Genome-wide analysis and expression profiling of the HD-ZIP gene family in kiwifruit.

The homeodomain-leucine zipper (HD-Zip) gene family plays a pivotal role in plant development and stress responses. Nevertheless, a comprehensive characterization of the HD-Zip gene family in kiwifruit has been lacking. In this study, we have systematically identified 70 HD-Zip genes in the Actinidia chinensis (Ac) genome and 55 in the Actinidia eriantha (Ae) genome. These genes have been categorized into four subfamilies (HD-Zip I, II, III, and IV) through rigorous phylogenetic analysis. Analysis of synteny patterns and selection pressures has provided insights into how whole-genome duplication (WGD) or segmental may have contributed to the divergence in gene numbers between these two kiwifruit species, with duplicated gene pairs undergoing purifying selection. Furthermore, our study has unveiled tissue-specific expression patterns among kiwifruit HD-Zip genes, with some genes identified as key regulators of kiwifruit responses to bacterial canker disease and postharvest processes. These findings not only offer valuable insights into the evolutionary and functional characteristics of kiwifruit HD-Zips but also shed light on their potential roles in plant growth and development.

A Report of Pectobacterium carotovorum subsp. actinidiae Causing Kiwifruit Bacterial Canker in Guangxi, China.

A bacterial pathogen strain was isolated from susceptible tissue of Hongyang variety kiwifruit in Zhongfeng Town, Ziyuan County, Guilin City, Guangxi, China. Due to the relatively single variety of kiwifruit in Guangxi, the control technology of fruit farmers is backward, and the climate is humid, which is suitable for the growth of pathogenic bacteria, resulting in frequent occurrence of diseases. In this study, the pathogen strain was identified based on morphological, physiological, and biochemical tests; 16S rRNA gene; PCR detection with specific primers; and Biolog analysis. The results showed that a tobacco allergic reaction could be induced by inoculation with the pathogenic bacteria. Additionally, brown necrotic plaques appeared on kiwifruit leaves, necrotic phloem lesions appeared, and wounds on kiwifruit branches turned brown. The characteristics identified by morphological, physiological, biochemical, and Biolog identification were similar to those caused by Pectobacterium sp. Through 16S rRNA gene sequence analysis and PCR identification with specific primers, bands with a size corresponding to target bands indicated that the pathogen was Pectobacterium carotovorum subsp. actinidiae. This is the first report of kiwifruit canker disease caused by P. carotovorum subsp. actinidiae in Guangxi, China. In addition, through this study, a preliminary understanding of the pathogen has been obtained, which will lay the foundation for the prevention and control of the disease in the future.

Complete genome sequence of the kiwifruit bacterial canker pathogen Pseudomonas savastanoi strain MHT1.

BACKGROUND: Pseudomonas savastanoi is an important plant pathogen that infects and causes symptoms in a variety of economically important crops, causing considerable loss of yield and quality. Because there has been no research reported to date on bacterial canker of kiwifruit (Actinidia chinensis) plants caused by P. savastanoi and, in particular, no in-depth studies of the complete genome sequence or pathogenic mechanism, long-lasting and environmentally friendly control measures against this pathogen in kiwifruit are lacking. This study therefore has both theoretical value and practical significance. RESULTS: We report the complete genome sequence of P. savastanoi strain MHT1, which was first reported as the pathogen causing bacterial canker in kiwifruit plants. The genome consists of a 6.00-Mb chromosome with 58.5% GC content and 5008 predicted genes. Comparative genome analysis of four sequenced genomes of representative P. savastanoi strains revealed that 230 genes are unique to the MHT1 strain and that these genes are enriched in antibiotic metabolic processes and metabolic pathways, which may be associated with the drug resistance and host range observed in this strain. MHT1 showed high syntenic relationships with different P. savastanoi strains. Furthermore, MHT1 has eight conserved effectors that are highly homologous to effectors from P. syringae, Pseudomonas amygdali, and Ralstonia solanacearum strains. The MHT1 genome contains six genomic islands and two prophage sequences. In addition, 380 genes were annotated as antibiotic resistance genes and another 734 as encoding carbohydrate-active enzymes. CONCLUSION: The whole-genome sequence of this kiwifruit bacterial canker pathogen extends our knowledge of the P. savastanoi genome, sets the stage for further studies of the interaction between kiwifruit and P. savastanoi, and provides an important theoretical foundation for the prevention and control of bacterial canker.

Defence-related pathways, phytohormones and primary metabolism are key players in kiwifruit plant tolerance to Pseudomonas syringae pv. actinidiae.

