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Scientific Reports Mar 2015In the track of new biopesticides, four genes namely cytA, cytB, cytC and cytD encoding proteins homologous to Bacillus thuringiensis (Bt) Cyt toxins have been...
In the track of new biopesticides, four genes namely cytA, cytB, cytC and cytD encoding proteins homologous to Bacillus thuringiensis (Bt) Cyt toxins have been identified in the plant pathogenic bacteria Dickeya dadantii genome. Here we show that three Cyt-like δ-endotoxins from D. dadantii (CytA, CytB and CytC) are toxic to the pathogen of the pea aphid Acyrthosiphon pisum in terms of both mortality and growth rate. The phylogenetic analysis of the comprehensive set of Cyt toxins available in genomic databases shows that the whole family is of limited taxonomic occurrence, though in quite diverse microbial taxa. From a structure-function perspective the 3D structure of CytC and its backbone dynamics in solution have been determined by NMR. CytC adopts a cytolysin fold, structurally classified as a Cyt2-like protein. Moreover, the identification of a putative lipid binding pocket in CytC structure, which has been probably maintained in most members of the Cyt-toxin family, could support the importance of this lipid binding cavity for the mechanism of action of the whole family. This integrative approach provided significant insights into the evolutionary and functional history of D. dadantii Cyt toxins, which appears to be interesting leads for biopesticides.
Topics: Amino Acid Sequence; Bacterial Proteins; Endotoxins; Enterobacteriaceae; Models, Molecular; Molecular Sequence Data; Multigene Family; Nuclear Magnetic Resonance, Biomolecular; Phylogeny; Protein Conformation; Sequence Alignment; Solutions
PubMed: 25740111
DOI: 10.1038/srep08791 -
Molecular Plant Pathology Feb 2019The identification of the virulence factors of plant-pathogenic bacteria has relied on the testing of individual mutants on plants, a time-consuming process. Transposon...
The identification of the virulence factors of plant-pathogenic bacteria has relied on the testing of individual mutants on plants, a time-consuming process. Transposon sequencing (Tn-seq) is a very powerful method for the identification of the genes required for bacterial growth in their host. We used this method in a soft-rot pathogenic bacterium to identify the genes required for the multiplication of Dickeya dadantii in chicory. About 100 genes were identified showing decreased or increased fitness in the plant. Most had no previously attributed role in plant-bacterium interactions. Following our screening, in planta competition assays confirmed that the uridine monophosphate biosynthesis pathway and the purine biosynthesis pathway were essential to the survival of D. dadantii in the plant, as the mutants ∆carA, ∆purF, ∆purL, ∆guaB and ∆pyrE were unable to survive in the plant in contrast with the wild-type (WT) bacterium. This study also demonstrated that the biosynthetic pathways of leucine, cysteine and lysine were essential for bacterial survival in the plant and that RsmC and GcpA were important in the regulation of the infection process, as the mutants ∆rsmC and ∆gcpA were hypervirulent. Finally, our study showed that D. dadantii flagellin was glycosylated and that this modification conferred fitness to the bacterium during plant infection. Assay by this method of the large collections of environmental pathogenic strains now available will allow an easy and rapid identification of new virulence factors.
Topics: Bacterial Proteins; Cichorium intybus; DNA Transposable Elements; Enterobacteriaceae; Gene Expression Regulation, Bacterial; Glycosylation; Virulence
PubMed: 30267562
DOI: 10.1111/mpp.12754 -
Pharmaceutics Mar 2022L-asparaginases (EC 3.5.1.1) are a family of enzymes that catalyze the hydrolysis of L-asparagine to L-aspartic acid and ammonia. These proteins with different... (Review)
Review
L-asparaginases (EC 3.5.1.1) are a family of enzymes that catalyze the hydrolysis of L-asparagine to L-aspartic acid and ammonia. These proteins with different biochemical, physicochemical and pharmacological properties are found in many organisms, including bacteria, fungi, algae, plants and mammals. To date, asparaginases from and (formerly known as ) are widely used in hematology for the treatment of lymphoblastic leukemias. However, their medical use is limited by side effects associated with the ability of these enzymes to hydrolyze L-glutamine, as well as the development of immune reactions. To solve these issues, gene-editing methods to introduce amino-acid substitutions of the enzyme are implemented. In this review, we focused on molecular analysis of the mechanism of enzyme action and to optimize the antitumor activity.
