The potential range of west Asian apple species Malus orientalis Uglitzk. under climate change.

The wild relatives of cultivated apples would be an ideal source of diversity for breeding new varieties, which could potentially grow in diverse habitats shaped by climate change. However, there is still a lack of knowledge about the potential distribution of these species. The aim of the presented work was the understand the impacts of climate change on the potential distribution and habitat fragmentation of Caucasian crab apple (Malus orientalis Uglitzk.) and the designation of areas of high interest according to climatic conditions. We used the MaxEnt models and Morphological-Spatial Analysis (MSPA) to evaluate the potential distribution, suitability changes, habitat fragmentation, and connectivity throughout the species range in Turkey, Armenia, Georgia, Russia, and Iran. The results revealed that the potentially suitable range of M. orientalis encompasses 858,877 km², 635,279 km² and 456,795 km² under the present, RCP4.5 and RCP8.5 scenario, respectively. The range fragmentation analysis demonstrated a notable shift in the edge/core ratio, which increased from 50.95% in the current scenario to even 67.70% in the future. The northern part of the range (Armenia, northern Georgia, southern Russia), as well as the central and western parts of Hyrcania will be a core of the species range with suitable habitats and a high connectivity between M. orientalis populations and could work as major refugia for the studied species. However, in the Zagros and central Turkey, the potential range will shrink due to the lack of suitable climatic conditions, and the edge/core ratio will grow. In the southern part of the range, a decline of M. orientalis habitats is expected due to changing climatic conditions. The future outlook suggests that the Hyrcanian forest and the Caucasus region could serve as important refuges for M. orientalis. This study helps to understand spatial changes in species' range in response to climate change and can help develop conservation strategies. This is all the more important given the species' potential use in future breeding programs aimed at enriching the gene pool of cultivated apple varieties.

Molecular evolution of Phytocyanin gene and analysis of expression at different coloring periods in apple (Malus domestica).

BACKGROUND: PC (phytocyanin) is a class of copper-containing electron transfer proteins closely related to plant photosynthesis, abiotic stress responses growth and development in plants, and regulation of the expression of some flavonoids and phenylpropanoids, etc., however, compared with other plants, the PC gene family has not been systematically characterized in apple. RESULTS: A total of 59 MdPC gene members unevenly distributed across 12 chromosomes were identified at the genome-wide level. The proteins of the MdPC family were classified into four subfamilies based on differences in copper binding sites and glycosylation sites: Apple Early nodulin-like proteins (MdENODLs), Apple Uclacyanin-like proteins (MdUCLs), Apple Stellacyanin-like proteins (MdSCLs), and Apple Plantacyanin-like proteins (MdPLCLs). Some MdPC members with similar gene structures and conserved motifs belong to the same group or subfamily. The internal collinearity analysis revealed 14 collinearity gene pairs among members of the apple MdPC gene. Interspecific collinearity analysis showed that apple had 31 and 35 homologous gene pairs with strawberry and grape, respectively. Selection pressure analysis indicated that the MdPC gene was under purifying selection. Prediction of protein interactions showed that MdPC family members interacted strongly with the Nad3 protein. GO annotation results indicated that the MdPC gene also regulated the biosynthesis of phenylpropanoids. Chip data analysis showed that (MdSCL3, MdSCL7 and MdENODL27) were highly expressed in mature fruits and peels. Many cis-regulatory elements related to light response, phytohormones, abiotic stresses and flavonoid biosynthetic genes regulation were identified 2000 bp upstream of the promoter of the MdPC gene, and qRT-PCR results showed that gene members in Group IV (MdSCL1/3, MdENODL27) were up-regulated at all five stages of apple coloring, but the highest expression was observed at the DAF13 (day after fruit bag removal) stage. The gene members in Group II (MdUCL9, MdPLCL3) showed down-regulated or lower expression in the first four stages of apple coloring but up-regulated and highest expression in the DAF 21 stage. CONCLUSION: Herein, one objective of these findings is to provide valuable information for understanding the structure, molecular evolution, and expression pattern of the MdPC gene, another major objective in this study was designed to lay the groundwork for further research on the molecular mechanism of PC gene regulation of apple fruit coloration.

Inhibition of PbeXTH1 and PbeSEOB1 is required for the Valsa canker resistance contributed by Wall-associated kinase gene MbWAK1.

