RESUMO
Benanti et al. report that Burkholderia pseudomallei and Burkholderia mallei bacteria express proteins that mimic Ena/Vasp family proteins to polymerize actin, thereby inducing actin-based motility. Thus, bacteria can use the various cellular actin polymerization mechanisms for intra- and inter-cellular dissemination.
Assuntos
Actinas/metabolismo , Infecções por Burkholderia/microbiologia , Burkholderia/fisiologia , Burkholderia/patogenicidade , Moléculas de Adesão Celular/metabolismo , Proteínas dos Microfilamentos/metabolismo , Fosfoproteínas/metabolismo , Animais , HumanosRESUMO
Burkholderia pseudomallei and B. mallei are bacterial pathogens that cause melioidosis and glanders, whereas their close relative B. thailandensis is non-pathogenic. All use the trimeric autotransporter BimA to facilitate actin-based motility, host cell fusion, and dissemination. Here, we show that BimA orthologs mimic different host actin-polymerizing proteins. B. thailandensis BimA activates the host Arp2/3 complex. In contrast, B. pseudomallei and B. mallei BimA mimic host Ena/VASP actin polymerases in their ability to nucleate, elongate, and bundle filaments by associating with barbed ends, as well as in their use of WH2 motifs and oligomerization for activity. Mechanistic differences among BimA orthologs resulted in distinct actin filament organization and motility parameters, which affected the efficiency of cell fusion during infection. Our results identify bacterial Ena/VASP mimics and reveal that pathogens imitate the full spectrum of host actin-polymerizing pathways, suggesting that mimicry of different polymerization mechanisms influences key parameters of infection.
Assuntos
Actinas/metabolismo , Infecções por Burkholderia/microbiologia , Burkholderia/fisiologia , Burkholderia/patogenicidade , Moléculas de Adesão Celular/metabolismo , Proteínas dos Microfilamentos/metabolismo , Fosfoproteínas/metabolismo , Sequência de Aminoácidos , Animais , Burkholderia/classificação , Burkholderia/enzimologia , Células COS , Fusão Celular , Linhagem Celular Tumoral , Chlorocebus aethiops , Células HEK293 , Humanos , Dados de Sequência Molecular , Alinhamento de SequênciaRESUMO
Burkholderia glumae causes bacterial leaf blight in rice, and its global spread has been exacerbated by climate change. To understand the genetic diversity and virulence of B. glumae strains isolated from rice cultivars in Perú, 47 isolates were obtained from infected rice fields, all belonging to B. glumae, and confirmed by recA and toxB sequences. The BOX-PCR typing group has 38 genomic profiles, and these turn into seven variable number tandem repeats (VNTR) haplotypes. There was no correlation between clustering and geographical origin. Nineteen strains were selected for phenotypic characterization and virulence, using both the maceration level of the onion bulb proxy and inoculation of seeds of two rice cultivars. Several strains produced pigments other than toxoflavin, which correlated with onion bulb maceration. In terms of virulence at the seed level, all strains produced inhibition at the root and coleoptile level, but the severity of symptoms varied significantly between strains, revealing significant differences in pathogenicity. There is no correlation between maceration and virulence scores, probably reflecting different virulence mechanisms depending on the host infection stage. This is the first study to evaluate the VNTR diversity and virulence of Peruvian strains of B. glumae in two commercial cultivars.
Assuntos
Burkholderia , Variação Genética , Oryza , Doenças das Plantas , Oryza/microbiologia , Burkholderia/genética , Burkholderia/patogenicidade , Burkholderia/isolamento & purificação , Doenças das Plantas/microbiologia , Virulência/genética , Filogenia , Repetições MinissatélitesRESUMO
Interbacterial antagonism and communication are driving forces behind microbial community development. In many Gram-negative bacteria, contact-dependent growth inhibition (CDI) systems contribute to these microbial interactions. CDI systems deliver the toxic C-terminus of a large surface exposed protein to the cytoplasm of neighboring bacteria upon cell-contact. Termed the BcpA-CT, import of this toxic effector domain is mediated by specific, yet largely unknown receptors on the recipient cell outer and inner membranes. In this study, we demonstrated that cytoplasmic membrane proteins GltJK, components of a predicted ABC-type transporter, are required for entry of CDI system protein BcpA-2 into Burkholderia multivorans recipient cells. Consistent with current CDI models, gltJK were also required for recipient cell susceptibility to a distinct BcpA-CT that shared sequences within the predicted "translocation domain" of BcpA-2. Strikingly, this translocation domain showed low sequence identity to the analogous region of an Escherichia coli GltJK-utilizing CDI system protein. Our results demonstrated that recipient bacteria expressing E. coli gltJK were resistant to BcpA-2-mediated interbacterial antagonism, suggesting that BcpA-2 specifically recognizes Burkholderia GltJK. Using a series of chimeric proteins, the specificity determinant was mapped to Burkholderia-specific sequences at the GltK C-terminus, providing insight into BcpA transport across the recipient cell cytoplasmic membrane.