PubMed: 35335974
DOI: 10.3390/pharmaceutics14030599 -
Biomolecules Dec 2019Edible plant fruits are safe raw materials free of toxicants and rich in biomolecules for reducing metal ions and stabilizing nanoparticles. Zinc oxide nanoparticles...
Edible plant fruits are safe raw materials free of toxicants and rich in biomolecules for reducing metal ions and stabilizing nanoparticles. Zinc oxide nanoparticles (ZnONPs) and titanium dioxide nanoparticles (TiONPs) are the most produced consumer nanomaterials and have known antibacterial activities but have rarely been used against phytopathogenic bacteria. Here, we synthesized ZnONPs and TiONPs simply by mixing ZnO or TiO solution with a lemon fruit extract at room temperature and showed their antibacterial activities against Dickeya dadantii, which causes sweet potato stem and root rot disease occurring in major sweet potato planting areas in China. Ultraviolet-visible spectrometry and energy dispersive spectroscopy determined their physiochemical characteristics. Transmission electron microscopy, scanning electron microscopy, and X-ray diffraction spectroscopy revealed the nanoscale size and polymorphic crystalline structures of the ZnONPs and TiONPs. Fourier-transform infrared spectroscopy revealed their surface stabilization groups from the lemon fruit extract. In contrast to ZnO and TiO, which had no antibacterial activity against D. dadantii, ZnONPs and TiONPs showed inhibitions on D. dadantii growth, swimming motility, biofilm formation, and maceration of sweet potato tuber slices. ZnONPs and TiONPs showed similar extents of antibacterial activities, which increased with the increase of nanoparticle concentrations, and inhibited about 60% of activities at the concentration of 50 µg∙mL. The green synthetic ZnONPs and TiONPs can be used to control the sweet potato soft rot disease by control of pathogen contamination of seed tubers.
Topics: Anti-Bacterial Agents; Citrus; Dickeya; Fruit; Gammaproteobacteria; Microbial Sensitivity Tests; Nanoparticles; Plant Extracts; Titanium; Zinc Oxide
PubMed: 31835898
DOI: 10.3390/biom9120863 -
Frontiers in Microbiology 2020Currently, the poultry industry has been faced with consumer pressure to utilize only vegetable feedstuffs in poultry diets, eliminate antibiotics from poultry...
Currently, the poultry industry has been faced with consumer pressure to utilize only vegetable feedstuffs in poultry diets, eliminate antibiotics from poultry production, and rear poultry in free range systems. To maintain current production standards, the industry must determine ways to enhance nutrient uptake and utilization further. One possible solution is the supplementation of pectinase, an enzyme that degrades pectin within the cell walls of plants, in poultry diets. Therefore, the objective of the current study was to determine the potential role of a pectinase producer, DSM 18020, as a commercially utilized pectinase producer in poultry diets against other known pectinase producers, . In the current study, whole genomes of DSM 18020 (Dd18020), 3937 (Dd3937), IPO 2222 (Ds2222), C-125 (BhC125), and subsp. str. 168 (Bs168) were compared using bioinformatic approaches to compare the chromosomal genome size, GC content, protein coding genes (CDS), total genes, average protein length (a.a.) and determine the predicted metabolic pathways, predicted pectin degrading enzymes, and pectin-degradation pathways across pectinase producers. Due to insufficient information surrounding the genome of Dd18020 (lack of annotation), the genome of Dd3937, a 99% identical genome to Dd18020, was utilized to compare pectinase-associated enzymes and pathways. The results from the current study demonstrated that Dd3937 possessed the most significant proportion of pathways presented and the highest number of pathways related to degradation, assimilation, and utilization of pectin. Also, Dd18020 exhibited a high number of pectinase-related enzymes. Both Dd3937 and Dd2222 shared the pectin degradation I pathway via the EC 3.1.1.11, EC 3.2.1.82, and EC 4.2.2.- enzymes, but did not share this pathway with either species. In conclusion, Dd18020 demonstrated the genetic potential to produce multiple pectinase enzymes that could be beneficial to the degradation of pectin in poultry diets. However, for Dd18020 to become a commercially viable enzyme producer for the poultry industry, further research quantifying the pectinase production and determining the stability of the produced pectinases during feed manufacturing are necessary.
PubMed: 32390987
DOI: 10.3389/fmicb.2020.00751 -
Frontiers in Microbiology 2018is a bacterial plant pathogen that causes soft rot disease on a wide range of host plants. The type III secretion system (T3SS) is an important virulence factor in ....
