogenesis on bayberry.

Genetic Dissection of Extreme Seed-Flooding Tolerance in a Wild Soybean PI342618B by Linkage Mapping and Candidate Gene Analysis.

Seed-flooding stress is one of major abiotic constraints that adversely affects soybean production worldwide. Identifying tolerant germplasms and revealing the genetic basis of seed-flooding tolerance are imperative goals for soybean breeding. In the present study, high-density linkage maps of two inter-specific recombinant inbred line (RIL) populations, named NJIRNP and NJIR4P, were utilized to identify major quantitative trait loci (QTLs) for seed-flooding tolerance using three parameters viz., germination rate (GR), normal seedling rate (NSR), and electrical conductivity (EC). A total of 25 and 18 QTLs were detected by composite interval mapping (CIM) and mixed-model-based composite interval mapping (MCIM), respectively, and 12 common QTLs were identified through both methods. All favorable alleles for the tolerance are notably from the wild soybean parent. Moreover, four digenic epistatic QTL pairs were identified, and three of them showed no main effects. In addition, the pigmented soybean genotypes exhibited high seed-flooding tolerance compared with yellow seed coat genotypes in both populations. Moreover, out of five identified QTLs, one major region containing multiple QTLs associated with all three traits was identified on Chromosome 8, and most of the QTLs within this hotspot were major loci (R2 > 10) and detectable in both populations and multiple environments. Based on the gene expression and functional annotation information, 10 candidate genes from QTL "hotspot 8-2" were screened for further analysis. Furthermore, the results of qRT-PCR and sequence analysis revealed that only one gene, GmDREB2 (Glyma.08G137600), was significantly induced under flooding stress and displayed a TTC tribasic insertion mutation of the nucleotide sequence in the tolerant wild parent (PI342618B). GmDREB2 encodes an ERF transcription factor, and the subcellular localization analysis using green fluorescent protein (GFP) revealed that GmDREB2 protein was localized in the nucleus and plasma membrane. Furthermore, overexpression of GmDREB2 significantly promoted the growth of soybean hairy roots, which might indicate its critical role in seed-flooding stress. Thus, GmDREB2 was considered as the most possible candidate gene for seed-flooding tolerance.

Conserved hierarchical gene regulatory networks for drought and cold stress response in Myrica rubra.

Stress response in plant is regulated by a large number of genes co-operating in diverse networks that serve multiple adaptive process. To understand how gene regulatory networks (GRNs) modulating abiotic stress responses, we compare the GRNs underlying drought and cold stresses using samples collected at 4 or 6 h intervals within 48 h in Chinese bayberry (Myrica rubra). We detected 7,583 and 8,840 differentially expressed genes (DEGs) under drought and cold stress respectively, which might be responsive to environmental stresses. Drought- and cold-responsive GRNs, which have been built according to the timing of transcription under both abiotic stresses, have a conserved trans-regulator and a common regulatory network. In both GRNs, basic helix-loop-helix family transcription factor (bHLH) serve as central nodes. MrbHLHp10 transcripts exhibited continuous increase in the two abiotic stresses and acts upstream regulator of ASCORBATE PEROXIDASE (APX) gene. To examine the potential biological functions of MrbHLH10, we generated a transgenic Arabidopsis plant that constitutively overexpresses the MrbHLH10 gene. Compared to wild-type (WT) plants, overexpressing transgenic Arabidopsis plants maintained higher APX activity and biomass accumulation under drought and cold stress. Consistently, RNAi plants had elevated susceptibility to both stresses. Taken together, these results suggested that MrbHLH10 mitigates abiotic stresses through the modulation of ROS scavenging.

Molecular cloning and functional analysis of Chinese bayberry MrSPL4 that enhances growth and flowering in transgenic tobacco.

