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Given the severe impact of Citrus Huanglongbing on orchard production, accurate detection of the disease is crucial in orchard management. In the natural environments, due to factors such as varying light intensities, mutual occlusion of citrus leaves, the extremely small size of Huanglongbing leaves, and the high similarity between Huanglongbing and other citrus diseases, there remains an issue of low detection accuracy when using existing mainstream object detection models for the detection of citrus Huanglongbing. To address this issue, we propose YOLO-EAF (You Only Look Once–Efficient Asymptotic Fusion), an improved model based on YOLOv8n. Firstly, the Efficient Multi-Scale Attention Module with cross-spatial learning (EMA) is integrated into the backbone feature extraction network to enhance the feature extraction and integration capabilities of the model. Secondly, the adaptive spatial feature fusion (ASFF) module is used to enhance the feature fusion ability of different levels of the model so as to improve the generalization ability of the model. Finally, the focal and efficient intersection over union (Focal–EIOU) is utilized as the loss function, which accelerates the convergence process of the model and improves the regression precision and robustness of the model. In order to verify the performance of the YOLO-EAF method, we tested it on the self-built citrus Huanglongbing image dataset. The experimental results showed that YOLO-EAF achieved an 8.4% higher precision than YOLOv8n on the self-built dataset, reaching 82.7%. The F1-score increased by 3.33% to 77.83%, and the mAP (0.5) increased by 3.3% to 84.7%. Through experimental comparisons, the YOLO-EAF model proposed in this paper offers a new technical route for the monitoring and management of Huanglongbing in smart orange orchards. [ABSTRACT FROM AUTHOR]

The citrus industry is of great importance in Mexico, with an estimated production value of USD 2.4 billion and the potential to generate up to 28 million employees per year. Approximately 69,000 Mexican families depend on this sector. However, it is under serious threat from the disease known as Huanglongbing (HLB). The rapid spread of this disease has caused significant economic losses, impacted the livelihoods of citrus growers, and led to substantial job losses. Currently, HLB is not under control in Mexico, and as the management strategies applied have proven to be ineffective, yields have not been recovered to pre-HLB levels, and production costs have significantly increased. Therefore, it remains the most destructive citrus disease. This review paper describes the current situation of the disease, its economic and social impact, and the measures adopted for its management in Mexico. Future perspectives highlight advances in research based on new biotechnological tools as an eco-friendly management alternative. Practical suggestions to address HLB are also mentioned in our analysis. [ABSTRACT FROM AUTHOR]

Background: With the rapid development of single-cell sequencing technology, histological studies are no longer limited to conventional homogenized tissues. Laser microdissection enables the accurate isolation of specific tissues or cells, and when combined with next-generation sequencing, it can reveal important biological processes at the cellular level. However, traditional laser microdissection techniques have often been complicated and time-consuming, and the quality of the RNA extracted from the collected samples has been inconsistent, limiting follow-up studies. Therefore, an improved, simple, and efficient laser microdissection method is urgently needed. Results: We omitted the sample fixation and cryoprotectant addition steps. Instead, fresh samples were embedded in Optimal Cutting Temperature medium within 1.5 ml centrifuge tube caps, rapidly frozen with liquid nitrogen, and immediately subjected to cryosectioning. A series of section thicknesses of citrus rind were tested for RNA extraction, which showed that 18 μm thickness yielded the highest quality RNA. By shortening the dehydration time to one minute per ethanol gradient and omitting the tissue clearing step, the resulting efficient dehydration and preserved morphology ensured high-quality RNA extraction. We also propose a set of laser microdissection parameters by adjusting the laser power to optimal values, reducing the aperture size, and lowering the pulse frequency. Both the epidermal and subepidermal cells from the citrus rind were collected, and RNA extraction was completed within nine hours. Using this efficient method, the transcriptome sequencing of the isolated tissues generated high-quality data with average Q30 values and mapping rates exceeding 91%. Moreover, the transcriptome analysis revealed significant differences between the cell layers, further confirming the effectiveness of our isolation approach. Conclusions: We developed a simple and rapid laser microdissection method and demonstrated its effectiveness through a study based on citrus rind, from which we generated high-quality transcriptomic data. This fast and efficient method of cell isolation, combined with transcriptome sequencing not only contributes to precise histological studies at the cellular level in citrus but also provides a promising approach for cell-specific transcriptome analysis in a broader range of other plant tissues. [ABSTRACT FROM AUTHOR]

