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Abstract Plants possess the ability to induce programmed cell death (PCD) in response to abiotic and biotic stresses; nevertheless, the evidence on PCD initiation during pear scald development and the involvement of the scald trigger 6-methyl-5-hepten-2-one (MHO) in this process is rudimentary. Pyrus bretschneideri Rehd. cv. ‘Dangshansuli’ pear was used to validate such hypothesis. The results showed that superficial scald occurred after 120-d chilling exposure, which accompanied by typical PCD-associated morphological alterations, such as plasmolysis, cell shrinkage, cytosolic and nuclear condensation, vacuolar collapse, tonoplast disruption, subcellular organelle swelling, and DNA fragmentation. These symptoms were aggravated after MHO fumigation but alleviated by diphenylamine (DPA) dipping. Through transcriptome assay, 24 out of 146 PCD-related genes, which were transcribed during cold storage, were identified as the key candidate members responsible for these cellular biological alternations upon scald development. Among these, PbrCNGC1, PbrGnai1, PbrACD6, and PbrSOBIR1 were implicated in the MHO signaling pathway. Additionally, PbrWRKY2, 34 and 39 could bind to the W-box element in the promoter of PbrGnai1 or PbrSOBIR1 and activate their transcription, as confirmed by dual-luciferase, yeast one-hybrid, and transient overexpression assays. Hence, our study confirms the PCD initiation during scald development and explores the critical role of MHO in this process.

Abstract Background PSEUDO RESPONSE REGULATOR (PRR) genes are essential components of circadian clock, playing vital roles in multiple processes including plant growth, flowering and stress response. Nonetheless, little is known about the evolution and function of PRR family in Rosaceae species. Results In this study, a total of 43 PRR genes in seven Rosaceae species were identified through comprehensive analysis. The evolutionary relationships were analyzed with phylogenetic tree, duplication events and synteny. PRR genes were classified into three groups (PRR1, PRR5/9, PRR3/7). The expansion of PRR family was mainly derived from dispersed and whole-genome duplication events. Purifying selection was the major force for PRR family evolution. Synteny analysis indicated the existence of multiple orthologous PRR gene pairs between pear and other Rosaceae species. Moreover, the conserved motifs of eight PbPRR proteins supported the phylogenetic relationship. PRR genes showed diverse expression pattern in various tissues of pear (Pyrus bretschneideri). Transcript analysis under 12-h light/ dark cycle and constant light conditions revealed that PRR genes exhibited distinct rhythmic oscillations in pear. PbPRR59a and PbPRR59b highly homologous to AtPRR5 and AtPRR9 were cloned for further functional verification. PbPRR59a and PbPRR59b proteins were localized in the nucleus. The ectopic overexpression of PbPRR59a and PbPRR59b significantly delayed flowering in Arabidopsis transgenic plants by repress the expression of AtGI, AtCO and AtFT under long-day conditions. Conclusions These results provide information for exploring the evolution of PRR genes in plants, and contribute to the subsequent functional studies of PRR genes in pear and other Rosaceae species.

Abstract Pathogenesis-related (PR) genes are key regulators of plant adaptation responses to biotic and abiotic stresses. Family 10 PRs (PR10s, also known as major latex proteins) are usually induced by pathogens and environmental stresses. However, the evolutionary trajectory and functional divergence of the PR10 gene family in Chinese white pear (Pyrus bretschneideri ‘Dangshan Suli’) remain uncharacterized. The presence of 61 PR10s was detected across six Rosaceae species. The PR10 gene family was classified into two distinct groups by employing phylogenetic analysis and the taxonomic criteria of model plants. Interspecies synchrony revealed an ancient origin of the PR10 family in the six Rosaceae species, with 18 synchronic gene pairs. The expansion and evolution of the PR10 family were driven by various types of gene duplication events, with whole-genome duplication (WGD) being the primary mechanism. A candidate Colletotrichum fructicola (C. fructicola) resistance gene Pyr us bretschneideri major latex-like proteins (PbrMLP) belonging to the PR10 family was screened through transcriptomics and qRT-PCR. In addition, PbrMLP-silenced pear seedlings were more sensitive to C. fructicola than the controls. These results showed that PbrMLP is a candidate gene vital for anthracnose resistance in pears. These findings offer novel insights into the molecular mechanisms underlying the resistance to pear anthracnose infection in Rosaceae species and identify potential target genes for developing disease-resistant cultivars through genetic engineering.

Abstract Background LACS (long-chain acyl-CoA synthetase) genes are widespread in organisms and have multiple functions in plants, especially in lipid metabolism. However, the origin and evolutionary dynamics of the LACS gene family remain largely unknown. Results Here, we identified 1785 LACS genes in the genomes of 166 diverse plant species and identified the clades (I, II, III, IV, V, VI) of six clades for the LACS gene family of green plants through phylogenetic analysis. Based on the evolutionary history of plant lineages, we found differences in the origins of different clades, with Clade IV originating from chlorophytes and representing the origin of LACS genes in green plants. The structural characteristics of different clades indicate that clade IV is relatively independent, while the relationships between clades (I, II, III) and clades (V, VI) are closer. Dispersed duplication (DSD) and transposed duplication (TRD) are the main forces driving the evolution of plant LACS genes. Network clustering analysis further grouped all LACS genes into six main clusters, with genes within each cluster showing significant co-linearity. Ka/Ks results suggest that LACS family genes underwent purifying selection during evolution. We analyzed the phylogenetic relationships and characteristics of six clades of the LACS gene family to explain the origin, evolutionary history, and phylogenetic relationships of different clades and proposed a hypothetical evolutionary model for the LACS family of genes in plants. Conclusions Our research provides genome-wide insights into the evolutionary history of the LACS gene family in green plants. These insights lay an important foundation for comprehensive functional characterization in future research.

Abstract Background The homodomain-leucine zipper (HD-Zip) is a conserved transcription factor family unique to plants that regulate multiple developmental processes including lignificaion. Stone cell content is a key determinant negatively affecting pear fruit quality, which causes a grainy texture of fruit flesh, because of the lignified cell walls. Results In this study, a comprehensive bioinformatics analysis of HD-Zip genes in Chinese white pear (Pyrus bretschneideri) (PbHBs) was performed. Genome-wide identification of the PbHB gene family revealed 67 genes encoding PbHB proteins, which could be divided into four subgroups (I, II, III, and IV). For some members, similar intron/exon structural patterns support close evolutionary relationships within the same subgroup. The functions of each subgroup of the PbHB family were predicted through comparative analysis with the HB genes in Arabidopsis and other plants. Cis-element analysis indicated that PbHB genes might be involved in plant hormone signalling and external environmental responses, such as light, stress, and temperature. Furthermore, RNA-sequencing data and quantitative real-time PCR (RT-qPCR) verification revealed the regulatory roles of PbHB genes in pear stone cell formation. Further, co-expression network analysis revealed that the eight PbHB genes could be classified into different clusters of co-expression with lignin-related genes. Besides, the biological function of PbHB24 in promoting stone cell formation has been demonstrated by overexpression in fruitlets. Conclusions This study provided the comprehensive analysis of PbHBs and highlighted the importance of PbHB24 during stone cell development in pear fruits.

The Arabidopsis Tóxicos en Levadura (ATL) protein is a subfamily of the E3 ubiquitin ligases, which exists widely in plants and is extensively involved in plant growth and development. Although the ATL family has been identified in other species, such as Arabidopsis, Oryza sativa, and grapevine, few reports on pear ATL gene families have been reported. In this study, 92 PbrATL genes were identified and analyzed from the Pyrus breschneideri genome. Motif analysis and phylogenetic tree generation divided them into nine subgroups, and chromosome localization analysis showed that the 92 PbrATL genes were distributed in 16 of 17 pear chromosomes. Transcriptome data and quantitative real-time polymerase chain reaction (qRT-PCR) experiments demonstrated that PbrATL18, PbrATL41, and PbrATL88 were involved in both pear drought resistance and Colletotrichum fructicola infection. In addition, Arabidopsis thaliana overexpressing PbrATL18 showed greater resistance to drought stress than the wild type (WT), and PbrATL18-silenced pear seedlings showed greater sensitivity to drought and C. fructicola infection than the controls. PbrATL18 regulated plant resistance by regulating chitinase (CHI), phenylalanine ammonia-lyase (PAL), polyphenol oxidase (PPO), catalase (CAT), peroxidase (POD), and superoxide dismutase (SOD) activities. This study provided a reference for further exploring the functions of the PbrATL gene in drought resistance and C. fructicola infection.