Assuntos
Proteínas de Bactérias/fisiologia , Burkholderia/fisiologia , Proteínas de Membrana/fisiologia , Interações Microbianas , Aderência Bacteriana , Fenômenos Fisiológicos Bacterianos , Biofilmes/crescimento & desenvolvimento , Burkholderia/patogenicidade , Escherichia coli/genética , Proteínas de Escherichia coli/genética , Humanos , Mutagênese Insercional/métodos , Domínios Proteicos , Especificidade da EspécieRESUMO
One of the most commonly prescribed antibiotics against Burkholderia infections is co-trimoxazole, a cocktail of trimethoprim and sulfamethoxazole. Trimethoprim elicits an upregulation of the mal gene cluster, which encodes proteins involved in synthesis of the cytotoxic polyketide malleilactone; trimethoprim does so by increasing expression of the malR gene, which encodes the activator MalR. We report that B. thailandensis grown on trimethoprim exhibited increased virulence against Caenorhabditis elegans. This enhanced virulence correlated with an increase in expression of the mal gene cluster. Notably, inhibition of xanthine dehydrogenase by addition of allopurinol led to similar upregulation of malA and malR, with addition of trimethoprim or allopurinol also resulting in an equivalent intracellular accumulation of xanthine. Xanthine is a ligand for the transcription factor MftR that leads to attenuated DNA binding, and we show using chromatin immunoprecipitation that MftR binds directly to malR. Our gene expression data suggest that malR expression is repressed by both MftR and by a separate transcription factor, which also responds to a metabolite that accumulates on exposure to trimethoprim. Since allopurinol elicits a similar increase in malR/malA expression as trimethoprim, we suggest that impaired purine homeostasis plays a primary role in trimethoprim-mediated induction of malR and in turn malA.
Assuntos
Proteínas de Bactérias/fisiologia , Burkholderia/efeitos dos fármacos , Burkholderia/fisiologia , Caenorhabditis elegans/microbiologia , Regulação Bacteriana da Expressão Gênica , Purinas/metabolismo , Proteínas Repressoras/fisiologia , Trimetoprima/farmacologia , Animais , Antibacterianos/farmacologia , Burkholderia/patogenicidade , Infecções por Burkholderia/microbiologia , Homeostase , Família Multigênica , Sulfametoxazol/farmacologia , Fatores de Transcrição/metabolismo , Combinação Trimetoprima e Sulfametoxazol/farmacologia , Virulência , Xantina/metabolismoRESUMO
Bacterial bloodstream infections (BSI) are a major health concern and can cause up to 40% mortality. Pseudomonas aeruginosa BSI is often of nosocomial origin and is associated with a particularly poor prognosis. The mechanism of bacterial persistence in blood is still largely unknown. Here, we analyzed the behavior of a cohort of clinical and laboratory Pseudomonas aeruginosa strains in human blood. In this specific environment, complement was the main defensive mechanism, acting either by direct bacterial lysis or by opsonophagocytosis, which required recognition by immune cells. We found highly variable survival rates for different strains in blood, whatever their origin, serotype, or the nature of their secreted toxins (ExoS, ExoU or ExlA) and despite their detection by immune cells. We identified and characterized a complement-tolerant subpopulation of bacterial cells that we named "evaders". Evaders shared some features with bacterial persisters, which tolerate antibiotic treatment. Notably, in bi-phasic killing curves, the evaders represented 0.1-0.001% of the initial bacterial load and displayed transient tolerance. However, the evaders are not dormant and require active metabolism to persist in blood. We detected the evaders for five other major human pathogens: Acinetobacter baumannii, Burkholderia multivorans, enteroaggregative Escherichia coli, Klebsiella pneumoniae, and Yersinia enterocolitica. Thus, the evaders could allow the pathogen to persist within the bloodstream, and may be the cause of fatal bacteremia or dissemination, in particular in the absence of effective antibiotic treatments.
