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1.
PLoS One ; 17(8): e0273481, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36037153

RESUMO

Dickeya solani is a soft rot bacterium with high virulence. In potato, D. solani, like the other potato-infecting soft rot bacteria, causes rotting and wilting of the stems and rotting of tubers in the field and in storage. Latent, asymptomatic infections of potato tubers are common in harvested tubers, and if the storage conditions are not optimal, the latent infection turns into active rotting. We characterized potato gene expression in artificially inoculated tubers in nonsymptomatic, early infections 1 and 24 hours post-inoculation (hpi) and compared the results to the response in symptomatic tuber tissue 1 week (168 hpi) later with RNA-Seq. In the beginning of the infection, potato tubers expressed genes involved in the detection of the bacterium through pathogen-associated molecular patterns (PAMPs), which induced genes involved in PAMPs-triggered immunity, resistance, production of pathogenesis-related proteins, ROS, secondary metabolites and salicylic acid (SA) and jasmonic acid (JA) biosynthesis and signaling genes. In the symptomatic tuber tissue one week later, the PAMPs-triggered gene expression was downregulated, whereas primary metabolism was affected, most likely leading to free sugars fueling plant defense but possibly also aiding the growth of the pathogen. In the symptomatic tubers, pectic enzymes and cell wall-based defenses were activated. Measurement of hormone production revealed increased SA concentration and almost no JA in the asymptomatic tubers at the beginning of the infection and high level of JA and reduced SA in the symptomatic tubers one week later. These findings suggest that potato tubers rely on different defense strategies in the different phases of D. solani infection even when the infection takes place in fully susceptible plants incubated in conditions leading to rotting. These results support the idea that D. solani is a biotroph rather than a true necrotroph.


Assuntos
Solanum tuberosum , Dickeya , Enterobacteriaceae/genética , Expressão Gênica , Padrões Moleculares Associados a Patógenos , Doenças das Plantas/genética , Doenças das Plantas/microbiologia , Reguladores de Crescimento de Plantas , Ácido Salicílico , Solanum tuberosum/microbiologia
2.
Microbiol Res ; 263: 127147, 2022 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-35914414

RESUMO

A variety of replication fork traps have recently been characterised in Enterobacterales, unveiling two different types of architecture. Of these, the degenerate type II fork traps are commonly found in Enterobacteriaceae such as Escherichia coli. The newly characterised type I fork traps are found almost exclusively outside Enterobacteriaceae within Enterobacterales and include several archetypes of possible ancestral architectures. Dickeya paradisiaca harbours a somewhat degenerate type I fork trap with a unique Ter1 adjacent to tus gene on one side of the circular chromosome and three putative Ter2-4 sites on the other side of the fork trap. The two innermost Ter1 and Ter2 sites are only separated by 18 kb, which is the shortest distance between two innermost Ter sites of any chromosomal fork trap identified so far. Of note, the dif site is located between these two sites, coinciding with a sharp GC-skew flip. Here we examined and compared the binding modalities of E. coli and D. paradisiaca Tus proteins for these Ter sites. Surprisingly, while Ter1-3 were functional, no significant Tus binding was observed for Ter4 even in low salt conditions, which is in stark contrast with the significant non-specific protein-DNA interactions that occur with E. coli Tus. Even more surprising was the finding that D. paradisiaca Tus has a relatively moderate binding affinity to double-stranded Ter while retaining an extremely high affinity to Ter-lock sequences. Our data revealed major differences in the salt resistance and stability between the D. paradisiaca and E. coli Tus protein complexes, suggesting that while Tus protein evolution can be quite flexible regarding the initial Ter binding step, it requires a highly stringent purifying selection for its final locked complex formation.


Assuntos
Replicação do DNA , Dickeya/metabolismo , Proteínas de Escherichia coli , Escherichia coli , Cromossomos/metabolismo , DNA Bacteriano/genética , DNA Bacteriano/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Escherichia coli/genética , Escherichia coli/metabolismo , Proteínas de Escherichia coli/metabolismo
3.
Mol Plant Pathol ; 23(10): 1487-1507, 2022 10.
Artigo em Inglês | MEDLINE | ID: mdl-35819797