Abstract In pears, the presence of stone cells adversely affects fruit quality. Pectin methylesterase (PME) plays various roles in plant biology, including lignin biosynthesis. However, only a limited fraction has been functionally characterized, and the distribution and function of PME in many Rosaceae trees remain unexplored. In this study, we identified 396 putative PME family candidate genes, with 81 in Pyrus bretschneideri, 92 in Malus domestica, 62 in Fragaria vesca, 65 in Prunus mume, 15 in Pyrus communis, and 81 in Pyrus pyrifolia. Leveraging insights from model plants, we categorized PME family genes into four groups. Additionally, the evolution of the PME gene family was shaped by various gene duplication events, primarily dispersed duplication, influenced by purifying selection. A specific gene, Pbr031522.1, designated PbPME35, emerged as a candidate associated with lignin biosynthesis in pear fruits, supported by RNA-seq data. The role of PbPME35 in repressing lignification was validated through its overexpression in pear callus and Arabidopsis. Overall, our findings highlight the ability of PbPME35 to reduce lignin content in pear fruit by downregulating the expression levels of lignin biosynthesis genes. These findings provide new insights into the characteristics of PME genes and their role in regulating lignification in pear fruits.

Abstract Background Extensive research has been conducted on fruit development in crops, but the metabolic regulatory networks underlying perennial fruit trees remain poorly understood. To address this knowledge gap, we conduct a comprehensive analysis of the metabolome, proteome, transcriptome, DNA methylome, and small RNAome profiles of pear fruit flesh at 11 developing stages, spanning from fruitlet to ripening. Here, we systematically investigate the metabolic landscape and regulatory network involved. Results We generate an association database consisting of 439 metabolites and 14,399 genes to elucidate the gene regulatory network of pear flesh metabolism. Interestingly, we detect increased DNA methylation in the promoters of most genes within the database during pear flesh development. Application of a DNA methylation inhibitor to the developing fruit represses chlorophyll degradation in the pericarp and promotes xanthophyll, β-carotene, and abscisic acid (ABA) accumulation in the flesh. We find the gradual increase in ABA production during pear flesh development is correlated with the expression of several carotenoid pathway genes and multiple transcription factors. Of these transcription factors, the zinc finger protein PbZFP1 is identified as a positive mediator of ABA biosynthesis in pear flesh. Most ABA pathway genes and transcription factors are modified by DNA methylation in the promoters, although some are induced by the DNA methylation inhibitor. These results suggest that DNA methylation inhibits ABA accumulation, which may delay fruit ripening. Conclusion Our findings provide insights into epigenetic regulation of metabolic regulatory networks during pear flesh development, particularly with regard to DNA methylation.

Genomic selection (GS) has the potential to improve selection efficiency and shorten the breeding cycle in fruit tree breeding. In this study, we evaluated the effect of prediction methods, marker density and the training population (TP) size on pear GS for improving its performance and reducing cost. We evaluated GS under two scenarios: (1) five-fold cross-validation in an interspecific pear family; (2) independent validation. Based on the cross-validation scheme, the prediction accuracy (PA) of eight fruit traits varied between 0.33 (fruit core vertical diameter) and 0.65 (stone cell content). Except for single fruit weight, a slightly better prediction accuracy (PA) was observed for the five parametrical methods compared with the two non-parametrical methods. In our TP of 310 individuals, 2 000 single nucleotide polymorphism (SNP) markers were sufficient to make reasonably accurate predictions. PAs for different traits increased by 18.21%–46.98% when the TP size increased from 50 to 100, but the increment was smaller (−4.13%–33.91%) when the TP size increased from 200 to 250. For independent validation, the PAs ranged from 0.11 to 0.45 using rrBLUP method. In summary, our results showed that the TP size and SNP numbers had a greater impact on the PA than prediction methods. Furthermore, relatedness among the training and validation sets, and the complexity of traits should be considered when designing a TP to predict the test panel.

The pear is an economic fruit that is widely planted around the world and is loved by people for its rich nutritional value. Autophagy is a self-protection mechanism in eukaryotes, and its occurrence often accompanied by the degradation of damaged substances in cells and the recycling of nutrients. Autophagy is one of the mechanisms through which plants respond to environmental stress and plays an important role in plant development and stress resistance. Functional studies of autophagy-related genes (ATGs) have been performed on a variety of plant species, but little information is available on the ATG family in pear (Pyrus bretschneideri Rehd). Therefore, we analyzed the evolutionary dynamics and performed a genome-wide characterization of the PbrATG gene family. A total of 28 PbrATG members were identified. Phylogenetic analysis showed that PbrATGs were more closely related to ATGs of European pear and apple. Evolutionary analysis revealed that whole-genome duplication (WGD) and dispersed duplication events were the main driving forces of PbrATG family expansion. Expression analysis of different pear tissues showed that all the genes were expressed in different pear tissues, and different PbrATGs are expressed at different times and in different locations. Moreover, all PbrATGs also responded to different abiotic stresses, especially salt and drought stress, which elicited the highest expression levels. Pear seedlings were subsequently infected with Botryosphaeria dothidea (B. dothidea). The results showed that different PbrATGs had different expression patterns at different infection stages. According to the gene expression data, PbrATG1a was selected as a key autophagy gene for further analysis. Silencing of PbrATG1a reduced the resistance of pear to B. dothidea, which resulted in increased lesions, reactive oxygen species (ROS) contents, antioxidant enzyme activity, and gene expression levels in the silenced pear seedlings after B. dothidea inoculation. In this study, a comprehensive bioinformatic analysis of ATGs was conducted, and the functions of PbrATGs in pear development and in response to stress were elucidated, which laid a foundation for further study of the molecular mechanism of autophagy and a new strategy for pear resistance breeding.

Abstract Background ATP-binding cassette (ABC) transporter proteins constitute a plant gene superfamily crucial for growth, development, and responses to environmental stresses. Despite their identification in various plants like maize, rice, and Arabidopsis, little is known about the information on ABC transporters in pear. To investigate the functions of ABC transporters in pear development and abiotic stress response, we conducted an extensive analysis of ABC gene family in the pear genome. Results In this study, 177 ABC transporter genes were successfully identified in the pear genome, classified into seven subfamilies: 8 ABCAs, 40 ABCBs, 24 ABCCs, 8 ABCDs, 9 ABCEs, 8 ABCFs, and 80 ABCGs. Ten motifs were common among all ABC transporter proteins, while distinct motif structures were observed for each subfamily. Distribution analysis revealed 85 PbrABC transporter genes across 17 chromosomes, driven primarily by WGD and dispersed duplication. Cis-regulatory element analysis of PbrABC promoters indicated associations with phytohormones and stress responses. Tissue-specific expression profiles demonstrated varied expression levels across tissues, suggesting diverse functions in development. Furthermore, several PbrABC genes responded to abiotic stresses, with 82 genes sensitive to salt stress, including 40 upregulated and 23 downregulated genes. Additionally, 91 genes were responsive to drought stress, with 22 upregulated and 36 downregulated genes. These findings highlight the pivotal role of PbrABC genes in abiotic stress responses. Conclusion This study provides evolutionary insights into PbrABC transporter genes, establishing a foundation for future research on their functions in pear. The identified motifs, distribution patterns, and stress-responsive expressions contribute to understanding the regulatory mechanisms of ABC transporters in pear. The observed tissue-specific expression profiles suggest diverse roles in developmental processes. Notably, the significant responses to salt and drought stress emphasize the importance of PbrABC genes in mediating adaptive responses. Overall, our study advances the understanding of PbrABC transporter genes in pear, opening avenues for further investigations in plant molecular biology and stress physiology.

