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Frontiers in Microbiology 2023For a coherent response to environmental changes, bacterial evolution has formed a complex transcriptional regulatory system comprising classical DNA binding proteins...
For a coherent response to environmental changes, bacterial evolution has formed a complex transcriptional regulatory system comprising classical DNA binding proteins sigma factors and modulation of DNA topology. In this study, we investigate replication-induced gene copy numbers - a regulatory concept that is unlike the others not based on modulation of promoter activity but on replication dynamics. We show that a large fraction of genes are predominantly affected by transient copy numbers and identify cellular functions and central pathways governed by this mechanism in . Furthermore, we show quantitatively that the previously observed spatio-temporal expression pattern between different growth phases mainly emerges from transient chromosomal copy numbers. We extend the analysis to the plant pathogen and the biotechnologically relevant organism . The analysis reveals a connection between growth phase dependent gene expression and evolutionary gene migration in these species. A further extension to the bacterial kingdom indicates that chromosome evolution is governed by growth rate related transient copy numbers.
PubMed: 37152747
DOI: 10.3389/fmicb.2023.1119878 -
Frontiers in Plant Science 2016The negative impact of conventional farming on environment and human health make improvements on farming management mandatory. Imaging techniques are implemented in...
The negative impact of conventional farming on environment and human health make improvements on farming management mandatory. Imaging techniques are implemented in remote sensing for monitoring crop fields and plant phenotyping programs. The increasingly large size and complexity of the data obtained by these techniques, makes the implementation of powerful mathematical tools necessary in order to identify informative parameters and to apply them in precision agriculture. Multicolor fluorescence imaging is a useful approach for the study of plant defense responses to stress factors at bench scale. However, it has not been fully applied to plant phenotyping. This work evaluates the possible application of multicolor fluorescence imaging in combination with thermography for the particular case of zucchini plants affected by soft-rot, caused by . Several statistical models -based on logistic regression analysis (LRA) and artificial neural networks (ANN)- were obtained for the experimental system zucchini-, which classify new samples as "healthy" or "infected." The LRA worked best in identifying high dose-infiltrated leaves (in infiltrated and non-infiltrated areas) whereas ANN offered a higher accuracy at identifying low dose-infiltrated areas. To assess the applicability of these results to cucurbits in a more general way, these models were validated for melon infected by the same pathogen, achieving accurate predictions for the infiltrated areas. The values of accuracy achieved are comparable to those found in the literature for classifiers identifying other infections based on data obtained by different techniques. Thus, MCFI in combination with thermography prove useful at providing data at lab scale that can be analyzed by machine learning. This approach could be scaled up to be applied in plant phenotyping.
PubMed: 27994607
DOI: 10.3389/fpls.2016.01790 -
Research in Microbiology May 2016Dickeya dadantii is a pathogen infecting a wide range of plant species. Soft rot, the visible symptom, is mainly due to production of pectate lyases (Pels) that can...
Regulation of pel genes, major virulence factors in the plant pathogen bacterium Dickeya dadantii, is mediated by cooperative binding of the nucleoid-associated protein H-NS.
Dickeya dadantii is a pathogen infecting a wide range of plant species. Soft rot, the visible symptom, is mainly due to production of pectate lyases (Pels) that can destroy plant cell walls. Previously, we found that nucleoid-associated protein (NAP) H-NS is a key regulator of pel gene expression. The primary binding sites of this NAP have been determined here by footprinting experiments on the pelD gene, encoding an essential virulence factor. Quantitative analysis of DNAse I footprints and surface plasmon resonance imagery experiments further revealed that high-affinity binding sites initiate cooperative binding to establish the nucleoprotein structure required for gene expression silencing. Mutations in the primary binding sites resulted in reduction or loss of repression by H-NS. Overall, these data suggest that H-NS represses pelD, and by inference, other pel genes, by a cooperative binding mechanism, through oligomerization of H-NS molecules.
Topics: Bacterial Proteins; Binding Sites; DNA Footprinting; DNA, Bacterial; DNA-Binding Proteins; Gammaproteobacteria; Gene Expression Regulation, Bacterial; Mutation; Polysaccharide-Lyases; Protein Binding; Surface Plasmon Resonance; Virulence Factors
PubMed: 26912324
DOI: 10.1016/j.resmic.2016.02.001 -
Frontiers in Plant Science 2018Cyclotides are cyclic plant polypeptides of 27-37 amino acid residues. They have been extensively studied in bioengineering and drug development contexts. However, less...