Wall-associated kinases (WAKs) have been determined to recognize pathogenic signals and initiate plant immune responses. However, the roles of the family members in host resistance against Valsa canker, a serious fungal disease of apples and pears, are largely unknown. Here, we identified MbWAK1 in Malus baccata, a resistant germplasm differentially expressed during infection by Valsa mali (Vm). Over-expression of MbWAK1 enhanced the Valsa canker resistance of apple and pear fruits and 'Duli-G03' (Pyrus betulifolia) suspension cells. A large number of phloem, cell wall, and lipid metabolic process-related genes were differentially expressed in overexpressed suspension cell lines in response to Valsa pyri (Vp) signals. Among these, the expression of xyloglucan endotransglucosylase/hydrolase (XTH) gene PbeXTH1 and sieve element occlusion B-like (SEOB) gene PbeSEOB1 were significantly inhibited. Transient expression of PbeXTH1 or PbeSEOB1 compromised the expressional induction of MbWAK1 and the resistance contributed by MbWAK1. In addition, PbeXTH1 and PbeSEOB1 suppressed the immune response induced by MbWAK1. Our results enriched the molecular mechanisms for MbWAK1 against Valsa canker and resistant breeding.

Assessment and Partial Characterization of Candidate Genes in Dihydrochalcone and Arbutin Biosynthesis in an Apple-Pear Hybrid by De Novo Transcriptome Assembly.

Apples (Malus × domestica Borkh.) and pears (Pyrus communis L.) are valuable crops closely related within the Rosaceae family with reported nutraceutical properties derived from secondary metabolites including phloridzin and arbutin, which are distinctive phenolic metabolites characterizing apples and pears, respectively. Here, we generated a de novo transcriptome assembly of an intergeneric hybrid between apple and pear, accumulating intermediate levels of phloridzin and arbutin. Combining RNA-seq, in silico functional annotation prediction, targeted gene expression analysis, and expression-metabolite correlations, we identified candidate genes for functional characterization, resulting in the identification of active arbutin synthases in the hybrid and parental genotypes. Despite exhibiting an active arbutin synthase in vitro, the natural lack of arbutin in apples is reasoned by the absence of the substrate and broad substrate specificity. Altogether, our study serves as the basis for future assessment of potential physiological roles of identified genes by genome editing of hybrids and pears.

Evolution of the SPX gene family and its role in the response mechanism to low phosphorus stress in self-rooted apple stock.

BACKGROUND: Phosphorus plays a key role in plant adaptation to adversity and plays a positive role in the yield and quality formation of apples. Genes of the SPX domain-containing family are widely involved in the regulation of phosphorus signalling networks. However, the mechanisms controlling phosphorus deficiency are not completely understood in self-rooted apple stock. RESULTS: In this study, 26 members of the apple SPX gene family were identified by genome-wide analysis, and further divided into four subfamilies (SPX, SPX-MFS, SPX-EXS, and SPX-RING) based on their structural features. The chromosome distribution and gene duplications of MdSPXs were also examined. The promoter regions of MdSPXs were enriched for multiple biotic/abiotic stresses, hormone responses and typical P1BS-related elements. Analysis of the expression levels of 26 MdSPXs showed that some members were remarkably induced when subjected to low phosphate (Pi) stress, and in particular MdSPX2, MdSPX3, and MdPHO1.5 exhibited an intense response to low Pi stress. MdSPX2 and MdSPX3 showed significantly divergent expression levels in low Pi sensitive and insensitive apple species. Protein interaction networks were predicted for 26 MdSPX proteins. The interaction of MdPHR1 with MdSPX2, MdSPX3, MdSPX4, and MdSPX6 was demonstrated by yeast two-hybrid assay, suggesting that these proteins might be involved in the Pi-signaling pathway by interacting with MdPHR1. CONCLUSION: This research improved the understanding of the apple SPX gene family and contribute to future biological studies of MdSPX genes in self-rooted apple stock.

Apple-arbuscular mycorrhizal symbiosis confers resistance to Fusarium solani by inducing defense response and elevating nitrogen absorption.

