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Xanthomonas spp. are plant pathogens known for significantly impacting crop yields. Among them, Xanthomonas albilineans (Xal) is notable for colonizing the xylem and causing sugarcane leaf scald disease. This study employed homologous recombination to mutate quorum sensing (QS) regulatory genes (rpf) to investigate their role in Xal pathogenicity. Deletions of rpfF (ΔrpfF), rpfC (ΔrpfC), and rpfG (ΔrpfG) led to reduced swarming, growth, and virulence. However, DSF supplementation restored swarming and growth in the ΔrpfF mutant. Deleting rpfC, rpfG, and rpfF also reduced twitching motility and affected Type IV Pilus (T4P) expression. Transcriptomic analysis revealed that ΔrpfF positively regulates flagellar genes. DSF supplementation in ΔrpfF (ΔrpfF-DSF) modulated the expression of flagellar, chemotaxis, and T4P genes. These findings elucidate the DSF-mediated swarming pathway in Xal and provide valuable insights into its regulatory mechanisms.
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A homeobox transcription factor is a conserved transcription factor, ubiquitous in eukaryotes, that regulates the tissue formation of structure, cell differentiation, proliferation, and cancer. This study identified the homeobox transcription factor family and its distribution in Phoma sorghina var. saccharum at the whole genome level. It elucidated the gene structures and evolutionary characteristics of this family. Additionally, knockout experiments were carried out and the preliminary function of these transcription factors was studied. Through bioinformatics approaches, nine homeobox transcription factors (PsHOX1-PsHOX9) were identified in P. sorghina var. saccharum, and these contained HOX-conserved domains and helix-turn-helix secondary structures. Nine homeobox gene deletion mutants were obtained using the homologous recombinant gene knockout technique. Protoplast transformation was mediated by polyethylene glycol (PEG) and the transformants were identified using PCR. The knockouts of PsHOX1, PsHOX2, PsHOX3, PsHOX4, PsHOX6, PsHOX8, and PsHOX9 genes resulted in a smaller growth diameter in P. sorghina var. saccharum. In contrast, the knockouts of the PsHOX3, PsHOX6, and PsHOX9 genes inhibited the formation of conidia and led to a significant decrease in the pathogenicity. This study's results will provide insights for understanding the growth and development of P. sorghina var. saccharum. The pathogenic mechanism of the affected sugarcane will provide an essential theoretical basis for preventing and controlling sugarcane twisted leaf disease.
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Proteínas de Homeodomínio , Doenças das Plantas , Saccharum , Saccharum/genética , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Doenças das Plantas/microbiologia , Doenças das Plantas/genética , Ascomicetos/patogenicidade , Ascomicetos/genética , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Folhas de Planta/genética , FilogeniaRESUMO
Pokkah boeng disease (PBD), a sugarcane foliar disease, is caused by various Fusarium spp. within the Fusarium fujikuroi species complex (FFSC). In the current study, we investigated the diversity of Fusarium spp. associated with PBD in China. In total, 320 leaf samples displaying PBD symptoms were collected over 10 consecutive years (2012 to 2021), during winter and summer, from six various sugarcane-growing regions (Guangxi, Yunnan, Guangdong, Zhejiang, Hainan, and Fujian) in China. Phylogenetic analysis of Fusarium spp. was reconstructed using translation elongation factor 1-α, and DNA-directed RNA polymerase II largest subunit and second-largest subunit multigene sequences. Evolutionary studies of these regions categorized the isolates into four FFSC species (F. sacchari, F. proliferatum, F. verticillioides, and F. andiyazi). The identified isolates, which developed irregular necrotic patches and rotting symptoms on the sugarcane plant after approximately 30 days were tested for their pathogenicity. Symptoms that appeared during pathogenicity testing were consistent with those observed under field conditions. Each strain of the pathogenic Fusarium spp. belonged to different vegetative compatibility groups (VCGs), and there was no affinity between VCGs. Our results contribute to understanding FFSC and accurately identifying Fusarium spp. associated with the sugarcane crop.
