Ascorbic acid releases dormancy and promotes germination by an integrated regulation of abscisic acid and gibberellin in Pyrus betulifolia seeds.

Seed dormancy is an important life history state in which intact viable seeds delay or prevent germination under suitable conditions. Ascorbic acid (AsA) acts as a small molecule antioxidant, and breaking seed dormancy and promoting subsequent growth are among its numerous functions. In this study, a germination test using Pyrus betulifolia seeds treated with exogenous AsA or AsA synthesis inhibitor lycorine (Lyc) and water absorption was conducted. The results indicated that AsA released dormancy and increased germination and 20 mmol L-1 AsA promoted cell division, whereas Lyc reduced germination. Seed germination showed typical three phases of water absorption; and seeds at five key time points were sampled for transcriptome analysis. It revealed that multiple pathways were involved in breaking dormancy and promoting germination through transcriptome data, and 12 differentially expressed genes (DEGs) related to the metabolism and signal transduction of abscisic acid (ABA) and gibberellins (GA) were verified by subsequent RT-qPCR. For metabolites, exogenous AsA increased endogenous AsA and GA3 but reduced ABA and the ABA/GA3 ratio. In addition, three genes regulating ABA synthesis were downregulated by AsA, while five genes mediating ABA degradation were upregulated. Taken together, AsA regulates the pathways associated with ABA and GA synthesis, catalysis, and signal transduction, with subsequent reduction in ABA and increase in GA and further the balance of ABA/GA, ultimately releasing dormancy and promoting germination.

Increased DNA methylation contributes to the early ripening of pear fruits during domestication and improvement.

BACKGROUND: DNA methylation is an essential epigenetic modification. However, its contribution to trait changes and diversity in the domestication of perennial fruit trees remains unknown. RESULTS: Here, we investigate the variation in DNA methylation during pear domestication and improvement using whole-genome bisulfite sequencing in 41 pear accessions. Contrary to the significant decrease during rice domestication, we detect a global increase in DNA methylation during pear domestication and improvement. We find this specific increase in pear is significantly correlated with the downregulation of Demeter-like1 (DML1, encoding DNA demethylase) due to human selection. We identify a total of 5591 differentially methylated regions (DMRs). Methylation in the CG and CHG contexts undergoes co-evolution during pear domestication and improvement. DMRs have higher genetic diversity than selection sweep regions, especially in the introns. Approximately 97% of DMRs are not associated with any SNPs, and these DMRs are associated with starch and sucrose metabolism and phenylpropanoid biosynthesis. We also perform correlation analysis between DNA methylation and gene expression. We find genes close to the hypermethylated DMRs that are significantly associated with fruit ripening. We further verify the function of a hyper-DMR-associated gene, CAMTA2, and demonstrate that overexpression of CAMTA2 in tomato and pear callus inhibits fruit ripening. CONCLUSIONS: Our study describes a specific pattern of DNA methylation in the domestication and improvement of a perennial pear tree and suggests that increased DNA methylation plays an essential role in the early ripening of pear fruits.

Endoglucanase H from Aspergillus westerdijkiae Plays an Important Role in the Virulence on Pear Fruits.

Aspergillus westerdijkiae can infect many agricultural products including cereals, grapes, and pear. Pathogenic fungi secrete diverse effectors as invasive weapons for successful invasion the host plant. During the pathogen-host interaction, 4486 differentially expressed genes were observed in A. westerdijkiae with 2773 up-regulated and 1713 down-regulated, whereas 8456 differentially expressed genes were detected in pear fruits with 4777 up-regulated and 3679 down-regulated. A total of 309 effector candidate genes were identified from the up-regulated genes in A. westerdijkiae. Endoglucanase H (AwEGH) was significantly induced during the pathogen-host interaction. Deletion of AwEGH resulted in altered fungal growth and morphology and reduced conidia production and germination compared to the wild-type. Further experiments demonstrated that AwEGH plays a role in cell wall integrity. Importantly, disruption of AwEGH significantly reduced the fungal virulence on pear fruits, and this defect can be partly explained by the impaired ability of A. westerdijkiae to penetrate host plants.

