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This study describes the first genome sequence and analysis of Coniella granati, a fungal pathogen with a broad host range, which is responsible for postharvest crown rot, shoot blight, and canker diseases in pomegranates. C. granati is a geographically widespread pathogen which has been reported across Europe, Asia, the Americas, and Africa. Our analysis revealed a 46.8 Mb genome with features characteristic of hemibiotrophic fungi. Approximately one third of its genome was compartmentalised within ‘AT-rich’ regions exhibiting a low GC content (30 to 45%). These regions primarily comprised transposable elements that are repeated at a high frequency and interspersed throughout the genome. Transcriptome-supported gene annotation of the C. granati genome revealed a streamlined proteome, mirroring similar observations in other pathogens with a latent phase. The genome encoded a relatively compact set of 9568 protein-coding genes with a remarkable 95% having assigned functional annotations. Despite this streamlined nature, a set of 40 cysteine-rich candidate secreted effector-like proteins (CSEPs) was predicted as well as a gene cluster involved in the synthesis of a pomegranate-associated toxin. These potential virulence factors were predominantly located near repeat-rich and AT-rich regions, suggesting that the pathogen evades host defences through Repeat-Induced Point mutation (RIP)-mediated pseudogenisation. Furthermore, 23 of these CSEPs exhibited homology to known effector and pathogenicity genes found in other hemibiotrophic pathogens. The study establishes a foundational resource for the study of the genetic makeup of C. granati, paving the way for future research on its pathogenicity mechanisms and the development of targeted control strategies to safeguard pomegranate production.

Pomegranate cultivation faces significant challenges due to fruit diseases that significantly impact crop yield and farmer income. Traditional methods for disease detection are often slow and prone to errors, delaying timely intervention. This paper proposes a deep learning-based system for automatic, multi-class disease classification in pomegranates using transfer learning. A dataset comprising 5099 annotated images was used to train and evaluate several CNN models, including DenseNet121, EfficientNetB0V2, MobileNetV2, ResNet50, VGG16, and InceptionV3. DenseNet121 emerged as the top performer, achieving an accuracy of 99.35%. To enhance practical value, a novel Healthy-Based Deviation Scoring (HBDS) method was developed to estimate disease severity using Grad-CAM ++ for lesion localization and Mahalanobis distance-based scoring, followed by Gaussian Mixture Model clustering. The severity predictions of the system were verified against manually labeled images, and the system has shown superior accuracy compared to pixel-based methods. Also, a recommendation module was integrated using a retrieval-augmented language model, which provides disease-specific treatment suggestions based on the predicted severity. The complete pipeline is implemented as a user-friendly web application that delivers real-time diagnosis, severity estimation, and actionable treatment plans, which offer a practical and scalable solution for modern precision agriculture.

Bacterial blight, caused by Xanthomonas citri pv. punicae (Xcp) , severely impacts the global pomegranate industry, with alarming yield losses up to 80%. This pathogen becomes a glitch in the pomegranate industry, leading to reduced commercial pockets globally, especially in Pakistan. The objective of this study was to conduct biochemical characterisation and in vitro evaluation of copper compounds and antibiotics against Xcp , for optimal fruit quality and yield. The pathogen was isolated from infected tissues of the pomegranate, and various biochemical tests were performed to identify the bacteria. Pathogenicity was confirmed using the pin-prick and infiltration method on cv. Golden. The in vitro efficacy of antibiotics and copper compounds was assessed using the disk diffusion method against Xcp. The pathogen was aseptically cultured via the streak plate method, and disks impregnated with different chemical concentrations were placed on solidified media. A control disk was treated with sterile water containing 1% v/v surfactant. Plates were incubated at 28 ± 2 °C for 48 and 72 h to observe bacterial growth inhibition. Biochemical analysis showed positive results for the oxidase test, catalase test, KOH solubility test, gelatin liquefaction, and starch hydrolysis test, while Gram staining was negative. Antibacterial efficacy was determined by measuring inhibition zones after 48 and 72 h of incubation. The highest inhibition zones, measuring 29.55 mm and 37.73 mm at 48 and 72 h, respectively, were recorded with 550 µg/ml oxytetracycline hydrochloride, followed by streptomycin sulfate (550 µg/ml) with zones of 29.12 mm and 32.42 mm. In contrast, copper-based compounds, particularly copper sulfate (2000 µg/ml), exhibited the lowest inhibition, with zones of 8.85 mm and 8.86 mm at the respective time intervals. A comparative analysis demonstrated that antibiotics showed outstanding and prodigious results compared to copper compounds. The results of this experiment demonstrate the effectiveness of oxytetracycline hydrochloride and streptomycin sulfate in inhibiting the radial growth of Xcp under controlled conditions. Furthermore, changes in dosage/concentration, time of application, and field application method may help researchers with field-oriented results in the future.