RESUMO

Dickeya zeae is an aggressive bacterial phytopathogen that infects a wide range of host plants. It has been reported that integration host factor (IHF), a nucleoid-associated protein consisting of IHFα and IHFß subunits, regulates gene expression by influencing nucleoid structure and DNA bending. To define the role of IHF in the pathogenesis of D. zeae MS2, we deleted either and both of the IHF subunit encoding genes ihfA and ihfB, which significantly reduced the production of cell wall-degrading enzymes (CWDEs), an unknown novel phytotoxin and the virulence factor-modulating (VFM) quorum-sensing (QS) signal, cell motility, biofilm formation, and thereafter the infection ability towards both potato slices and banana seedlings. To characterize the regulatory pathways of IHF protein associated with virulence, IHF binding sites (consensus sequence 5'-WATCAANNNNTTR-3') were predicted and 272 binding sites were found throughout the genome. The expression of 110 tested genes was affected by IHF. Electrophoretic mobility shift assay (EMSA) showed direct interaction of IhfA protein with the promoters of vfmE, speA, pipR, fis, slyA, prtD, hrpL, hecB, hcp, indA, hdaA, flhD, pilT, gcpJ, arcA, arcB, and lysR. This study clarified the contribution of IHF in the pathogenic process of D. zeae by controlling the production of VFM and putrescine QS signals, phytotoxin, and indigoidine, the luxR-solo system, Fis, SlyA, and FlhD transcriptional regulators, and secretion systems from type I to type VI. Characterization of the regulatory networks of IHF in D. zeae provides a target for prevention and control of plant soft rot disease.


Assuntos
Dickeya , Enterobacteriaceae , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Dickeya/genética , Regulação Bacteriana da Expressão Gênica , Fatores Hospedeiros de Integração/genética , Fatores Hospedeiros de Integração/metabolismo , Virulência/genética , Fatores de Virulência/genética
4.
Water Res ; 220: 118724, 2022 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-35696807

RESUMO

Irrigation with surface water carrying plant pathogens poses a risk for agriculture. Managed aquifer recharge enhances fresh water availability while simultaneously it may reduce the risk of plant diseases by removal of pathogens during aquifer passage. We compared the transport of three plant pathogenic bacteria with Escherichia coli WR1 as reference strain in saturated laboratory column experiments filled with quartz sand, or sandy aquifer sediments. E. coli showed the highest removal, followed by Pectobacterium carotovorum, Dickeya solani and Ralstonia solanacearum. Bacterial and non-reactive tracer breakthrough curves were fitted with Hydrus-1D and compared with colloid filtration theory (CFT). Bacterial attachment to fine and medium aquifer sand under anoxic conditions was highest with attachment rates of max. katt1 = 765 day-1 and 355 day-1, respectively. Attachment was the least to quartz sand under oxic conditions (katt1 = 61 day-1). In CFT, sticking efficiencies were higher in aquifer than in quartz sand but there was no differentiation between fine and medium aquifer sand. Overall removal ranged between < 6.8 log10 m-1 in quartz and up to 40 log10 m-1 in fine aquifer sand. Oxygenation of the anoxic aquifer sediments for two weeks with oxic influent water decreased the removal. The results highlight the potential of natural sand filtration to sufficiently remove plant pathogenic bacteria during aquifer storage.


Assuntos
Filtração , Água Subterrânea , Quartzo , Dickeya/isolamento & purificação , Escherichia coli , Filtração/métodos , Sedimentos Geológicos , Água Subterrânea/microbiologia , Pectobacterium carotovorum/isolamento & purificação , Ralstonia solanacearum/isolamento & purificação , Areia , Água
5.
Sci Rep ; 12(1): 10725, 2022 06 24.
Artigo em Inglês | MEDLINE | ID: mdl-35750797

RESUMO

Resistance to bacteriophage infections protects bacteria in phage-replete environments, enabling them to survive and multiply in the presence of their viral predators. However, such resistance may confer costs for strains, reducing their ecological fitness as expressed as competitiveness for resources or virulence or both. There is limited knowledge about such costs paid by phage-resistant plant pathogenic bacteria in their natural habitats. This study analyzed the costs of phage resistance paid by the phytopathogenic pectinolytic bacterium Dickeya solani both in vitro and in potato (Solanum tuberosum L.) plants. Thirteen Tn5 mutants of D. solani IPO 2222 were identified that exhibited resistance to infection by lytic bacteriophage vB_Dsol_D5 (ΦD5). The genes disrupted in these mutants encoded proteins involved in the synthesis of bacterial envelope components (viz. LPS, EPS and capsule). Although phage resistance did not affect most of the phenotypes of ΦD5-resistant D. solani such as growth rate, production of effectors, swimming and swarming motility, use of various carbon and nitrogen sources and biofilm formation evaluated in vitro, all phage resistant mutants were significantly compromised in their ability to survive on leaf surfaces as well as to grow within and cause disease symptoms in potato plants.