Chinese bayberry (Myrica rubra) is an important tree in South China, with its fruit being of nutritional and high economic value. In this study, early ripening (ZJ), medium ripening (BQ) and late ripening (DK) varieties were used as test materials. Young leaves of ZJ, BQ and DK in the floral bud morphological differentiation periods were selected for transcriptome sequencing to excavate earliness related genes. A total of 4,538 differentially expressed genes were detected. Based on clustering analysis and comparisons with genes reportedly related to flowering in Arabidopsis thaliana, 25 homologous genes were identified. Of these, one gene named MrSPL4 was determined, with its expression down-regulated in DK but up-regulated in ZJ and BQ. MrSPL4 contained SBP domain and the target site of miR156, and its total and CDS length were 1,664 bp and 555 bp respectively. The overexpression vector of MrSPL4 (35S::35S::MrSPL4-pCambia2301-KY) was further constructed and successfully transfected into tobacco to obtain MrSPL4-positive plants. Based on the results of qRT-PCR, the relative expression of MrSPL4 was up regulated by 3,862.0-5,938.4 times. Additionally, the height of MrSPL4-positive plants was also significantly higher than that of wild-type (WT), with the bud stage occurring 12 days earlier. Altogether, this study identified an important gene -MrSPL4 in Chinese bayberry, which enhanced growth and flowering, which provided important theoretical basis for early-mature breeding of Chinese bayberry.

Alcohol extracts of Chinese bayberry branch induce S-phase arrest and apoptosis in HepG2 cells.

The alcohol extracts of Chinese bayberry (Myrica rubra) branches (MRBE) are rich in flavonoids which have a variety of medicinal benefits, but their effects on human HepG2 were unknown. In this study, the effects of MRBE on HepG2 cell growth and its potential for inhibiting cancer were explored. The results displayed that MRBE inhibited HepG2 proliferation both by arresting cells in S phase and promoting apoptosis. Quantitative reverse-transcription PCR (qRT-PCR), western blotting, and immunofluorescence showed that MRBE induced S-phase arrest by upregulating p21, which in turn downregulated cyclin and cyclin-dependent kinase messenger RNA (mRNA) and protein. Apoptosis was induced by blocking the expression of BCL-2 and suppression of the Raf/ERK1 signaling pathways. These results indicated that MRBE may have the potential for treatment of human liver cancer, highlighting novel approaches for developing new pharmacological tools for the treatment of this deadly type cancer. Meanwhile, it provides a new direction for the medicinal added values of Chinese bayberry, which helped to broaden the diversified development of its industry chain.

Effects of Enhanced Resistance and Transcriptome Analysis of Twig Blight Disease by Exogenous Brassinolide in Myrica rubra.

Twig blight disease is the primary disease that affects the production of Myrica rubra in China. It was reported that exogenous brassinolide (BL) can improve disease resistance in plants. Here, we examined the effects of exogenous BL on disease resistance, chlorophyll contents, antioxidant enzyme activity, ROS accumulation, and key gene expression of M. rubra to analyze the mechanism of BR-induced resistance of twig blight disease in M. rubra. The results demonstrated that 2.0 mg·L-1 of BL could significantly lessen the severity of twig blight disease in M. rubra. Exogenous BL increased the contents of chlorophyll a, chlorophyll b, carotenoids, and total chlorophyll. Moreover, exogenous BL also significantly enhanced the activity of antioxidant enzymes such as superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT), and decreased malondialdehyde (MDA) content and reactive oxygen species (ROS) accumulation in leaves, such as H2O2 and O2·-. Additionally, exogenous BL dramatically up-regulated the expression of pathogenesis-related (PR) genes such as MrPR1, MrPR2, and MrPR10, as well as important genes such as MrBAK1, MrBRI1, and MrBZR1 involved in brassinosteroid (BR) signaling pathway. The transcriptome analysis revealed that a total of 730 common differentially expressed genes (DEGs) under BL treatment were found, and these DEGs were primarily enriched in four Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways. Based on these findings, nine important candidate genes related to the resistance of twig blight disease under BL treatment were further identified. In this study, we elucidated the effects of exogenous BL on enhancing the resistance of M. rubra to twig blight disease and preliminary analyzed the potential mechanism of resistance induction, which will provide a crucial foundation for the management and prevention of twig blight disease in M. rubra.

An overview of the nutritional value, health properties, and future challenges of Chinese bayberry.

