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Fruit aroma is an important organoleptic quality, which influences consumer preference and market competitiveness. Aroma compound synthesis pathways in plants have been widely identified, among the lipoxygenase pathway is crucial for fatty acid catabolism to form esters in apple. However, the regulatory mechanism of this pathway remains elusive. In this study, linear regression analysis and transgene verification revealed that the lipoxygenase MdLOX1a is involved in ester biosynthesis. Yeast one-hybrid library screening indicated that a protein, MdASG1 (ABIOTIC STRESS GENE 1), was a positive regulator of MdLOX1a and ester production based on yeast one-hybrid and dual-luciferase assays, as well as correlation analysis among eight different apple cultivars. Overexpression of MdASG1 in apple and tomato stimulated the lipoxygenase pathway and increased the fatty acid-derived volatile content, whereas the latter was decreased by MdASG1 silencing and CRISPR/Cas9 knockout. Furthermore, MdASG1 overexpression enhanced the salt-stress tolerance of tomato and apple 'Orin' calli accompanied by a higher content of fatty acid-derived volatiles compared to that of non-stressed transgenic tomato fruit, while MdASG1 -Cas9 knockdown calli do not respond to salt stress and promote the biosynthesis of fatty acid-derived volatiles. Collectively, these findings indicate that MdASG1 activates MdLOX1a expression and participates in the lipoxygenase pathway, subsequently increasing the accumulation of aroma compounds, especially under moderate salt stress treatment. The results also provide insight into the theory for improving fruit aroma quality in adversity., Competing Interests: The authors declare no conflict of interests., (© The Author(s) 2024. Published by Oxford University Press on behalf of Nanjing Agricultural University.)

Malus sieversii is a precious apple germplasm resource. Browning of explants is one of the most important factors limiting the survival rate of plant tissue culture. In order to explore the molecular mechanism of the browning degree of different strains of Malus sieversii, we compared the dynamic changes of Malus sieversii and Malus robusta Rehd. during the whole browning process using a multi-group method. A total of 44 048 differentially expressed genes (DEGs) were identified by transcriptome analysis on the DNBSEQ-T7 sequencing platform. KEGG enrichment analysis showed that the DEGs were significantly enriched in the flavonoid biosynthesis pathway. In addition, metabonomic analysis showed that (-)-epicatechin, astragalin, chrysin, irigenin, isoquercitrin, naringenin, neobavaisoflavone and prunin exhibited different degrees of free radical scavenging ability in the tissue culture browning process, and their accumulation in different varieties led to differences in the browning degree among varieties. Comprehensive transcriptome and metabonomics analysis of the data related to flavonoid biosynthesis showed that PAL, 4CL, F3H, CYP73A, CHS, CHI, ANS, DFR and PGT1 were the key genes for flavonoid accumulation during browning. In addition, WGCNA analysis revealed a strong correlation between the known flavonoid structure genes and the selected transcriptional genes. Protein interaction predictions demonstrated that 19 transcription factors (7 MYBs and 12 bHLHs) and 8 flavonoid structural genes had targeted relationships. The results show that the interspecific differential expression of flavonoid genes is the key influencing factor of the difference in browning degree between Malus sieversii and Malus robusta Rehd., providing a theoretical basis for further study on the regulation of flavonoid biosynthesis., (© 2024 Scandinavian Plant Physiology Society.)

Apple replant disease (ARD) is common in apple production, which seriously affects the growth and development of apples. In this study, hydrogen peroxide with a bactericidal effect was used to treat the replanted soil, and the effects of different concentrations of hydrogen peroxide on replanted seedlings and soil microbiology were investigated in order to seek a green, clean way to control ARD. Five treatments were set up in this study: replanted soil (CK1), replanted soil with methyl bromide fumigation (CK2), replanted soil + 1.5% hydrogen peroxide (H1), replanted soil + 3.0% hydrogen peroxide (H2), and replanted soil + 4.5% hydrogen peroxide (H3). The results showed that hydrogen peroxide treatment improved replanted seedling growth and also inactivated a certain number of Fusarium , while the Bacillus , Mortierella , and Guehomyces also became more abundant in relative terms. The best results were obtained with replanted soil + 4.5% hydrogen peroxide (H3). Consequently, hydrogen peroxide applied to the soil can effectively prevent and control ARD., Competing Interests: The authors declare no competing financial interest., (© 2023 The Authors. Published by American Chemical Society.)