Fusarium solani exerts detrimental effects on plant growth, which is one of the reasons for the incidence of apple replant disease. Arbuscular mycorrhizal fungi (AMF) enhance plant resistance to Fusarium wilt; however, the mechanism remains poorly understood. Therefore, the present study investigated the symbiosis between apple and AMF and explored the physiology, especially nitrate metabolism, antioxidant defense, and photosynthetic performance, when infected by F. solani. The experiment was carried out with four treatments, namely -AMF - F. solani, -AMF + F. solani, -AMF + F. solani, and + AMF + F. solani. In this study, the -AMF + F. solani treatment increased the activity of enzymes associated with nitrogen metabolism, such as the nitrate and nitrite reductases, in the apple root system. The +AMF + F. solani treatment showed higher antioxidant enzyme activities than the -AMF + F. solani by F. solani infection. The apple seedlings of the +AMF + F. solani treatment decreased reactive oxygen accumulation and reduced the oxidative damages triggered by F. solani infection. The improvement in antioxidant capacity due to the +AMF + F. solani treatment was closely associated with the upregulation of genes related to the antioxidant system. The F. solani infection greatly damaged the photosynthetic process, while the +AMF + F. solani treatment significantly improved it compared to the -AMF + F. solani treatment. In conclusion, the study demonstrated that the apple-AMF symbiosis plays an active role in regulating the resistance against F. solani infection by enhancing defense response and nitrogen metabolism.

MdTPR16, an apple tetratricopeptide repeat (TPR)-like superfamily gene, positively regulates drought stress in apple.

The Tetratricopeptide repeat (TPR)-like superfamily with TPR conserved domains is widely involved in the growth and abiotic stress in many plants. In this report, the gene MdTPR16 belongs to the TPR family in apple (Malus domestica). Promoter analysis reveal that MdTPR16 incorporated various stress response elements, including the drought stress response elements. And different abiotic stress treatments, drought especially, significantly induce the response of MdTPR16. Overexpression of MdTPR16 result in better drought tolerance in apple and Arabidopsis by up-regulating the expression levels of drought stress-related genes, achieving a higher chlorophyll content level, more material accumulation, and overall better growth compared to WT in the drought. Under drought stress, the overexpressed MdTPR16 also mitigate the oxidative damage in cells by reducing the electrolyte leakage, malondialdehyde content, and the H2O2 and O2- accumulation in apples and Arabidopsis. In conclusion, MdTPR16 act as a beneficial regulator of drought stress response by regulating the expression of related genes and the cumulation of reactive oxygen species (ROS).

Self-(in)compatibility in Tunisian apple accessions [Malus domestica. Borkh]: S-genotypes identification and pollen tube growth analysis.

MAIN CONCLUSION: Self-incompatibility studies have revealed a potential use of Tunisian apple resources for crop improvement and modern breeding programs and a likely correlation between the pollen tube growth and flowering period. Apples [Malus domestica. Borkh] exhibit an S-RNase-based gametophytic self-incompatibility (GSI) system. Four primer combinations were used to S-genotype eighteen Tunisian local apple accessions and twelve introduced accessions that served as references. Within the Tunisian local accessions, S2, S3, S7, and S28 S-alleles were the most frequent and were assigned to 14 S-genotypes; among them, S7S28, S3S7, S2S5, and S2S3 were the most abundant. PCA plot showed that population structuring was affected by the S-alleles frequencies and revealed a modern origin of the Tunisian varieties rather than being ancient ones. Nonetheless, the results obtained with 17 SSR markers showed a separate grouping of local Tunisian accessions that calls into question the hypothesis discussed. Pollination experiments showed that the pollen started to germinate within 24 h of pollination but 48 h after pollination in the "El Fessi" accession. The first pollen tubes arrived in the styles within 36 h of pollination in two early flowering accessions known as "Arbi" and "Bokri", and after 72 h of pollination in late flowering "El Fessi" and 48 h after pollination in remaining accessions. The first pollen tube arrests were observed in accessions "Arbi" and "Bokri" within 84 h of pollination, within 108 h of pollination in "El Fessi" and within 108 h of pollination in remaining accessions. In the apple accession called "Boutabgaya," the pollen tubes reached the base of the style within 120 h of pollination without being aborted. Nevertheless, the self-compatible nature of "Boutabgaya" needs more studies to be confirmed. However, our results revealed the malfunction of the female component of the GSI in this accession. To conclude, this work paved the path for further studies to enhance the insight (i) into the relation between the flowering period and the pollen tube growth, (ii) self-compatible nature of "Boutabgaya", and (iii) the origin of the Tunisian apple.

Molecular Mechanism of Resistance to Alternaria alternata Apple Pathotype in Apple by Alternative Splicing of Transcription Factor MdMYB6-like.