Cyclotides are cyclic plant polypeptides of 27-37 amino acid residues. They have been extensively studied in bioengineering and drug development contexts. However, less is known about the relevance of cyclotides for the plants producing them. The anti-insect larvae effects of kB1 and antibacterial activity of cyO2 suggest that cyclotides are a part of plant host defense. The sweet violet ( L.) produces a wide array of cyclotides, including kB1 (kalata B1) and cyO2 (cycloviolacin O2), with distinct presumed biological roles. Here, we evaluate V. odorata cyclotides' potency against plant pathogens and their mode of action using bioassays, liposome experiments and immunogold labeling for transmission electron microscopy (TEM). We explore the link between the biological activity and distribution in plant generative, vegetative tissues and seeds, depicted by immunohistochemistry and matrix assisted laser desorption ionization mass spectrometry imaging (MALDI-MSI). Cyclotides cyO2, cyO3, cyO13, and cyO19 are shown to have potent activity against model fungal plant pathogens () and fungi isolated from violets (), with minimal inhibitory concentrations (MICs) ranging from 0.8 μM to 25 μM. Inhibition of phytopathogenic bacteria - pv. and - is also observed with MIC = 25-100 μM. A membrane-disrupting antifungal mode of action is shown. Finding cyO2 inside the fungal spore cells in TEM images may indicate that other, intracellular targets may be involved in the mechanism of toxicity. Fungi can not break down cyclotides in the course of days. varv A (kalata S) and kB1 show little potency against pathogenic fungi when compared with the tested cycloviolacins. cyO2, cyO3, cyO19 and kB1 are differentially distributed and found in tissues vulnerable to pathogen (epidermis, rizodermis, vascular bundles, protodermis, procambium, ovary walls, outer integuments) and pest (ground tissues of leaf and petiole) attacks, respectively, indicating a link between the cyclotides' sites of accumulation and biological role. Cyclotides emerge as a comprehensive defense system in , in which different types of peptides have specific targets that determine their distribution in plant tissues.
PubMed: 30254654
DOI: 10.3389/fpls.2018.01296 -
Frontiers in Plant Science 2018The bacterium is responsible of important economic losses in crop yield worldwide. In melon leaves, produced multiple necrotic spots surrounded by a chlorotic halo,...
The bacterium is responsible of important economic losses in crop yield worldwide. In melon leaves, produced multiple necrotic spots surrounded by a chlorotic halo, followed by necrosis of the whole infiltrated area and chlorosis in the surrounding tissues. The extent of these symptoms, as well as the day of appearance, was dose-dependent. Several imaging techniques (variable chlorophyll fluorescence, multicolor fluorescence, and thermography) provided spatial and temporal information about alterations in the primary and secondary metabolism, as well as the stomatal activity in the infected leaves. Detection of diseased leaves was carried out by using machine learning on the numerical data provided by these imaging techniques. Mathematical algorithms based on data from infiltrated areas offered 96.5 to 99.1% accuracy when classifying them as mock vs. bacteria-infiltrated. These algorithms also showed a high performance of classification of whole leaves, providing accuracy values of up to 96%. Thus, the detection of disease on whole leaves by a model trained on infiltrated areas appears as a reliable method that could be scaled-up for use in plant breeding programs or precision agriculture.
PubMed: 29491881
DOI: 10.3389/fpls.2018.00164 -
Molecular Plant Pathology May 2018Bacteria from the genus Dickeya cause severe symptoms on numerous economically important plants. Dickeya solani is the Dickeya species most frequently found on infected...