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Fusarium , Saccharum , Filogenia , Virulência/genética , China , Grão Comestível , Variação GenéticaRESUMO
Nitrogen availability might play an essential role in plant diseases by enhancing fungal cell growth and influencing the expression of genes required for successful pathogenesis. Nitrogen availability could modulate secondary metabolic pathways as evidenced by the significant differential expression of several core genes involved in mycotoxin biosynthesis and genes encoding polyketide synthase/nonribosomal peptide synthetases, cytochrome P450 and carbohydrate-active enzymes in Fusarium sacchari, grown on different nitrogen sources. A combined analysis was carried out on the transcript and metabolite profiles of regulatory metabolic processes and the virulence of Fusarium sacchari grown on various nitrogen sources. The nitrogen regulation of the gibberellin gene cluster included the metabolic flux and multiple steps of gibberellin synthesis. UHPLC-MS/MS-based metabolome analysis revealed the coordination of these related transcripts and the accumulation of gibberellin metabolites. This integrated analysis allowed us to uncover additional information for a more comprehensive understanding of biological events relevant to fungal secondary metabolic regulation in response to nitrogen availability.
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Fusarium , Transcriptoma , Metabolismo Secundário/genética , Nitrogênio/metabolismo , Espectrometria de Massas em Tandem , Giberelinas/metabolismo , Regulação Fúngica da Expressão GênicaRESUMO
BACKGROUND: Xanthomonas is a genus of gram-negative bacterium containing more than 35 species. Among these pathogenic species, Xanthomonas albilineans (Xal) is of global interest, responsible for leaf scald disease in sugarcane. Another notable Xanthomonas species is Xanthomonas sachari (Xsa), a sugarcane-associated agent of chlorotic streak disease. RESULT: The virulence of 24 Xanthomonas strains was evaluated by disease index (DI) and Area Under Disease Progress Curve (AUDPC) in the susceptible inoculated plants (GT 46) and clustered into three groups of five highly potent, seven mild virulent, and twelve weak virulent strains. The highly potent strain (X. albilineans, Xal JG43) and its weak virulent related strain (X. sacchari, Xsa DD13) were sequenced, assembled, and annotated in the circular genomes. The genomic size of JG43 was smaller than that of DD13. Both strains (JG43 and DD13) lacked a Type III secretory system (T3SS) and T6SS. However, JG43 possessed Salmonella pathogenicity island-1 (SPI-1). More pathogen-host interaction (PHI) genes and virulent factors in 17 genomic islands (GIs) were detected in JG43, among which six were related to pathogenicity. Albicidin and a two-component system associated with virulence were also detected in JG43. Furthermore, 23 Xanthomonas strains were sequenced and classified into three categories based on Single Nucleotide Polymorphism (SNP) mutation loci and pathogenicity, using JG43 as a reference genome. Transitions were dominant SNP mutations, while structural variation (SV) is frequent intrachromosomal rearrangement (ITX). Two essential genes (rpfC/rpfG) of the two-component system and another gene related to SNP were mutated to understand their virulence effect. The mutation of rpfG resulted in a decrease in pathogenicity. CONCLUSION: These findings revealed virulence of 24 Xanthomonas strains and variations by 23 Xanthomonas strains. We sequenced, assembled, and annotated the circular genomes of Xal JG43 and Xsa DD13, identifying diversity detected by pathogenic factors and systems. Furthermore, complete genomic sequences and sequenced data will provide a theoretical basis for identifying pathogenic factors responsible for sugarcane leaf scald disease.
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Saccharum , Xanthomonas , Doenças das Plantas/microbiologia , Folhas de Planta/genética , Saccharum/microbiologia , Virulência/genética , Fatores de Virulência/genética , Xanthomonas/genéticaRESUMO
Sugarcane pokkah boeng disease (PBD) is emerging as a prevalent foliar disease in China. This airborne disease is caused by the Fusarium species complex. To investigate the diversity and evolution of Fusarium spp., we performed whole-genome sequencing of Fusarium andiyazi YN28 using a combination of Oxford Nanopore and Illumina technology. The F. andiyazi YN28 genome was sequenced, assembled, and annotated. A high-quality genome was assembled into 24 contigs with an N50 of 2.80 Mb. The genome assembly generated a total size of 44.1 Mb with a GC content of 47.64%. In total, 15,508 genes were predicted, including 794 genes related to the carbohydrate-active enzymes, 397 noncoding RNA, 155 genes associated with transporter classification, 4,550 genes linked to pathogen-host interactions, and 269 genes involved in effector proteins. Collectively, our results will provide insight into the host-pathogen interactions and will facilitate the breeding of new varieties of sugarcane resistant to PBD.[Formula: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.