Assuntos
Infecções Bacterianas/sangue , Infecções Bacterianas/imunologia , Ativação do Complemento/imunologia , Acinetobacter baumannii/crescimento & desenvolvimento , Acinetobacter baumannii/patogenicidade , Bacteriemia/sangue , Bacteriemia/imunologia , Bacteriemia/microbiologia , Bactérias , Burkholderia/crescimento & desenvolvimento , Burkholderia/patogenicidade , Proteínas do Sistema Complemento/imunologia , Escherichia coli/crescimento & desenvolvimento , Escherichia coli/patogenicidade , Humanos , Klebsiella pneumoniae/crescimento & desenvolvimento , Klebsiella pneumoniae/patogenicidade , Testes de Sensibilidade Microbiana , Infecções por Pseudomonas/sangue , Infecções por Pseudomonas/imunologia , Infecções por Pseudomonas/microbiologia , Pseudomonas aeruginosa/crescimento & desenvolvimento , Pseudomonas aeruginosa/patogenicidade , Yersinia enterocolitica/crescimento & desenvolvimento , Yersinia enterocolitica/patogenicidadeRESUMO
Burkholderia cepacia complex bacteria comprise opportunistic pathogens causing chronic respiratory infections in cystic fibrosis (CF) patients. These microorganisms produce an exopolysaccharide named cepacian, which is considered a virulence determinant. To find genes implicated in the regulation of cepacian biosynthesis, we characterized an evolved nonmucoid variant (17616nmv) derived from the ancestor, Burkholderia multivorans ATCC 17616, after prolonged stationary phase. Lack of cepacian biosynthesis was correlated with downregulation of the expression of bce genes implicated in its biosynthesis. Furthermore, genome sequencing of the variant identified the transposition of the mobile element IS406 upstream of the coding sequence of an hns-like gene (Bmul_0158) encoding a histone-like nucleoid structuring (H-NS) protein, a known global transcriptional repressor. This insertion sequence (IS) element upregulated the expression of Bmul_0158 by 4-fold. Transcriptome analysis identified the global effects of this mutation on gene expression, with major changes in genes implicated in motility, pilus synthesis, type VI secretion, and chromosome-associated functions. Concomitant with these differences, the nonmucoid variant displays reduced adherence to a CF lung bronchial cell line and reduced surface hydrophobicity and forms smaller cellular aggregates but has an increase in swimming and swarming motilities. Finally, analysis of the GC content of the upstream region of differentially expressed genes led to the identification of various genomic regions, possibly acquired by horizontal gene transfer, which were transcriptionally repressed by the increased expression of the Bmul_0158 gene in the 17616nmv strain. Taken together, the results revealed a significant role for this H-NS protein in the regulation of B. multivorans persistence- and virulence-associated genes. IMPORTANCE Members of the histone-like nucleoid structuring (H-NS) family of proteins, present in many bacteria, are important global regulators of gene expression. Many of the regulated genes were acquired horizontally and include pathogenicity islands and prophages, among others. Additionally, H-NS can play a structural role by bridging and compacting DNA, fulfilling a crucial role in cell physiology. Several virulence phenotypes have been frequently identified in several bacteria as dependent on H-NS activity. Here, we describe an H-NS-like protein of the opportunistic pathogen Burkholderia multivorans, a species commonly infecting the respiratory tract of cystic fibrosis patients. Our results indicate that this protein is involved in regulating virulence traits such as exopolysaccharide biosynthesis, adhesion to biotic surfaces, cellular aggregation, and motility. Furthermore, this H-NS-like protein is one out of eight orthologs present in the B. multivorans ATCC 17616 genome, posing relevant questions to be investigated on how these proteins coordinate the expression of virulence traits.