Bacteria from the genus Dickeya cause severe symptoms on numerous economically important plants. Dickeya solani is the Dickeya species most frequently found on infected potato plants in Europe. D. solani strains from different countries show high genetic homogeneity, but significant differences in their virulence level. Dickeya species possess two quorum sensing (QS) mechanisms: the Exp system based on classic N-acyl-homoserine lactone (AHL) signals and a specific system depending on the production and perception of a molecule of unknown structure, Virulence Factor Modulating (VFM). To study the interplay between these two QS systems, five D. solani strains exhibiting different virulence levels were selected. Mutants were constructed by inactivating genes coding for each QS system. Double mutants were obtained by simultaneous inactivation of genes coding for both QS systems. Most of the D. solani mutants showed an attenuation of chicory maceration and a decreased production of plant cell wall-degrading enzymes (PCWDEs) and motility, but to different degrees depending on the strain. The VFM-QS system seems to regulate virulence in both D. solani and Dickeya dadantii, but the AHL-QS system has greater effects in D. solani than in D. dadantii. The inactivation of both QS systems in D. solani did not reveal any additive effect on the tested features. The inactivation of vfm genes generally has a more dominant effect relative to that of exp genes. Thus, VFM- and AHL-QS systems do not work in synergy to modulate the production of diverse virulence factors and the ability to macerate plant tissue.
Topics: 4-Butyrolactone; Cichorium intybus; Enterobacteriaceae; Genes, Bacterial; Mutation; Phenotype; Plant Diseases; Plant Leaves; Plant Tubers; Quorum Sensing; Solanum tuberosum; Virulence; Virulence Factors
PubMed: 28921772
DOI: 10.1111/mpp.12614 -
Frontiers in Microbiology 2024The type VI secretion system (T6SS) assembles into a contractile nanomachine to inject effectors across bacterial membranes for secretion. The species complex is a...
The type VI secretion system (T6SS) assembles into a contractile nanomachine to inject effectors across bacterial membranes for secretion. The species complex is a group of soil inhabitants and phytopathogens that deploys T6SS as an antibacterial weapon against bacterial competitors at both inter-species and intra-species levels. The strain 1D1609 genome encodes one main T6SS gene cluster and four genes (i.e., ), each encoding a spike protein as an effector carrier. A previous study reported that associated gene 2, named encodes a His-Me finger nuclease toxin (also named HNH/ENDO VII nuclease), contributing to DNase-mediated antibacterial activity. However, the functions and roles of other putative effectors remain unknown. In this study, we identified associated gene 2 () that encodes another His-Me finger nuclease but with a distinct Serine Histidine Histidine (SHH) motif that differs from the AHH motif of V2a. We demonstrated that the ectopic expression of V2c caused growth inhibition, plasmid DNA degradation, and cell elongation in using DNAse activity assay and fluorescence microscopy. The cognate immunity protein, V3c, neutralizes the DNase activity and rescues the phenotypes of growth inhibition and cell elongation. Ectopic expression of V2c DNase-inactive variants retains the cell elongation phenotype, while V2a induces cell elongation in a DNase-mediated manner. We also showed that the amino acids of conserved SHH and HNH motifs are responsible for the V2c DNase activity and . Notably, V2c also mediated the DNA degradation and cell elongation of the target cell in the context of interbacterial competition. Importantly, V2a and V2c exhibit different capacities against different bacterial species and function synergistically to exert stronger antibacterial activity against the soft rot phytopathogen, .
PubMed: 38426053
DOI: 10.3389/fmicb.2024.1351590 -
International Journal of Molecular... Nov 2019The prominent antibacterial and quorum sensing (QS) inhibition activity of aromatic plants can be used as a novel intervention strategy for attenuating bacterial...
The prominent antibacterial and quorum sensing (QS) inhibition activity of aromatic plants can be used as a novel intervention strategy for attenuating bacterial pathogenicity. In the present work, a total of 29 chemical components were identified in the essential oil (EO) of leaves by gas chromatography-mass spectrometry (GC-MS). The principal component was methyleugenol, followed by methyl trans-cinnamate, with relative contents of 90.46% and 4.25%, respectively. Meanwhile, the antibacterial activity and the QS inhibitory activity of EO were first evaluated here. Antibacterial activity assay and MIC detection against seven pathogens ( Onc5, ATCC25933, spp., ATCC25922, MG1, PAO1 and ATCC31532) demonstrated that ATCC25933 and MG1 had the higher sensitivity to EO, while PAO1 displayed the strongest resistance to EO. An anti-QS (anti-quorum sensing) assay revealed that at sub-minimal inhibitory concentrations (sub-MICs), EO strongly interfered with the phenotype, including violacein production, biofilm biomass, and swarming motility, as well as -hexanoyl-L-homoserine lactone (C6-HSL) production (i.e., a signaling molecule in ATCC31532) of . Detection of C6-HSL indicated that EO was capable of not only inhibiting C6-HSL production in , but also degrading the C6-HSL. Importantly, changes of exogenous C6-HSL production in CV026 revealed a possible interaction between EO and a regulatory protein (cviR). Additionally, quantitative real-time polymerase chain reaction (RT-qPCR) analysis demonstrated that the expression of QS-related genes (, , , , , , , and ) was significantly suppressed. Conclusively, these results indicated that EO can act as a potential antibacterial agent and QS inhibitor (QSI) against pathogens, preventing and controlling bacterial contamination.