The reasons underlying the differential tolerance of Actinidia spp. to the pandemic pathogen Pseudomonas syringae pv. actinidiae (Psa) have not yet been elucidated. We hypothesized that differential plant-defence strategies linked to transcriptome regulation, phytohormones and primary metabolism might be key and that Actinidia chinensis susceptibility results from an inefficient activation of defensive mechanisms and metabolic impairments shortly following infection. Here, 48 h postinoculation bacterial density was 10-fold higher in A. chinensis var. deliciosa than in Actinidia arguta, accompanied by significant increases in glutamine, ornithine, jasmonic acid (JA) and salicylic acid (SA) (up to 3.2-fold). Actinidia arguta showed decreased abscisic acid (ABA) (0.7-fold), no changes in primary metabolites, and 20 defence-related genes that were only differentially expressed in this species. These include GLOX1, FOX1, SN2 and RBOHA, which may contribute to its higher tolerance. Results suggest that A. chinensis' higher susceptibility to Psa is due to an inefficient activation of plant defences, with the involvement of ABA, JA and SA, leading to impairments in primary metabolism, particularly the ammonia assimilation cycle. A schematic overview on the interaction between Psa and genotypes with distinct tolerance is provided, highlighting the key transcriptomic and metabolomic aspects contributing to the different plant phenotypes after infection.

Characterization of Pseudomonas syringae pv. actinidiae biovar 3 on kiwifruit in north-west Portugal.

AIMS: Bacterial kiwifruit canker disease, caused by Pseudomonas syringae pv. actinidiae (Psa) was detected in north-west Portugal in 2010, and has since caused significant losses. The objectives of this work were to characterize the Portuguese population(s) of Psa and to define the actual prevalence of Psa biovars in the most productive kiwifruit region in Portugal. METHODS AND RESULTS: Isolates obtained from Actinidia deliciosa orchards were characterized by morphological, biochemical, physiological, fatty acids and molecular tests (PCR, BOX-PCR, duplex-PCR, multiplex-PCR and RFLP), phaseolotoxin, housekeeping and effector genes and pathogenicity. Results established that only Psa biovar 3 is present in the north-west of Portugal, despite phenotypic and genetic variability among the isolates. CONCLUSIONS: This work provides new information on P. syringae pv. actinidiae (Psa) genetic profile in Portugal, indicating for the first time, that two genetically different subpopulations of Psa biovar 3 are present. SIGNIFICANCE AND IMPACT OF THE STUDY: A new subpopulation of Psa biovar 3 was found for the first time in Portugal, contributing to increase knowledge about this population worldwide and to support further understanding of the impact of Psa.

Streptothricin-F Inhibition of FtsZ Function: A Promising Approach for Controlling Pseudomonas syringae pv. actinidiae.

Pseudomonas syringae pv. actinidiae (Psa) is a significant pathogenic bacterium affecting the kiwifruit industry. This study investigated the target sites of streptothricin-F (ST-F), produced by Streptomyces lavendulae gCLA4. The inhibition of ST-F on Psa was examined by the microscopic structural differences of Psa before and after treatment with ST-F, as well as the interaction between ST-F and cell division-related proteins. The results revealed filamentation of Psa after ST-F treatment, and fluorescence microscopy showed that ST-F inhibited the formation of the Z-ring composed of FtsZ protein. In vitro experiments and molecular docking demonstrated that ST-F can bind to FtsZ with a binding energy of 0.4 µM and inhibit FtsZ's GTP-dependent polymerization reaction. In addition, ST-F does not exert inhibitory effects on cell division in Psa strains overexpressing ftsZ. In conclusion, FtsZ is one of the target sites for ST-F inhibition of Psa, highlighting its potential as a therapeutic target for controlling Psa-induced kiwifruit bacterial canker.

Enhancing Pseudomonas syringae pv. Actinidiae sensitivity in kiwifruit by repressing the NBS-LRR genes through miRNA-215-3p and miRNA-29-3p identification.

Kiwifruit bacterial canker, caused by Pseudomonas syringae pv. actinidiae (PSA), poses a grave threat to the global kiwifruit industry. In this study, we examined the role of microRNAs (miRNAs) in kiwifruit's response to PSA. Kiwifruit seedlings subjected to PSA treatment showed significant changes in both miRNA and gene expression compared to the control group. We identified 364 differentially expressed miRNAs (DEMs) and 7170 differentially expressed genes (DEGs). Further analysis revealed 180 miRNAs negatively regulating 641 mRNAs. Notably, two miRNAs from the miRNA482 family, miRNA-215-3p and miRNA-29-3p, were found to increase kiwifruit's sensitivity to PSA when overexpressed. These miRNAs were linked to the regulation of NBS-LRR target genes, shedding light on their role in kiwifruit's defence against PSA. This study offers insights into the miRNA482-NBS-LRR network as a crucial component in enhancing kiwifruit bioresistance to PSA infestation and provides promising candidate genes for further research.