Assuntos
Proteínas de Bactérias/genética , Burkholderia/genética , Burkholderia/patogenicidade , Virulência/genética , Aderência Bacteriana , Burkholderia/fisiologia , Agregação Celular , Linhagem Celular , Regulação Bacteriana da Expressão Gênica , Genes Bacterianos , Genoma Bacteriano , Histonas , Humanos , Interações Hidrofóbicas e Hidrofílicas , Fenótipo , Polissacarídeos Bacterianos/biossínteseRESUMO
Rice is an important source of food for more than half of the world's population. Bacterial panicle blight (BPB) is a disease of rice characterized by grain discoloration or sheath rot caused mainly by Burkholderia glumae. B. glumae synthesizes toxoflavin, an essential virulence factor that is required for symptoms of the disease. The products of the tox operons, ToxABCDE and ToxFGHI, are responsible for the synthesis and the proton motive force (PMF)-dependent secretion of toxoflavin, respectively. The DedA family is a highly conserved membrane protein family found in most bacterial genomes that likely function as membrane transporters. Our previous work has demonstrated that absence of certain DedA family members results in pleiotropic effects, impacting multiple pathways that are energized by PMF. We have demonstrated that a member of the DedA family from Burkholderia thailandensis, named DbcA, is required for the extreme polymyxin resistance observed in this organism. B. glumae encodes a homolog of DbcA with 73% amino acid identity to Burkholderia thailandensis DbcA. Here, we created and characterized a B. glumae ΔdbcA strain. In addition to polymyxin sensitivity, the B. glumae ΔdbcA strain is compromised for virulence in several BPB infection models and secretes only low amounts of toxoflavin (â¼15% of wild-type levels). Changes in membrane potential in the B. glumae ΔdbcA strain were reproduced in the wild-type strain by the addition of subinhibitory concentrations of sodium bicarbonate, previously demonstrated to cause disruption of PMF. Sodium bicarbonate inhibited B. glumae virulence in rice, suggesting a possible non-toxic chemical intervention for bacterial panicle blight. IMPORTANCE Bacterial panicle blight (BPB) is a disease of rice characterized by grain discoloration or sheath rot caused mainly by Burkholderia glumae. The DedA family is a highly conserved membrane protein family found in most bacterial genomes that likely function as membrane transporters. Here, we constructed a B. glumae mutant with a deletion in a DedA family member named dbcA and report a loss of virulence in models of BPB. Physiological analysis of the mutant shows that the proton motive force is disrupted, leading to reduction of secretion of the essential virulence factor toxoflavin. The mutant phenotypes are reproduced in the virulent wild-type strain without an effect on growth using sodium bicarbonate, a nontoxic buffer that has been reported to disrupt the PMF. The results presented here suggest that bicarbonate may be an effective antivirulence agent capable of controlling BPB without imposing an undue burden on the environment.
Assuntos
Burkholderia , Oryza/microbiologia , Doenças das Plantas/microbiologia , Força Próton-Motriz , Antibacterianos/farmacologia , Proteínas de Bactérias/genética , Burkholderia/efeitos dos fármacos , Burkholderia/genética , Burkholderia/metabolismo , Burkholderia/patogenicidade , Carbonil Cianeto m-Clorofenil Hidrazona/farmacologia , Cebolas/microbiologia , Pirimidinonas/metabolismo , Bicarbonato de Sódio/farmacologia , Triazinas/metabolismo , Virulência , Fatores de Virulência/metabolismoRESUMO
Rice is often infected by bacterial panicle blight disease caused by Burkholderia glumae. Since most studies have assessed the transcriptome of the plant when it is exposed to bacteria, the gene expression of the phytopathogenic bacteria have not been well elaborated during the infection process or in the host cell. Recently, a few researches were conducted to evaluate the in vivo transcriptome of bacteria during the infective process. Most bacterial cells do not express genes involved in pathogenicity in culture medium making it difficult to investigate gene expression of bacterial cells in plant cells. Here, we sought a simulated patho-system that would allow bacterial cells to express their pathogenic genes. Thus, rice root exudates (RE) and bacterial N-acyl homoserine lactone (AHL) were used and their effects on bacterial gene expression were assessed. Transcription patterns of B. glumae virulence determinants showed that enrichment medium (LB + RE + C8-HSL) could significantly induce virulence factor genes compared with Luria Bertani (LB; control) medium. The data indicate that the artificial environment is similar to the real patho-system, and that this induced maximum relevant gene expression. In this model system, bacterial gene expression changes are traceable in the infection process. Bacterial cells exposed to either an artificial environment or LB + RE + C8-HSL behaved similarly to the natural environment in situ.