An improved pear disease classification approach using cycle generative adversarial network.

A large number of countries worldwide depend on the agriculture, as agriculture can assist in reducing poverty, raising the country's income, and improving the food security. However, the plan diseases usually affect food crops and hence play a significant role in the annual yield and economic losses in the agricultural sector. In general, plant diseases have historically been identified by humans using their eyes, where this approach is often inexact, time-consuming, and exhausting. Recently, the employment of machine learning and deep learning approaches have significantly improved the classification and recognition accuracy for several applications. Despite the CNN models offer high accuracy for plant disease detection and classification, however, the limited available data for training the CNN model affects seriously the classification accuracy. Therefore, in this paper, we designed a Cycle Generative Adversarial Network (CycleGAN) to overcome the limitations of over-fitting and the limited size of the available datasets. In addition, we developed an efficient plant disease classification approach, where we adopt the CycleGAN architecture in order to enhance the classification accuracy. The obtained results showed an average enhancement of 7% in the classification accuracy.

Identification of Mycoviruses in the Pathogens of Fragrant Pear Valsa Canker from Xinjiang in China.

As a common disease, canker seriously affects the yield and quality of fragrant pear due to the lack of effective control measures. Some fungi have been reported to harbor rich reservoirs of viral resources, and some mycoviruses can be used as biocontrol agents against plant diseases. In this study, 199 isolates were obtained from diseased branches of fragrant pear in the main production areas of Xinjiang. Among them, 134 belonged to Valsa spp., identified using morphological and molecular biological techniques, in which V. mali was the dominant species. The mycoviruses in Valsa spp. were further identified using metatranscriptomic sequencing and RT-PCR. The results revealed that a total of seven mycoviruses were identified, belonging to Botourmiaviridae, Endornaviridae, Fusariviridae, Hypoviridae, Mitoviridae, and Narnaviridae, among which Phomopsis longicolla hypovirus (PlHV) was dominant in all the sample collection regions. The Cryphonectria hypovirus 3-XJ1 (CHV3-XJ1), Botourmiaviridae sp.-XJ1 (BVsp-XJ1), and Fusariviridae sp.-XJ1 (Fvsp-XJ1) were new mycoviruses discovered within the Valsa spp. More importantly, compared with those in the virus-free Valsa spp. strain, the growth rate and virulence of the VN-5 strain co-infected with PlHV and CHV3-XJ1 were reduced by 59% and 75%, respectively, and the growth rate and virulence of the VN-34 strain infected with PlHV were reduced by 42% and 55%, respectively. On the other hand, the horizontal transmission efficiency of PlHV decreased when PlHV was co-infected with CHV3-XJ1, indicating that PlHV and CHV3-XJ1 were antagonistic. In summary, the mycoviruses in Valsa spp. were identified in Xinjiang for the first time, and three of them were newly discovered mycoviruses, with two strains yielding good results. These results will offer potential biocontrol resources for managing pear canker disease and provide a theoretical basis for the control of fruit tree Valsa canker disease.

The seed dispersal syndrome hypothesis in ungulate-dominated landscapes.

The Seed Dispersal Syndrome Hypothesis (SDSH) posits that fruit traits predict the main dispersers interacting with plant species. Mammalian dispersers, relying heavily on olfactory cues, are expected to select dull-colored, scented, and larger fruits compared to birds. However, challenges like overabundant seed predators and context-dependency of frugivore-plant interactions complicate SDSH expectations. We studied the Iberian pear, Pyrus bourgaeana, an expected mammal-dispersed tree based on its fruit traits. Extensive camera-trapping data (over 35,000 records) from several tree populations and years revealed visits from seven frugivore groups, with ungulate fruit predators (59-97%) and carnivore seed dispersers (1-20%) most frequent, while birds, lagomorphs, and rodents were infrequent (0-10%). Red deer and wild boar were also the main fruit removers in all sites and years but acted as fruit and seed predators, and thus likely exert conflicting selection pressures to those exerted by seed dispersers. Although, as predicted by the SDSH, most Iberian pear fruits were consumed by large and medium-sized mammals, the traits of Iberian pear fruits likely reflect selection pressures from dispersal vectors in past times. Our results do not challenge the SDHS but do reveal the importance of considering frugivore functional roles for its adequate evaluation.