Organic acids secreted by plants, such as p-hydroxybenzoic acid, ferulic acid, cinnamic acid, and benzoic acid, can inhibit seed germination and root growth. The effects of root and soil leaching liquor from orchards on the growth of M. hupehensis Rehd. seedlings under sand culture are studied; the seedlings are sampled at 15, 30, 45, and 60 d. Changes in the amount of root exudates are determined using HPLC. Low concentrations of root leaching liquor (A1) and soil leaching liquor (B1) significantly promoted plant growth and chlorophyll synthesis; high concentrations of root leaching liquor (A6) and soil leaching liquor (B4-6) inhibited growth. Low concentrations of soil leaching liquor had no significant effect on the POD, SOD, and CAT activities. A5-6 and B5-6 significantly decreased Fv / Fm and qP values, respectively, and increased NPQ values. All root and soil leaching liquor treatments inhibited the secretion of gallic acid, hydroxybenzoic acid, benzoic acid, and phloridzin, and promoted the secretion of caffeic acid. The root leaching liquor treatments inhibited the secretion of catechin and promoted the secretion of phloretin. The soil leaching liquor treatments promoted the secretion of cinnamic acid. The secretion of other phenolic acids is likely associated with the different concentrations of leaching liquor.

Exogenous application of nitrogen (N) and phosphate (P) has been demonstrated to alleviate apple replant disease (ARD). Yet, the effect of controlled-release diammonium phosphate (C-DAP), which continuously supply N and P for ARD control, is still poorly understood. Applying C-DAP markedly alleviated the typical symptoms of ARD. C-DAP maintained soil N and P at relatively high and stable levels during the entire growth period of the replanted seedlings, thus, limiting the copy number of the four key pathogenic Fusarium species that cause ARD. Particularly, continuously supplying N and P by C-DAP established a higher fungal diversity than that of conventional diammonium phosphate and induced the fungal community to be more similar to fumigated soil. The positive effect of C-DAP originated from the synergistic effects of regulating microorganisms and enhancing the resistance of the plant caused by a continuous nutrient supply. These findings provide a new perspective in the management of soil-borne diseases.

Identifying the genetic variation characteristics of phenotypic traits is important for fruit tree breeding. During the long-term evolution of fruit trees, gene recombination and natural mutation have resulted in a high degree of heterozygosity. Apple ( Malus × domestica Borkh.) shows strong ecological adaptability and is widely cultivated, and is among the most economically important fruit crops worldwide. However, the high level of heterozygosity and large genome of apple, in combination with its perennial life history and long juvenile phase, complicate investigation of the genetic basis of fruit quality traits. With continuing augmentation in the apple genomic resources available, in recent years important progress has been achieved in research on the genetic variation of fruit quality traits. This review focuses on summarizing recent genetic studies on apple fruit quality traits, including appearance, flavor, nutritional, ripening, and storage qualities. In addition, we discuss the mapping of quantitative trait loci, screening of molecular markers, and mining of major genes associated with fruit quality traits. The overall aim of this review is to provide valuable insights into the mechanisms of genetic variation and molecular breeding of important fruit quality traits in apple., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Liu, Chen, Jiang, Wang, Fang, Zhang, Chen and Wang.)