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Fusarium , Saccharum , Fusarium/genética , Melhoramento Vegetal , Doenças das PlantasRESUMO
Phoma sorghina var. saccharum is a fungal pathogen that causes sugarcane twisted leaf disease in China. Here, we report complete genome assemblies of the Phoma sorghina var. saccharum isolate BS2-1, generated using single-molecule real-time sequencing. We present a high-quality genome sequence of a Phoma isolate that was assembled into 22 contigs with an N50 length of 1.92 Mb, a total length of 33.12 Mb, and a GC content of 52.12%. A total of 7,870 genes were annotated, using a combination of gene prediction tools, including 281 noncoding RNAs, 515 genes encoding carbohydrate-active enzymes, 2,440 genes associated with pathogen-host interactions, and 583 genes encoding secreted proteins. The complete genome sequence will be useful for understanding host-pathogen interaction and for improving disease management strategies.
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Ascomicetos , Genoma Fúngico , Doenças das Plantas/microbiologia , Saccharum/microbiologia , Ascomicetos/genética , China , Folhas de PlantaRESUMO
BACKGROUND: Pokkah boeng is one of the most serious and devastating diseases of sugarcane and causes significant loss in cane yield and sugar content. Although carbendazim is widely used to prevent fungal diseases, the molecular basis of Fusarium species complex (FSC) resistance to carbendazim remains unknown. RESULTS: The EC50 (fungicide concentration that inhibits 50% of mycelial growth) values of carbendazim for 35 FSC isolates collected in cane growing regions of China were ranged from 0.5097 to 0.6941 µg mL- 1 of active ingredient (a.i.), in an average of 0.5957 µg a.i. mL- 1. Among carbendazim-induced mutant strains, SJ51M (F. verticillioides) had a CTG rather than CAG codon (Q134L) at position 134 of the FVER_09254 gene, whereas in the mutant strain HC30M (F. proliferatum) codon ACA at position 351 of the FPRO_07779 gene was replaced by ATA (T351I). Gene expression profiling analysis was performed for SJ51M and its corresponding wild type strain SJ51, with and without carbendazim treatment. The gene expression patterns in SJ51 and SJ51M changed greatly as evidenced by the detection of 850 differentially expressed genes (DEGs). Functional categorization indicated that genes associated with oxidation-reduction process, ATP binding, integral component of membrane, transmembrane transport and response to stress showed the largest expression changes between SJ51M and SJ51. The expression levels of many genes involved in fungicide resistance, such as detoxification enzymes, drug efflux transporters and response to stress, were up-regulated in SJ51M compared to SJ51 with and without carbendazim treatment. CONCLUSION: FSC was sensitive to carbendazim and had the potential for rapid development of carbendazim resistance. The transcriptome data provided insight into the molecular pathways involved in FSC carbendazim resistance.
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Benzimidazóis/farmacologia , Carbamatos/farmacologia , Farmacorresistência Fúngica/genética , Fusarium/efeitos dos fármacos , Fusarium/fisiologia , Doenças das Plantas/microbiologia , Saccharum/microbiologia , Fusarium/genética , Genes Fúngicos/genética , Mutação , Temperatura , Transcriptoma/efeitos dos fármacosRESUMO
Sugarcane is a critical sugar and bioenergy crop in China. However, numerous factors, including root rot disease, hamper its yield. Root rot disease is a severe agricultural issue, reducing yield and threatening sustainable crop production. The current study aimed to explore the fungal community structure, identify and characterize the primary pathogen for sugarcane root rot in Guangzhou, China. Eighty-nine samples of sugarcane root, stalk, rhizosphere soil, and irrigation water were collected from five sites in Guangzhou, China. Subsequently, 276 fungal strains were isolated to identify the primary pathogens. The five most common genera identified were Penicillium, Fusarium, Gongronella, Trichoderma, and Cladosporium. Fusarium was more prevalent in the infected soil samples than in healthy ones. Pathogenic assays of the strains revealed that the strain GX4-46 caused 80% of the disease. The strain was confirmed as Fusarium commune through phylogenetic and genome sequence analysis. Rhizosphere soil samples from different regional crops were collected to better understand the fungal community structure and the primary pathogen. We observed a significant presence of Fusarium in irrigation water, indicating that the root rot disease could originate from the irrigation water and then spread as a soil-borne disease. This research is pioneering and one of the most comprehensive investigations on the occurrence and prevalence of sugarcane root rot disease. This study will serve as a reference for expanding the sugarcane industry and a foundation for further exploration and control of root rot.IMPORTANCESugarcane, a significant economic crop, faces challenges due to root rot pathogens that accumulate each year in plants and soil through ratoon planting. This disrupts soil microbial balance and greatly impedes sugarcane industry growth. Symptoms range from wilting and yellowing leaves to stunted growth and reduced seedling tillers. The rhizosphere microbiota plays an important role in plant development and soil health. Little is known about root rot fungal community structure, especially in sugarcane. Here, we focused on exploring the main causative pathogen of root rot in the area alongside a detailed survey of the rhizosphere soil of different severity sugarcane cultivars and rotation crops of the region. To validate the findings, we also investigated the irrigation water of the area. Our study revealed Fusarium commune as the causative pathogen of root rot in the area, primarily originating from water and later as soil-borne. Using Trichoderma can control the disease effectively.