Assuntos
Bacteriófagos , Solanum tuberosum , Bacteriófagos/genética , Dickeya , Enterobacteriaceae/genética , Doenças das Plantas/microbiologia , Solanum tuberosum/microbiologia
7.
mBio ; 13(3): e0052422, 2022 06 28.
Artigo em Inglês | MEDLINE | ID: mdl-35491820

RESUMO

Dickeya dadantii is a phytopathogenic bacterium that causes soft rot in a wide range of plant hosts worldwide and a model organism for studying virulence gene regulation. The present study provides a comprehensive and annotated transcriptomic map of D. dadantii obtained by a computational method combining five independent transcriptomic data sets: (i) paired-end RNA sequencing (RNA-seq) data for a precise reconstruction of the RNA landscape; (ii) DNA microarray data providing transcriptional responses to a broad variety of environmental conditions; (iii) long-read Nanopore native RNA-seq data for isoform-level transcriptome validation and determination of transcription termination sites; (iv) differential RNA sequencing (dRNA-seq) data for the precise mapping of transcription start sites; (v) in planta DNA microarray data for a comparison of gene expression profiles between in vitro experiments and the early stages of plant infection. Our results show that transcription units sometimes coincide with predicted operons but are generally longer, most of them comprising internal promoters and terminators that generate alternative transcripts of variable gene composition. We characterize the occurrence of transcriptional read-through at terminators, which might play a basal regulation role and explain the extent of transcription beyond the scale of operons. We finally highlight the presence of noncontiguous operons and excludons in the D. dadantii genome, novel genomic arrangements that might contribute to the basal coordination of transcription. The highlighted transcriptional organization may allow D. dadantii to finely adjust its gene expression program for a rapid adaptation to fast-changing environments. IMPORTANCE This is the first transcriptomic map of a Dickeya species. It may therefore significantly contribute to further progress in the field of phytopathogenicity. It is also one of the first reported applications of long-read Nanopore native RNA-seq in prokaryotes. Our findings yield insights into basal rules of coordination of transcription that might be valid for other bacteria and may raise interest in the field of microbiology in general. In particular, we demonstrate that gene expression is coordinated at the scale of transcription units rather than operons, which are larger functional genomic units capable of generating transcripts with variable gene composition for a fine-tuning of gene expression in response to environmental changes. In line with recent studies, our findings indicate that the canonical operon model is insufficient to explain the complexity of bacterial transcriptomes.


Assuntos
Enterobacteriaceae , Regulação Bacteriana da Expressão Gênica , Bactérias , Dickeya , Enterobacteriaceae/metabolismo
8.
mBio ; 13(3): e0025322, 2022 06 28.
Artigo em Inglês | MEDLINE | ID: mdl-35546537

RESUMO

The phytopathogenic proteobacterium Dickeya dadantii secretes an array of plant cell wall-degrading enzymes and other virulence factors via the type 2 secretion system (T2SS). T2SSs are widespread among important plant, animal, and human bacterial pathogens. This multiprotein complex spans the double membrane cell envelope and secretes fully folded proteins through a large outer membrane pore formed by 15 subunits of the secretin GspD. Secretins are also found in the type 3 secretion system and the type 4 pili. Usually, specialized lipoproteins termed pilotins assist the targeting and assembly of secretins into the outer membrane. Here, we show that in D. dadantii, the pilotin acts in concert with the scaffolding protein GspB. Deletion of gspB profoundly impacts secretin assembly, pectinase secretion, and virulence. Structural studies reveal that GspB possesses a conserved periplasmic homology region domain that interacts directly with the N-terminal secretin domain. Site-specific photo-cross-linking unravels molecular details of the GspB-GspD complex in vivo. We show that GspB facilitates outer membrane targeting and assembly of the secretin pores and anchors them to the inner membrane while the C-terminal extension of GspB provides a scaffold for the secretin channel in the peptidoglycan cell wall. Phylogenetic analysis shows that in other bacteria, GspB homologs vary in length and domain composition and act in concert with either a cognate ATPase GspA or the pilotin GspS. IMPORTANCE Gram-negative bacteria have two cell membranes sandwiching a peptidoglycan net that together form a robust protective cell envelope. To translocate effector proteins across this multilayer envelope, bacteria have evolved several specialized secretion systems. In the type 2 secretion system and some other bacterial machineries, secretins form large multimeric pores that allow transport of effector proteins or filaments across the outer membrane. The secretins are essential for nutrient acquisition and pathogenicity and constitute a target for development of new antibacterials. Targeting of secretin subunits into the outer membrane is often facilitated by a special class of lipoproteins called pilotins. Here, we show that in D. dadantii and some other bacteria, the scaffolding protein GspB acts in concert with pilotin, facilitating the assembly of the secretin pore and its anchoring to both the inner membrane and the bacterial cell wall. GspB homologs of varied domain composition are present in many other T2SSs.