Abstract Putrescine plays a role in superficial scald development during the cold storage of pear fruit. However, the molecular mechanism behind this phenomenon has not been un-fully clarified until recently. In this study, a conjoint analysis of metabolites and gene expression profiles in the putrescine-metabolic pathway of P. bretschneideri Rehd. fruit followed by experimental validation revealed that PbrADC1, forming a homodimer in the chloroplast, was involved in putrescine biosynthesis and thus fruit chilling resistance. Additionally, the substrate-binding residue Cys546 in PbrADC1, whose activity was modified by H2O2, played a crucial role in arginine decarboxylation into agmatine. Through a combined analysis of the distribution of cis-acting elements in the PbrADC1 promoter as well as the expression profiles of related transcription factors (TFs), several TFs were identified as upstream regulators of PbrADC1 gene. Further investigation revealed that the nuclear PbrWRKY62 could directly bind to the W-box elements in the PbrADC1 promoter, activate its expression, enhance putrescine accumulation, and thus increase fruit chilling tolerance. In conclusion, our results suggest that the PbrWRKY62-PbrADC1 module is involved in the development of superficial scald in P. bretschneideri Rehd. fruit via regulating putrescine biosynthesis. Consequently, these findings could serve as valuable genetic resources for breeding scald-resistant pear fruit. Graphical Abstract

Abstract Pear anthracnose, caused by the fungus Colletotrichum fructicola, is a devastating disease for the pear industry. The apoplast, an extracellular compartment outside the plasma membrane, plays a crucial role in water and nutrient transport, as well as plant-microbe interactions. This study aimed to uncover the molecular mechanism of pear leaf apoplastic protein-mediated resistance to C. fructicola. Apoplast fluid was isolated using the vacuum infiltration method, and defence-related apoplastic proteins were identified through protein mass spectrometry and transcriptome sequencing. We found 213 apoplastic proteins in the leaf apoplast fluid during early C. fructicola infection, with the majority (74.64%) being enzymes, including glycosidases, proteases, and oxidoreductases. Gene Ontology analysis revealed their involvement in defence response, enzyme inhibition, carbohydrate metabolism, and phenylpropanoid biosynthesis. Transcriptome analysis showed the infection induced expression of certain apoplast proteins, potentially contributing to pear leaf resistance. Notably, the expression of PbrGlu1, an endo-β-1,3-glucanase from the glycoside hydrolase 17 family, was significantly higher in infected leaves. Silencing of the PbrGlu1 gene increased pear leaf susceptibility to C. fructicola, leading to more severe symptoms and higher reactive oxygen species content. Overall, our study provides insights into the apoplast space interaction between pear leaves and C. fructicola, identifies a key gene in infected pears, and offers a foundation and new strategy for understanding the molecular mechanisms underlying pear anthracnose and breeding disease-resistant pears.

Abstract Due to self-incompatibility (SI) prevents self-fertilization, natural or artificial cross-pollination has been conducted in many orchards to stabilize fruit yield. However, it is still puzzled which routes of self S-RNase arresting pollen tube growth. Herein, 17 COBRA genes were isolated from pear genome. Of these genes, the pollen-specifically expressed PbCOB.A.1 and PbCOB.A.2 positively mediates pollen tube growth. The promoters of PbCOB.A.1 and/or PbCOB.A.2 were bound and activated by PbABF.E.2 (an ABRE-binding factor) and PbC2H2.K16.2 (a C2H2-type zinc finger protein). Notably, the expressions of PbCOB.A.1, PbCOB.A.2, and PbC2H2.K16.2 were repressed by self S-RNase, suggesting that self S-RNase reduces the expression of PbCOB.A.1 and PbCOB.A.2 by decreasing the expression of their upstream factors, such as PbC2H2.K16.2, to arrest pollen tube growth. PbCOB.A.1 or PbCOB.A.2 accelerates the growth of pollen tubes treated by self S-RNase, but can hardly affect level of reactive oxygen species and deploymerization of actin cytoskeleton in pollen tubes and cannot physically interact with any reported proteins involved in SI. These results indicate that PbCOB.A.1 and PbCOB.A.2 may not relieve S-RNase toxicity in incompatible pollen tube. The information provides a new route to elucidate the arresting pollen tube growth during SI reaction. Graphical Abstract

Abstract Pear (Pyrus bretschneideri), a valuable widely cultivated fruit, faces significant economic losses due to black spot disease caused by Alternaria alternate (Fr.) Keissl. Trihelix transcription factors (TFs) are crucial in regulating plant defense and autoimmunity. This study aimed to analyze the trihelix transcription factor (GT) genes within pear through genome-wide identification, phylogenetic, gene structure, synteny, and cis-acting elements analyses. Among the 31 trihelix genes, 28 were on 12 known chromosomes, while the remaining 3 were located on unknown chromosomes. These genes were categorized into five clades: SIP1, GTγ, GT1, GT2 and SH4, containing 7, 2, 9, 11 and 2 genes, respectively. Synteny analysis indicated eight duplicated gene pairs. Based on the expression pattern of PbGT genes in seven tissues from the database, the PbGT genes of the GT2 clade were selected for further investigation. The quantitative reverse transcriptase–polymerase chain reaction confirmed that PbrGT5, PbrGT6, PbrGT15 and PbrGT16 correlated with black spot disease resistance. Notably, the salicylic acid (SA) treatment significantly upregulated the expression levels of PbrGT10, PbrGT13, PbrGT15 and PbrGT23. Among these, PbrGT15 showed the highest induction to both SA and black spot infection. Subcellular localization demonstrated that PbrGT15 functions as a nuclear protein. Virus-induced gene silencing of PbrGT15 increased pear plants' susceptibility to black spot disease, indicating its pivotal role in enhancing resistance. These results indicated that PbrGT15 positively regulated black spot disease resistance in pears.

Abstract Gametophytic self-incompatibility (SI) is regulated by S-allele recognition; that is, pollen in a style with the same S-genotype will undergo programmed cell death and stop growing so that it is unable to complete double fertilization, ultimately resulting in the SI response. S-RNase is the female determinant of SI in pear (Pyrus). In the Pyrus genome, there are two different S-RNase alleles at the S-locus, which generate two different S-RNase products in the pistil. The extracted S-glycoprotein is actually a protein complex. In this study, artificial self-pollination was conducted at the bud stage to overcome SI in ‘Huanghua’ (S 1 S 2 ) pear. Seven plants homozygous for S 1 -RNase and four homozygous for S 2 -RNase were selected from the selfed progeny of ‘Huanghua’ by S-gene molecular identification biotechnology. We investigated the function of single S-RNases isolated from the pistils of S-gene homozygous Pyrus germplasm. The pollen of ‘Huanghua’ could smoothly pass through the style of the S-gene homozygous germplasm and complete fertilization. S-RNases were extracted from flower styles of different genotypes and used to treat different types of pollen. The S-RNase from ‘Huanghua’ completely inhibited the growth of S 1 S 2, S 1 S 1 , and S 2 S 2 pollen, while the S-RNase from homozygous germplasm allowed some S 1 S 2 pollen and different single genotypes of pollen to continue growing. These results further validate the core events of SI including cytoskeleton depolymerization and programmed cell death. By iTRAQ-based proteomic analysis of style proteins, a total of 13 S-RNase-related proteins were identified. In summary, we have created reliable S-RNase gene homozygous germplasm, which will play a crucial role in further research on SI in pear and in the development of the pear industry.

The microstructure evolution and corrosion mechanism of Mg–9Er–xGd–2Ni alloys, subjected to Gd addition and extrusion, were investigated in this study. Results shows that with the increase in Gd content from 1.0 wt.% to 5.0 wt.%, the second phase change from continuous network LPSO phase with intragranular γʹ phase to a monolithic continuous network LPSO phase, while the corresponding volume fraction of the second phase remains essentially unchanged. Moreover, it is noted that increasing Gd content contributed to a reduction in the corrosion rate of alloy by reducing galvanic corrosion couples and facilitating the formation of double Gd2O3 and Er2O3 corrosion-resistant films. Ultimately, what's most noteworthy is that compared with as-cast alloys, the corrosion rate of as-extruded alloys increased by 49.2%–125.3% with increasing Gd content, which is due to the streamlined LPSO phase distribution weakening the corrosion barrier effect of high-volume fraction network distribution LPSO phase alloy and grain refinement providing more corrosion interfaces that inhibited the formation of corrosion-resistant product films. This study provides new provides new insights for the development of highly degradable Mg–Er–Gd–Ni alloys that have high strength potential for applications in unconventional oil and gas extraction.