Brown spot of citrus caused by Alternaria citri is one of the emerging threats to the successful production of citrus crops. The present study, conducted with a substantial sample size of 50 leaf samples for statistical reliability, aimed to determine the change in mineral content in citrus leaves after brown spot disease attack. Leaf samples from a diverse range of susceptible citrus varieties (Valentia late, Washington navel, and Kinnow) and resistant varieties (Citron, Eruka lemon, and Mayer lemon) were analyzed. Significant variations (p ≤ 0.05) in mineral contents were observed across reaction groups (inoculated and un-inoculated), types (resistant and susceptible), and varieties of citrus in response to infection of Alternaria citri. The analysis of variance showed significant changes in mineral levels of citrus leaves, including nitrogen (N), phosphorus (P), potassium (K), calcium (Ca), magnesium (Mg), zinc (Zn), sodium (Na), iron (Fe), and copper (Cu). The results indicate that the concentration of N and P differed by 6.63% and 1.44%, respectively, in resistant plants, while susceptible plants showed a difference of 6.07% and 1.19%. Moreover, resistant plants showed a higher concentrations of K, Ca, Mg, Zn, Na, Fe, and Cu at 8.40, 2.1, 1.83, 2.21, 1.58, 2.89, and 0.36 ppm respectively, compared to susceptible plants which showed concentrations of 5.99, 1.93, 1.47, 1.09, 1.24, 1.81, and 0.31 ppm respectively. Amounts of mineral contents were reduced in both resistant as well as susceptible plants of citrus after inoculation. Amount of N (8.56), P (1.87) % while K (10.74), Ca (2.71), Mg (2.62), Zn (2.20), Na (2.08), Fe (3.57) and Cu (0.20) ppm were recorded in un-inoculated group of citrus plants that reduced to 3.15 and 0.76% and 3.66, 1.40, 0.63,0.42, 0.74, 1.13 and 0.13 ppm in inoculated group respectively. It was accomplished that susceptible varieties contained lower ionic contents than resistant varieties. The higher concentrations of ionic contents in resistant citrus varieties build up the biochemical and physiological processes of the citrus plant, which help to restrict spread of pathogens. Further research could explore the interplay between mineral nutrition and disease resistance in citrus, potentially leading to the development of new disease-resistant varieties. [ABSTRACT FROM AUTHOR]

Bacterial canker disease caused by Clavibacter michiganensis is a substantial threat to the cultivation of tomatoes, leading to considerable economic losses and global food insecurity. Infection is characterized by white raised lesions on leaves, stem, and fruits with yellow to tan patches between veins, and marginal necrosis. Several agrochemical substances have been reported in previous studies to manage this disease but these were not ecofriendly. Thus present study was designed to control the bacterial canker disease in tomato using green fabricated silver nanoparticles (AgNps). Nanosilver particles (AgNPs) were synthesized utilizing Moringa oleifera leaf extract as a reducing and stabilizing agent. Synthesized AgNPs were characterized using UV–visible spectroscopy, scanning electron microscopy (SEM), X-ray diffraction (XRD), energy-dispersive X-ray (EDX), and Fourier transform infrared spectrometry (FTIR). FTIR showed presence of bioactive compounds in green fabricated AgNPs and UV-visible spectroscopy confirmed the surface plasmon resonance (SPR) band in the range of 350 nm to 355 nm. SEM showed the rectangular segments fused together, and XRD confirmed the crystalline nature of the synthesized AgNPs. The presence of metallic silver ions was confirmed by an EDX detector. Different concentrations (10, 20, 30, and 40 ppm) of the green fabricated AgNPs were exogenously applied on tomato before applying an inoculum of Clavibacter michigensis to record the bacterial canker disease incidence at different day intervals. The optimal concentration of AgNPs was found to be 30 µg/mg that exhibited the most favorable impact on morphological (shoot length, root length, plant fresh and dry weights, root fresh and dry weights) and physiological parameters (chlorophyll contents, membrane stability index, and relative water content) as well as biochemical parameters (proline, total soluble sugar and catalase activity). These findings indicated a noteworthy reduction in biotic stress through the increase of both enzymatic and non-enzymatic activities by the green fabricated AgNPs. This study marks a first biocompatible approach in assessing the potential of green fabricated AgNPs in enhancing the well-being of tomato plants that affected with bacterial canker and establishing an effective management strategy against Clavibacter michiganensis. This is the first study suggests that low concentration of green fabricated nanosilvers (AgNPs) from leaf extract of Moringa oleifera against Clavibacter michiganensis is a promisingly efficient and eco-friendly alternative approach for management of bacterial canker disease in tomato crop. [ABSTRACT FROM AUTHOR]