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Fusarium , Micobioma , Saccharum , Trichoderma , Raízes de Plantas/microbiologia , Filogenia , Trichoderma/genética , Solo/química , Produtos Agrícolas , Surtos de Doenças , ÁguaRESUMO
Fusarium sacchari is one of the primary pathogens causing pokkah boeng disease, which impairs the yield and quality of sugarcane around the world. Understanding the molecular mechanisms of the F. sacchari effectors that regulate plant immunity is of great importance for the development of novel strategies for the persistent control of pokkah boeng disease. In a previous study, Fs00367 was identified to inhibit BAX-induced cell death. In this study, Fs00367nsp (without signal peptide) was found to suppress BAX-induced cell death, reactive oxygen species bursts and callose accumulation. The amino acid region 113-142 of Fs00367nsp is the functional region. Gene mutagenesis indicated that Fs00367 is important for the full virulence of F. sacchari. A yeast two-hybrid assay revealed an interaction between Fs00367nsp and sugarcane ScPi21 in yeast that was further confirmed using bimolecular fluorescence complementation, pull-down assay and co-immunoprecipitation. ScPi21 can induce plant immunity, but this effect could be blunted by Fs00367nsp. These results suggest that Fs00367 is a core pathogenicity factor that suppresses plant immunity through inhibiting ScPi21-induced cell death. The findings of this study provide new insights into the molecular mechanisms of effectors in regulating plant immunity.
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Fusarium , Saccharum , Proteína X Associada a bcl-2/metabolismo , Proteína X Associada a bcl-2/farmacologia , Imunidade Vegetal/genética , Saccharum/genética , Saccharum/metabolismo , Morte Celular , Doenças das PlantasRESUMO
Pokkah Boeng disease (PBD), caused by Fusarium sacchari, severely affects sugarcane yield and quality. Necrosis-inducing secreted protein 1 (Nis1) is a fungal secreted effector that induces necrotic lesions in plants. It interacts with host receptor-like kinases and inhibits their kinase activity. FsNis1 contains the Nis1 structure and triggered a pathogen-associated molecular pattern-triggered immune response in Nicotiana benthamiana, as reflected by causing reactive oxygen species production, callose accumulation, and the upregulated expression of defense response genes. Knockout of this gene in F. sacchari revealed a significant reduction in its pathogenicity, whereas the pathogenicity of the complementary mutant recovered to the wild-type levels, making this gene an important virulence factor for F. sacchari. In addition, the signal peptide of FsNis1 was required for the induction of cell death and PTI response in N. benthamiana. Thus, FsNis1 may not only be a key virulence factor for F. sacchari but may also induce defense responses in plants. These findings provide new insights into the function of Nis1 in host-pathogen interactions.