Assuntos
Sistemas de Secreção Tipo II , Proteínas da Membrana Bacteriana Externa/metabolismo , Proteínas de Bactérias/metabolismo , Parede Celular/metabolismo , Dickeya , Enterobacteriaceae/metabolismo , Lipoproteínas/genética , Lipoproteínas/metabolismo , Peptidoglicano/metabolismo , Filogenia , Secretina/genética , Secretina/metabolismo , Sistemas de Secreção Tipo II/metabolismo
9.
PLoS One ; 17(4): e0265075, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35417462

RESUMO

Dickeya are plant pathogenic bacteria able to provoke disease on a wide range of plants. A type 2 secretion system (T2SS) named Out is necessary for Dickeya virulence. Previous studies showed that the D. dadantii T2SS secretes a wide range of plant cell wall degrading enzymes, including pectinases and a cellulase. However, the full repertoire of exoproteins it can secrete has probably not yet been identified. Secreted proteins possess a signal peptide and are first addressed to the periplasm before their recruitment by Out. T2SS-specific secretion signals remain unknown which prevents in silico identification of T2SS substrates. To identify new Out substrates, we analyzed D. dadantii transcriptome data obtained in plant infection condition and searched for genes strongly induced and encoding proteins with a signal sequence. We identified four new Out-secreted proteins: the expansin YoaJ, the putative virulence factor VirK and two proteins of the DUF 4879 family, SvfA and SvfB. We showed that SvfA and SvfB are required for full virulence of D. dadantii and that svf genes are present in a variable number of copies in other Pectobacteriaceae, up to three in D. fanghzongdai. This work opens the way to the study of the role of non-pectinolytic proteins secreted by the Out pathway in Pectobacteriaceae.


Assuntos
Gammaproteobacteria , Sistemas de Secreção Tipo II , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Dickeya , Enterobacteriaceae , Gammaproteobacteria/metabolismo , Doenças das Plantas/microbiologia , Sistemas de Secreção Tipo II/metabolismo , Fatores de Virulência/genética , Fatores de Virulência/metabolismo
10.
Mol Plant Pathol ; 23(8): 1187-1199, 2022 08.
Artigo em Inglês | MEDLINE | ID: mdl-35460168

RESUMO

Bacteria use signal transduction systems to sense and respond to their external environment. The two-component system CpxA/CpxR senses misfolded envelope protein stress and responds by up-regulating envelope protein factors and down-regulating virulence factors in several animal pathogens. Dickeya dadantii is a phytopathogen equipped with a type III secretion system (T3SS) for manipulating the host immune response. We found that deletion of cpxR enhanced the expression of the T3SS marker gene hrpA in a designated T3SS-inducing minimal medium (MM). In the ∆cpxR mutant, multiple T3SS and c-di-GMP regulators were also up-regulated. Subsequent analysis revealed that deletion of the phosphodiesterase gene egcpB in ∆cpxR abolished the enhanced T3SS expression. This suggested that CpxR suppresses EGcpB levels, causing low T3SS expression in MM. Furthermore, we found that the ∆cpxR mutant displayed low c-di-GMP phenotypes in biofilm formation and swimming. Increased production of cellular c-di-GMP by in trans expression of the diguanylate cyclase gene gcpA was negated in the ∆cpxR mutant. Here, we propose that CpxA/CpxR regulates T3SS expression by manipulating the c-di-GMP network, in turn modifying the multiple physiological activities involved in the response to environmental stresses in D. dadantii.


Assuntos
Proteínas de Bactérias , Regulação Bacteriana da Expressão Gênica , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , GMP Cíclico/análogos & derivados , GMP Cíclico/metabolismo , Dickeya , Enterobacteriaceae , Virulência/genética
11.
Microbiol Spectr ; 10(2): e0180521, 2022 04 27.
Artigo em Inglês | MEDLINE | ID: mdl-35352959