Introduction: The white-spotted flower chafer (Protaetia brevitarsis seulensis), which is widely distributed in Asian countries, is traditionally used in oriental medicine. However, its larvae are prone to severe damage by green muscardine disease (caused by Metarhizium anisopliae) during breeding. The aim of this study was to characterize Bacillus velezensis TJS119, which has been isolated from freshwater, and investigate its potential as a biocontrol agent against M. anisopliae in insects. Methods: TJS119 was obtained from freshwater samples in the Republic of Korea and was classified as B. velezensis. We evaluated its in vitro antifungal effect, sequenced the bacterial whole genome, mined genes responsible for the synthesis of secondary metabolites, performed secondary metabolite analysis Ultra performance liquid chromatography-mass spectrometry (UPLC-MS/MS), and conducted bioassays for determining green muscardine disease control ability. Results: Bacillus velezensis TJS119 inhibited the mycelial growth of M. anisopliae in vitro. The size of the B. velezensis TJS119 genome was estimated to be 3,890,913 bp with a GC content of 46.67% and 3,750 coding sequences. Biosynthetic gene clusters for secondary metabolites with antifungal activity were identified in the genome. Lipopeptides, including fengycin secreted by TJS119 exhibit antifungal activity. Application of TJS119 for the biocontrol against green muscardine disease increased the viability of white-spotted flower chafer by 94.7% compared to the control. Discussion: These results indicate that B. velezensis TJS119 is a potential biocontrol agent for insect pathogens. [ABSTRACT FROM AUTHOR]

Background: The finger lime (Citrus australasica), one of six Australian endemic citrus species shows a high natural phenotypic diversity and novel characteristics. The wide variation and unique horticultural features have made this lime an attractive candidate for domestication. Currently no haplotype resolved genome is available for this species. Here we present a high quality, haplotype-resolved reference genome for this species using PacBio HiFi and Hi-C sequencing. Results: Hifiasm assembly and SALSA scaffolding resulted in a collapsed genome size of 344.2 Mb and 321.1 Mb and 323.2 Mb size for the two haplotypes. The nine pseudochromosomes of the collapsed genome had an N50 of 35.2 Mb, 99.1% genome assembly completeness and 98.9% gene annotation completeness (BUSCO). A total of 41,304 genes were predicted in the nuclear genome. Comparison with C. australis revealed that 13,661 genes in pseudochromosomes were unique in C. australasica. These were mainly involved in plant-pathogen interactions, stress response, cellular metabolic and developmental processes, and signal transduction. The two genomes showed a syntenic arrangement at the chromosome level with large structural rearrangements in some chromosomes. Genetic variation among five C. australasica cultivars was analysed. Genes related to defense, synthesis of volatile compounds and red/yellow coloration were identified in the genome. A major expansion of genes encoding thylakoid curvature proteins was found in the C. australasica genome. Conclusions: The genome of C. australasica present in this study is of high quality and contiguity. This genome helps deepen our understanding of citrus evolution and reveals disease resistance and quality related genes with potential to accelerate the genetic improvement of citrus. [ABSTRACT FROM AUTHOR]

Sustainable and effective means to control flying insect vectors are critically needed, especially with widespread insecticide resistance and global climate change. Understanding and controlling vectors requires accurate information about their movement and activity, which is often lacking. The Photonic Fence (PF) is an optical system that uses machine vision, infrared light, and lasers to identify, track, and interdict vectors in flight. The PF examines an insect's outline, flight speed, and other flight parameters and if these match those of a targeted vector species, then a low-power, retina-safe laser kills it. We report on proof-of-concept tests of a large, field-sized PF (30 mL × 3 mH) conducted with Aedes aegypti, a mosquito that transmits dangerous arboviruses, and Diaphorina citri, a psyllid which transmits the fatal huanglongbing disease of citrus. In tests with the laser engaged, < 1% and 3% of A. aegypti and D. citri, respectfully, were recovered versus a 38% and 19% recovery when the lacer was silenced. The PF tracked, but did not intercept the orchid bee, Euglossa dilemma. The system effectively intercepted flying vectors, but not bees, at a distance of 30 m, heralding the use of photonic energy, rather than chemicals, to control flying vectors. [ABSTRACT FROM AUTHOR]