Abstract Background The cytochrome P450 (CYP) superfamily is the largest enzyme metabolism family in plants identified to date, and it is involved in many biological processes, including secondary metabolite biosynthesis, hormone metabolism and stress resistance. However, the P450 gene superfamily has not been well studied in pear (Pyrus spp.). Results Here, the comprehensive identification and a comparative analysis of P450 superfamily members were conducted in cultivated and wild pear genomes. In total, 338, 299 and 419 P450 genes were identified in Chinese white pear, European pear and the wild pear, respectively. Based on the phylogenetic analyses, pear P450 genes were divided into ten clans, comprising 48 families. The motif and gene structure analyses further supported this classification. The expansion of the pear P450 gene family was attributed to whole-genome and single-gene duplication events. Several P450 gene clusters were detected, which have resulted from tandem and proximal duplications. Purifying selection was the major force imposed on the long-term evolution of P450 genes. Gene dosage balance, subfunctionalization and neofunctionalization jointly drove the retention and functional diversification of P450 gene pairs. Based on the association analysis between transcriptome expression profiles and flavonoid content during fruit development, three candidate genes were identified as being closely associated with the flavonoid biosynthesis, and the expression of one gene was further verified using qRT-PCR and its function was validated through transient transformation in pear fruit. Conclusions The study results provide insights into the evolution and biological functions of P450 genes in pear.

Bananas are one of the most important cash crops in the tropics and subtropics. Drought and low-temperature stress affect the growth of banana. The DREB (dehydration responsive element binding protein) gene family, as one of the major transcription factor families, plays crucial roles in defense against abiotic stress. Currently, systematic analyses of the banana DREB (MaDREB) gene family have not yet been reported. In this study, 103 members of the MaDREB gene family were identified in the banana genome. In addition, transcriptomic analysis results revealed that MaDREBs responded to drought and cold stress. The expression of MaDREB14/22/51 was induced by drought and cold stress; these geneswere selected for further analysis. The qRT-PCR validation results confirmed the transcriptome results. Additionally, transgenic Arabidopsis plants overexpressing MaDREB14/22/51 exhibited enhanced resistance to drought and cold stress by reducing MDA content and increasing PRO and soluble sugar content. This study enhances our understanding of the function of the MaDREB gene family, provides new insights into their regulatory role under abiotic stress, and lays a good foundation for improving drought and cold stress-tolerant banana verities.

The leaf area-to-fruit ratio (LAFR) is an important factor affecting fruit quality. Previous studies on LAFR have provided some recommendations for optimal values. However, these recommendations have been quite broad and lack effectiveness during the fruit thinning period. In this study, data on the LAFR and fruit quality of pears at 5 stages were collected by continuously girdling bearing branches throughout the entire fruit development process. Five different clustering algorithms, including KMeans, Agglomerative clustering, Spectral clustering, Birch, and Spectral biclustering, were employed to classify the fruit quality data. Agglomerative clustering yielded the best results when the dataset was divided into 4 clusters. The least squares method was utilized to fit the LAFR corresponding to the best quality cluster, and the optimal LAFR values for 28, 42, 63, 91, and 112 days after flowering were 12.54, 18.95, 23.79, 27.06, and 28.76 dm2 (the corresponding leaf-to-fruit ratio values were 19, 29, 36, 41, and 44, respectively). Furthermore, field verification experiments demonstrated that the optimal LAFR contributed to improving pear fruit quality, and a relatively high LAFR beyond the optimum value did not further increase quality. In summary, we optimized the LAFR of pear trees at different stages and confirmed the effectiveness of the optimal LAFR in improving fruit quality. Our research provides a theoretical basis for managing pear tree fruit load and achieving high-quality, clean fruit production.

Pear fruit senescence under high- and low-temperature conditions has been reported to be mediated by microRNAs. Long non-coding RNAs (lncRNAs), which can function as competing endogenous RNAs that interact with microRNAs, may also be involved in temperature-affected fruit senescence. Based on the transcriptome and microRNA sequencings, in this study, 3 330 lncRNAs were isolated from Pyrus pyrifolia fruit. Of these lncRNAs, 2 060 and 537 were responsive to high- and low-temperature conditions, respectively. Of these differentially expressed lncRNAs, 82 and 24 correlated to the mRNAs involved in fruit senescence under high- and low-temperature conditions, respectively. Moreover, three lncRNAs were predicted to be competing endogenous RNAs (ceRNAs) that interact with the microRNAs involved in fruit senescence, while one and two ceRNAs were involved in fruit senescence under high- and low-temperature conditions, respectively. A dual-luciferase assay showed that the interaction of an lncRNA with a microRNA disrupts the action of the microRNA on the expression of its target mRNA(s). Furthermore, four alternative splicing-derived lncRNAs interacted with miR172i homologies (Novel_88 and Novel_69) to relieve the repressed expression of their target and produce an miR172i precursor. Correlation analysis of microRNA expression suggested that Novel_69 is likely involved in the cleavage of the pre-miR172i hairpin to generate mature miR172i. Taken together, lncRNAs are involved in pear fruit senescence under high- or low-temperature conditions through ceRNAs and the production of microRNA.

Abstract Through agronomic traits and sequencing data, the cultivated and wild varieties of grapes and peaches were analyzed and compared in terms of fruit size, fruit flavor, fruit resistance, and fruit color. Cultivated grapes and peaches have advantages in fruit size, soluble sugar content, sugar and acid ratio, etc. Wild grapes and peaches have utility value in resistance. The results showed that there were 878 and 301 differentially expressed genes in cultivated and wild grapes and peaches in the three growth stages, respectively based on the next-generation sequencing study. Ten and twelve genes related to the differences between cultivated and wild grapes and peaches were found respectively. Among them, three genes, namely chalcone synthase (CHS), glutathione S-transferase (GST) and malate dehydrogenase (MDH1) were present in both cultivated and wild grapes and peaches.

In the past two decades, many studies on piezoelectric nanofibers (NFs) prepared from poly(vinylidene fluoride) (PVDF) and its copolymers, including single NFs, randomly oriented nonwoven mats, and aligned NFs, have been reported. However, studies on the relationships between the PVDF NF diameter, the orientation of the β-phase crystals inside NFs, and the piezoelectric properties of the NFs are still limited. In this study, the effect of the fiber diameter on the internal molecular packing/orientation and piezoelectric properties of aligned PVDF NF thin films was investigated. Herein, piezoelectric thin films composed of densely packed, uniaxially aligned, PVDF NFs with diameters ranging from 228 to 1315 nm were prepared by means of electrospinning with a rotating collector and successive hot-pressing and poling. The effect of the diameters of PVDF NFs on their internal structures, as well as the piezoelectric properties of the thin films, was investigated. All prepared NFs mainly contained β-phase crystals with a similar total crystallinity. The orientation of the β-phase crystals inside the NFs increased with an increase in the fiber diameter, resulting in an improved transverse piezoelectric coefficient (d31) for the thin films. The output voltage of the prepared thin films reached a maximum of 2.7 V at 104 Hz.

Mango fruits are susceptible to diseases, such as anthracnose, during fruit development, leading to yield reduction. Epicuticular wax is closely related to resistance of plants to pathogenic bacterial invasion. In this study, the effect of mango fruit epicuticular wax on the invasion of Colletotrichum gloeosporioides was investigated, followed by to understand the changes of wax chemical composition and crystal morphology during mango fruit development using GC-MS and SEM. Results showed that the epicuticular wax of mango fruits can prevent the invasion of C. gloeosporioides, and ‘Renong’ showed the strongest resistance to C. gloeosporioides. The wax content of four mango varieties first increased and then decreased from 40 days after full bloom (DAFB) to 120 DAFB. In addition, 95 compounds were detected in the epicuticular wax of the four mango varieties at five developmental periods, in which primary alcohols, terpenoids and esters were the main wax chemical composition. Furthermore, the surface wax structure of mango fruit changed dynamically during fruit development, and irregular platelet-like crystals were the main wax structure. The present study showed the changes of wax content, chemical composition and crystal morphology during mango fruit development, and the special terpenoids (squalene, farnesyl acetate and farnesol) and dense crystal structure in the epicuticular wax of ‘Renong’ fruit may be the main reason for its stronger resistance to C. gloeosporioides than other varieties. Therefore, these results provide a reference for the follow-up study of mango fruit epicuticular wax synthesis mechanism and breeding.