As a fruit tree with great economic value, apple is widely cultivated in China. However, apple leaf spot disease causes significant damage to apple quality and economic value. In our study, we found that MdMYB6-like is a transcription factor without auto-activation activity and with three alternative spliced variants. Among them, MdMYB6-like-ß responded positively to the pathogen infection. Overexpression of MdMYB6-like-ß increased the lignin content of leaves and improved the pathogenic resistance of apple flesh callus. In addition, all three alternative spliced variants of MdMYB6-like could bind to the promoter of MdBGLU H. Therefore, we believe that MdMYB6-like plays an important role in the infection process of the pathogen and lays a solid foundation for breeding disease-resistant cultivars of apple in the future.

Identification of Hsp20 gene family in Malus domestica and functional characterization of Hsp20 class I gene MdHsp18.2b.

Heat shock protein 20 (Hsp20) is a small molecule heat shock protein that plays an important role in plant growth, development, and stress resistance. Little is known about the function of Hsp20 family genes in apple (Malus domestica). Here, we performed a genome-wide analysis of the apple Hsp20 gene family, and a total of 49 Hsp20s genes were identified from the apple genome. Phylogenetic analysis revealed that the 49 genes were divided into 11 subfamilies, and MdHsp18.2b, a member located in the CI branch, was selected as a representative member for functional characterization. Treatment with NaCl and Botryosphaeria dothidea (B. dothidea), the causal agent of apple ring rot disease, significantly induced MdHsp18.2b transcription level. Further analysis revealed that overexpressing MdHsp18.2b reduced the resistance to salt stress but enhanced the resistance to B. dothidea infection in apple calli. Moreover, MdHsp18.2b positively regulated anthocyanin accumulation in apple calli. Physiology assays revealed that MdHsp18.2b promoted H2O2 production, even in the absence of stress factors, which might contribute to its functions in response to NaCl and B. dothidea infection. Hsps usually function as homo- or heterooligomers, and we found that MdHsp18.2b could form a heterodimer with MdHsp17.9a and MdHsp17.5, two members from the same branch with MdHsp18.2b in the phylogenetic tree. Therefore, we identified 49 Hsp20s genes from the apple genome and found that MdHsp18.2b was involved in regulating plant resistance to salt stress and B. dothidea infection, as well as in regulating anthocyanin accumulation in apple calli.

Penicillium-Infected Apples Benefit Larval Development of Conogethes punctiferalis via Alterations of Their Gut Bacteria Community and Gene Expression.

Pathogenic microorganisms can impact the behavior and physiology of herbivores by direct or indirect means. This study demonstrated that yellow peach moth Conogethes punctiferalis larvae feeding on Penicillium-infected apples exhibited significantly longer body length and weight parameters compared to the control group. The sequencing of gut 16S rRNA showed a significant increase in the diversity and abundance of bacteria in the larvae feeding on Penicillium-infected apples. Additionally, transcriptomic sequencing of the larval gut indicated significant upregulation of genes related to digestion and cuticle formation after consuming Penicillium-infected apples. Furthermore, enzyme activity assays revealed notable changes in the trypsin and lipase activity. Consequently, these alterations in gut microbiota structure, diversity, and gene expression levels may underlie the observed growth and developmental variations in C. punctiferalis larvae mediated by pathogenic microorganisms. This study holds theoretical significance for a deeper understanding of the tripartite interaction among microorganisms, insects, and plants as well as for the development of novel pest control measures based on gut microbiota.

MdWRKY71 promotes the susceptibility of apple to Glomerella leaf spot by controlling salicylic acid degradation.

Glomerella leaf spot (GLS), a fungal disease caused by Colletotrichum fructicola, severely affects apple (Malus domestica) quality and yield. In this study, we found that the transcription factor MdWRKY71 was significantly induced by C. fructicola infection in the GLS-susceptible apple cultivar Royal Gala. The overexpression of MdWRKY71 in apple leaves resulted in increased susceptibility to C. fructicola, whereas RNA interference of MdWRKY71 in leaves showed the opposite phenotypes. These findings suggest that MdWRKY71 functions as a susceptibility factor for the apple-C. fructicola interaction. Furthermore, MdWRKY71 directly bound to the promoter of the salicylic acid (SA) degradation gene Downy Mildew Resistant 6 (DMR6)-Like Oxygenase 1 (DLO1) and promoted its expression, resulting in a reduced SA level. The sensitivity of 35S:MdWRKY71 leaves to C. fructicola can be effectively alleviated by knocking down MdDLO1 expression, confirming the critical role of MdWRKY71-mediated SA degradation via regulating MdDLO1 expression in GLS susceptibility. In summary, we identified a GLS susceptibility factor, MdWRKY71, that targets the apple SA degradation pathway to promote fungal infection.