Self-incompatibility (SI) is a genetic mechanism common in flowering plants to prevent self-fertilization. Among citrus species, several pummelo, mandarin, and mandarin-like accessions show SI behavior. In these species, SI is coupled with a variable degree of parthenocarpy ensuring the production of seedless fruits, a trait that is highly appreciated by consumers. In Citrus, recent evidences have shown the presence of a gametophytic SI system based on S-ribonucleases (SRNases) ability to impair self-pollen tube growth in the upper/middle part of the style. In the present study, we combined PCR analysis and next-generation sequencing technologies, to define the presence of S7- and S11-Rnases in the Sgenotype of the Citrus clementina (Hort. ex Tan.), the self-incompatible ‘Comune’ clementine and its self-compatible natural mutant ‘Monreal’. The reference genome of ‘Monreal’ clementine is presented for the first time, providing more robust results on the genetic sequence of the newly discovered S7-RNase. SNP discovery analysis coupled with the annotation of the variants detected enabled the identification of 7,781 SNPs effecting 5,661 genes in ‘Monreal’ compared to the reference genome of C. clementina. Transcriptome analysis of unpollinated pistils at the mature stage from both clementine genotypes revealed the lack of expression of S7-RNase in ‘Monreal’ suggesting its involvement in the loss of the SI response. RNA-seq analysis followed by gene ontology studies enabled the identification of 2,680 differentially expressed genes (DEGs), a significant number of those is involved in oxidoreductase and transmembrane transport activity. Merging of DNA sequencing and RNA data led to the identification of 164 DEGs characterized by the presence of at least one SNP predicted to induce mutations with a high effect on their amino acid sequence. Among them, four candidate genes referring to two Agamous-like MADS-box proteins, to MYB111 and to MLO-like protein 12 were validated. Moreover, the transcription factor MYB111 appeared to contain a binding site for the 2.0-kb upstream sequences of the S7- and S11- RNase genes. These results provide useful information about the genetic bases of SI indicating that SNPs present in their sequence could be responsible for the differential expression and the regulation of S7-RNase and consequently of the SI mechanism. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Elymus dahucirus is an essential plant for ecological restoration in fragile ecological areas and mining area restoration. As lawn grass, it can quickly cover soil and prevent soil erosion, so it is commonly used as a pioneer grass for lawn greening and slope protection. In recent years, with the development of mineral resources, Qinghai–Tibet Plateau soil is facing the threat of heavy metal cadmium (Cd) pollution. E. dahuricus can host the filamentous fungus Epichloë bromicola. To make better use of the advantages that Epichloë bring to host plants to alleviate heavy metal pollution in soil, plant growth and antioxidant capacity effects on E. bromicola infected (E+) and uninfected (E−) E. dahuricus were determined under Cd stress. During Cd treatment, plant growth was decreased by Cd stress, while E+ plants exhibited equal or better growth compared to E− plants. Cd treatment induces a proline and antioxidant enzyme burst in infected plants, while malondialdehyde (MDA) increases. E. bromicola improved plant growth and antioxidant capacity. E. dahuricus breeding strategies could use the information here in efforts to improve the performance of E. dahuricus in both environmental protection and agronomic contexts. [ABSTRACT FROM AUTHOR]