Contribution of Sucrose Metabolism in Phloem to Kiwifruit Bacterial Canker Resistance.

Kiwifruit bacterial canker, caused by Pseudomonas syringae pv. actinidiae (Psa), is a catastrophic disease affecting kiwifruit worldwide. As no effective cure has been developed, planting Psa-resistant cultivars is the best way to avoid bacterial canker in kiwifruit cultivation. However, the differences in the mechanism of resistance between cultivars is poorly understood. In the present study, five local kiwifruit cultivars were used for Psa resistance evaluation and classified into different resistance categories, tolerant (T), susceptible (S), and highly susceptible (HS), based on their various symptoms of lesions on the cane. Susceptible and highly susceptible varieties had a higher sucrose concentration, and a greater decrease in sucrose content was observed after Psa inoculation in phloem than in tolerant varieties. Three invertase activities and their corresponding gene expressions were detected in the phloem with lesions and showed the same trends as the variations in sucrose concentration. Meanwhile, after Psa inoculation, enzyme activities involved in antioxidant defense responses, such as PAL, POD, and CAT, were also altered in the phloem of the lesion position. With no differences among cultivars, PAL and POD activities in phloem first increased and then decreased after Psa inoculation. However, great differences in CAT activities were observed between T and S/HS categories. Our results demonstrate that sucrose content was negatively correlated with the disease resistance of different cultivars and that the increase in immune response enzymes is likely caused by increased sucrose metabolism in the phloem.

Paenibacillus polymyxa YLC1: a promising antagonistic strain for biocontrol of Pseudomonas syringae pv. actinidiae, causing kiwifruit bacterial canker.

BACKGROUND: Kiwifruit bacterial canker (KBC) caused by Pseudomonas syringae pv. actinidiae (Psa) is the main limiting factor in the kiwifruit industry. This study aimed to identify bacterial strains with antagonistic activity against Psa, analyze antagonistically active substances and provide a new basis for the biological control of KBC. RESULTS: A total of 142 microorganisms were isolated from the rhizosphere soil of asymptomatic kiwifruit. Among them, an antagonistic bacterial strain was identified as Paenibacillus polymyxa YLC1 by 16S rRNA sequencing. KBC control by strain YLC1 (85.4%) was comparable to copper hydroxide treatment (81.8%) under laboratory conditions and field testing. Active substances of strain YLC1 were identified by genetic sequence analysis using antiSMASH. Six biosynthetic active compound gene clusters were identified as encoding ester peptide synthesis, such as polymyxins. An active fraction was purified and identified as polymyxin B1 using chromatography, hydrogen nuclear magnetic resonance (NMR), and liquid chromatography-mass spectrometry. In addition, polymyxin B1 also was found significantly to suppress the expression of T3SS-related genes, but did not affect the growth of Psa at low concentrations. CONCLUSION: In this study, a biocontrol strain P. polymyxa YLC1 obtained from kiwifruit rhizosphere soil exhibited excellent control effects on KBC in vitro and in field tests. Its active compound was identified as polymyxin B1, which inhibits a variety of pathogenic bacteria. We conclude that P. polymyxa YLC1 is a biocontrol strain with excellent prospects for development and application. © 2023 Society of Chemical Industry.

Real-Time Detection Algorithm for Kiwifruit Canker Based on a Lightweight and Efficient Generative Adversarial Network.

Disease diagnosis and control play important roles in agriculture and crop protection. Traditional methods of identifying plant disease rely primarily on human vision and manual inspection, which are subjective, have low accuracy, and make it difficult to estimate the situation in real time. At present, an intelligent detection technology based on computer vision is becoming an increasingly important tool used to monitor and control crop disease. However, the use of this technology often requires the collection of a substantial amount of specialized data in advance. Due to the seasonality and uncertainty of many crop pathogeneses, as well as some rare diseases or rare species, such data requirements are difficult to meet, leading to difficulties in achieving high levels of detection accuracy. Here, we use kiwifruit trunk bacterial canker (Pseudomonas syringae pv. actinidiae) as an example and propose a high-precision detection method to address the issue mentioned above. We introduce a lightweight and efficient image generative model capable of generating realistic and diverse images of kiwifruit trunk disease and expanding the original dataset. We also utilize the YOLOv8 model to perform disease detection; this model demonstrates real-time detection capability, taking only 0.01 s per image. The specific contributions of this study are as follows: (1) a depth-wise separable convolution is utilized to replace part of ordinary convolutions and introduce noise to improve the diversity of the generated images; (2) we propose the GASLE module by embedding a GAM, adjust the importance of different channels, and reduce the loss of spatial information; (3) we use an AdaMod optimizer to increase the convergence of the network; and (4) we select a real-time YOLOv8 model to perform effect verification. The results of this experiment show that the Fréchet Inception Distance (FID) of the proposed generative model reaches 84.18, having a decrease of 41.23 compared to FastGAN and a decrease of 2.1 compared to ProjectedGAN. The mean Average Precision (mAP@0.5) on the YOLOv8 network reaches 87.17%, which is nearly 17% higher than that of the original algorithm. These results substantiate the effectiveness of our generative model, providing a robust strategy for image generation and disease detection in plant kingdoms.