RESUMO

Dickeya dadantii is a phytopathogenic bacterium that causes diseases on a wide range of host plants. The pathogen secretes pectate lyases (Pel) through the type II secretion system (T2SS) that degrades the cell wall in host plants. The virulence of D. dadantii is controlled by the second messenger cyclic diguanylate monophosphate (c-di-GMP), and the homeostasis of c-di-GMP is maintained by a number of diguanylate cyclases and phosphodiesterases. Deletion of a phosphodiesterase ecpC repressed pelD transcription, and such repression can be suppressed by an additional deletion in vfmE. VfmE is an AraC type of transcriptional regulator in the Vfm quorum-sensing system. Our results suggest that VfmE is a c-di-GMP effector that functions as an activator of pel at low c-di-GMP concentrations and a repressor of pel at high c-di-GMP concentrations through regulation of the transcriptional activator SlyA. Multiple sequence alignment with known c-di-GMP effectors identified an RWIWR motif in VfmE that we demonstrate is required for the c-di-GMP binding. Mutation of R93D in the RxxxR motif eliminates the c-di-GMP-related phenotypes in Pel activity. Our results show that VfmE is not only a quorum-sensing regulator but also a c-di-GMP effector, suggesting that D. dadantii integrates the c-di-GMP signaling network with the Vfm quorum-sensing pathway during environmental adaptation. IMPORTANCE How bacteria integrate environmental cues from multiple sources to appropriately regulate adaptive phenotypes is a central question in microbiology. In Dickeya dadantii, the quorum-sensing regulator VfmE controls the key virulence factor pectate lyase (Pel). Here, we demonstrate that VfmE also binds to c-di-GMP, resulting in VfmE functioning as an activator of pel at low c-di-GMP concentrations and repressor of pel at high c-di-GMP concentrations. The RWIWR motif in VfmE is required for c-di-GMP binding, and mutation of the motif in the mutant R93D eliminates the c-di-GMP-related phenotypes in Pel activity. We propose that VfmE is an important mediator to integrate quorum-sensing signals with c-di-GMP to collectively regulate D. dadantii pathogenesis.


Assuntos
Proteínas de Bactérias , Regulação Bacteriana da Expressão Gênica , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , GMP Cíclico/análogos & derivados , GMP Cíclico/metabolismo , Dickeya , Enterobacteriaceae/metabolismo , Polissacarídeo-Liases
12.
Mol Plant Pathol ; 23(6): 870-884, 2022 06.
Artigo em Inglês | MEDLINE | ID: mdl-35254732

RESUMO

Dickeya oryzae is a bacterial pathogen causing the severe rice stem rot disease in China and other rice-growing countries. We showed recently that the universal bacterial second messenger c-di-GMP plays an important role in modulation of bacterial motility and pathogenicity, but the mechanism of regulation remains unknown. In this study, bioinformatics analysis of the D. oryzae EC1 genome led to the identification of two proteins, YcgR and BcsA, both of which contain a conserved c-di-GMP receptor domain, known as the PilZ-domain. By deleting all the genes encoding c-di-GMP-degrading enzymes in D. oryzae EC1, the resultant mutant 7ΔPDE with high c-di-GMP levels became nonmotile, formed hyperbiofilm, and lost the ability to colonize and invade rice seeds. These phenotypes were partially reversed by deletion of ycgR in the mutant 7ΔPDE, whereas deletion of bcsA only reversed the hyperbiofilm phenotype of mutant 7ΔPDE. Significantly, double deletion of ycgR and bcsA in mutant 7ΔPDE rescued its motility, biofilm formation, and virulence to levels of wild-type EC1. In vitro biochemical experiments and in vivo phenotypic assays further validated that YcgR and BcsA proteins are the receptors for c-di-GMP, which together play a critical role in regulating the c-di-GMP-associated functionality. The findings from this study fill a gap in our understanding of how c-di-GMP modulates bacterial motility and biofilm formation, and provide useful clues for further elucidation of sophisticated virulence regulatory mechanisms in this important plant pathogen.


Assuntos
Dickeya , Oryza , Proteínas de Bactérias/metabolismo , Proteínas de Transporte/metabolismo , GMP Cíclico/análogos & derivados , GMP Cíclico/metabolismo , Regulação Bacteriana da Expressão Gênica , Oryza/microbiologia , Fenótipo , Virulência
13.
PLoS One ; 17(2): e0263124, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35192621

RESUMO

Dickeya solani is a pathogen most frequently responsible for infecting potato plants in Europe. As in the case of most plant pathogens, its ability to colonize and invade the host depends on chemotaxis and motility. The coordinated movement of Dickeya over solid surfaces is governed by a quorum sensing mechanism. In D. solani motility is regulated by ExpI-ExpR proteins, homologous to luxI-luxR system from Vibrio fisheri, in which N-acyl-homoserine lactones (AHLs) serve as signaling molecules. Moreover, in many Gram-negative bacteria motility is coupled with central metabolism via carbon catabolite repression. This enables them to reach more nutrient-efficient niches. The aim of this study was to analyze the swarming motility of D. solani depending on the volume of the medium in the cultivation plate and glucose content. We show that the ability of this bacterium to move is strictly dependent on both these factors. Moreover, we analyze the production of AHLs and show that the quorum sensing mechanism in D. solani is also influenced by the availability of glucose in the medium and that the distribution of these signaling molecules are different depending on the volume of the medium in the plate.