Phenomics and transcriptomic profiling of fruit development in distinct apple varieties.

Apple is one of the most economically important and popular temperate fruit trees. The domestication of apple has resulted in substantial phenotypic differences, particularly between wild and cultivated varieties. However, the relationship between gene expression and phenotypic variations in apple remains poorly understood. Here, we present a comprehensive dataset featuring five distinct apple varieties, including two wild varieties and three representative cultivated varieties. The dataset comprises of both phenomics data, encompassing twelve fruit quality-related traits continuously measured over two years, and transcriptomic data obtained at different developmental stages with three biological replicates. We performed basic quality control process, gene expression normalization and differential gene expression analysis to demonstrate the utility and reliability of the dataset. Our findings indicate that gene expression strongly related with phenotypic variations in apple. This dataset serves as a valuable resource, encompassing phenomics and transcriptomic data in multiple formats, thereby facilitating further exploration of the relationships between gene expression and phenotypic traits in apple.

Epigenomic mechanism regulating the quality and ripeness of apple fruit with differing harvest maturity.

Harvest maturity significantly affects the quality of apple fruit in post-harvest storage process. Although the regulatory mechanisms underlying fruit ripening have been studied, the associated epigenetic modifications remain unclear. Thus, we compared the DNA methylation changes and the transcriptional responses of mature fruit (MF) and immature fruit (NF). There were significant correlations between DNA methylation and gene expression. Moreover, the sugar contents (sucrose, glucose, and fructose) were higher in MF than in NF, whereas the opposite pattern was detected for the starch content. The expression-level differences were due to DNA methylations and ultimately resulted in diverse fruit textures and ripeness. Furthermore, the higher ethylene, auxin, and abscisic acid levels in MF than in NF, which influenced the fruit texture and ripening, were associated with multiple differentially expressed genes in hormone synthesis, signaling, and response pathways (ACS, ACO, ZEP, NCED, and ABA2) that were regulated by DNA methylations. Multiple transcription factor genes involved in regulating fruit ripening and quality via changes in DNA methylation were identified, including MIKCC-type MADS-box genes and fruit ripening-related genes (NAP, SPL, WRKY, and NAC genes). These findings reflect the diversity in the epigenetic regulation of gene expression and may be relevant for elucidating the epigenetic regulatory mechanism underlying the ripening and quality of apple fruit with differing harvest maturity.

Assessment of genetic homogeneity of in-vitro propagated apple root stock MM 104 using ISSR and SCoT primers.

Apple is an important fruit crop that is always in demand due to its commercial and nutraceutical value. Also, the requirement for quality planting material for this fruit crop for new plantations is increasing continuously. In-vitro propagation is an alternative approach, which may help to produce genetically identical high grade planting material. In this study, for the first time, an efficient and reproducible propagation protocol has been established for apple root stock MM 104 via axillary bud. Culturing axillary buds on Murashige and Skoog apple rootstock (MM 104) resulted in better in-vitro propagation. (MS) basal medium supplemented with 3.0% (w/v) sucrose and 0.8% (w/v) agar. The axillary buds were established in MS basal medium with BA (5.0 µM), NAA (1.0 µM) and further used to establish invitro propagation protocol. Plant Growth Regulators (PGRs), BA (1.0 µM) in combination with NAA (1.0 µM) was found most efficient for shoot multiplication (100%) and produced 9.8 shoots/explants with an average shoot length of (2.4 ± cm). All the shoots produced roots in 0.1 µM IBA with a 5-day dark period. Acclimatization of in-vitro raised plantlets was obtained with vermiculite: perlite: sand: soil (2:2:1:1) resulting in 76% survival under field conditions. The study showed that the use of axillary bud is efficient for multiple-shoot production of apple rootstock (MM 104). This is the first comprehensive report on in-vitro growth of apple root stock MM 104 with an assessment of genetic stability using DNA fingerprinting profiles based on Inter Simple Sequence Repeats (ISSR) and Start Codon Targeted (SCoT). The genetic stability of in-vitro-produced plants, as determined by SCoT and ISSR primers, demonstrated genetic closeness to the mother plant.