Response of Soil Microbial Community Structure Mediated by Sulfur-Induced Resistance Against Kiwifruit Bacterial Canker.

Kiwifruit bacterial canker caused by Pseudomonas syringae pv. actinidiae (Psa) is a major threat to kiwifruit worldwide, and effective control measures are still lacking. Sulfur, as a mineral, has been proved to improve plants' resistance to pathogens. It is of great significance to study the effect of sulfur on rhizosphere microorganisms in kiwifruit planting areas infected by Psa for controlling kiwifruit canker. In this study, the sulfur powder and organic fertilizer were mixed as base fertilizer to treat the soil in the area where kiwifruit bacterial canker occurs. We investigated the incidence of kiwifruit bacterial canker in 2018 and 2019 after sulfur application and the changes in microbial characteristics and community composition structure in the kiwifruit rhizosphere by using the plate-counting method and high-throughput sequencing technology. Fertilization treatments of kiwifruit roots with sulfur and organic fertilizer reduced kiwifruit bacterial canker severity. The diversity of soil microbial communities increased significantly after sulfur application in the range of 1.0~2.0 kg/m3. In particular, the bacterial genera level showed a high diversity after 2 years of sulfur application, reaching more than 516 genera. Furthermore, sulfur treatment resulted in a significant increase in specific microbial taxa, including members of the Acidothermus, norank_f__HSB_OF53-F07, and norank_f __Acidobacteriaceae__Subgroup_1. Moreover, the proportion of the dominant bacteria Acidothermus in the population showed an increasing trend. Altogether, the sulfur application is the key factor leading to microbial differences in kiwifruit rhizosphere soil. Appropriate sulfur can improve microbial structure characteristics of kiwifruit rhizosphere soil, increase bacterial diversity index, and change bacterial community composition structure.

Evaluation of the Abilities of Three Kinds of Copper-Based Nanoparticles to Control Kiwifruit Bacterial Canker.

Kiwifruit bacterial canker caused by Pseudomonas syringae pv. actinidiae reduces kiwifruit crop yield and quality, leading to economic losses. Unfortunately, few agents for its control are available. We prepared three kinds of copper-based nanoparticles and applied them to control kiwifruit bacterial canker. The successful synthesis of Cu(OH)2 nanowires, Cu3(PO4)2 nanosheets, and Cu4(OH)6Cl2 nanoparticles were confirmed by transmission and scanning electron microscopy, energy dispersive spectroscopy, X-ray diffraction analysis, and X-ray photoelectron spectroscopy. The minimum bactericidal concentrations (MBCs) of the three nanoparticles were 1.56 µg/mL, which exceeded that of the commercial agent thiodiazole copper (MBC > 100 µg/mL). The imaging results indicate that the nanoparticles could interact with bacterial surfaces and kill bacteria by inducing reactive oxygen species' accumulation and disrupting cell walls. The protective activities of Cu(OH)2 nanowires and Cu3(PO4)2 nanosheets were 59.8% and 63.2%, respectively, similar to thiodiazole copper (64.4%) and better than the Cu4(OH)6Cl2 nanoparticles (40.2%). The therapeutic activity of Cu4(OH)6Cl2 nanoparticles (67.1%) bested that of Cu(OH)2 nanowires (43.9%), Cu3(PO4)2 nanosheets (56.1%), and thiodiazole copper (53.7%). Their therapeutic and protective activities for control of kiwifruit bacterial canker differed in vivo, which was related to their sizes and morphologies. This study suggests these copper-based nanoparticles as alternatives to conventional bactericides for controlling kiwifruit diseases.

Transcriptome Analysis on the Mechanism of Ethylicin Inhibiting Pseudomonas syringae pv. actinidiae on Kiwifruit.