Assuntos
Burkholderia/genética , Burkholderia/patogenicidade , Oryza/microbiologia , Transcriptoma , Fatores de Virulência/genética , Acil-Butirolactonas , Técnicas de Cultura , Genes Bacterianos/genética , Exsudatos de PlantasRESUMO
The nonpathogenic soil saprophyte Burkholderia thailandensis is a member of the Burkholderia pseudomallei/B. thailandensis/B. mallei group, which also comprises the closely related human pathogens B. pseudomallei and Burkholderia mallei responsible for the melioidosis and glanders diseases, respectively. ScmR, a recently identified LysR-type transcriptional regulator in B. thailandensis, acts as a global transcriptional regulator throughout the stationary phase and modulates the production of a wide range of secondary metabolites, including N-acyl-l-homoserine lactones and 4-hydroxy-3-methyl-2-alkylquinolines and virulence in the Caenorhabditis elegans nematode worm host model, as well as several quorum sensing (QS)-dependent phenotypes. We have investigated the role of ScmR in B. thailandensis strain E264 during the exponential phase. We used RNA sequencing transcriptomic analyses to identify the ScmR regulon, which was compared to the QS-controlled regulon, showing a considerable overlap between the ScmR-regulated genes and those controlled by QS. We characterized several genes modulated by ScmR using quantitative reverse transcription-PCR or mini-CTX-lux transcriptional reporters, including the oxalate biosynthetic gene obc1 required for pH homeostasis, the orphan LuxR-type transcriptional regulator BtaR5-encoding gene, and the bsa (Burkholderia secretion apparatus) type III secretion system genes essential for both B. pseudomallei and B. mallei pathogenicity, as well as the scmR gene itself. We confirmed that the transcription of scmR is under QS control, presumably ensuring fine-tuned modulation of gene expression. Finally, we demonstrated that ScmR influences virulence using the fruit fly model host Drosophila melanogaster We conclude that ScmR represents a central component of theB. thailandensis QS regulatory network.IMPORTANCE Coordination of the expression of genes associated with bacterial virulence and environmental adaptation is often dependent on quorum sensing (QS). The QS circuitry of the nonpathogenic bacterium Burkholderia thailandensis, widely used as a model system for the study of the human pathogen Burkholderia pseudomallei, is complex. We found that the LysR-type transcriptional regulator, ScmR, which is highly conserved and involved in the control of virulence/survival factors in the Burkholderia genus, is a global regulator mediating gene expression through the multiple QS systems coexisting in B. thailandensis, as well as QS independently. We conclude that ScmR represents a key QS modulatory network element, ensuring tight regulation of the transcription of QS-controlled genes, particularly those required for acclimatization to the environment.
Assuntos
Proteínas de Bactérias/metabolismo , Infecções por Burkholderia/microbiologia , Burkholderia/metabolismo , Burkholderia/patogenicidade , Regulação Bacteriana da Expressão Gênica , Percepção de Quorum , Ácidos/metabolismo , Acil-Butirolactonas/metabolismo , Animais , Proteínas de Bactérias/genética , Burkholderia/genética , Caenorhabditis elegans , Drosophila melanogaster , Genes Reguladores , Homeostase , Humanos , Concentração de Íons de Hidrogênio , Masculino , VirulênciaRESUMO
The regulation and timely expression of bacterial genes during infection is critical for a pathogen to cause an infection. Bacteria have multiple mechanisms to regulate gene expression in response to their environment, one of which is two-component systems (TCS). TCS have two components. One component is a sensory histidine kinase (HK) that autophosphorylates when activated by a signal. The activated sensory histidine kinase then transfers the phosphoryl group to the second component, the response regulator, which activates transcription of target genes. The genus Burkholderia contains members that cause human disease and are often extensively resistant to many antibiotics. The Burkholderia cepacia complex (BCC) can cause severe lung infections in patients with cystic fibrosis (CF) or chronic granulomatous disease (CGD). BCC members have also recently been associated with several outbreaks of bacteremia from contaminated pharmaceutical products. Separate from the BCC is Burkholderia pseudomallei, which is the causative agent of melioidosis, a serious disease that occurs in the tropics, and a potential bioterrorism weapon. Bioinformatic analysis of sequenced Burkholderia isolates predicts that most strains have at least 40 TCS. The vast majority of these TCS are uncharacterized both in terms of the signals that activate them and the genes that are regulated by them. This review will highlight TCS that have been described to play a role in virulence in either the BCC or B. pseudomallei Since many of these TCS are involved in virulence, TCS are potential novel therapeutic targets, and elucidating their function is critical for understanding Burkholderia pathogenesis.
Assuntos
Fenômenos Fisiológicos Bacterianos , Infecções por Burkholderia/microbiologia , Burkholderia/fisiologia , Interações Hospedeiro-Patógeno , Burkholderia/patogenicidade , Complexo Burkholderia cepacia/fisiologia , Regulação Bacteriana da Expressão Gênica , Interações Hospedeiro-Patógeno/imunologia , Humanos , Transdução de Sinais , Virulência , Fatores de Virulência/genéticaRESUMO
Melioidosis is a neglected tropical disease with an estimated annual mortality rate of 89,000 in 45 countries across tropical regions. The causative agent is Burkholderia pseudomallei, a gram-negative soil-dwelling bacterium. In Thailand, B. pseudomallei can be found across multiple regions, along with the low-virulence B. thailandensis and the recently discovered B. thailandensis variant (BTCV), which expresses B. pseudomallei-like capsular polysaccharide. Comprehensive studies of human immune responses to B. thailandensis variants and cross-reactivity to B. pseudomallei are not complete. We evaluated human immune responses to B. pseudomallei, B. thailandensis, and BTCV in melioidosis patients and healthy persons in B. pseudomallei-endemic areas using a range of humoral and cellular immune assays. We found immune cross-reactivity to be strong for both humoral and cellular immunity among B. pseudomallei, B. thailandensis, and BTCV. Our findings suggest that environmental exposure to low-virulence strains may build cellular immunity to B. pseudomallei.