Chinese bayberry (CB) is among the most popular and valuable fruits in China owing to its attractive color and unique sweet/sour taste. Recent studies have highlighted the nutritional value and health-related benefits of CB. CB has special biological characteristics of evergreen, special aroma, dioecious, nodulation, nitrogen fixation. Moreover, the fruits, leaves, and bark of CB plants harbor a number of bioactive compounds including proanthocyanidins, flavonoids, vitamin C, phenolic acids, and anthocyanins that have been linked to the anti-cancer, anti-oxidant, anti-inflammatory, anti-obesity, anti-diabetic, and neuroprotective properties and to the treatment of cardiovascular and cerebrovascular diseases. The CB fruits have been used to produce a range of products: beverages, foods, and washing supplies. Future CB-related product development is thus expected to further leverage the health-promoting potential of this valuable ecological resource. The present review provides an overview of the botanical characteristics, processing, nutritional value, health-related properties, and applications of CB in order to provide a foundation for further research and development.

A novel mitochondrial protein is required for cell wall integrity, auxin accumulation and root elongation in Arabidopsis under salt stress.

Maintenance of root elongation is beneficial for the growth and survival of plants under salt stress, but currently the cellular components involved in the regulation of root growth under high salinity are not fully understood. In this study, we identified an Arabidopsis mutant, rres1, which exhibited reduced root elongation under treatment of a variety of salts, including NaCl, NaNO3, KCl, and KNO3. RRES1 encodes a novel mitochondrial protein and its molecular function is still unknown. Under salt stress, the root meristem length was shorter in the rres1 mutant compared to the wild type, which was correlated with a reduced auxin accumulation in the mutant. Reactive oxygen species (ROS), as important signals that regulate root elongation, were accumulated to higher levels in the rres1 mutant than the wild type after salt treatment. Measurement of monosaccharides in the cell wall showed that arabinose and xylose contents were decreased in the rres1 mutant under salt stress, and application of boric acid, which is required for the crosslinking of pectic polysaccharide rhamnogalacturonan-II (RG-II), largely rescued the root growth arrest of the rres1 mutant, suggesting that RRES1 participates in the maintenance of cell wall integrity under salt stress. GUS staining assay indicated that the RRES1 gene was expressed in leaves and weakly in root tip under normal conditions, but its expression was dramatically increased in leaves and roots after salt treatment. Together, our study reveals a novel mitochondrial protein that regulates root elongation under salt stress via the modulation of cell wall integrity, auxin accumulation, and ROS homeostasis.

Effect of Two Kinds of Fertilizers on Growth and Rhizosphere Soil Properties of Bayberry with Decline Disease.

Decline disease causes severe damage to bayberry. However, the cause of this disease remains unclear. Interestingly, our previous studies found that the disease severity is related with the level of soil fertilizer. This study aims to explore the effect and mechanism of compound fertilizer (CF) and bio-organic fertilizer (OF) in this disease by investigating the vegetative growth, fruit characters, soil property, rhizosphere microflora and metabolites. Results indicated that compared with the disease control, CF and OF exhibited differential effect in plant healthy and soil quality, together with the increase in relative abundance of Burkholderia and Mortierella, and the reduction in that of Rhizomicrobium and Acidibacter, Trichoderma, and Cladophialophora reduced. The relative abundance of Geminibasidium were increased by CF (251.79%) but reduced by OF (13.99%). In general, the composition of bacterial and fungal communities in rhizosphere soil was affected significantly at genus level by exchangeable calcium, available phosphorus, and exchangeable magnesium, while the former two variables had a greater influence in bacterial communities than fungal communities. Analysis of GC-MS metabonomics indicated that compared to the disease control, CF and OF significantly changed the contents of 31 and 45 metabolites, respectively, while both fertilizers changed C5-branched dibasic acid, galactose, and pyrimidine metabolic pathway. Furthermore, a significant correlation was observed at the phylum, order and genus levels between microbial groups and secondary metabolites of bayberry rhizosphere soil. In summary, the results provide a new way for rejuvenation of this diseased bayberry trees.