Pear is one of the most economically important fruit crops widely distributed around the world. To enrich the pear germplasm resources for obtaining the polyploid plants, the seeds from pear cultivar of 'Dangshansuli' (Pyrus bretschneideri) were treated with colchicine solution under concentration gradients of 0.1%, 0.5% and 1.0% for 12-, 24- and 48-h, respectively. Our results showed that colchicine could inhibit seed germination, and the inhibitory effect was positive with the treated concentration of colchicine. We detected that the 0.1% colchicine solution for 12 h was the most effective treatment for inducing pear polyploidy. Furthermore, compared with the wild diploid pear plants, a significant morphological variation, such as shorter plants, smaller and oval leaves were observed in the induced tetraploid pear plants. Significant changes in leaf anatomy, including larger and sparse stomata and the epidermal cells with irregularly or polygonal-shape were also detected in tetraploid pear plants. In addition, the contents of 10 cuticular wax compounds from the leaves of diploid and tetraploid pear plants were also detected with significant difference. Taken together, this study lays the foundation for polyploid breeding of pear for contributing diverse characteristics.

Abstract Background The circadian clock integrates endogenous and exogenous signals and regulates various physiological processes in plants. REVEILLE (RVE) proteins play critical roles in circadian clock system, especially CCA1 (CIRCADIAN CLOCK ASSOCIATED 1) and LHY (LATE ELONGATED HYPOCOTYL), which also participate in flowering regulation. However, little is known about the evolution and function of the RVE family in Rosaceae species, especially in Pyrus bretschneideri. Results In this study, we performed a genome-wide analysis and identified 51 RVE genes in seven Rosaceae species. The RVE family members were classified into two groups based on phylogenetic analysis. Dispersed duplication events and purifying selection were the main drivers of evolution in the RVE family. Moreover, the expression patterns of ten PbRVE genes were diverse in P. bretschneideri tissues. All PbRVE genes showed diurnal rhythms under light/dark cycles in P. bretschneideri leaves. Four PbRVE genes also displayed robust rhythms under constant light conditions. PbLHY, the gene with the highest homology to AtCCA1 and AtLHY in P. bretschneideri, is localized in the nucleus. Ectopic overexpression of PbLHY in Arabidopsis delayed flowering time and repressed the expression of flowering time-related genes. Conclusion These results contribute to improving the understanding and functional research of RVE genes in P. bretschneideri.

The Chinese pear (Pyrus spp.) exhibits typical gametophytic self-incompatibility (GSI), which inhibits self-crossing and promotes out-crossing, similar to other fruit species in the Rosaceae family. Thus, S-compatible cultivars are required in pear orchards to ensure successful pollination and stable yields. In this study, 84 native Chinese pear accessions were genotyped by allele-specific PCR using one pair allele consensus primers and 29 pairs of S-allele-specific primers that were designed in this study. After cloning and sequencing the PCR products, the S-genotypes of all 84 pear accessions, including wild and cultivated accessions, were determined. The reported 34 S-alleles and a novel S-allele were isolated from these pear accessions. These S-alleles were expressed specifically in the style. Sequence analysis identified that six pear cultivars originated in China shared the same S-RNases with P. communis (Pc). These findings supported the hypothesis that oriental and occidental Pyrus spp. may share the same pool of alleles at the S-locus. A novel S-RNase was isolated in ‘Putiandouli’, ‘Daguoshanli’, ‘Yunhongli 1′, and ‘Dianli’ and deposited as S67-RNase under accession number MT773568. Furthermore, the deduced amino acid sequences exhibited high similarity (99.56%) to S32-RNase in Malus. The high similarity between S-RNase in Pyrus and Malus indicated that the existence of S-RNase could have predated speciation between Pyrus and Malus. Furthermore, S-allele information was rearranged in Asian and European pears to provide information for selecting the best pollinator for widely cultivated pear cultivars in China. This information is useful for pear production, cross-breeding, and understanding the mechanism of the self-incompatibility reaction.

Abstract In contrast to most land plant species, sorbitol, instead of sucrose, is the major photosynthetic product in many Rosaceae species. It has been well illustrated that three key functional genes encoding sorbitol-6-phosphate dehydrogenase (S6PDH), sorbitol dehydrogenase (SDH), and sorbitol transporter (SOT), are mainly responsible for the synthesis, degradation and transportation of sorbitol. In this study, the genome-wide identification of S6PDH, SDH and SOT genes was conducted in four Rosaceae species, peach, mei, apple and pear, and showed the sorbitol bio-pathway to be dominant (named sorbitol present group, SPG); another three related species, including tomato, poplar and Arabidopsis, showed a non-sorbitol bio-pathway (named sorbitol absent group, SAG). To understand the evolutionary differences of the three important gene families between SAG and SPG, their corresponding gene duplication, evolutionary rate, codon bias and positive selection patterns have been analyzed and compared. The sorbitol pathway genes in SPG were found to be expanded through dispersed and tandem gene duplications. Branch-specific model analyses revealed SDH and S6PDH clade A were under stronger purifying selection in SPG. A higher frequency of optimal codons was found in S6PDH and SDH than that of SOT in SPG, confirming the purifying selection effect on them. In addition, branch-site model analyses revealed SOT genes were under positive selection in SPG. Expression analyses showed diverse expression patterns of sorbitol-related genes. Overall, these findings provide new insights in the evolutionary characteristics for the three key sorbitol metabolism-related gene families in Rosaceae and other non-sorbitol dominant pathway species.

Abstract Background Glucose can be involved in metabolic activities as a structural substance or signaling molecule and plays an important regulatory role in fruit development. Glucose metabolism is closely related to the phenylpropanoid pathway, but the specific role of glucose in regulating lignin biosynthesis in pear fruit is still unclear. The transcriptome of pear calli generated from fruit and treated with glucose was analyzed to investigate the role of glucose in lignin biosynthesis. Results The treatment of exogenous glucose significantly enhanced the accumulation of lignin in pear calli. A total of 6566 differentially expressed genes were obtained by transcriptome sequencing. Glycolysis was found to be the pathway with significant changes. Many differentially expressed genes were enriched in secondary metabolic pathways, especially the phenylpropanoid pathway. Expression of structural genes (PbPAL, PbHCT, PbCOMT, PbPRX) in lignin biosynthesis was up-regulated after glucose treatment. In addition, glucose might regulate lignin biosynthesis through interactions with ABA, GA, and SA signaling. Several candidate MYB transcription factors involved in glucose-induced lignin biosynthesis have also been revealed. The qRT-PCR analyses showed that the expression pattern of PbPFP at early developmental stage in ‘Dangshansuli’ fruits was consistent with the trend of lignin content. Transient expression of PbPFP resulted in a significant increase of lignin content in ‘Dangshansuli’ fruits at 35 days after full bloom (DAB) and tobacco leaves, indicating that PbPFP (Pbr015118.1) might be associated with the enhancement of lignin biosynthesis in response to glucose treatment. Conclusions PbPFP plays a positive role in regulating lignin biosynthesis in response to glucose treatment. This study may reveal the regulatory pathway related to lignin accumulation in pear calli induced by glucose.

ObjectivesResection of pheochromocytoma and paraganglioma (PPGL) carries risks with perioperative hemodynamic instability. Phenoxybenzamine (PXB) is a commonly used α-blockade to prevent it. It is unclear whether lengthening the preoperative duration of PXB is better for hemodynamic stability and postoperative outcomes. Furthermore, different types of catecholamines have varying effects on perioperative hemodynamics. Thus, our study aimed to investigate the impact of the duration of preoperative preparation with PXB and secretory phenotypes of the patients on intraoperative hemodynamic stability and postoperative complications in PPGL.MethodsBetween Dec 2014 and Jan 2022, 166 patients with PPGL were operated on by the same team at Sun Yat-sen Memorial Hospital. They were divided into group A(1-14d), Group B(15-21d), and Group C(>21d) based on the duration of management with PXB and into the adrenergic and the noradrenergic phenotype group based on secretory profiles. Data on intraoperative hemodynamics and postoperative outcomes were collected and compared among groups.ResultsA total of 96 patients occurred intraoperative hemodynamic instability, and 24 patients had 29 postoperative complications related to the surgery. Among the 145 patients treated with PXB, no significant differences were found in the cumulative time outside the target blood pressure(6.67%[0-17.16%] vs. 5.97%[0-23.08%] vs. 1.22%[0-17.27%], p=0.736) or in the median total HI-score(42.00[30.00-91.00] vs. 89.00[30.00-113.00] vs. 49.00[30.00-93.00], p=0.150) among group A(n=45), B(n=51) and C(n=49). Multivariate analysis demonstrated that the level of plasma-free metanephrine(MN) was an independent risk factor for intraoperative hemodynamic instability. And the median cumulative time outside of the target blood pressure in the adrenergic phenotype group was significantly greater than that in the noradrenergic phenotype group(8.17%[0-26.22%] vs. 1.86%[0-11.74%], p=0.029). However, the median total HI-score(99.50[85.00-113.25] vs. 90.00[78.00-105.00], p=0.570) and postoperative outcomes showed no differences between the two groups.ConclusionsA preoperative duration of nearly 14 days with PXB is sufficient for ensuring intraoperative hemodynamic stability in PPGL. And lengthening the preparation duration may not provide additional benefits in the era of widespread application and advanced techniques of laparoscopic surgery. Additionally, patients with the adrenergic phenotype are more prone to intraoperative hemodynamic instability than the noradrenergic phenotype. Thus, more attention should be given to the adrenergic phenotype during surgery.