Assuntos
Acil-Butirolactonas/farmacologia , Proteínas de Bactérias/genética , Dickeya/efeitos dos fármacos , Glucose/farmacologia , Solanum tuberosum/microbiologia , Fatores de Virulência/genética , Acil-Butirolactonas/metabolismo , Proteínas de Bactérias/metabolismo , Quimiotaxia/efeitos dos fármacos , Quimiotaxia/genética , Meios de Cultura/química , Meios de Cultura/farmacologia , Dickeya/genética , Dickeya/metabolismo , Dickeya/patogenicidade , Regulação Bacteriana da Expressão Gênica , Glucose/metabolismo , Doenças das Plantas/microbiologia , Percepção de Quorum/efeitos dos fármacos , Percepção de Quorum/genética , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Transativadores/genética , Transativadores/metabolismo , Fatores de Virulência/metabolismo
14.
Microbiol Spectr ; 10(1): e0159021, 2022 02 23.
Artigo em Inglês | MEDLINE | ID: mdl-35107329

RESUMO

Dickeya zeae is a worldwide destructive pathogen that causes soft rot diseases on various hosts such as rice, maize, banana, and potato. The strain JZL7 we recently isolated from clivia represents the first monocot-specific D. zeae and also has reduced pathogenicity compared to that of other D. zeae strains (e.g., EC1 and MS2). To elucidate the molecular mechanisms underlying its more restricted host range and weakened pathogenicity, we sequenced the complete genome of JZL7 and performed comparative genomic and functional analyses of JZL7 and other D. zeae strains. We found that, while having the largest genome among D. zeae strains, JZL7 lost almost the entire type III secretion system (T3SS), which is a key component of the virulence suite of many bacterial pathogens. Importantly, the deletion of T3SS in MS2 substantially diminished the expression of most type III secreted effectors (T3SEs) and MS2's pathogenicity on both dicots and monocots. Moreover, although JZL7 and MS2 share almost the same repertoire of cell wall-degrading enzymes (CWDEs), we found broad reduction in the production of CWDEs and expression levels of CWDE genes in JZL7. The lower expression of CWDEs, pectin lyases in particular, would probably make it difficult for JZL7 to break down the cell wall of dicots, which is rich in pectin. Together, our results suggest that the loss of T3SS and reduced CWDE activity together might have contributed to the host specificity and virulence of JZL7. Our findings also shed light on the pathogenic mechanism of Dickeya and other soft rot Pectobacteriaceae species in general. IMPORTANCE Dickeya zeae is an important, aggressive bacterial phytopathogen that can cause severe diseases in many crops and ornamental plants, thus leading to substantial economic losses. Strains from different sources showed significant diversity in their natural hosts, suggesting complicated evolution history and pathogenic mechanisms. However, molecular mechanisms that cause the differences in the host range of D. zeae strains remain poorly understood. This study carried out genomic and functional dissections of JZL7, a D. zeae strain with restricted host range, and revealed type III secretion system (T3SS) and cell wall-degrading enzymes (CWDEs) as two major factors contributing to the host range and virulence of D. zeae, which will provide a valuable reference for the exploration of pathogenic mechanisms in other bacteria and present new insights for the control of bacterial soft rot diseases on crops.


Assuntos
Proteínas de Bactérias/metabolismo , Parede Celular/microbiologia , Produtos Agrícolas/microbiologia , Dickeya/genética , Dickeya/patogenicidade , Especificidade de Hospedeiro , Sistemas de Secreção Tipo III/metabolismo , Proteínas de Bactérias/genética , Parede Celular/metabolismo , Produtos Agrícolas/metabolismo , Dickeya/enzimologia , Dickeya/fisiologia , Genoma Bacteriano , Filogenia , Doenças das Plantas/microbiologia , Sistemas de Secreção Tipo III/genética , Virulência
15.
Mol Plant Microbe Interact ; 35(5): 369-379, 2022 May.
Artigo em Inglês | MEDLINE | ID: mdl-35100009