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Fusarium , Fusarium/genética , Imunidade Vegetal/genética , Virulência/genética , Fatores de Virulência/genética , Doenças das Plantas/genética , Doenças das Plantas/microbiologiaRESUMO
Sugarcane is a vital crop with significant economic and industrial value. However, the cultivated sugarcane's ultra-complex genome still needs to be resolved due to its high ploidy and extensive recombination between the two subgenomes. Here, we generate a chromosomal-scale, haplotype-resolved genome assembly for a hybrid sugarcane cultivar ZZ1. This assembly contains 10.4 Gb genomic sequences and 68,509 annotated genes with defined alleles in two sub-genomes distributed in 99 original and 15 recombined chromosomes. RNA-seq data analysis shows that sugar accumulation-associated gene families have been primarily expanded from the ZZSO subgenome. However, genes responding to pokkah boeng disease susceptibility have been derived dominantly from the ZZSS subgenome. The region harboring the possible smut resistance genes has expanded significantly. Among them, the expansion of WAK and FLS2 families is proposed to have occurred during the breeding of ZZ1. Our findings provide insights into the complex genome of hybrid sugarcane cultivars and pave the way for future genomics and molecular breeding studies in sugarcane.
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Saccharum , Saccharum/genética , Melhoramento Vegetal , Genômica , Haplótipos/genética , CromossomosRESUMO
Introduction: Plant-specific Class III peroxidases (PRXs) play a crucial role in lignification, cell elongation, seed germination, and biotic and abiotic stresses. Methods: The class III peroxidase gene family in sugarcane were identified by bioinformatics methods and realtime fluorescence quantitative PCR. Results: Eighty-two PRX proteins were characterized with a conserved PRX domain as members of the class III PRX gene family in R570 STP. The ShPRX family genes were divided into six groups by the phylogenetic analysis of sugarcane, Saccharum spontaneum, sorghum, rice, and Arabidopsis thaliana. The analysis of promoter cis-acting elements revealed that most ShPRX family genes contained cis-acting regulatory elements involved in ABA, MeJA, light responsiveness, anaerobic induction, and drought inducibility. An evolutionary analysis indicated that ShPRXs was formed after Poaceae and Bromeliaceae diverged, and tandem duplication events played a critical role in the expansion of ShPRX genes of sugarcane. Purifying selection maintained the function of ShPRX proteins. SsPRX genes were differentially expressed in stems and leaves at different growth stages in S. spontaneum. However, ShPRX genes were differentially expressed in the SCMV-inoculated sugarcane plants. A qRT-PCR analysis showed that SCMV, Cd, and salt could specifically induce the expression of PRX genes of sugarcane. Discussion: These results help elucidate the structure, evolution, and functions of the class III PRX gene family in sugarcane and provide ideas for the phytoremediation of Cd-contaminated soil and breeding new sugarcane varieties resistant to sugarcane mosaic disease, salt, and Cd stresses.
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Fusarium sacchari is one of the primary pathogens causing Pokkah Boeng disease (PBD) in sugarcane in China. Pectate lyases (PL), which play a critical role in pectin degradation and fungal virulence, have been extensively studied in major bacterial and fungal pathogens of a wide range of plant species. However, only a few PLs have been functionally investigated. In this study, we analyzed the function of the pectate lyase gene, FsPL, from F. sacchari. FsPL is a key virulence factor of F. sacchari and can induce plant cell death. FsPL also triggers the pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI) response in Nicotiana benthamiana, as reflected by increases in reactive oxygen species (ROS) production, electrolyte leakage, and callose accumulation, as well as the upregulation of defense response genes. In addition, our study also found that the signal peptide of FsPL was necessary for induced cell death and PTI responses. Virus-induced gene silencing showed that FsPL-induced cell death in Nicotiana benthamiana was mediated by leucine-rich repeat (LRR) receptor-like kinases BAK1 and SOBIR1. Thus, FsPL may not only be a critical virulence factor for F. sacchari but may also induce plant defense responses. These findings provide new insights into the functions of pectate lyase in host-pathogen interactions. IMPORTANCE Pokkah Boeng disease (PBD) is one of the main diseases affecting sugarcane in China, seriously damaging sugarcane production and economic development. Therefore, it is important to clarify the pathogenic mechanisms of this disease and to provide a theoretical basis for the breeding of PBD-resistant sugarcane strains. The present study aimed to analyze the function of FsPL, a recently identified pectate lyase gene from F. sacchari. FsPL is a key virulence factor of F. sacchari that induces plant cell death. Our results provide new insights into the function of pectate lyase in host-pathogen interactions.