Sucrose, cell wall, and polyamine metabolisms involve in preserving postharvest quality of 'Zaosu' pear fruit by L-glutamate treatment.

'Zaosu' pear fruit is prone to yellowing of the surface and softening of the flesh after harvest. This work was performed to assess the influences of L-glutamate treatment on the quality of 'Zaosu' pears and elucidate the underlying mechanisms involved. Results demonstrated that L-glutamate immersion reduced ethylene release, respiratory intensity, weight loss, brightness (L*), redness (a*), yellowness (b*), and total coloration difference (ΔE); enhanced ascorbic acid, soluble solids, and soluble sugar contents; maintained chlorophyll content and flesh firmness of pears. L-glutamate also restrained the activities of neutral invertase and acid invertase, while enhancing sucrose phosphate synthetase and sucrose synthase activities to facilitate sucrose accumulation. The transcriptions of PbSGR1, PbSGR2, PbCHL, PbPPH, PbRCCR, and PbNYC were suppressed by L-glutamate, resulting in a deceleration of chlorophyll degradation. L-glutamate concurrently suppressed the transcription levels and enzymatic activities of polygalacturonases, pectin methylesterases, cellulase, and ß-glucosidase. It restrained polygalacturonic acid trans-eliminase and pectin methyl-trans-eliminase activities as well as inhibited the transcription levels of PbPL and Pbß-gal. Moreover, the gene transcriptions and enzymatic activities of arginine decarboxylase, ornithine decarboxylase, S-adenosine methionine decarboxylase, glutamate decarboxylase, γ-aminobutyric acid transaminase, glutamine synthetase along with the PbSPDS transcription was promoted by L-glutamate. L-glutamate also resulted in the down-regulation of PbPAO, PbDAO, PbSSADH, PbGDH, and PbGOGAT transcription levels, while enhancing γ-aminobutyric acid, glutamate, and pyruvate acid contents in pears. These findings suggest that L-glutamate immersion can effectively maintain the storage quality of 'Zaosu' pears via modulating key enzyme activities and gene transcriptions involved in sucrose, chlorophyll, cell wall, and polyamine metabolism.

Multi-omics provide insights into the regulation of DNA methylation in pear fruit metabolism.

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, ß-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.

Genome-wide identification of GA2ox genes family and analysis of PbrGA2ox1-mediated enhanced chlorophyll accumulation by promoting chloroplast development in pear.

BACKGROUND: Chlorophyll (Chl) is an agronomic trait associated with photosynthesis and yield. Gibberellin 2-oxidases (GA2oxs) have previously been shown to be involved in Chl accumulation. However, whether and how the PbrGA2ox proteins (PbrGA2oxs) mediate Chl accumulation in pear (Pyrus spp.) is scarce. RESULTS: Here, we aimed to elucidate the role of the pear GA2ox gene family in Chl accumulation and the related underlying mechanisms. We isolated 13 PbrGA2ox genes (PbrGA2oxs) from the pear database and identified PbrGA2ox1 as a potential regulator of Chl accumulation. We found that transiently overexpressing PbrGA2ox1 in chlorotic pear leaves led to Chl accumulation, and PbrGA2ox1 silencing in normal pear leaves led to Chl degradation, as evident by the regreening and chlorosis phenomenon, respectively. Meanwhile, PbrGA2ox1-overexpressing (OE) tobacco plants discernably exhibited Chl built-up, as evidenced by significantly higher Pn and Fv/Fm. In addition, RNA sequencing (RNA-seq), physiological and biochemical investigations revealed an increase in abscisic acid (ABA), methyl jasmonate (MeJA), and salicylic acid (SA) concentrations and signaling pathways; a marked elevation in reducing and soluble sugar contents; and a marginal decline in the starch and sucrose levels in OE plants. Interestingly, PbrGA2ox1 overexpression did not prominently affect Chl synthesis. However, it indeed facilitated chloroplast development by increasing chloroplast number per cell and compacting the thylakoid granum stacks. These findings might jointly contribute to Chl accumulation in OE plants. CONCLUSION: Overall, our results suggested that GA2oxs accelerate Chl accumulation by stimulating chloroplast development and proved the potential of PbrGA2ox1 as a candidate gene for genetically breeding biofortified pear plants with a higher yield.