RESUMO

The GacS-GacA type two-component system (TCS) positively regulates pathogenicity-related phenotypes in many plant pathogens. In addition, Dickeya oryzae EC1, the causative agent of soft rot disease, produces antibiotic-like toxins called zeamines as one of the major virulence factors that inhibit the germination of rice seeds. The present study identified a GacS-GacA type TCS, named TzpS-TzpA, that positively controls the virulence of EC1, mainly by regulating production of the toxin zeamines. RNA-seq analysis of strain EC1 and its tzpA mutant showed that the TCS regulated a wide range of virulence genes, especially those encoding zeamines. Protein-protein interaction was detected between TzpS and TzpA through the bacterial two-hybrid system and pull-down assay. In trans expression of tzpA failed to rescue the defective phenotypes in both the ΔtzpS and ΔtzpSΔtzpA mutants. Furthermore, TzpA controls target gene expression by direct binding to DNA promoters that contain a Gac-box motif, including a regulatory RNA rsmB and the vfm quorum-sensing system regulator vfmE. These findings therefore suggested that the EC1 TzpS-TzpA TCS system mediates the pathogenicity of Dickeya oryzae EC1 mainly by regulating the production of zeamines.[Formula: see text] Copyright © 2022 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.


Assuntos
Proteínas de Bactérias , Dickeya , Proteínas de Bactérias/metabolismo , Regulação Bacteriana da Expressão Gênica , Macrolídeos , Doenças das Plantas/microbiologia , Poliaminas , Virulência/genética
16.
Int J Mol Sci ; 23(2)2022 Jan 09.
Artigo em Inglês | MEDLINE | ID: mdl-35054882

RESUMO

In bacteria, the DsbA oxidoreductase is a crucial factor responsible for the introduction of disulfide bonds to extracytoplasmic proteins, which include important virulence factors. A lack of proper disulfide bonds frequently leads to instability and/or loss of protein function; therefore, improper disulfide bonding may lead to avirulent phenotypes. The importance of the DsbA function in phytopathogens has not been extensively studied yet. Dickeya solani is a bacterium from the Soft Rot Pectobacteriaceae family which is responsible for very high economic losses mainly in potato. In this work, we constructed a D. solani dsbA mutant and demonstrated that a lack of DsbA caused a loss of virulence. The mutant bacteria showed lower activities of secreted virulence determinants and were unable to develop disease symptoms in a potato plant. The SWATH-MS-based proteomic analysis revealed that the dsbA mutation led to multifaceted effects in the D. solani cells, including not only lower levels of secreted virulence factors, but also the induction of stress responses. Finally, the outer membrane barrier seemed to be disturbed by the mutation. Our results clearly demonstrate that the function played by the DsbA oxidoreductase is crucial for D. solani virulence, and a lack of DsbA significantly disturbs cellular physiology.


Assuntos
Dickeya/enzimologia , Isomerases de Dissulfetos de Proteínas , Virulência , Proteínas de Bactérias , Dickeya/patogenicidade , Oxirredutases , Proteínas Periplásmicas , Proteômica
17.
Environ Microbiol ; 24(3): 1467-1483, 2022 03.
Artigo em Inglês | MEDLINE | ID: mdl-35014170

RESUMO

The Vfm quorum sensing (QS) system is preponderant for the virulence of different species of the bacterial genus Dickeya. The vfm gene cluster encodes 26 genes involved in the production, sensing or transduction of the QS signal. To date, the Vfm QS signal has escaped detection by analytical chemistry methods. However, we report here a strain-specific polymorphism in the biosynthesis genes vfmO and vfmP, which is predicted to be related to the production of different analogues of the QS signal. Consequently, the Vfm communication could be impossible between strains possessing different variants of the genes vfmO/P. We constructed three Vfm QS biosensor strains possessing different vfmO/P variants and compared these biosensors for their responses to samples prepared from 34 Dickeya strains possessing different vfmO/P variants. A pattern of specificity was demonstrated, providing evidence that the polymorphism in the genes vfmO/P determines the biosynthesis of different analogues of the QS signal. Unexpectedly, this vfmO/P-dependent pattern of specificity is linked to a polymorphism in the ABC transporter gene vfmG, suggesting an adaptation of the putative permease VfmG to specifically bind different analogues of the QS signal. Accordingly, we discuss the possible involvement of VfmG as co-sensor of the Vfm two-component regulatory system.


Assuntos
Proteínas de Bactérias , Percepção de Quorum , Bactérias/metabolismo , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Dickeya , Regulação Bacteriana da Expressão Gênica , Polimorfismo Genético , Percepção de Quorum/genética
18.
J Biol Chem ; 298(1): 101446, 2022 01.
Artigo em Inglês | MEDLINE | ID: mdl-34826421