Pilidiella granati, also known as Coniella granati, is the etiological agent of pomegranate fruit dry rot. This fungal pathogen is also well-known as responsible for both plant collar rot and leaf spot. Because of its aggressiveness and the worldwide diffusion of pomegranate crops, the selection of cultivars less susceptible to this pathogen might represent an interesting preventive control measure. In the present investigation, the role of polyphenols in the susceptibility to P. granati of the two royalties-free pomegranate cultivars Wonderful and Mollar de Elche was compared. Pomegranate fruit were artificially inoculated and lesion diameters were monitored. Furthermore, pathogen DNA was quantified at 12–72 h post-inoculation within fruit rind by a real time PCR assay setup herein, and host total RNA was used in expression assays of genes involved in host-pathogen interaction. Similarly, protein extracts were employed to assess the specific activity of enzymes implicated in defense mechanisms. Pomegranate phenolic compounds were evaluated by HPLC-ESI-MS and MS2. All these data highlighted ‘Wonderful’ as less susceptible to P. granati than ‘Mollar de Elche’. In the first cultivar, the fungal growth seemed controlled by the activation of the phenylpropanoid pathway, the production of ROS, and the alteration of fungal cell wall. Furthermore, antifungal compounds seemed to accumulate in ‘Wonderful’ fruit following inoculation. These data suggest that pomegranate polyphenols have a protective effect against P. granati infection and their content might represent a relevant parameter in the selection of the most suitable cultivars to reduce the economic losses caused by this pathogen.

The pomegranate ( Punica granatum L.) is highly valued for its significant health benefits, adaptability, cold resistance, and ability to thrive in low-fertility soils. Iran, a major producer, contributes nearly one-fifth of global pomegranate production, with Fars Province being the leading region. Recently, there has been a notable increase in pomegranate diseases, including decline, dieback, and canker. This investigation aimed to identify the fungal pathogens responsible for these diseases in Fars Province. Morphological and molecular analyses identified several fungal isolates belonging to Paramicrosphaeropsis eriobotryae , an emerging canker-associated ascomycete pathogen. The most aggressive isolate was identified through inoculation on detached branches of the ‘Rabab-e-Neyriz’ cultivar and subsequently used in pathogenicity assessments on five commercially important cultivars. The ‘Malas-e-Saveh’ and ‘Wonderful’ cultivars exhibited higher susceptibility, while ‘Malas-e-Danesiyah-e-Esfahani’ cultivar demonstrated relative resistance. Cross-pathogenicity assessments with isolates from pomegranate and other natural hosts, including fig, loquat, almond, and olive indicated diverse levels of host susceptibility and isolate aggressiveness. This study represents the first global report of P. eriobotryae causing branch canker and dieback on pomegranate trees.

Bacterial blight caused by Xanthomonas axonopodis pv. punicae is a major disease of pomegranate. Bacterial blight drastically reduces the yield and quality of fruits, which are critical for pomegranate production. Precise and early diagnosis of bacterial blight is crucial for active surveillance and effective management of the disease. Symptoms based disease diagnostic methods are labor-intensive, time-consuming and may not detect disease on asymptomatic plants. DNA-based disease diagnostics using polymerase chain reaction (PCR) are reliable, precise, accurate and quick. PCR coupled with agarose gel electrophoresis (PCR-AGE), PCR coupled with capillary electrophoresis (PCR-CE) and real-time PCR (qPCR) were applied for the early and accurate diagnosis of bacterial blight in pomegranate. PCR-CE and qPCR were capable of diagnosing bacterial blight 6 to 10 days before symptom appearance, with detection limits of 100 fg and 10 fg of bacterial DNA respectively. However, conventional PCR-AGE detected pathogen at the onset of disease symptoms with a detection limit of 10 pg of bacterial DNA. qPCR detected bacterial blight in orchards that did not show any disease symptoms. Our data demonstrate that qPCR is more sensitive than other PCR methods along with being reliable for early diagnosis.

Pomegranate (Punica granatum L.) belongs to the Punicaceae plant family. It is an important fruit due to its nutritional and medicinal properties. Pomegranates are widely distributed around the world and, therefore, have a broad genetic diversity, resulting in differences in their phytochemical composition. The scientific community has focused on the positive health effects of pomegranate as a whole, but the different varieties have rarely been compared according to their bioactive compounds and bioactivity. This review aims to provide a holistic overview of the current knowledge on the bioactivity of pomegranate trees, with an emphasis on differentiating both the varieties and the different plant parts. This review intends to provide a general and organized overview of the accumulated knowledge on pomegranates, the identification of the most bioactive varieties, their potential consumption pathways and seeks to provide knowledge on the present gaps to guide future research.