Abstract Abscisic acid (ABA) is a phytohormone that plays important roles in the regulation of plant growth, seed germination, and stress responses. The pyrabactin resistance 1-like (PYR/PYL) protein, an ABA receptor, was involved in the initial step in ABA signal transduction. However, the evolutionary history and characteristics of PYL genes expression remain unclear in Chinese white pear (Pyrus bretschneideri) or other Rosaceae species. In this study, 67 PYL genes were identified in eight Rosaceae species, and have been classified into three subgroups based on specific motifs and phylogenetic analysis. Intriguingly, we observed that whole-genome duplication (WGD) and dispersed duplication (DSD) have a major contribution to PYL family expansion. Purifying selection was the major force in PYL genes evolution. Expression analysis finds that PYL genes may function in multiple pear tissues. qRT-PCR validation of 11 PbrPYL genes indicates their roles in seed germination and abiotic stress responses. Our study provides a basis for further elucidation of the function of PYL genes and analysis of their expansion, evolution and expression patterns, which helps to understand the molecular mechanism of pear response to seed germination and seedling abiotic stress.

To realize autonomous navigation and intelligent management in orchards, vehicles require real-time positioning and globally consistent mapping of surroundings with sufficient information. However, the unstructured and unstable characteristics of orchards present challenges for accurate and stable localization and mapping. This study proposes a framework fusing LiDAR, visual, and inertial data by using the extended Kalman filter (EKF) to achieve real-time localization and colorful LiDAR point-cloud mapping in orchards. First, the multi-sensor data were integrated into a loosely-coupled framework based on the EKF to improve the pose estimation, with the pose estimation from LiDAR and gyroscope acting as the predictions, while that from visual-inertial odometry acting as the observations. Then, the Loam_Livox algorithm was enhanced by incorporating color from the image into the LiDAR point cloud, enabling the real-time construction of a three-dimensional colorful map of the orchard. The method demonstrates a high accuracy for localization in different motion trajectories (average RMSE: 0.3436) and different scenarios (average RMSE: 0.1230) and clear and efficient construction of three-dimensional colorful mapping, taking only 75.01 ms in localization and mapping for a frame of LiDAR point cloud. This indicates the proposed method has a great potential for the autonomous navigation of agricultural vehicles.

Whole-genome duplication (WGD) has occurred repeatedly during plant evolution and diversification, providing genetic layers for evolving new functions and phenotypes. Advances in long-read sequencing technologies have enabled sequencing and assembly of over 1000 plant genomes spanning nearly 800 species, in which a large set of ancient WGDs has been uncovered. Here, we review the recently reported WGDs that occurred in major plant lineages and key evolutionary positions, and highlight their contributions to morphological innovation and adaptive evolution. Current gaps and challenges in integrating enormous volumes of sequenced plant genomes, accurately inferring WGDs, and developing web-based analysis tools are emphasized. Looking to the future, ambitious genome sequencing projects and global efforts may substantially recapitulate the plant tree of life based on broader sampling of phylogenetic diversity, reveal much of the timetable of ancient WGDs, and address the biological significance of WGDs in plant adaptation and radiation.

Crop breeding programs generate large datasets. Thus, it is difficult to ensure the accuracy and integrity of all the collected data in the breeding process. To improve breeding efficiency, we established an open source and free breeding evaluation information system (BreedingEIS). The full system is composed of a web client and a mobile client. The web client is used to name the individual breeding offspring plants and analyze data. The mobile client is based on the technology of widely used smartphones and is suitable for Android and iOS systems. Its functions focus on field evaluation, including quick response code recognition, evaluation data entry, and real-time viewing. In addition, near-field communication technology and portable label machines are introduced to enable breeders to quickly locate each individual plant and accurately label any samples collected from it. Generally, BreedingEIS enables users to accurately and conveniently register phenotypic data and quickly lock target individual plants from large volumes of data. The system provides a low-cost and highly efficient solution for crop information evaluation and enables breeders to better collect, manage, and use breeding data for decision making, which is a valuable resource for crop breeding.

Abstract Background Transcription factors (TFs) are involved in many important biological processes, including cell stretching, histological differentiation, metabolic activity, seed storage, gene regulation, and response to abiotic and biotic stresses. Little is known about the functions, evolutionary history, and expression patterns of basic region-leucine zipper TF family genes in pear, despite the release of the genome of Chinese white pears (“Dangshansuli”). Results Overall, 92 bZIP genes were identified in the pear genome (Pyrus breschneideri). Of these, 83 were randomly distributed on all 17 chromosomes except chromosome 4, and the other 9 genes were located on loose scaffolding. The genes were divided into 14 subgroups. Whole-genome duplications, dispersed duplication, and purifying selection for whole-genome duplications are the main reasons for the expansion of the PbrbZIP gene family. The analysis of functional annotation enrichment indicated that most of the functions of PbrbZIP genes were enriched in Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathways involved in the abiotic stress response. Next, expression analysis and virus-induced gene silencing results indicated that PbrbZIP genes might play critical roles in response to drought and cold stresses, especially for the genes from subgroups A, C, G, I, and S. Conclusions Ninety-two PbrbZIP genes were identified from the pear genome and classified into 14 subgroups. PbrbZIP genes were mainly expanded from whole-genome duplications and dispersed duplications and retained by purifying selection. PbrbZIP genes were induced by cold and drought stresses and played important roles in drought and cold tolerance. These results provided useful information for further increasing the tolerance of pears to stresses and a foundation to study the cold and drought tolerance mechanism of PbrbZIP genes.

Abstract Background The content of stone cells in pears has a great influence on taste. Stone cells are formed by the accumulation of lignin. The treatment of exogenous calcium can affect the lignin synthesis, but this Ca-mediated mechanism is still unclear. In this study, the author performed a comparative transcriptomic analysis of callus of pears (Pyrus x bretschneideri) treated with calcium nitrate Ca (NO3)2 to investigate the role of calcium in lignin synthesis. Results There were 2889 differentially expressed genes (DEGs) detected between the Control and Ca (NO3)2 treatment in total. Among these 2889 DEGs, not only a large number of genes related to Ca single were found, but also many genes were enriched in secondary metabolic pathway, especially in lignin synthesis. Most of them were up-regulated during the development of callus after Ca (NO3)2 treatment. In order to further explore how calcium nitrate treatment affects lignin synthesis, the author screened genes associated with transduction of calcium signal in DEGs, and finally found CAM, CML, CDPK, CBL and CIPK. Then the author identified the PbCML3 in pears and conducted relevant experiments finding the overexpression of PbCML3 would increase the content of pear stone cells, providing potential insights into how Ca treatment enhances the stone cell in pears. Conclusions Our deep analysis reveals the effects of exogenous calcium on calcium signal and lignin biosynthesis pathway. The function of PbCML3 on stone cells formation was verified in pear.