RESUMO

The catabolism of pectin from plant cell walls plays a crucial role in the virulence of the phytopathogen Dickeya dadantii. In particular, the timely expression of pel genes encoding major pectate lyases is essential to circumvent the plant defense systems and induce massive pectinolytic activity during the maceration phase. Previous studies identified the role of a positive feedback loop specific to the pectin-degradation pathway, whereas the precise signals controlling the dynamics of pectate lyase expression were unclear. Here, we show that the latter is controlled by a metabolic switch involving both glucose and pectin. We measured the HPLC concentration profiles of the key metabolites related to these two sources of carbon, cAMP and 2-keto-3-deoxygluconate, and developed a dynamic and quantitative model of the process integrating the associated regulators, cAMP receptor protein and KdgR. The model describes the regulatory events occurring at the promoters of two major pel genes, pelE and pelD. It highlights that their activity is controlled by a mechanism of carbon catabolite repression, which directly controls the virulence of D. dadantii. The model also shows that quantitative differences in the binding properties of common regulators at these two promoters resulted in a qualitatively different role of pelD and pelE in the metabolic switch, and also likely in conditions of infection, justifying their evolutionary conservation as separate genes in this species.


Assuntos
Repressão Catabólica , Dickeya , Pectinas , Proteínas de Bactérias/metabolismo , Dickeya/metabolismo , Digestão , Enterobacteriaceae/metabolismo , Regulação Bacteriana da Expressão Gênica , Pectinas/metabolismo , Polissacarídeo-Liases/química
19.
Mol Plant Pathol ; 23(1): 45-59, 2022 01.
Artigo em Inglês | MEDLINE | ID: mdl-34693617

RESUMO

Dickeya zeae is the causal agent of rice foot rot disease. The pathogen is known to rely on a range of virulence factors, including phytotoxin zeamines, extracellular enzymes, cell motility, and biofilm, which collectively contribute to the establishment of infections. Phytotoxin zeamines play a critical role in bacterial virulence; signalling pathways and regulatory mechanisms that govern bacterial virulence remain unclear. In this study, we identified a transcriptional regulator OhrR (organic hydroperoxide reductase regulator) that is involved in the regulation of zeamine production in D. zeae EC1. The OhrR null mutant was significantly attenuated in its virulence against rice seed, potato tubers and radish roots. Phenotype analysis showed that OhrR was also involved in the regulation of other virulence traits, including the production of extracellular cellulase, biofilm formation, and swimming/swarming motility. DNA electrophoretic mobility shift assay showed that OhrR directly regulates the transcription of key virulence genes and genes encoding bis-(3'-5')-cyclic dimeric guanosine monophosphate synthetases. Furthermore, OhrR positively regulates the transcription of regulatory genes slyA and fis through binding to their promoter regions. Our findings identify a key regulator of the virulence of D. zeae and add new insights into the complex regulatory network that modulates the physiology and virulence of D. zeae.


Assuntos
Proteínas de Bactérias , Doenças das Plantas , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Dickeya , Enterobacteriaceae/metabolismo , Regulação Bacteriana da Expressão Gênica , Virulência/genética
20.
Int J Syst Evol Microbiol ; 71(11)2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34726587

RESUMO

The genus Dickeya comprises plant pathogens that cause diseases in a large range of economically important crops and ornamentals. Strains previously assigned to the species Dickeya zeae are major pathogens attacking vital crops such as maize and rice. They are also frequently isolated from surface water. The newly described species Dickeya oryzae is closely related to D. zeae members, so that the limit between the two species can be difficult to define. In order to clearly distinguish the two species, globally described by the term 'D. zeae complex', we sequenced the genome of four new water isolates and compared them to 14 genomes available in databases. Calculation of average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) values confirmed the phylogenomic classification into the two species D. zeae and D. oryzae. It also allowed us to propose a new species, Dickeya parazeae sp. nov., to characterize a clade distinct from those containing the D. zeae type strain NCPPB2538T. Strain S31T (CFBP 8716T=LMG 32070T) isolated from water in France is proposed as the type strain of the new species. Phenotypic analysis of eight publically available strains revealed traits common to the five tested D. oryzae members but apparently not shared by the D. oryzae type strain. Genomic analyses indicated that a simple distinction between the species D. zeae, D. parazeae and D. oryzae can be obtained on the basis of the recA sequence. D. oryzae can be distinguished from the two other species by growth on l-tartaric acid. Based on the recA marker, several strains previously identified as D. zeae were re-assigned to the species D. parazeae or D. oryzae. This study also highlighted the broad host range diversity of these three species.


Assuntos
Dickeya , Filogenia , Doenças das Plantas/microbiologia , Técnicas de Tipagem Bacteriana , Composição de Bases , DNA Bacteriano/genética , Dickeya/classificação , Dickeya/isolamento & purificação , França , Hibridização de Ácido Nucleico , RNA Ribossômico 16S/genética , Análise de Sequência de DNA
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