Development of a Method to Monitor Gene Expression in Single Bacterial Cells During the Interaction With Plants and Use to Study the Expression of the Type III Secretion System in Single Cells of in Potato.
is a bacterial plant pathogen that causes soft rot disease on a wide range of host plants. The type III secretion system (T3SS) is an important virulence factor in . Expression of the T3SS is induced in the plant apoplast or in -inducing minimal medium (hrp-MM), and is repressed in nutrient-rich media. Despite the understanding of induction conditions, how individual cells in a clonal bacterial population respond to these conditions and modulate T3SS expression is not well understood. In our previous study, we reported that in a clonal population, only a small proportion of bacteria highly expressed T3SS genes while the majority of the population did not express T3SS genes under hrp-MM condition. In this study, we developed a method that enabled observation and quantification of gene expression in single bacterial cells . Using this technique, we observed that the expression of the T3SS genes and is restricted to a small proportion of cells during the infection of potato. We also report that the expression of T3SS genes is higher at early stages of infection compared to later stages. This expression modulation is achieved through adjusting the ratio of T3SS and T3SS cells and the expression intensity of T3SS cells. Our findings not only shed light into how bacteria use a bi-stable gene expression manner to modulate an important virulence factor, but also provide a useful tool to study gene expression in individual bacterial cells .
PubMed: 30002651
DOI: 10.3389/fmicb.2018.01429 -
International Journal of Molecular... May 2021Type II toxin-antitoxin (TA) systems are genetic elements usually encoding two proteins: a stable toxin and an antitoxin, which binds the toxin and neutralizes its toxic...
Type II toxin-antitoxin (TA) systems are genetic elements usually encoding two proteins: a stable toxin and an antitoxin, which binds the toxin and neutralizes its toxic effect. The disturbance in the intracellular toxin and antitoxin ratio typically leads to inhibition of bacterial growth or bacterial cell death. Despite the fact that TA modules are widespread in bacteria and archaea, the biological role of these systems is ambiguous. Nevertheless, a number of studies suggests that the TA modules are engaged in such important processes as biofilm formation, stress response or virulence and maintenance of mobile genetic elements. The 3937 strain serves as a model for pathogens causing the soft-rot disease in a wide range of angiosperm plants. Until now, several chromosome-encoded type II TA systems were identified in silico in the genome of this economically important bacterium however so far only one of them was experimentally validated. In this study, we investigated three putative type II TA systems in 3937: , and , which represents a novel toxin/antitoxin superfamily. We provide an experimental proof for their functionality in vivo both in and . Finally, we examined the prevalence of those systems across the Pectobacteriaceae family by a phylogenetic analysis.
Topics: Bacterial Proteins; Bacterial Toxins; Dickeya; Gene Expression Regulation, Bacterial; Plant Diseases; Toxin-Antitoxin Systems; Virulence
PubMed: 34073004
DOI: 10.3390/ijms22115932 -
Infection and Immunity May 2017Type II secretion (T2S) is one means by which Gram-negative pathogens secrete proteins into the extracellular milieu and/or host organisms. Based upon recent genome... (Review)
Review
Type II secretion (T2S) is one means by which Gram-negative pathogens secrete proteins into the extracellular milieu and/or host organisms. Based upon recent genome sequencing, it is clear that T2S is largely restricted to the , occurring in many, but not all, genera in the , , , and classes. Prominent human and/or animal pathogens that express a T2S system(s) include , , , , , , , , , and T2S-expressing plant pathogens include , , , , , , and T2S also occurs in nonpathogenic bacteria, facilitating symbioses, among other things. The output of a T2S system can range from only one to dozens of secreted proteins, encompassing a diverse array of toxins, degradative enzymes, and other effectors, including novel proteins. Pathogenic processes mediated by T2S include the death of host cells, degradation of tissue, suppression of innate immunity, adherence to host surfaces, biofilm formation, invasion into and growth within host cells, nutrient assimilation, and alterations in host ion flux. The reach of T2S is perhaps best illustrated by those bacteria that clearly use it for both environmental survival and virulence; e.g., employs T2S for infection of amoebae, growth within lung cells, dampening of cytokines, and tissue destruction. This minireview provides an update on the types of bacteria that have T2S, the kinds of proteins that are secreted via T2S, and how T2S substrates promote infection.