Simple Summary: Our study shows that the African citrus psyllid, a vector of Huanglongbing disease, prefers to develop at mild temperatures and is not well adapted to dry regions, where relative humidity is around or below 40%. Its life cycle is not completed or is severely delayed at temperatures above 30 °C and below 10 °C. Eggs are unable to hatch at constant temperatures below 8 °C or above 33 °C. Thus, T. erytreae establishment and spread will be maximum in regions with a temperate and humid climate, being rare in regions where dry weather conditions predominate. The African citrus psyllid, Trioza erytreae, is one of the two vectors of Huanglongbing, the most serious citrus disease worldwide. The first detection of T. erytreae in the European mainland was on the northwest of the Iberian Peninsula in 2014. Since then, the pest has spread throughout northern Spain (Galicia, Asturias, Cantabria, País Vasco) and along the western Atlantic coast of Portugal (from the Douro e Minho region to the Algarve). We conducted a series of laboratory experiments on lemon plants at different temperatures (from 8 to 34 °C) and humidity conditions (from 40 to 90%) to find out the influence of extreme temperatures and relative humidities (RHs) on the mortality, development and reproduction of T. erytreae. Our results show that temperatures above 30 °C and below 10 °C are very detrimental for nymphal development and nymphs were unable to reach the adult stage. Furthermore, eggs were unable to hatch under temperatures above 33 °C and below 8 °C. Adult mortality was highest at 34 °C and killed more than 50% of the population. We also found that relative humidity is crucial for the development and survival of T. erytreae. Nymphs were unable to reach the adult stage at an RH of 90% and 40%. Also, fecundity was significantly reduced at 90 and 40% RH, and fertility was lowest at 40% RH. Nymphal mortality was highest at an RH of 40%, which was the most detrimental humidity among all tested for the survival and development of T. erytreae. Our work concludes that T. erytreae establishment and spread will be maximum in regions with a temperate and humid climate, being rare in regions where dry and hot weather conditions predominate. [ABSTRACT FROM AUTHOR]

The Asian Citrus Psyllid (ACP), Diaphorina citri, is a vector of the pathological bacterium Candidatus Liberibacter asiaticus (CLas), which causes the most devastating disease to the citrus industry worldwide, known as greening or huanglongbing (HLB). Earlier field tests with an acetic acid-based lure in greening-free, 'Valencia' citrus orange groves in California showed promising results. The same type of lures tested in São Paulo, Brazil, showed unsettling results. During the unsuccessful trials, we noticed a relatively large proportion of females in the field, ultimately leading us to test field-collected males and females for Wolbachia and CLas. The results showed high rates of Wolbachia and CLas infection in field populations. We then compared the olfactory responses of laboratory-raised, CLas-free, and CLas-infected males to acetic acid. As previously reported, CLas-uninfected males responded to acetic acid at 1 µg. Surprisingly, CLas-infected males required 50 × higher doses of the putative sex pheromone, thus explaining the failure to capture CLas-infected males in the field. CLas infection was also manifested in electrophysiological responses. Electroantennogram responses from CLas-infected ACP males were significantly higher than those obtained with uninfected males. To the best of our knowledge, this is the first report of a pathogen infection affecting a vector's response to a sex attractant. [ABSTRACT FROM AUTHOR]

Due to human activities and changes in land use, the spatiotemporal pattern of citrus has undergone significant changes after the outbreak of Huanglongbing (HLB). We selected time-series Sentinel-2 images to delineate citrus orchard areas following the onset of HLB. This was conducted to extract citrus orchards in Xunwu County between 2017 and 2022. The spatial and temporal patterns and their influencing factors were investigated by spatial analysis. Results show (1) a notable decline in total citrus orchard area by 216.70 km2, primarily witnessed in orchards without insect-proof screens (IPS), shifting towards cropland, bush, and IPS areas. Contrastingly, citrus orchards with IPS exhibited a modest increase from 7.82 km2 to 111.39 km2, predominantly converting from areas lacking IPS, cropland, and bare land. (2) Spatial distribution patterns revealing a "cold in the south and hot in the north" trend. Orchards without IPS are concentrated in central and northern regions, while those with IPS are clustered predominantly in the north, with a recent shift towards the northeast. (3) Landscape analysis indicating a trend of fragmentation of citrus orchards, while a gradual dispersion of orchards without IPS and those with IPS showcased enhanced concentration and aggregation. (4) Orchards with IPS predominantly occupy regions characterized by an elevation ranging between 300 m and 400 m, primarily in the southeast, southwest, and southern directions. These areas exhibit slopes averaging between 10° and 15°, with surface temperatures ranging from 18 °C to 26 °C. Additionally, these orchards tend to be situated in proximity to impervious surfaces and roads. [ABSTRACT FROM AUTHOR]