Assuntos
Burkholderia/imunologia , Melioidose/epidemiologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Burkholderia/patogenicidade , Estudos de Coortes , Reações Cruzadas , Feminino , Humanos , Imunidade , Masculino , Melioidose/microbiologia , Pessoa de Meia-Idade , Estudos Prospectivos , Tailândia/epidemiologia , Virulência , Adulto JovemRESUMO
Bacteria produce a diverse array of secondary metabolites that have been invaluable in the clinic and in research. These metabolites are synthesized by dedicated biosynthetic gene clusters (BGCs), which assemble architecturally complex molecules from simple building blocks. The majority of BGCs in a given bacterium are not expressed under normal laboratory growth conditions, and our understanding of how they are silenced is in its infancy. Here, we have addressed this question in the Gram-negative model bacterium Burkholderia thailandensis E264 using genetic, transcriptomic, metabolomic, and chemical approaches. We report that a previously unknown, quorum-sensing-controlled LysR-type transcriptional regulator, which we name ScmR (for secondary metabolite regulator), serves as a global gatekeeper of secondary metabolism and a repressor of numerous BGCs. Transcriptionally, we find that 13 of the 20 BGCs in B. thailandensis are significantly (threefold or more) up- or down-regulated in a scmR deletion mutant (ΔscmR) Metabolically, the ΔscmR strain displays a hyperactive phenotype relative to wild type and overproduces a number of compound families by 18- to 210-fold, including the silent virulence factor malleilactone. Accordingly, the ΔscmR mutant is hypervirulent both in vitro and in a Caenorhabditis elegans model in vivo. Aside from secondary metabolism, ScmR also represses biofilm formation and transcriptionally activates ATP synthesis and stress response. Collectively, our data suggest that ScmR is a pleiotropic regulator of secondary metabolism, virulence, biofilm formation, and other stationary phase processes. A model for how the interplay of ScmR with pathway-specific transcriptional regulators coordinately silences virulence factor production is proposed.
Assuntos
Proteínas de Bactérias/genética , Burkholderia/metabolismo , Burkholderia/patogenicidade , Metabolismo Secundário/genética , Animais , Proteínas de Bactérias/metabolismo , Biofilmes , Burkholderia/genética , Infecções por Burkholderia/microbiologia , Caenorhabditis elegans/microbiologia , Regulação Bacteriana da Expressão Gênica , Genes Reguladores , Lactonas/metabolismo , Família Multigênica , Virulência/genéticaRESUMO
Various diseases, including bacterial panicle blight (BPB) and sheath rot, threaten rice production. It has been established that Burkholderia glumae (B. glumae) is the causative agent of the above mentioned pathologies. In the present study, antagonistic activity, growth promotion, and the metabolite profiles of two rhizobacteria, isolated in different paddy fields, were assessed against B. glumae. Strains were identified based on 16S rRNA gene sequences, and the phylogenetic analyses showed that both strains belong to the genus Enterobacter, with high similarity to the strain Enterobacter tabaci NR146667.2 (99%). The antagonistic activity was assessed with the disc diffusion method. Active fractions were isolated through a liquid/liquid extraction with ethyl acetate (EtOAc) from the fermentation media, and their antibacterial activities were evaluated following the Clinical and Laboratory Standards Institute (CLSI) guidelines. The Pikovskaya modified medium was used to test the ability of in vitro inorganic phosphorus solubilization, and BSB1 proved to be the best inorganic phosphorus solubilizer, with a solubilization index (SI) of 4.5 ± 0.2. The glass-column fractionation of the EtOAc extracted from BCB11 produced an active fraction (25.9 mg) that inhibited the growth of five B. glumae strains by 85-95%. Further, metabolomic analysis, based on GC-MS, showed 3-phenylpropanoic acid (3-PPA) to be the main compound both in this fraction (46.7%), and in the BSB1 extract (28.6%). This compound showed antibacterial activity against all five strains of B. glumae with a minimum inhibitory concentration (MIC) of 1000 mg/L towards all of them. The results showed that rice rhizosphere microorganisms are a source of compounds that inhibit B. glumae growth and are promising plant growth promoters (PGP).