Topics: Anti-Bacterial Agents; Bacteria; Biofilms; Gas Chromatography-Mass Spectrometry; Melaleuca; Microbial Sensitivity Tests; Oils, Volatile; Phenotype; Plant Extracts; Plant Leaves; Quorum Sensing
PubMed: 31739398
DOI: 10.3390/ijms20225696 -
Environmental Microbiology Aug 2019Dickeya dadantii is a plant pathogen that causes soft rot disease on vegetable and potato crops. To successfully cause infection, this pathogen needs to coordinately...
Dickeya dadantii is a plant pathogen that causes soft rot disease on vegetable and potato crops. To successfully cause infection, this pathogen needs to coordinately modulate the expression of genes encoding several virulence determinants, including plant cell wall degrading enzymes (PCWDEs), type III secretion system (T3SS) and flagellar motility. Here, we uncover a novel feed-forward signalling circuit for controlling virulence. Global RNA chaperone Hfq interacts with an Hfq-dependent sRNA ArcZ and represses the translation of pecT, encoding a LysR-type transcriptional regulator. We demonstrate that the ability of ArcZ to be processed to a 50 nt 3'- end fragment is essential for its regulation of pecT. PecT down-regulates PCWDE and the T3SS by repressing the expression of a global post-transcriptional regulator- (RsmA-) associated sRNA encoding gene rsmB. In addition, we show that the protein levels of two cyclic di-GMP (c-di-GMP) diguanylate cyclases (DGCs), GcpA and GcpL, are repressed by Hfq. Further studies show that both DGCs are essential for the Hfq-mediated post-transcriptional regulation on RsmB. Overall, our report provides new insights into the interplays between ubiquitous signalling transduction systems that were most studied independently and sheds light on multitiered regulatory mechanisms for a precise disease regulation in bacteria.
Topics: Bacterial Proteins; Cell Wall; Cyclic GMP; Enterobacteriaceae; Gene Expression Regulation, Bacterial; Plant Diseases; RNA, Bacterial; RNA, Small Untranslated; RNA-Binding Proteins; Signal Transduction; Type III Secretion Systems; Virulence; Virulence Factors
PubMed: 30895662
DOI: 10.1111/1462-2920.14603 -
ACS Chemical Biology Oct 2019Quaternary distance restraints are essential to define the three-dimensional structures of protein assemblies. These distances often fall within a range of 10-18 Å,...
Quaternary distance restraints are essential to define the three-dimensional structures of protein assemblies. These distances often fall within a range of 10-18 Å, which challenges the high and low measurement limits of conventional nuclear magnetic resonance (NMR) and double electron-electron resonance electron spin resonance spectroscopies. Here, we report the use of F paramagnetic relaxation enhancement (PRE) NMR in combination with F/paramagnetic labeling to equivalent sites in different subunits of a protein complex in micelles to determine intersubunit distances. The feasibility of this strategy was evaluated on a pentameric ligand-gated ion channel, for which we found excellent agreement of the F PRE NMR results with previous structural information. The study suggests that F PRE NMR is a viable tool in extracting distance restraints to define quaternary structures.
Topics: Animals; Bacterial Proteins; Dickeya; Fluorine; Gammaproteobacteria; Ion Channels; Mice; Nuclear Magnetic Resonance, Biomolecular; Protein Structure, Quaternary; Protein Subunits
PubMed: 31525026
DOI: 10.1021/acschembio.9b00692