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Twig blight is a serious disease of bayberry, which limits bayberry production. In order to prevent and manage the disease, we used high-throughput sequencing technology to analyze bacterial and fungal community richness and diversity in various organs of the tree, bulk and rhizosphere soil of healthy and diseased bayberry. The results showed significant differences in richness and diversity of bacteria and fungi in bulk soil, rhizosphere soil, roots, trunks, barks, and leaves between twig blight diseased trees and healthy trees. In bulk soil, the richness and diversity of bacteria significantly decreased, while that of fungi significantly increased. In barks of diseased trees, the richness and diversity of bacteria significantly increased, and those of fungi significantly decreased. The relative abundance of dominant bacteria and fungi in various organs, bulk soil, and root surface soil changed significantly at phylum, class, and genus levels in the diseased trees. The relative abundance of Pseudomonas sp. on the trunks, roots, and root surface soils of diseased trees significantly decreased, and Fusarium sp. of the diseased root surface and bulk soils also significantly decreased, while the relative abundance of Penicillium sp. on the diseased root surface and bulk soils significantly increased. Pestalotiopsis sp., from the same genus as the twig blight pathogen, was less abundant in the roots but more abundant in the leaves, trunks, barks as well as root surface soils and bulk soils of the diseased bayberry trees than those of the health trees. The relative abundance of Pestalotiopsis sp. was positively correlated with those of most of the fungi. Our results might provide useful theoretical basis for the development of ecological improvement and healthy-tree cultivation technology, and biological control of bayberry twig blight disease.

The bayberry database: a multiomic database for Myrica rubra, an important fruit tree with medicinal value.

BACKGROUND: Chinese bayberry (Myrica rubra Sieb. & Zucc.) is an important fruit tree in China, and has high medicinal value. At present, the genome, transcriptome and germplasm resources of bayberry have been reported. In order to make more convenient use of these data, the Bayberry Database was established. RESULTS: The Bayberry Database is a comprehensive and intuitive data platform for examining the diverse annotated genome and germplasm resources of this species. This database contains nine central functional domains to interact with multiomic data: home, genome, germplasm, markers, tools, map, expression, reference, and contact. All domains provide pathways to a variety of data types composed of a reference genome sequence, transcriptomic data, gene patterns, phenotypic data, fruit images of Myrica rubra varieties, gSSR data, gene maps with annotation and evolutionary analyses. The tools module includes BLAST search, keyword search, sequence fetch and enrichment analysis functions. CONCLUSIONS: The web address of the database is as follows http://www.bayberrybase.cn/ . The Myrica rubra database is an intelligent, interactive, and user-friendly system that enables researchers, breeders and horticultural personnel to browse, search and retrieve relevant and useful information and thus facilitate genomic research and breeding efforts concerning Myrica rubra. This database will be of great help to bayberry research and breeding in the future.

The Damage Caused by Decline Disease in Bayberry Plants through Changes in Soil Properties, Rhizosphere Microbial Community Structure and Metabolites.

Decline disease causes serious damage and rapid death in bayberry, an important fruit tree in south China, but the cause of this disease remains unclear. The aim of this study was to investigate soil quality, microbial community structure and metabolites of rhizosphere soil samples from healthy and diseased trees. The results revealed a significant difference between healthy and diseased bayberry in soil properties, microbial community structure and metabolites. Indeed, the decline disease caused a 78.24% and 78.98% increase in Rhizomicrobium and Cladophialophora, but a 28.60%, 57.18%, 38.84% and 68.25% reduction in Acidothermus, Mortierella, Trichoderma and Geminibasidium, respectively, compared with healthy trees, based on 16S and ITS amplicon sequencing of soil microflora. Furthermore, redundancy discriminant analysis of microbial communities and soil properties indicated that the main variables of bacterial and fungal communities included pH, organic matter, magnesium, available phosphorus, nitrogen and calcium, which exhibited a greater influence in bacterial communities than in fungal communities. In addition, there was a high correlation between the changes in microbial community structure and secondary metabolites. Indeed, GC-MS metabolomics analysis showed that the healthy and diseased samples differed over six metabolic pathways, including thiamine metabolism, phenylalanine-tyrosine-tryptophan biosynthesis, valine-leucine-isoleucine biosynthesis, phenylalanine metabolism, fatty acid biosynthesis and fatty acid metabolism, where the diseased samples showed a 234.67% and 1007.80% increase in palatinitol and cytidine, respectively, and a 17.37-8.74% reduction in the other 40 metabolites compared to the healthy samples. Overall, these results revealed significant changes caused by decline disease in the chemical properties, microbiota and secondary metabolites of the rhizosphere soils, which provide new insights for understanding the cause of this bayberry disease.