Anthocyanin and proanthocyanidin play important roles in plant secondary metabolism. Although previous studies identified many transcription factors involved in anthocyanin and proanthocyanidin synthesis, the effects of MADS-box transcription factors are unclear in apple. Brassinosteroids (BRs) are steroid hormones that affect plant flavonoid biosynthesis, but the underlying regulatory mechanism is not yet well established. In this study, we identified a MADS-box transcription factor, MdJa2, which contained a highly conserved MADS-box domain and belonged to the STMADS11 subfamily. Additionally, MdJa2 was responsive to BR signal, and the overexpression of MdJa2 inhibited the synthesis of anthocyanin and proanthocyanidin. The silencing of MdJa2 in "Orin" calli promoted anthocyanin and proanthocyanidin accumulations. Moreover, MdJa2 interacted with MdBZR1. MdJa2 was revealed to independently regulate anthocyanin and proanthocyanidin synthesis pathways. The MdJa2-MdBZR1 complex enhanced the binding of MdJa2 to the promoters of downstream target genes. Our research provides new insights into how MADS-box transcription factors in the BR signaling pathway control the accumulations of anthocyanin and proanthocyanidin in red-fleshed apple., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Su, Wang, Liu, Yang, Zhang, Wang and Chen.)

In this study, an endophytic phlorizin-degrading Bacillus licheniformis XNRB-3 was isolated from the root tissue of healthy apple trees, and its control effect on apple replant disease (ARD) and how it alleviates the pathogen pressure via changes in soil microbiomes were studied. The addition of strain XNRB-3 in Fusarium infested soils significantly reduced the number of pathogens in the soil, thus resulting in a lower disease incidence, and the relative control effect on Fusarium oxysporum reached the highest of 66.11%. The fermentation broth can also protect the roots of the plants from Fusarium oxysporum , Fusarium moniliforme , Fusarium proliferatum , and Fusarium solani infection. These antagonistic effects were further validated using an in vitro assay in which the pathogen control was related to growth and spore germination inhibition via directly secreted antimicrobial substances and indirectly affecting the growth of pathogens. The secreted antimicrobial substances were identified using gas chromatography-mass spectrometry (GC-MS) technology. Among them, alpha-bisabolol and 2,4-di-tert-butylphenol had significant inhibitory effects on many planted pathogenic fungi. Butanedioic acid, monomethyl ester, and dibutyl phthalate promoted root development of Arabidopsis plants. Strain XNRB-3 has multifarious plant growth promoting traits and antagonistic potential. In pot and field experiments, the addition of strain XNRB-3 significantly promoted the growth of plants, and the activity of enzymes related to disease resistance [superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT)] was also significantly enhanced. It also reduced the abundance of four species of Fusarium and the content of phenolic acids in the rhizosphere soil, improved soil microbial community structure and nutritional conditions, and increased soil microbial diversity and activity, as well as the soil enzyme activity. The above results indicated that B. licheniformis XNRB-3 could be developed into a promising biocontrol and plant-growth-promoting agent., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Duan, Zhao, Jiang, Chen, Zhang, Chen, Yin and Mao.)