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Nicotiana , Imunidade Vegetal , Virulência , Fatores de Virulência/genética , Doenças das Plantas/microbiologiaRESUMO
The lemon (Citrus limon; family Rutaceae) is one of the most important and popular fruits worldwide. Lemon also tolerates huanglongbing (HLB) disease, which is a devastating citrus disease. Here we produced a gap-free and haplotype-resolved chromosome-scale genome assembly of the lemon by combining Pacific Biosciences circular consensus sequencing, Oxford Nanopore 50-kb ultra-long, and high-throughput chromatin conformation capture technologies. The assembly contained nine-pair chromosomes with a contig N50 of 35.6 Mb and zero gaps, while a total of 633.0 Mb genomic sequences were generated. The origination analysis identified 338.5 Mb genomic sequences originating from citron (53.5%), 147.4 Mb from mandarin (23.3%), and 147.1 Mb from pummelo (23.2%). The genome included 30 528 protein-coding genes, and most of the assembled sequences were found to be repetitive sequences. Several significantly expanded gene families were associated with plant-pathogen interactions, plant hormone signal transduction, and the biosynthesis of major active components, such as terpenoids and flavor compounds. Most HLB-tolerant genes were expanded in the lemon genome, such as 2-oxoglutarate (2OG)/Fe(II)-dependent oxygenase and constitutive disease resistance 1, cell wall-related genes, and lignin synthesis genes. Comparative transcriptomic analysis showed that phloem regeneration and lower levels of phloem plugging are the elements that contribute to HLB tolerance in lemon. Our results provide insight into lemon genome evolution, active component biosynthesis, and genes associated with HLB tolerance.
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IMPORTANCE: Common fungal extracellular membrane (CFEM) domain-containing protein has long been considered an essential effector, playing a crucial role in the interaction of pathogens and plant. Strategies aimed at understanding the pathogenicity mechanism of F. sacchari are eagerly anticipated to ultimately end the spread of pokkah boeng disease. Twenty FsCFEM proteins in the genome of F. sacchari have been identified, and four FsCFEM effector proteins have been found to suppress BCL2-associated X protein-triggered programmed cell death in N. benthamiana. These four effector proteins have the ability to enter plant cells and inhibit plant immunity. Furthermore, the expression of these four FsCFEM effector proteins significantly increases during the infection stage, with the three of them playing an essential role in achieving full virulence. These study findings provide a direction toward further exploration of the immune response in sugarcane. By applying these discoveries, we can potentially control the spread of disease through techniques such as host-induced gene silencing.
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Proteínas Fúngicas , Proteínas de Membrana , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Virulência , Imunidade Vegetal , Doenças das Plantas/microbiologiaRESUMO
One of the causative agents of pokkah boeng disease (PBD), which affects sugarcane crops globally, is the fungus Fusarium sacchari. These fungal infections reduce sugar quality and yield, resulting in severe economic losses. Effector proteins play important roles in the interactions between pathogenic fungi and plants. Here, we used bioinformatic prediction approaches to identify 316 candidate secreted effector proteins (CSEPs) in the complete genome of F. sacchari. In total, 95 CSEPs contained known conserved structures, representing 40 superfamilies and 18 domains, while an additional 91 CSEPs contained seven known motifs. Of the 130 CSEPs containing no known domains or motifs, 14 contained one of four novel motifs. A heterogeneous expression system in Nicotiana benthamiana was used to investigate the functions of 163 CSEPs. Seven CSEPs suppressed BAX-triggered programmed cell death in N. benthamiana, while four caused cell death in N. benthamiana. The expression profiles of these eleven CSEPs during F. sacchari infection suggested that they may be involved in sugarcane-F. sacchari interaction. Our results establish a basis for further studies of the role of effector molecules in pathogen-sugarcane interactions, and provide a framework for future predictions of pathogen effector molecules.