The genetic basis of apple shape and size unraveled by digital phenotyping.

Great diversity of shape, size, and skin color is observed among the fruits of different apple genotypes. These traits are critical for consumers and therefore interesting targets for breeding new apple varieties. However, they are difficult to phenotype and their genetic basis, especially for fruit shape and ground color, is largely unknown. We used the FruitPhenoBox to digitally phenotype 525 genotypes of the apple reference population (apple REFPOP) genotyped for 303,148 single nucleotide polymorphism (SNP) markers. From the apple images, 573 highly heritable features describing fruit shape and size as well as 17 highly heritable features for fruit skin color were extracted to explore genotype-phenotype relationships. Out of these features, seven principal components (PCs) and 16 features with the Pearson's correlation r < 0.75 (selected features) were chosen to carry out genome-wide association studies (GWAS) for fruit shape and size. Four PCs and eight selected features were used in GWAS for fruit skin color. In total, 69 SNPs scattered over all 17 apple chromosomes were significantly associated with round, conical, cylindrical, or symmetric fruit shapes and fruit size. Novel associations with major effect on round or conical fruit shapes and fruit size were identified on chromosomes 1 and 2. Additionally, 16 SNPs associated with PCs and selected features related to red overcolor as well as green and yellow ground color were found on eight chromosomes. The identified associations can be used to advance marker-assisted selection in apple fruit breeding to systematically select for desired fruit appearance.

miR164-MhNAC1 regulates apple root nitrogen uptake under low nitrogen stress.

Nitrogen is an essential nutrient for plant growth and serves as a signaling molecule to regulate gene expression inducing physiological, growth and developmental responses. An excess or deficiency of nitrogen may have adverse effects on plants. Studying nitrogen uptake will help us understand the molecular mechanisms of utilization for targeted molecular breeding. Here, we identified and functionally validated an NAC (NAM-ATAF1/2-CUC2) transcription factor based on the transcriptomes of two apple rootstocks with different nitrogen uptake efficiency. NAC1, a target gene of miR164, directly regulates the expression of the high-affinity nitrate transporter (MhNRT2.4) and citric acid transporter (MhMATE), affecting root nitrogen uptake. To examine the role of MhNAC1 in nitrogen uptake, we produced transgenic lines that overexpressed or silenced MhNAC1. Silencing MhNAC1 promoted nitrogen uptake and citric acid secretion in roots, and enhanced plant tolerance to low nitrogen conditions, while overexpression of MhNAC1 or silencing miR164 had the opposite effect. This study not only revealed the role of the miR164-MhNAC1 module in nitrogen uptake in apple rootstocks but also confirmed that citric acid secretion in roots affected nitrogen uptake, which provides a research basis for efficient nitrogen utilization and molecular breeding in apple.

DIA-Based Quantitative Proteomics in the Flower Buds of Two Malus sieversii (Ledeb.) M. Roem Subtypes at Different Overwintering Stages.

Malus sieversii is considered the ancestor of the modern cultivated apple, with a high value for apple tolerance breeding. Despite studies on the temperature adaptability of M. sieversii carried out at a physiological response and the genome level, information on the proteome changes of M. sieversii during dormancy is limited, especially about the M. sieversii subtypes. In this study, a DIA-based approach was employed to screen and identify differential proteins involved in three overwintering periods of flower buds in two M. sieversii subtypes (Malus sieversii f. luteolus, GL; Malus sieversii f. aromaticus, HC) with different overwintering adaptabilities. The proteomic analysis revealed that the number of the down-regulated differential expression proteins (DEPs) was obviously higher than that of the up-regulated DEPs in the HC vs. GL groups, especially at the dormancy stage and dormancy-release stage. Through functional classification of those DEPs, the majority of the DEPs in the HC vs. GL groups were associated with protein processing in the endoplasmic reticulum, oxidative phosphorylation, starch and sucrose metabolism and ribosomes. Through WGCNA analysis, tricarboxylic acid cycle and pyruvate metabolism were highly correlated with the overwintering stages; oxidative phosphorylation and starch and sucrose metabolism were highly correlated with the Malus sieversii subtypes. This result suggests that the down-regulation of DEPs, which are predominantly enriched in these pathways, could potentially contribute to the lower cold tolerance observed in HC during overwintering stage.