Topics: Bacterial Proteins; Gram-Negative Bacteria; Protein Transport; Type II Secretion Systems; Virulence Factors
PubMed: 28264910
DOI: 10.1128/IAI.00014-17 -
Innovation and Application of the Type III Secretion System Inhibitors in Plant Pathogenic Bacteria.Microorganisms Dec 2020Many Gram-negative pathogenic bacteria rely on a functional type III secretion system (T3SS), which injects multiple effector proteins into eukaryotic host cells, for... (Review)
Review
Many Gram-negative pathogenic bacteria rely on a functional type III secretion system (T3SS), which injects multiple effector proteins into eukaryotic host cells, for their pathogenicity. Genetic studies conducted in different host-microbe pathosystems often revealed a sophisticated regulatory mechanism of their T3SSs, suggesting that the expression of T3SS is tightly controlled and constantly monitored by bacteria in response to the ever-changing host environment. Therefore, it is critical to understand the regulation of T3SS in pathogenic bacteria for successful disease management. This review focuses on a model plant pathogen, , and summarizes the current knowledge of its T3SS regulation. We highlight the roles of several T3SS regulators that were recently discovered, including the transcriptional regulators: FlhDC, RpoS, and SlyA; the post-transcriptional regulators: PNPase, Hfq with its dependent sRNA ArcZ, and the RsmA/B system; and the bacterial second messenger cyclic-di-GMP (c-di-GMP). Homologs of these regulatory components have also been characterized in almost all major bacterial plant pathogens like , , spp., spp., and spp. The second half of this review shifts focus to an in-depth discussion of the innovation and development of T3SS inhibitors, small molecules that inhibit T3SSs, in the field of plant pathology. This includes T3SS inhibitors that are derived from plant phenolic compounds, plant coumarins, and salicylidene acylhydrazides. We also discuss their modes of action in bacteria and application for controlling plant diseases.
PubMed: 33317075
DOI: 10.3390/microorganisms8121956 -
Applied and Environmental Microbiology Nov 2016Modification of teichoic acid through the incorporation of d-alanine confers resistance in Gram-positive bacteria to antimicrobial peptides (AMPs). This process involves...
UNLABELLED
Modification of teichoic acid through the incorporation of d-alanine confers resistance in Gram-positive bacteria to antimicrobial peptides (AMPs). This process involves the products of the dltXABCD genes. These genes are widespread in Gram-positive bacteria, and they are also found in a few Gram-negative bacteria. Notably, these genes are present in all soft-rot enterobacteria (Pectobacterium and Dickeya) whose dltDXBAC operons have been sequenced. We studied the function and regulation of these genes in Dickeya dadantii dltB expression was induced in the presence of the AMP polymyxin. It was not regulated by PhoP, which controls the expression of some genes involved in AMP resistance, but was regulated by ArcA, which has been identified as an activator of genes involved in AMP resistance. However, arcA was not the regulator responsible for polymyxin induction of these genes in this bacterium, which underlines the complexity of the mechanisms controlling AMP resistance in D. dadantii Two other genes involved in resistance to AMPs have also been characterized, phoS and phoH dltB, phoS, phoH, and arcA but not dltD mutants were more sensitive to polymyxin than the wild-type strain. Decreased fitness of the dltB, phoS, and phoH mutants in chicory leaves indicates that their products are important for resistance to plant AMPs.
IMPORTANCE
Gram-negative bacteria can modify their lipopolysaccharides (LPSs) to resist antimicrobial peptides (AMPs). Soft-rot enterobacteria (Dickeya and Pectobacterium spp.) possess homologues of the dlt genes in their genomes which, in Gram-positive bacteria, are involved in resistance to AMPs. In this study, we show that these genes confer resistance to AMPs, probably by modifying LPSs, and that they are required for the fitness of the bacteria during plant infection. Two other new genes involved in resistance were also analyzed. These results show that bacterial resistance to AMPs can occur in bacteria through many different mechanisms that need to be characterized.
Topics: Anti-Bacterial Agents; Antimicrobial Cationic Peptides; Bacterial Proteins; Cichorium intybus; Drug Resistance, Bacterial; Enterobacteriaceae; Gene Expression Regulation, Bacterial; Mutation; Plant Leaves; Polymyxins; Repressor Proteins
PubMed: 27565623
DOI: 10.1128/AEM.01757-16