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Huanglongbing (HLB; greening disease), caused by Candidatus Liberibacter asiaticus (CLas), is the most damaging citrus disease worldwide. The disease has spread throughout the citrus-producing regions of Guangxi, Guangdong, Fujian, and others in China. A total of 1,788 HLB-like symptomatic or asymptomatic samples were collected from the Guangxi and Fujian provinces of China to decipher the genetic diversity of CLas and its correlation with geographic region and host plant. The disease was the most severe in orange and the least in pomelo. CLas bacteria associated with the specific geographical and citrus variety infected more than 50% of the HLB-like symptomatic samples. We identified 6,286 minor variations by comparing 35 published CLas genomes and observed a highly heterogeneous variation distribution across the genome, including four highly diverse nonprophages and three prophage segments. Four hypervariable genomic regions (HGRs) were identified to determine the genetic diversity among the CLas isolates collected from Guangxi and Fujian, China. A phylogenetic tree constructed from four HGRs showed that 100 CLas strains could be separated into four distinct clades. Ten new strains with high variations of prophage regions were identified in the mandarin and tangerine grown in new plantation areas of Guangxi. Characterizing these HGR variations in the CLas bacteria may provide insight into their evolution and adaptation to host plants and insects. IMPORTANCE The hypervariable genomic regions derived from 35 published CLas genomes were used to decipher the genetic diversity of CLas strains and identify 10 new strains with high variations in prophage regions. Characterizing these variations in the CLas bacteria might provide insight into their evolution and adaptation to host plants and insects in China.
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Liberibacter , Rhizobiaceae , Animais , Filogenia , Rhizobiaceae/genética , China , Insetos , Genômica , Variação Genética , Doenças das Plantas/microbiologiaRESUMO
Fusarium sacchari and Fusarium andiyazi are two devastating sugarcane pathogens that cause pokkah boeng disease (PBD) in China. RNA_Seq was conducted to identify mycoviruses in F. sacchari and F. andiyazi isolates collected from PBD symptom-showing sugarcane plants across China. Fifteen isolates with a normal, debilitated, or abnormal phenotype in colony morphology were screened out for the existence of dsRNA from 104 Fusarium isolates. By sequencing the mixed pool of dsRNA from these Fusarium isolates, a total of 26 contigs representing complete or partial genome sequences of ten mycoviruses and their strains were identified, including one virus belonging to Hypoviridae, two mitoviruses with seven strains belonging to Narnaviridae, one virus of Chrysoviridae, and one alphavirus-like virus. RT-PCR amplification with primers specific to individual mycoviruses revealed that mitoviruses were the most prevalent and the alphavirus-like virus and chrysovirus were the least prevalent. In terms of host preference, more mitoviruses were found in F. andiyazi than in F. sacchari. Fusarium sacchari hypovirus 1 with a 13.9 kb genome and a defective genome of 12.2 kb, shares 54% identity at the amino acid level to the Wuhan insect virus 14, which is an unclassified hypovirus identified from insect meta-transcriptomics. The alphavirus-like virus, Fusarium sacchari alphavirus-like virus 1 (FsALV1), seemed to hold a distinct status amid fungal alphavirus-like viruses, with the highest identity of 27% at the amino acid level to Sclerotium rolfsii alphavirus-like virus 3 and 29% to a hepevirus, Ferret hepatitis E virus. While six of the seven mitoviruses shared 72-94% identities to known mitoviruses, Fusarium andiyazi mitovirus 2 was most similar to Alternaria brassicicola mitovirus with an identity of only 49% between the two viruses. Transmission of FsALV1 and Fusarium sacchari chrysovirus 1 (FsCV1) from F. sacharri to F. commune was observed and the characterization of the four-segment dsRNA chrysovirus was performed with aid of electron microscopy and analysis of the encapsidated RNAs. These findings provide insight into the diversity and spectrum of mycoviruses in PBD pathogens and should be useful for exploring agents to control the disease.
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Pokkah boeng, caused by Fusarium verticillioides, is a serious disease in sugarcane industry. The disease severity is related to the sugarcane genotype as well as environmental considerations, such as nitrogen application. The impact of the nitrogen source (ammonium sulfate, urea, or sodium nitrate) on sugarcane pokkah boeng disease and its pathogen was investigated in planta and fungal growth and sporulation production was measured in vitro. The results showed that ammonium and nitrate were beneficial to fungal mycelium growth, cell densities, and sporulation, which enhanced the disease symptoms of sugarcane pokkah boeng compared to urea fertilization. A total of 1,779 transcripts out of 13,999 annotated genes identified from global transcriptomic analysis were differentially expressed in F. verticillioides CNO-1 grown in the different sources of nitrogen. These were found to be involved in nitrogen metabolism, transport, and assimilation. Many of these genes were also associated with pathogenicity based on the PHI-base database. Several transcription factors were found to be associated with specific biological processes related to nitrogen utilization. Our results further demonstrated that nitrogen availability might play an important role in disease development by increasing fungal cell growth as well as influencing the expression of genes required for successful pathogenesis.