Construction of a High-Density Genetic Map and Identification of Leaf Trait-Related QTLs in Chinese Bayberry (Myrica rubra).

Chinese bayberry (Myrica rubra) is an economically important fruit tree that is grown in southern China. Owing to its over 10-year seedling period, the crossbreeding of bayberry is challenging. The characteristics of plant leaves are among the primary factors that control plant architecture and potential yields, making the analysis of leaf trait-related genetic factors crucial to the hybrid breeding of any plant. In the present study, molecular markers associated with leaf traits were identified via a whole-genome re-sequencing approach, and a genetic map was thereby constructed. In total, this effort yielded 902.11 Gb of raw data that led to the identification of 2,242,353 single nucleotide polymorphisms (SNPs) in 140 F1 individuals and parents (Myrica rubra cv. Biqizhong × Myrica rubra cv. 2012LXRM). The final genetic map ultimately incorporated 31,431 SNPs in eight linkage groups, spanning 1,351.85 cM. This map was then used to assemble and update previous scaffold genomic data at the chromosomal level. The genome size of M. rubra was thereby established to be 275.37 Mb, with 94.98% of sequences being assembled into eight pseudo-chromosomes. Additionally, 18 quantitative trait loci (QTLs) associated with nine leaf and growth-related traits were identified. Two QTL clusters were detected (the LG3 and LG5 clusters). Functional annotations further suggested two chlorophyll content-related candidate genes being identified in the LG5 cluster. Overall, this is the first study on the QTL mapping and identification of loci responsible for the regulation of leaf traits in M. rubra, offering an invaluable scientific for future marker-assisted selection breeding and candidate gene analyses.

Identification of QTN and Candidate Gene for Seed-flooding Tolerance in Soybean [Glycine max (L.) Merr.] using Genome-Wide Association Study (GWAS).

Seed-flooding stress is one of the major abiotic constraints severely affecting soybean yield and quality. Understanding the molecular mechanism and genetic basis underlying seed-flooding tolerance will be of greatly importance in soybean breeding. However, very limited information is available about the genetic basis of seed-flooding tolerance in soybean. The present study performed Genome-Wide Association Study (GWAS) to identify the quantitative trait nucleotides (QTNs) associated with three seed-flooding tolerance related traits, viz., germination rate (GR), normal seedling rate (NSR) and electric conductivity (EC), using a panel of 347 soybean lines and the genotypic data of 60,109 SNPs with MAF > 0.05. A total of 25 and 21 QTNs associated with all three traits were identified via mixed linear model (MLM) and multi-locus random-SNP-effect mixed linear model (mrMLM) in three different environments (JP14, HY15, and Combined). Among these QTNs, three major QTNs, viz., QTN13, qNSR-10 and qEC-7-2, were identified through both methods MLM and mrMLM. Interestingly, QTN13 located on Chr.13 has been consistently identified to be associated with all three studied traits in both methods and multiple environments. Within the 1.0 Mb physical interval surrounding the QTN13, nine candidate genes were screened for their involvement in seed-flooding tolerance based on gene annotation information and available literature. Based on the qRT-PCR and sequence analysis, only one gene designated as GmSFT (Glyma.13g248000) displayed significantly higher expression level in all tolerant genotypes compared to sensitive ones under flooding treatment, as well as revealed nonsynonymous mutation in tolerant genotypes, leading to amino acid change in the protein. Additionally, subcellular localization showed that GmSFT was localized in the nucleus and cell membrane. Hence, GmSFT was considered as the most likely candidate gene for seed-flooding tolerance in soybean. In conclusion, the findings of the present study not only increase our knowledge of the genetic control of seed-flooding tolerance in soybean, but will also be of great utility in marker-assisted selection and gene cloning to elucidate the mechanisms of seed-flooding tolerance.