Assuntos
Antibacterianos/farmacologia , Burkholderia/patogenicidade , Enterobacter/metabolismo , Oryza/microbiologia , Doenças das Plantas/microbiologia , Doenças das Plantas/prevenção & controle , Acetatos/química , Enterobacter/classificação , Fermentação , Cromatografia Gasosa-Espectrometria de Massas , Espectroscopia de Ressonância Magnética , Metabolômica , Fenilpropionatos/química , Fosfatos/química , Filogenia , RNA Ribossômico 16S/genética , Metabolismo Secundário , Espectroscopia de Infravermelho com Transformada de Fourier , Virulência/genéticaRESUMO
Bacterial panicle blight caused by Burkholderia glumae is a major bacterial disease of rice. Our preliminary RNA-seq study showed that a serine metalloprotease gene, prtA, is regulated in a similar manner to the genes for the biosynthesis and transport of toxoflavin, which is a known major virulence factor of B. glumae. prtA null mutants of the virulent strain B. glumae 336gr-1 did not show a detectable extracellular protease activity, indicating that prtA is the solely responsible gene for the extracellular protease activity detected from this bacterium. In addition, inoculation of rice panicles with the prtA mutants resulted in a significant reduction of disease severity compared with the wild-type parent strain, suggesting the requirement of prtA for the full virulence of B. glumae. A double mutant deficient in both serine metalloprotease and toxoflavin (ΔtoxA/prtA-) exhibited a further numeric but not statistically significant decrease of disease development compared with the ΔtoxA strain. Both the prtA-driven extracellular protease activity and the toxoflavin production were dependent on both the tofI/tofR quorum-sensing and the global regulatory gene qsmR, indicating the important roles of the two global regulatory factors for the bacterial pathogenesis by this pathogen.
Assuntos
Burkholderia , Regulação Bacteriana da Expressão Gênica , Metaloendopeptidases , Virulência , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Burkholderia/enzimologia , Burkholderia/genética , Burkholderia/patogenicidade , Metaloendopeptidases/genética , Metaloendopeptidases/metabolismo , Virulência/genéticaRESUMO
Covering: up to February 2018 In recent years, genome sequencing revealed the full biosynthetic potential of bacteria causing plant diseases. Bioinformatics and advanced analytical techniques paved the way to clarify the structures of long-sought natural products with a role in virulence. Furthermore, several compounds without disease-associated function were discovered. The exploration of these molecules disclosed persistence strategies of plant pathogenic bacteria outside their hosts and provided access to new bioactive compounds with therapeutic potential. In this review, we will summarize some of the striking findings in the field, paying particular attention to unique natural product pathways and their unprecedented biosynthetic features as well as the biological activities of the retrieved compounds.
Assuntos
Bactérias/genética , Bactérias/patogenicidade , Produtos Biológicos/metabolismo , Plantas/microbiologia , Bactérias/metabolismo , Burkholderia/genética , Burkholderia/metabolismo , Burkholderia/patogenicidade , Genoma Bacteriano , Genômica/métodos , Doenças das Plantas/microbiologia , Ralstonia solanacearum/genética , Ralstonia solanacearum/metabolismo , Ralstonia solanacearum/patogenicidade , Streptomyces/genética , Streptomyces/metabolismo , Streptomyces/patogenicidade , Xanthomonas/genética , Xanthomonas/metabolismo , Xanthomonas/patogenicidadeRESUMO
Bacteria produce a vast range of exopolysaccharides (EPSs) to thrive in diverse environmental niches and often display a mucoid phenotype in solid media. One such exopolysaccharide, cepacian, is produced by bacteria of the genus Burkholderia and is of interest due to its role in pathogenesis associated with lung infections in cystic fibrosis (CF) patients. Cepacian is a repeat-unit polymer that has been implicated in biofilm formation, immune system evasion, interaction with host cells, resistance against antimicrobials, and virulence. Its biosynthesis proceeds through the Wzy-dependent polymerization and secretion mechanism, which requires a multienzymatic complex. Key aspects of its structure, genetic organization, and the regulatory network involved in mucoid switch and regulation of cepacian biosynthesis at transcriptional and posttranscriptional levels are reviewed. It is also evaluated the importance of cepacian biosynthesis/regulation key players as evolutionary targets of selection and highlighted the complexity of the regulatory network, which allows cells to coordinate the expression of metabolic functions to the ones of the cell wall, in order to be successful in ever changing environments, including in the interaction with host cells.