Bacterial canker disease caused by Pseudomonas syringae pv. actinidiae (Psa) is a devastating disease of kiwifruit, which is severely limiting the development of the kiwifruit industry. Ethylicin is a broad-spectrum plant biomimetic fungicide. However, its application in the control of kiwifruit bacterial canker is rarely reported, and the mechanism of ethylicin on Psa remains unknown. In this study, we investigated the effect of ethylicin on Psa in vitro and in vivo and found that ethylicin can inhibit the growth of Psa and prevent the cankering in the plant stem. Mechanism investigation indicated that ethylicin acted by limiting the movement of Psa, destroying the cell membrane of Psa, and inhibiting the formation of Psa biofilm. In addition, it was also found through transcriptomics research that ethylicin can up-regulate the expression of genes related to protein export and biofilm formation-Pseudomonas aeruginosa and down-regulate the expression of genes related to flagellar assembly in Psa. This study concluded that ethylicin can effectively inhibit Psa growth, and it could help to gain a better understanding of the mechanisms of ethylicin inhibiting Psa and provide practical data for the application of ethylicin as a highly potent agent for controlling the bacterial canker disease of kiwifruit.

Role of the Type VI Secretion System in the Pathogenicity of Pseudomonas syringae pv. actinidiae, the Causative Agent of Kiwifruit Bacterial Canker.

The type VI secretion system (T6SS), a macromolecular machine, plays an important role in the pathogenicity of many Gram-negative bacteria. However, the role of T6SS in the pathogenicity of Pseudomonas syringae pv. actinidiae (Psa), the pathogen of kiwifruit bacterial canker, is yet to be studied. Here, we found a T6SS gene cluster consisting of 13 core genes (A-J) in the genome of Psa M228 based on a genome-wide analysis. To determine whether the T6SS gene cluster affects the pathogenicity of Psa M228, T6SS and its 13 core gene deletion mutants were constructed and their pathogenicity was determined. The deletion mutants showed different degrees of reduction in pathogenicity compared with the wild-type strain M228; in tssM and tssJ mutants, pathogenicity was significantly reduced by 78.7 and 71.3%, respectively. The pathogenicity results were also confirmed by electron microscopy. To further confirm that the reduction in pathogenicity is related to the function of T6SS, we selected the T6SS gene cluster, comprising tssM and tssJ, for further analyses. Western blot results revealed that tssM and tssJ were necessary for hemolytic co-regulatory protein secretion, indicating that they encode a functional T6SS. Further, we explored the mechanism by which T6SS affects the pathogenicity of Psa M228. The ability of bacterial competition, biofilm formation, hydrogen peroxide tolerance, and proteolytic activity were all weakened in the deletion mutants M228ΔT6SS, M228ΔtssM, and M228ΔtssJ. All these properties of the two gene complementation mutants were restored to the same levels as those of the wild-type strain, M228. Quantitative real-time results showed that during the interaction between the deletion mutant M228ΔT6SS and the host, expression levels of T3SS transcriptional regulatory gene hrpR, structural genes hrpZ, hrcC, hopP1, and effector genes hopH1 and hopM1 were down-regulated at different levels. Taken together, our data provide evidence for the first time that the T6SS plays an important role in the pathogenicity of Psa, probably via effects on bacterial competition, biofilm formation, and environmental adaptability. Moreover, a complicated relationship exists between T6SS and T3SS.

Identification and Analysis of NBS-LRR Genes in Actinidia chinensis Genome.

Nucleotide-binding site and leucine-rich repeat (NBS-LRR) genes represent the most important disease resistance genes in plants. The genome sequence of kiwifruit (Actinidia chinensis) provides resources for the characterization of NBS-LRR genes and identification of new R-genes in kiwifruit. In the present study, we identified 100 NBS-LRR genes in the kiwifruit genome and they were grouped into six distinct classes based on their domain architecture. Of the 100 genes, 79 are truncated non-regular NBS-LRR genes. Except for 37 NBS-LRR genes with no location information, the remaining 63 genes are distributed unevenly across 18 kiwifruit chromosomes and 38.01% of them are present in clusters. Seventeen families of cis-acting elements were identified in the promoters of the NBS-LRR genes, including AP2, NAC, ERF and MYB. Pseudomonas syringae pv. actinidiae (pathogen of the kiwifruit bacterial canker) infection induced differential expressions of 16 detected NBS-LRR genes and three of them are involved in plant immunity responses. Our study provides insight of the NBS-LRR genes in kiwifruit and a resource for the identification of new R-genes in the fruit.