Introduction of a diverse genetic background of Pyrus into Malus through intergeneric hybridization.

Wide hybridizations across species and genera have been employed to enhance agriculturally important traits in crops. Within the tribe Maleae of the Rosaceae family, different genera and species exhibit several traits useful for increasing diversity and gene pool through hybridization. This study aimed to develop and characterize intergeneric hybrid individuals between Malus and Pyrus. Through seed germination, shoot multiplication, and rooting in vitro, acclimatized seedlings showing vegetative growth on their own roots were obtained from crosses of Malus × domestica pollinated by Pyrus communis, P. bretschneideri, and the Pyrus interspecific hybrid (P. communis × P. pyrifolia). Comparative analysis of leaf morphology, flow cytometry, and molecular genotyping confirmed the hybrid status of the individuals. Genome-wide genotyping revealed that all the hybrid individuals inherited genomic fragments symmetrically from the Malus and Pyrus parents. To the best of our knowledge, this is the first report on the development of intergeneric hybrid seedlings between Malus × domestica and P. bretschneideri. Furthermore, the Pyrus interspecific hybrid individual served as a bridge plant for introducing the genetic background of P. pyrifolia into Malus × domestica. The results of this study provided a crucial foundation for breeding through intergeneric hybridization between Malus and Pyrus, facilitating the incorporation of valuable traits from diverse gene pools.

A novel fluorescent probe with a large Stokes shift for colorimetric and selective detection of cysteine in water, milk, cucumber, pear and tomato.

Cysteine is an important amino acid that is related to human health and food safety. How to effectively detect Cys in food has received widespread attention. Compared with other methods, fluorescent probes have the advantages of simple operation, high sensitivity, and good selectivity. Therefore, a selective fluorescence probe 2 for Cys in food was designed and synthesized. Probe 2 employed the acrylate group as a thiol-recognition site for Cys, which endowed probe 2 with better selectivity for Cys over Hcy and GSH. The recognition pathway underwent Michael addition, intramolecular cyclization, and concomitant release of the piperideine-based fluorophore, along with a chromogenic change from yellow to orange. This pathway was supported by 1H NMR analysis and DFT calculations. In addition, probe 2 displays a linear response to Cys concentrations (0-30 µM), low detection limit (0.89 µM), and large Stokes shift (125 nm). Overall, probe 2 showed great application potential for the quantitative determination of Cys in water, milk, cucumber, pear and tomato.

Transcriptome and metabolome profiling reveal the effects of hormones on current-year shoot growth in Chinese 'Cuiguan' pear grafted onto vigorous rootstock 'Duli' and dwarf rootstock 'Quince A'.

BACKGROUND: Dwarf rootstocks have important practical significance for high-density planting in pear orchards. The shoots of 'Cuiguan' grafted onto the dwarf rootstock were shorter than those grafted onto the vigorous rootstock. However, the mechanism of shorter shoot formation is not clear. RESULTS: In this study, the current-year shoot transcriptomes and phytohormone contents of 'CG‒QA' ('Cuiguan' was grafted onto 'Quince A', and 'Hardy' was used as interstock) and 'CG‒DL' ('Cuiguan' was grafted onto 'Duli', and 'Hardy' was used as interstock) were compared. The transcriptome results showed that a total of 452 differentially expressed genes (DEGs) were identified, including 248 downregulated genes and 204 upregulated genes; the plant hormone signal transduction and zeatin biosynthesis pathways were significantly enriched in the top 20 KEGG enrichment terms. Abscisic acid (ABA) was the most abundant hormone in 'CG‒QA' and 'CG‒DL'; auxin and cytokinin (CTK) were the most diverse hormones; additionally, the contents of ABA, auxin, and CTK in 'CG‒DL' were higher than those in 'CG‒QA', while the fresh shoot of 'CG‒QA' accumulated more gibberellin (GA) and salicylic acid (SA). Metabolome and transcriptome co-analysis identified three key hormone-related DEGs, of which two (Aldehyde dehydrogenase gene ALDH3F1 and YUCCA2) were upregulated and one (Cytokinin oxidase/dehydrogenase gene CKX3) was downregulated. CONCLUSIONS: Based on the results of transcriptomic and metabolomic analysis, we found that auxin and CTK mainly regulated the shoot differences of 'CG-QA' and 'CG-DL', and other hormones such as ABA, GA, and SA synergistically regulated this process. Three hormone-related genes ALDH3F1, YUCCA2, and CKX3 were the key genes contributing to the difference in shoot growth between 'CG-QA' and 'CG-DL' pear. This research provides new insight into the molecular mechanism underlying shoot shortening after grafted onto dwarf rootstocks.