Assuntos
Variação Biológica da População , Burkholderia/metabolismo , Polissacarídeos Bacterianos/biossíntese , Fatores de Virulência/biossíntese , Vias Biossintéticas/genética , Burkholderia/patogenicidade , Regulação Bacteriana da Expressão Gênica , Polissacarídeos Bacterianos/química , Polissacarídeos Bacterianos/genética , Fatores de Virulência/químicaRESUMO
Burkholderia species such as B. mallei and B. pseudomallei are bacterial pathogens causing fatal infections in humans and animals (glanders and melioidosis), yet knowledge on their virulence factors is limited. While pathogenic effects have been linked to a highly conserved gene locus (bur/mal) in the B. mallei group, the metabolite associated to the encoded polyketide synthase, burkholderic acid (syn. malleilactone), could not explain the observed phenotypes. By metabolic profiling and molecular network analyses of the model organism B. thailandensis, the primary products of the cryptic pathway were identified as unusual cyclopropanol-substituted polyketides. First, sulfomalleicyprols were identified as inactive precursors of burkholderic acid. Furthermore, a highly reactive upstream metabolite, malleicyprol, was discovered and obtained in two stabilized forms. Cell-based assays and a nematode infection model showed that the rare natural product confers cytotoxicity and virulence.
Assuntos
Burkholderia/metabolismo , Éteres Cíclicos/metabolismo , Policetídeos/metabolismo , Fatores de Virulência/metabolismo , Animais , Burkholderia/genética , Burkholderia/patogenicidade , Caenorhabditis elegans/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Éteres Cíclicos/química , Éteres Cíclicos/farmacologia , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Humanos , Células K562 , Estrutura Molecular , Policetídeos/química , Policetídeos/farmacologia , Virulência , Fatores de Virulência/química , Fatores de Virulência/farmacologiaRESUMO
Fatty acid hydroperoxides are involved in host-pathogen interactions. In both plants and mammals, polyunsaturated fatty acids are liberated during infection and enzymatically oxidized to the corresponding toxic hydroperoxides during the defensive oxidative burst that is designed to thwart the infection. The bacterial transcription factor OhrR (organic hydroperoxide reductase regulator) is oxidized by organic hydroperoxides, as a result of which the ohr gene encoding organic hydroperoxide reductase is induced. This enzyme converts the hydroperoxides to less toxic alcohols. We show here that OhrR from Burkholderia thailandensis represses expression of ohr Gene expression is induced by cumene hydroperoxide and to a lesser extent by inorganic oxidants; however, Ohr contributes to degradation only of the organic hydroperoxide. B. thailandensis OhrR, which binds specific sites in both ohr and ohrR promoters, as evidenced by DNase I footprinting, belongs to the 2-Cys subfamily of OhrR proteins, and its oxidation leads to reversible disulfide bond formation between conserved N- and C-terminal cysteines in separate monomers. Oxidation of the N-terminal Cys is sufficient for attenuation of DNA binding in vitro, with complete restoration of DNA binding occurring on addition of a reducing agent. Surprisingly, both overexpression of ohr and deletion of ohr results in enhanced survival on exposure to organic hydroperoxide in vitro While Δohr cells are more virulent in a Caenorhabditis elegans model of infection, ΔohrR cells are less so. Taken together, our data suggest that B. thailandensis OhrR has several unconventional features and that both OhrR and organic hydroperoxides may contribute to virulence.
Assuntos
Proteínas de Bactérias/genética , Burkholderia/genética , Caenorhabditis elegans/microbiologia , Regulação Bacteriana da Expressão Gênica , Proteínas Repressoras/genética , Animais , Proteínas de Bactérias/química , Derivados de Benzeno/farmacologia , Burkholderia/efeitos dos fármacos , Burkholderia/patogenicidade , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/genética , Desoxirribonuclease I , Interações Hospedeiro-Patógeno , Peróxido de Hidrogênio/farmacologia , Oxidantes/farmacologia , Oxirredução , Proteínas Repressoras/químicaRESUMO
Covering: up to 2018 Burkholderia species are a vast group of human pathogenic, phytopathogenic, and plant- or environment-associated bacteria. B. pseudomallei, B. mallei, and B. cepacia complex are the causative agents of melioidosis, glanders, and cystic fibrosis-related infections, respectively, which are fatal diseases in humans and animals. Due to their high resistance to antibiotics, high mortality rates, and increased infectivity via the respiratory tract, B. pseudomallei and B. mallei have been listed as potential bioterrorism agents by the Centers for Disease Control and Prevention. Burkholderia species are able to produce a large network of surface-exposed polysaccharides, i.e., lipopolysaccharides, capsular polysaccharides, and exopolysaccharides, which are virulence factors, immunomodulators, major biofilm components, and protective antigens, and have crucial implications in the pathogenicity of Burkholderia-associated diseases. This review provides a comprehensive and up-to-date account regarding the structural elucidation and biological activities of surface polysaccharides produced by Burkholderia species. The chemical synthesis of oligosaccharides mimicking Burkholderia polysaccharides is described in detail. Emphasis is placed on the recent research efforts toward the development of glycoconjugate vaccines against melioidosis and glanders based on synthetic or native Burkholderia oligo/polysaccharides.