Methyl bromide has been banned worldwide because it causes damage to the ozone layer and the environment. To find a substitute for methyl bromide, the relationships among fumigation, plant growth, and the microbial community in replant soil require further study. We performed pot and field experiments to investigate the effects of dazomet fumigation on soil properties and plant performance. Changes in soil microbial community structure and diversity were assessed using high-throughput sequencing, and plant physiological performance and soil physicochemical properties were also measured. Dazomet fumigation enhanced photosynthesis and promoted plant growth in replant soil; it altered soil physical and chemical properties and reduced soil enzyme activities, although these parameters gradually recovered over time. After dazomet fumigation, the dominant soil phyla changed, microbial diversity decreased significantly, the relative abundance of biocontrol bacteria such as Mortierella increased, and the relative abundance of pathogenic bacteria such as Fusarium decreased. Over the course of the experiment, the soil microbial flora changed dynamically, and soil enzyme activities and other physical and chemical properties also recovered to a certain extent. This result suggested that the effect of dazomet on soil microorganisms was temporary. However, fumigation also led to an increase in some resistant pathogens, such as Trichosporon, that affect soil function and health. Therefore, it is necessary to consider potential negative impacts of dazomet on the soil environment and to perform active environmental risk management in China., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Anthocyanin biosynthesis and sugar metabolism are important processes during plant growth, but the molecular interactions underlying these pathways are still unclear. In this work, we analyzed the anthocyanin and soluble sugar contents, as well as the transcript levels of transcription factors that are known to be related to the biosynthesis of anthocyanin in 'Hongcui 1' apple flesh during fruit development. Overexpression of MdMYB6 in red-fleshed calli was found to reduce anthocyanin content and result in downregulated expression of the MdANS and MdGSTF12 proteins. Yeast one-hybrid and electrophoretic mobility shift analyses showed that MdMYB6 could directly bind to the promoters of MdANS and MdGSTF12 , indicating that MdMYB6 could inhibit anthocyanin biosynthesis by regulating MdANS and MdGSTF12 . Overexpression of MdTMT1 in the Arabidopsis tmt1 mutant restored the glucose and fructose contents to the wild-type levels, while overexpression of MdTMT1 in red-fleshed calli increased the contents of glucose and fructose but reduced the contents of UDP-glucose, UDP-galactose, and anthocyanin. Using a GUS reporter system, yeast one-hybrid, chromatin immunoprecipitation-PCR and electrophoretic mobility shift analyses, we found that MdMYB6 could bind to the promoter of MdTMT1 , resulting in increased promoter activity. Overexpression of MdMYB6 in calli overexpressing MdTMT1 increased the expression of MdTMT1 , which led to reduced contents of UDP-glucose and UDP-galactose and decreased anthocyanin content compared to those of the calli that overexpressed MdTMT1 . This finding suggested that MdMYB6 could also inhibit anthocyanin biosynthesis by regulating MdTMT1 to decrease the contents of UDP-glucose and UDP-galactose. Taken together, these results showed that MdMYB6 and MdTMT1 play key roles in both anthocyanin biosynthesis and sugar transport., Competing Interests: Conflict of interestThe authors declare that they have no conflict of interest., (© The Author(s) 2020.)

In plants, the vesicle fusion process plays a vital role in pathogen defence. However, the importance of the vesicle fusion process in apple ring rot has not been studied. Here, we isolated and characterised the apple syntaxin gene MdSYP121 . Silencing the MdSYP121 gene in transgenic apple calli increased tolerance to Botryosphaeria dothidea infection; this increased tolerance was correlated with salicylic acid (SA) synthesis-related and signalling-related gene transcription. In contrast, overexpressing MdSYP121 in apple calli resulted in the opposite phenotypes. In addition, the results of RNA sequencing (RNA-Seq) and quantitative real-time PCR (qRT-PCR) assays suggested that MdSYP121 plays an important role in responses to oxidation-reduction reactions. Silencing MdSYP121 in apple calli enhanced the expression levels of reactive oxygen species (ROS)-related genes and the activity of ROS-related enzymes. The enhanced defence response status in MdSYP121-RNAi lines suggests that syntaxins are involved in the defence response to B. dothidea . More importantly, we showed that MdSYP121 forms a soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complex with MdSNAP33, and the complex may participate in regulating resistance to B. dothidea . In conclusion, by regulating the interaction of SA pathway and oxidation-reduction process, MdSYP121 can influence the pathogen infection process in apple., Competing Interests: The authors declare that they have no conflict of interest.

The B-box proteins (BBXs) are a family of zinc finger proteins containing one/two B-box domain(s). Compared with intensive studies of animal BBXs, investigations of the plant BBX family are limited, though some specific plant BBXs have been demonstrated to act as transcription factors in the regulation of flowering and photomorphogenesis. In this study, using a global search of the apple (Malus domestica Borkh.) genome, a total of 64 members of BBX (MdBBX) were identified. All the MdBBXs were divided into five groups based on the phylogenetic relationship, numbers of B-boxes contained and whether there was with an additional CCT domain. According to the characteristics of organ-specific expression, MdBBXs were divided into three groups based on the microarray information. An analysis of cis-acting elements showed that elements related to the stress response were prevalent in the promoter sequences of most MdBBXs. Twelve MdBBX members from different groups were randomly selected and exposed to abiotic stresses. Their expressions were up-regulated to some extent in the roots and leaves. Six among 12 MdBBXs were sensitive to osmotic pressure, salt, cold stress and exogenous abscisic acid treatment, with their expressions enhanced more than 20-fold. Our results suggested that MdBBXs may take part in response to abiotic stress.