Comprehensive Evolutionary Analysis of the SMXL Gene Family in Rosaceae: Further Insights into Its Origin, Expansion, Diversification, and Role in Regulating Pear Branching.

SMXL genes constitute a conserved gene family that is ubiquitous in angiosperms and involved in regulating various plant processes, including branching, leaf elongation, and anthocyanin biosynthesis, but little is known about their molecular functions in pear branching. Here, we performed genome-wide identification and investigation of the SMXL genes in 16 angiosperms and analyzed their phylogenetics, structural features, conserved motifs, and expression patterns. In total, 121 SMXLs genes were identified and were classified into four groups. The number of non-redundant SMXL genes in each species varied from 3 (Amborella trichopoda Baill.) to 18 (Glycine max Merr.) and revealed clear gene expansion events over evolutionary history. All the SMXL genes showed conserved structures, containing no more than two introns. Three-dimensional protein structure prediction revealed distinct structures between but similar structures within groups. A quantitative real-time PCR analysis revealed different expressions of 10 SMXL genes from pear branching induced by fruit-thinning treatment. Overall, our study provides a comprehensive investigation of SMXL genes in the Rosaceae family, especially pear. The results offer a reference for understanding the evolutionary history of SMXL genes and provide excellent candidates for studying fruit tree branching regulation, and in facilitating pear pruning and planting strategies.

Protein/Lipid ratio of pollen biases the visitation of bumblebees (Bombus ignitus Smith) to male-fertile cultivars of the Japanese pear (Pyrus pyrifolia Nakai).

Bees have been known to visit the male-fertile cultivars of self-incompatible flowering plants more frequently than the male-sterile cultivars, but the origin of this preference is poorly understood. Here, we demonstrate that this preference is driven by the higher protein/lipid ratio of male-fertile pollen compared with male-sterile pollen by way of two caged-behavioral assays with six cultivars. In the first assay, flower-naïve bumblebees (Bombus ignitus Smith) showed a significantly higher flower-visitation rate to male-fertile cultivars (pollen germination rate > 55%; > 14 visits/10 min) of the Japanese pear (Pyrus pyrifolia Nakai) than male-sterile cultivars (pollen germination rate ≤ 20%; > 6 visits/10 min). In the second, bees still preferred the anthers of male-fertile cultivars (5-9 visits/10 min) more than those of male-sterile ones (less than 1 visit in 10 min) even in the absence of all other organs (i.e., petals, pistil, nectar), indicating that pollen is responsible for the preference. We then analyzed the macronutrient content of the pollen and its visual cues, and found that the bee preference was highly correlated with the protein/lipid ratio (0.3-1.6) but not color variables such as (a)chromatic contrast, intensity, and spectral purity. We conclude that the protein/lipid ratio influences the foraging behavior of the bumblebees likely by serving as (1) a chemotactile cue while antennating, (2) a gustatory cue after intake, and (3) an olfactory cue. In addition, the low bee visitation rate to poorly viable pollen could be due to its low protein/lipid ratio.

Identification and expression analysis of ATP-binding cassette (ABC) transporters revealed its role in regulating stress response in pear (Pyrus bretchneideri).

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.

Physicochemical Properties of Dried and Powdered Pear Pomace.