ObjectiveTo raise awareness of Cytochrome P450 Oxidoreductase Deficiency (PORD, a rare form of congenital adrenal hyperplasia (CAH), through a case of pregnant woman with virilization symptoms.Case descriptionA 30-year-old Chinese woman was referred to hospital after 7 years of presenting signs of virilization, including voice deepening, acromegaly, hirsutism, clitoromegaly, and acne. These symptoms appeared since her third gestation. Her second birth died 9 hours after birth and had signs of clitoris hypertrophy. Her third born was a son who presented with flat nose, radius and humerus bone malformation, and small penis at birth. Panel of POR-related genetic tests revealed that the patient carried c.1370 G>A (p.R457H), which is a POR heterozygous gene, while her husband carried a POR heterozygous gene as well, c.1379 C>A (p.S460Y). Two heterozygous mutations of the POR were found in her son: c.1370 G>A and c.1379 C>A. In PORD, c.1370 G>A (p.R457H) was reported as a susceptible gene, while c.1379 C>A (p.S460Y) has not been reported as responsible for the disease so far.Discussion and literature reviewPORD is a rare form of CAH and caused by POR gene mutations. Most PORD patients are identified and diagnosed in pediatrics department. Internal medicine and obstetrics physicians are unfamiliar with the disease. As clinical manifestations are diverse, PORD could be easy to miss or to be misdiagnosed. Typical clinical manifestation includes adrenal insufficiency-related symptoms, such as bone malformations and sexual development disorders. PORD is diagnosed through genetic testing. Investigations of steroid metabolic products in urine through gas chromatography-mass spectrometry or liquid chromatography-mass spectrometry are also helpful for the diagnosis, but neither of them are widely available in China. In this case, the patient had a history of infertility, and her third child was born with congenital defect and carried a PORD-related gene. In general clinical practice, if a pregnant woman presents with abnormal virilization symptoms, CAH possibilities should be considered, including rare causes such as PORD.ConclusionPORD is a rare autosomal recessive genetic disease. We summarised the clinical characteristics and genotypes that were previously reported in the Chinese population and identified a novel mutation.

The leaf phenotypic traits of plants have a significant impact on the efficiency of canopy photosynthesis. However, traditional methods such as destructive sampling will hinder the continuous monitoring of plant growth, while manual measurements in the field are both time-consuming and laborious. Nondestructive and accurate measurements of leaf phenotypic parameters can be achieved through the use of 3D canopy models and object segmentation techniques. This paper proposed an automatic branch–leaf segmentation pipeline based on lidar point cloud and conducted the automatic measurement of leaf inclination angle, length, width, and area, using pear canopy as an example. Firstly, a three-dimensional model using a lidar point cloud was established using SCENE software. Next, 305 pear tree branches were manually divided into branch points and leaf points, and 45 branch samples were selected as test data. Leaf points were further marked as 572 leaf instances on these test data. The PointNet++ model was used, with 260 point clouds as training input to carry out semantic segmentation of branches and leaves. Using the leaf point clouds in the test dataset as input, a single leaf instance was extracted by means of a mean shift clustering algorithm. Finally, based on the single leaf point cloud, the leaf inclination angle was calculated by plane fitting, while the leaf length, width, and area were calculated by midrib fitting and triangulation. The semantic segmentation model was tested on 45 branches, with a mean Precisionsem, mean Recallsem, mean F1-score, and mean Intersection over Union (IoU) of branches and leaves of 0.93, 0.94, 0.93, and 0.88, respectively. For single leaf extraction, the Precisionins, Recallins, and mean coverage (mCoV) were 0.89, 0.92, and 0.87, respectively. Using the proposed method, the estimated leaf inclination, length, width, and area of pear leaves showed a high correlation with manual measurements, with correlation coefficients of 0.94 (root mean squared error: 4.44°), 0.94 (root mean squared error: 0.43 cm), 0.91 (root mean squared error: 0.39 cm), and 0.93 (root mean squared error: 5.21 cm2), respectively. These results demonstrate that the method can automatically and accurately measure the phenotypic parameters of pear leaves. This has great significance for monitoring pear tree growth, simulating canopy photosynthesis, and optimizing orchard management.

Abstract Background Pears and apples are both perennial deciduous trees of the Rosaceae family, and both are important economic fruit trees worldwide. The emergence of many varieties in the market has been mostly domesticated from wild to cultivated and regulated by the differential expression of genes. However, the molecular process and pathways underlying this phenomenon remain unclear. Four typical wild and cultivar pear and apple trees at three developmental stages were used in our study to investigate the molecular process at the transcriptome level. Result Physiological observations indicated the obvious differences of size, weight, sugar acid content and peel color in wild and cultivar fruit among each developmental stage. Using next-generation sequencing based RNA-seq expression profiling technology, we produced a transcriptome in procession of a large fraction of annotated pear and apple genes, and provided a molecular basis underlying the phenomenon of wild and cultivar fruit tree differences. 5921 and 5744 differential expression genes were identified in pear and apple at three developmental stages respectively. We performed temporal and spatial differential gene expression profiling in developing fruits. Several key pathways such as signal transduction, photosynthesis, translation and many metabolisms were identified as involved in the differentiation of wild and cultivar fruits. Conclusion In this study, we reported on the next-generation sequencing study of the temporal and spatial mRNA expression profiling of pear and apple fruit trees. Also, we demonstrated that the integrated analysis of pear and apple transcriptome, which strongly revealed the consistent process of domestication in Rosaceae fruit trees. The results will be great influence to the improvement of cultivar species and the utilization of wild resources.

Abstract Background The plant U-box (PUB) proteins are a family of ubiquitin ligases (E3) enzymes that involved in diverse biological processes, as well as in responses to plant stress response. However, the characteristics and functional divergence of the PUB gene family have not yet been previously studied in the Chinese white pear (Pyrus bretschneideri). Results In the present study, we identified 62 PbrPUBs in Chinese white pear genome. Based on the phylogenetic relationship, 62 PUB genes were clustered into five groups. The results of conserved motif and gene structure analysis supported the classification phylogenetic tree. The PbrPUB genes were unevenly distribution on 17 pear chromosomes, chromosome 15 housed most member of PUB family, with eight PUB genes. Cis-acting element analysis indicated that PUB genes might participate in diverse biological processes, especially in the response to abiotic stresses. Based on RNA-data from ‘Dangshansuli’ at seven tissues, we found that PUB genes exhibited diverse of expression level in seven tissues, and qRT-PCR experiment further supported the reliable of RNA-Seq data. To identify candidate genes associated with resistance, we conducted qRT-PCR experiment the expression level of pear seed plant under four abiotic stresses, including: ABA, dehydration, salt and cold treatment. One candidate PUB gene associated with dehydration stress was selected to conduct further functional experiment. Subcellular localization revealed PbrPUB18 protein was located on cell nucleus. Furthermore, heterologous over-expression of PbrPUB18 in Arabidopsis indicated that the over-expression of PbrPUB18 could enhance resistance in drought treatment. In conclusions, we systematically identified the PUB genes in pear, and provided useful knowledge for functional identification of PUB genes in pear.

Abstract Pear [Pyrus bretschneideri cv. Dangshan Su] fruit quality is not always satisfactory owing to the presence of stone cells, and lignin is the main component of stone cells in pear fruits. Caffeoyl shikimate esterase (CSE) is a key enzyme in the lignin biosynthesis. Although CSE-like genes have been isolated from a variety of plant species, their orthologs are not characterized in pear. In this study, the CSE gene family (PbCSE) from P. bretschneideri was identified. According to the physiological data and quantitative RT-PCR (qRT-PCR), PbCSE1 was associated with lignin deposition and stone cell formation. The overexpression of PbCSE1 increased the lignin content in pear fruits. Relative to wild-type (WT) Arabidopsis, the overexpression of PbCSE1 delayed growth, increased the lignin deposition and lignin content in stems. Simultaneously, the expression of lignin biosynthetic genes were also increased in pear fruits and Arabidopsis. These results demonstrated that PbCSE1 plays an important role in cell lignification and will provide a potential molecular strategy to improve the quality of pear fruits.

PurposeThe association between primary aldosteronism (PA) and lower bone mineral density (BMD) has raised a concern, but the contributing factors remain unclear. We aim to explore the risk factors for lower BMD in PA patients.MethodsWe analyzed and compared the data of 60 PA patients with 60 matched essential hypertension (EH) patients. BMD, bone metabolites, and several oxidative stress and inflammation indicators—including C-reactive protein (CRP), superoxide dismutase (SOD), total bilirubin (TBIL), mean platelet volume (MPV), etc.—were assessed and compared in PA and EH patients. Bivariate correlation analysis and multivariate linear regression analysis were performed to explore the factors associated with BMD in PA patients.ResultsThe BMD measured by quantitative computed tomography in PA patients was lower than that in EH patients (141.9 ± 34.0 vs. 158.9 ± 55.9 g/cm3, p = 0.047), especially in patients less than 50 years old. BMD was independently negatively associated with age (standardized β = -0.581, p < 0.001), serum phosphorus (standardized β = -0.203, p = 0.008), urinary calcium excretion (standardized β = -0.185, p = 0.031), and MPV (standardized β = -0.172, p = 0.043) and positively associated with SOD (standardized β = 0.205, p = 0.011) and TBIL (standardized β = 0.212, p = 0.015).ConclusionsThe PA patients showed a lower BMD than the EH patients, which was associated with age, serum phosphorus, urinary calcium excretion, MPV, SOD, and TBIL. These variables might be potential markers for the assessment of bone loss and efficacy of treatments in PA patients.