Apple crown and collar canker and necrosis caused by Cytospora balanejica sp. nov. in Iran.

Apple is the most important fruit tree in West Azarbaijan province of Iran. In a survey of apple orchards, a disease with crown and collar canker and necrosis symptoms was observed in three young apple orchards in Urmia, affecting 15% and 1% of 'Red Delicious' and 'Golden Delicious' cultivars, respectively. A fungus with typical characteristics of the asexual morph of Cytospora was regularly isolated from the diseased tissues. Morphological characteristics and phylogenetic analyses inferred from the combined dataset of the ITS-rDNA, parts of LSU, tef1-α, rpb2, and act1 genes revealed that the isolates represent a new species of Cytospora, described herein as Cytospora balanejica sp. nov.. The pathogenicity of all isolates was confirmed on apple cv. 'Red Delicious' based on Koch's postulates. Also, the reaction of 12 other apple cultivars was assessed against five selected isolates with the highest virulence. The results showed that except for cv. 'Braeburn', which did not produce any symptoms of the disease, the other 11 cultivars showed characteristic disease symptoms including sunken and discolored bark and wood. The mean length of the discolored area was different among the 11 so-called susceptible cultivars, hence cvs. 'M4' and 'Golden Delicious' showed the highest and the lowest lesion length, respectively. Moreover, the aggressiveness of the five tested isolates was different, and the isolates BA 2-4 and BA 3-1 had the highest and lowest aggressiveness, respectively. Based on our observations on the potential ability of the fungus to cause disease on young and actively growing apple trees, it will be a serious threat to apple cultivation and industry.

Integrative physiological, metabolomic, and transcriptomic analysis reveals the drought responses of two apple rootstock cultivars.

BACKGROUND: Drought is considered the main environmental factor restricting apple production and thus the development of the apple industry. Rootstocks play an important role in enhancing the drought tolerance of apple plants. Studies of the physiology have demonstrated that 'ZC9-3' is a strong drought-resistant rootstock, whereas 'Jizhen-2' is a weak drought-resistant rootstock. However, the metabolites in these two apple rootstock varieties that respond to drought stress have not yet been characterized, and the molecular mechanisms underlying their responses to drought stress remain unclear. RESULTS: In this study, the physiological and molecular mechanisms underlying differences in the drought resistance of 'Jizhen-2' (drought-sensitive) and 'ZC9-3' (drought-resistant) apple rootstocks were explored. Under drought stress, the relative water content of the leaves was maintained at higher levels in 'ZC9-3' than in 'Jizhen-2', and the photosynthetic, antioxidant, and osmoregulatory capacities of 'ZC9-3' were stronger than those of 'Jizhen-2'. Metabolome analysis revealed a total of 95 and 156 differentially accumulated metabolites in 'Jizhen-2' and 'ZC9-3' under drought stress, respectively. The up-regulated metabolites in the two cultivars were mainly amino acids and derivatives. Transcriptome analysis revealed that there were more differentially expressed genes and transcription factors in 'ZC9-3' than in 'Jizhen-2' throughout the drought treatment. Metabolomic and transcriptomic analysis revealed that amino acid biosynthesis pathways play key roles in mediating drought resistance in apple rootstocks. A total of 13 metabolites, including L-α-aminoadipate, L-homoserine, L-threonine, L-isoleucine, L-valine, L-leucine, (2S)-2-isopropylmalate, anthranilate, L-tryptophan, L-phenylalanine, L-tyrosine, L-glutamate, and L-proline, play an important role in the difference in drought resistance between 'ZC9-3' and 'Jizhen-2'. In addition, 13 genes encoding O-acetylserine-(thiol)-lyase, S-adenosylmethionine synthetase, ketol-acid isomeroreductase, dihydroxyacid dehydratase, isopropylmalate isomerase, branched-chain aminotransferase, pyruvate kinase, 3-dehydroquinate dehydratase/shikimate 5-dehydrogenase, N-acetylglutamate-5-P-reductase, and pyrroline-5-carboxylate synthetase positively regulate the response of 'ZC9-3' to drought stress. CONCLUSIONS: This study enhances our understanding of the response of apple rootstocks to drought stress at the physiological, metabolic, and transcriptional levels and provides key insights that will aid the cultivation of drought-resistant apple rootstock cultivars. Especially, it identifies key metabolites and genes underlying the drought resistance of apple rootstocks.