Pear pomace, a byproduct of juice production, represents a valuable reservoir of bioactive compounds with potential health benefits for humans. This study aimed to evaluate the influence of drying method and temperature on pear pomace, specifically focusing on the drying kinetics, grinding characteristics, color, phenolic profile (LC-MS/MS), and antioxidant activities of the powder. Drying using the contact method at 40 °C with microwave assistance demonstrated the shortest duration, whereas freeze-drying was briefer compared to contact-drying without microwave assistance. Freeze-drying resulted in brighter and more easily comminuted pomace. Lyophilized samples also exhibited higher total phenolic compound levels compared to contact-dried ones, correlating with enhanced antioxidant activity. Twenty-one phenolic compounds were identified, with dominant acids being quinic, chlorogenic, and protocatechuic. Flavonoids, primarily isoquercitrin, and rutin, were also presented. Pear pomace dried via contact at 60 °C contained more quinic and protocatechuic acids, while freeze-dried pomace at the same temperature exhibited higher levels of chlorogenic acid, epicatechin, and catechin. The content of certain phenolic components, such as gallic acid and epicatechin, also varied depending on the applied drying temperature.

Genomic characterization of Pseudomonas syringae pv. syringae from Callery pear and the efficiency of associated phages in disease protection.

The Pseudomonas syringae species complex is a heterogeneous group of plant pathogenic bacteria associated with a wide distribution of plant species. Advances in genomics are revealing the complex evolutionary history of this species complex and the wide array of genetic adaptations underpinning their diverse lifestyles. Here, we genomically characterize two P. syringae isolates collected from diseased Callery pears (Pyrus calleryana) in Berkeley, California in 2019 and 2022. We also isolated a lytic bacteriophage, which we characterized and evaluated for biocontrol efficiency. Using a multilocus sequence analysis and core genome alignment, we classified the P. syringae isolates as members of phylogroup 2, related to other strains previously isolated from Pyrus and Prunus. An analysis of effector proteins demonstrated an evolutionary conservation of effectoromes across isolates classified in PG2 and yet uncovered unique effector profiles for each, including the two newly identified isolates. Whole-genome sequencing of the associated phage uncovered a novel phage genus related to Pseudomonas syringae pv. actinidiae phage PHB09 and the Flaumdravirus genus. Finally, using in planta infection assays, we demonstrate that the phage was equally useful in symptom mitigation of immature pear fruit regardless of the Pss strain tested. Overall, this study demonstrates the diversity of P. syringae and their viruses associated with ornamental pear trees, posing spill-over risks to commercial pear trees and the possibility of using phages as biocontrol agents to reduce the impact of disease.IMPORTANCEGlobal change exacerbates the spread and impact of pathogens, especially in agricultural settings. There is a clear need to better monitor the spread and diversity of plant pathogens, including in potential spillover hosts, and for the development of novel and sustainable control strategies. In this study, we characterize the first described strains of Pseudomonas syringae pv. syringae isolated from Callery pear in Berkeley, California from diseased tissues in an urban environment. We show that these strains have divergent virulence profiles from previously described strains and that they can cause disease in commercial pears. Additionally, we describe a novel bacteriophage that is associated with these strains and explore its potential to act as a biocontrol agent. Together, the data presented here demonstrate that ornamental pear trees harbor novel P. syringae pv. syringae isolates that potentially pose a risk to local fruit production, or vice versa-but also provide us with novel associated phages, effective in disease mitigation.

PbWRKY18 promotes resistance against black spot disease by activation of the chalcone synthase gene PbCHS3 in pear.

Flavonoids are plant pigments that play a major role in plant defense and have significant health benefits to humans. Chalcone synthase (CHS) is an important enzyme in flavonoid biosynthesis and investigation transcription factors (TFs) regulating its expression and downstream targets is critical to understanding its mechanism. Here, a novel TF, PbWRKY18, was isolated from the pear Pyrus betulaefolia. Its expression was evaluated in various tissues by RT-PCR, particularly in response to Alternaria alternata, the pathogen responsible for black spot disease, and exogenous hormone administration. The PbWRKY18 protein was primarily found in the nucleus where it regulated transcriptional activity. Yeast one-hybrid and dual-luciferase reporter assays showed a strong association between PbWRKY18 and the PbCHS3 promoter, which drives PbCHS3 expression. It was also found that PbCHS3 was critical for the development of resistance against black spot disease. In addition, PbWRKY18 was found to significantly increase the expression of PbCHS3 and salicylic acid-related genes, as well as defense enzyme activity and tolerance to black spot disease. PbWRKY18 or PbCHS3 knockdown in pear attenuates resistance to Alternaria alternata. In summary, the study identified a novel WRKY18-CHS3 axis involved in resistance against black spot disease in pear.