Stone cells have been described to substantially influence pear fruit quality, as lignin and cellulose are the main components of stone cells. However, there are limited studies on the relationship between the variation and molecular basis of stone cells, lignin and cellulose content among different pear varieties. Here, to reveal the variation of stone cell content within different cultivated species, we collected 236 germplasms of sand pear (Pyrus pyrifolia) at 50 days after flower blooming (DAFB), the key stage of stone cell formation. In our results, we measured the content of stone cells, lignin and cellulose and found that these contents ranged from 2.82% to 29.00%, 8.84% to 55.30% and 11.52% to 30.55%, respectively. Further analysis showed that the variation coefficient of stone cell, lignin and cellulose content was 39.10%, 28.03% and 16.71%, respectively. Additionally, a significant correlation between stone cell, lignin and cellulose content were detected, and the correlation coefficient between the contents of stone cell and lignin (0.912) was higher than between the contents of stone cell and cellulose (0.796). Moreover, the average lignin content (29.73%) was higher than the average cellulose content (18.03%) in stone cells in pear fruits, indicating that lignin is the main component of stone cell in pears. Finally, on the basic of the transcriptome data, we identified 10 transcription factors belonging to bHLH, ERF, MYB, and NAC transcript families, which might be involved in lignin formation in stone cells. qRT-PCR experiments verified coincident trends between expression of candidate genes and stone cell content. This research laid foundation for future studies on genetic variation of stone cells in pear fruits and provided important gene resources for stone cell regulation.

Abstract Background The basic helix-loop-helix (bHLH) transcription factors play important roles in many processes in plant growth, metabolism and responses to abiotic stresses. Although, the sequence of Chinese white pear genome (cv. ‘Dangshansuli’) has already been reported, there is still a lack of clarity regarding the bHLH family genes and their evolutionary history. Results In this work, a genome-wide identification of the bHLH genes in Chinese white pear was performed, and we characterized the functional roles of these PbrbHLH genes in response to abiotic stresses. Based on the phylogenetic analysis and structural characteristics, 197 identified bHLH genes could be well classified into 21 groups. Expansion of PbrbHLH gene family was mainly driven by WGD and dispersed duplication with the purifying selection from the recent WGD. The functional annotation enrichment showed that the majority of PbrbHLHs were enriched in the GO terms and KEGG pathways involved in responds to stress conditions as TFs. Transcriptomic profiles and qRT-PCR revealed that PbrbHLH7, PbrbHLH8, PbrbHLH128, PbrbHLH160, PbrbHLH161 and PbrbHLH195 were significantly up-regulated under cold and drought treatments. In addition, PbrbHLH195-silenced pear seedlings display significant reduced cold tolerance, exhibiting reduced chlorophyll content, as well as increased electrolyte leakage and concentrations of malondialdehyde and H2O2. Conclusion For the first time, a comprehensive analysis identified the bHLH genes in Chinese white pear and demonstrated that PbrbHLH195 is involved in the production of ROS in response to cold stress, suggesting that members of the PbrbHLH family play an essential role in the stress tolerance of pear.

Studies of fruit quality traits in pears are lagging behind the other major fruit trees. Here, the authors conduct GWAS of fruit quality and phenological traits in a panel of 312 sand pear accessions using SNPs called from resequencing data, and reveal the involvement of a lignin formation-related protein in regulating stone cell development.

N6-methylated adenine (m6A) is the most prevalent modification of mRNA methylation and can regulate many biological processes in plants, such as mRNA processing, development, and stress response. Some studies have increased our understanding of its various roles in model plants in recent years. Nevertheless, the distribution of m6A and the impact of m6A on the regulation of plant defense responses against pathogen inoculation are virtually unknown in pear. In this study, MeRIP-seq and RNA-seq data from healthy and inoculated plants were analyzed to assess the changes in the transcript levels and posttranscriptional modification of pear in response to the fire blight pathogen Erwinia amylovora. Following the analysis of 97,261 m6A peaks, we found that m6A preferred to modify duplicate genes rather than singleton genes and that m6A-methylated genes underwent stronger purifying selection. A total of 2,935 specific m6A sites were detected at the transcriptome level after inoculation, which may increase defense-related transcript abundance to enhance pear resistance. In addition, 1,850 transcripts were detected only in the mock-inoculated groups. The hypomethylated transcripts were mainly related to transcriptional regulation and various biological processes, such as chloroplast organization and sucrose biosynthetic processes. In addition, we found that the extent of m6A methylation was significantly positively correlated with the transcript level, suggesting a regulatory role for m6A in the plant response.

Abstract Background The lateral organ boundaries domain (LBD) gene is a plant-specific transcription factor that plays a critical role in diverse biological processes. However, the evolution and functional divergence of the LBD gene family has not yet been characterized for the Chinese White Pear. Results In our study, a total of 60 PbrLBDs were identified in the pear genome. The PbrLBD gene family was divided into two classes based on gene structure and phylogenetic analysis: class I (53) and class II (7). Cis-acting element analysis results suggested that PbrLBDs may participate in various biological processes, such as flavonoid biosynthetic and stress response. Synteny analysis results indicated that segmental duplication played a key role in the expansion of the PbrLBD gene family. The mean Ks and 4DTv values showed that the PbrLBD gene family had undergone only one recent whole-genome duplication event occurring at 30–45 MYA. Purifying selection was a primary force during the PbrLBD gene family evolution process. Transcriptome data analysis revealed that 10 PbrLBDs were expressed in all six examined tissues, and 73.33% of members in the PbrLBD gene family were expressed in pear sepal. qRT-PCR was conducted to verify the expression levels of 11 PbrLBDs in these six tissues. Specifically, PbrLBD20, PbrLBD35 and PbrLBD53 genes were down-regulated when anthocyanin concentrations were high, whereas PbrLBD33 was significantly up-regulated in pear when anthocyanin concentrations were high. Furthermore, PbrLBD20, one of the candidate genes related to anthocyanins was localized in the nucleus. Conclusions Our analysis provides valuable information for understanding the evolution of the PbrLBD gene family, and provides new insights into the regulation of pear pigment metabolism and lays a foundation for the future disclosure of the molecular mechanism of LBD gene regulating flavonoid metabolism.

Abstract Background Lysin motif-containing proteins (LYP), which act as pattern-recognition receptors, play central roles in growth, node formation, and responses to biotic stresses. The sequence of Chinese white pear genome (cv. ‘Dangshansuli’) along with the seven other species of Rosaceae has already been reported. Although, in these fruit crops, there is still a lack of clarity regarding the LYP family genes and their evolutionary history. Results In the existing study, eight Rosaceae species i.e., Pyrus communis, Prunus persica, Fragaria vesca, Pyrus bretschneideri, Prunus avium, Prunus mume, Rubus occidentalis, and Malus × domestica were evaluated. Here, we determined a total of 124 LYP genes from the underlined Rosaceae species. While eighteen of the genes were from Chinese white pear, named as PbrLYPs. According to the LYPs structural characteristics and their phylogenetic analysis, those genes were classified into eight groups (group LYK1, LYK2, LYK3, LYK4/5, LYM1/3, LYM2, NFP, and WAKL). Dispersed duplication and whole-genome duplication (WGD) were found to be the most contributing factors of LYP family expansion in the Rosaceae species. More than half of the duplicated PbrLYP gene pairs were dated back to the ancient WGD (~ 140 million years ago (MYA)), and PbrLYP genes have experienced long-term purifying selection. The transcriptomic results indicated that the PbrLYP genes expression was tissue-specific. Most PbrLYP genes showed differential expression in leaves under fungal pathogen infection with two of them located in the plasmalemma. Conclusion A comprehensive analysis identified 124 LYP genes in eight Rosaceae species. Our findings have provided insights into the functions and characteristics of the Rosaceae LYP genes and a guide for the identification of other candidate LYPs for further genetic improvements for pathogen-resistance in higher plants.