Background The pomegranate is cultivated across a wide range of geographical regions worldwide for ornamental purposes, fruit aril for the food industry, and juice production. The pomegranate genome is reported to consist of 2n = 16 chromosomes in some cultivars and 2n = 18 in others. Clustering assembled contigs using Hi-C technology resulted in eight pseudochromosomes, alternatively, anchoring these contigs onto a genetic map produced nine pseudochromosomes. The origin of the domesticated pomegranate is unknown. Previous studies have suggested that the variant Daru, which is grown in the region of India and Nepal, is the origin of the wild stock. Results Here we explored the phylogeny, genetic structure, and substitution pattern of P. granatum . Phylogenetic analysis of Punica granatum accessions revealed that Daru seedlings constitute a distinct clade, which is a sister to the clade of Asian accessions, rather than an outgroup. No evidence of introgression was observed between Daru and cultivated accessions. Analysis of the non-synonymous to synonymous substitutions ratio in genic regions outlines a similar profile of the Daru and the domesticated pomegranates with overall positive selection, while the wild type P. protopunica illustrated a different profile with positive selection only in some regions. Domesticated pomegranate variants show low heterozygosity and nucleotide diversity, particularly on chromosome 1. Conclusions Our findings suggest that differences in chromosome numbers may be associated with the formation of chromosome quadrivalents and their impact on the arrangement of contigs into pseudochromosomes. The profile of Kn to Ks ratio in domesticated pomegranates likely reflects positive selection. Genetic profiling of Daru, domesticated P. granatum , and wild-type P. protopunica revealed that Daru more closely resembles domesticated plants, challenging the notion that it represents the wild pomegranate.

Summary Pomegranate is an important perennial fruit tree distributed worldwide. Reference genomes with gaps and limit gene identification controlling important agronomic traits hinder its functional genomics and genetic improvements. Here, we reported a telomere‐to‐telomere (T2T) gap‐free genome assembly of the distinctive cultivar ‘Moshiliu’. The Moshiliu reference genome was assembled into eight chromosomes without gaps, totalling ~366.71 Mb, with 32 158 predicted protein‐coding genes. All 16 telomeres and eight centromeres were characterized; combined with FISH analysis, we revealed the atypical telomere units in pomegranate as TTTTAGGG. Furthermore, a total of 16 loci associated with 15 important agronomic traits were identified based on GWAS of 146 accessions. Gene editing and biochemical experiments demonstrated that a 37.2‐Kb unique chromosome translocation disrupting the coding domain sequence of PgANS was responsible for anthocyanin‐less, knockout of PgANS in pomegranate exhibited a defect in anthocyanin production; a unique repeat expansion in the promoter of PgANR may affected its expression, resulting in black peel; notably, the G → A transversion located at the 166‐bp coding domain of PgNST3, which caused a E56K mutation in the PgNST3 protein, closely linked with soft‐seed trait. Overexpression of PgNST3A in tomato presented smaller and softer seed coats. The E56K mutation in PgNST3 protein, eliminated the binding ability of PgNST3 to the PgMYB46 promoter, which subsequently affected the thickness of the inner seed coat of soft‐seeded pomegranates. Collectively, the validated gap‐free genome, the identified genes controlling important traits and the CRISPR‐Cas9‐mediated gene knockout system all provided invaluable resources for pomegranate precise breeding.

The combination of plant extract and antibiotic represents a template for developing of antibiofilm drugs. This study investigated the synergistic effects of pomegranate/rosemary/antibiotic combinations against antibiotic resistance and biofilm formation of Pseudomonas aeruginosa . The results showed that 17 (85%) of total P. aeruginosa isolates were biofilm producers; however, 5 (25%) isolates were demonstrated as a strong biofilm producer. The highest MIC level (1024 μg/ml) of tested antibiotics against strong biofilm producer isolates was observed with piperacillin, however the MIC ranges of ceftazidime, gentamycin, imipenem, and levofloxacin against these isolates were reached to (256–1024 μg/ml), (32–1024 μg/ml), (8–1024 μg/ml), and (8–512 μg/ml), respectively. PS-1 was the representative isolate for strong biofilm formation and high antibiotic resistance. 16S rRNA gene analysis suggested that PS-1 (accession No. MN619678) was identified as a strain of P. aeruginosa POA1. Pomegranate and rosemary extracts were the most effective extracts in biofilm inhibition, which significantly inhibited 91.93 and 90.83% of PS-1 biofilm, respectively. Notably, the synergism between both plant extracts and antibiotics has significantly reduced the MICs of used antibiotics at the level lower than the susceptibility breakpoints. Pomegranate/rosemary/antibiotic combinations achieved the highest biofilm eradication, which ranging from 90.0 to 99.6%, followed by the eradication ranges of pomegranate/rosemary combination, rosemary, and pomegranate extracts, which reached to (76.5–85.4%), (53.1–73.7%), and (41.2–71.5%), respectively. The findings suggest that pomegranate/rosemary/antibiotic combinations may be an effective therapeutic agent for antibiotic resistance and biofilm formation of P. aeruginosa.