Improved Dosimetry with Daily Online Adaptive Radiotherapy for Cervical Cancer: Waltzing the Pear.

AIMS: Standard of care radiotherapy for locally advanced cervical cancer includes large margins to ensure the uterocervix remains within the treatment fields over the course of treatment. Daily online cone-beam adaptive radiotherapy corrects for interfractional changes by adjusting the plan to match the target position during each treatment session, thus allowing for significantly reduced clinical target volume (CTV) to planning target volume (PTV) margins. We hypothesise that reduced margins from daily online adaptive radiotherapy will reduce organ at risk dose without compromising target coverage. MATERIALS AND METHODS: Ten patients with cervical cancer (stage IIB-IIIC2) were treated with definitive chemoradiation using daily online cone-beam adaptive radiotherapy in 25-27 fractions. Initial and all adapted treatment plans were generated with CTV to PTV margins versus standard of care image-guided radiotherapy (IGRT) plans as follows: cervix/uterus/gross tumour volume (0.5 versus 1.5 cm), parametria/vagina (0.5 versus 1.0 cm) and nodal chains and gross nodes (0.5 versus 0.5 cm). IGRT plans were created and copied to synthetic computed tomography scans and contours generated from each daily adapted fraction. The dosimetry of each clinically treated online adapted fraction was compared with emulated IGRT plans. Statistical significance was defined as P < 0.05. RESULTS: Daily online cone-beam adaptive radiotherapy significantly improves bowel bag dosimetry compared with IGRT, with a reduction in V40 by an average of 91.3 cm3 [V40 (-6.2%) and V45 (-6.1%)]. The daily adapted plans showed significant improvements in bladder and rectum [V40 (-25.2% and -36.0%) and V30 (-9.7% and -17.1%), respectively]. Additionally, bone marrow had a significantly reduced dose [V10 (-2.7%) and V20 (-3.3%)]. Daily online cone-beam adaptive radiotherapy improved uterocervix CTV coverage and reduced hotspots compared with IGRT [D95% (+1.6%) and Dmax (-0.9%)]. CONCLUSIONS: Reduced CTV to PTV margins achievable with daily online adaptive radiotherapy improves organ at risk dosimetry and target coverage when compared with standard of care IGRT for locally advanced cervical cancer. The clinical impact of improved dosimetry is currently undergoing investigation.

PbRbohH/J mediates ROS generation to regulate the growth of pollen tube in pear.

Respiratory burst oxidase homolog (Rboh) family genes play crucial functions in development and growth. However, comprehensive and systematic investigation of Rboh family members in Rosaceae and their specific functions during pear pollen development are still limited. In the study, 63 Rboh genes were identified from eight Rosaceae genomes (Malus domestica, Pyrus bretschneideri, Pyrus communis, Prunus persica, Rubus occidentalis, Fragaria vesca, Prunus mume and Prunus avium) and divided into seven main subfamilies (I-VII) according to phylogenetic and structural features. Different modes of gene duplication led to the expansion of Rboh family, with purifying selection playing a vital role in the evolution of Rboh genes. In addition, RNA sequencing and qRT-PCR results indicated that PbRbohH and PbRbohJ were specifically high-expressed in pear pollen. Subsequently, subcellular localization revealed that PbRbohH/J distributed at the plasma membrane. Furthermore, by pharmacological analysis and antisense oligodeoxynucleotide assay, PbRbohH/J were demonstrated to mediate the formation of reactive oxygen species (ROS) to manage pollen tube growth. In conclusion, our results provide useful insights into the functions, expression patterns, evolutionary history of the Rboh genes in pear and other Rosaceae species.