scholarly journals The carbon source-dependent pattern of antimicrobial activity and gene expression in Pseudomonas donghuensis P482

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Marta Matuszewska ◽  
Tomasz Maciąg ◽  
Magdalena Rajewska ◽  
Aldona Wierzbicka ◽  
Sylwia Jafra
Keyword(s):  
Gene Expression ◽  
Antimicrobial Activity ◽  
Carbon Source ◽  
Reference Genes ◽  
Plant Pathogens ◽  
Plant Protection ◽  
Carbon Sources ◽  
Unknown Compound ◽  
Fungal Plant Pathogens ◽  
The Impact

AbstractPseudomonas donghuensis P482 is a tomato rhizosphere isolate with the ability to inhibit growth of bacterial and fungal plant pathogens. Herein, we analysed the impact of the carbon source on the antibacterial activity of P482 and expression of the selected genes of three genomic regions in the P482 genome. These regions are involved in the synthesis of pyoverdine, 7-hydroxytropolone (7-HT) and an unknown compound (“cluster 17”) and are responsible for the antimicrobial activity of P482. We showed that the P482 mutants, defective in these regions, show variations and contrasting patterns of growth inhibition of the target pathogen under given nutritional conditions (with glucose or glycerol as a carbon source). We also selected and validated the reference genes for gene expression studies in P. donghuensis P482. Amongst ten candidate genes, we found gyrB, rpoD and mrdA the most stably expressed. Using selected reference genes in RT-qPCR, we assessed the expression of the genes of interest under minimal medium conditions with glucose or glycerol as carbon sources. Glycerol was shown to negatively affect the expression of genes necessary for 7-HT synthesis. The significance of this finding in the light of the role of nutrient (carbon) availability in biological plant protection is discussed.

scholarly journals Necessity requires no decision: carbon source-dependent pattern of antimicrobial activity and gene expression in Pseudomonas donghuensis P482

2021 ◽  
Author(s):  
Marta Matuszewska ◽  
Tomasz Maciag ◽  
Magdalena Rajewska ◽  
Aldona Wierzbicka ◽  
Sylwia Jafra
Keyword(s):  
Gene Expression ◽  
Antimicrobial Activity ◽  
Carbon Source ◽  
Reference Genes ◽  
Plant Pathogens ◽  
Plant Protection ◽  
Carbon Sources ◽  
Unknown Compound ◽  
Fungal Plant Pathogens ◽  
The Impact

Abstract Pseudomonas donghuensis P482 is a tomato rhizosphere isolate with the ability to inhibit growth of bacterial and fungal plant pathogens. Herein, we analysed the impact of the carbon source on the antibacterial activity of P482 and expression of the selected genes of three genomic regions in the P482 genome. These regions are involved in the synthesis of pyoverdine, 7‑hydroxytropolone (7-HT) and an unknown compound (“cluster 17”) and are responsible for antimicrobial activity of P482. We showed that the P482 mutants, defective in these regions, show variations and contrasting patterns of growth inhibition of the target pathogen under given nutritional conditions (with glucose or glycerol as a carbon source). We also selected and validated the reference genes for gene expression studies in P. donghuensis P482. Amongst ten candidate genes, we found gyrB, rpoD and mrdA the most stably expressed. Using selected reference genes in RT-qPCR, we assessed the expression of the genes of interest under minimal medium conditions with glucose or glycerol as carbon sources. Glycerol was shown to negatively affect the expression of genes necessary for 7-HT synthesis. The significance of this finding in the light of the role of nutrient (carbon) availability in biological plant protection is discussed.

scholarly journals The Cyclic AMP Receptor Protein Regulates Quorum Sensing and Global Gene Expression in Yersinia pestis during Planktonic Growth and Growth in Biofilms

mBio ◽  
2019 ◽  
Vol 10 (6) ◽  
Author(s):  
Jeremy T. Ritzert ◽  
George Minasov ◽  
Ryan Embry ◽  
Matthew J. Schipma ◽  
Karla J. F. Satchell
Keyword(s):  
Gene Expression ◽  
Quorum Sensing ◽  
Carbon Source ◽  
Cyclic Amp ◽  
Yersinia Pestis ◽  
Carbon Sources ◽  
Transcriptional Regulator ◽  
Receptor Protein ◽  
Content Type ◽  
Bacterial Physiology

ABSTRACT Cyclic AMP (cAMP) receptor protein (Crp) is an important transcriptional regulator of Yersinia pestis. Expression of crp increases during pneumonic plague as the pathogen depletes glucose and forms large biofilms within lungs. To better understand control of Y. pestis Crp, we determined a 1.8-Å crystal structure of the protein-cAMP complex. We found that compared to Escherichia coli Crp, C helix amino acid substitutions in Y. pestis Crp did not impact the cAMP dependency of Crp to bind DNA promoters. To investigate Y. pestis Crp-regulated genes during plague pneumonia, we performed RNA sequencing on both wild-type and Δcrp mutant bacteria growing in planktonic and biofilm states in minimal media with glucose or glycerol. Y. pestis Crp was found to dramatically alter expression of hundreds of genes in a manner dependent upon carbon source and growth state. Gel shift assays confirmed direct regulation of the malT and ptsG promoters, and Crp was then linked to Y. pestis growth on maltose as a sole carbon source. Iron regulation genes ybtA and fyuA were found to be indirectly regulated by Crp. A new connection between carbon source and quorum sensing was revealed as Crp was found to regulate production of acyl-homoserine lactones (AHLs) through direct and indirect regulation of genes for AHL synthetases and receptors. AHLs were subsequently identified in the lungs of Y. pestis-infected mice when crp expression was highest in Y. pestis biofilms. Thus, in addition to the well-studied pla gene, other Crp-regulated genes likely have important functions during plague infection. IMPORTANCE Bacterial pathogens have evolved extensive signaling pathways to translate environmental signals into changes in gene expression. While Crp has long been appreciated for its role in regulating metabolism of carbon sources in many bacterial species, transcriptional profiling has revealed that this protein regulates many other aspects of bacterial physiology. The plague pathogen Y. pestis requires this global regulator to survive in blood, skin, and lungs. During disease progression, this organism adapts to changes within these niches. In addition to regulating genes for metabolism of nonglucose sugars, we found that Crp regulates genes for virulence, metal acquisition, and quorum sensing by direct or indirect mechanisms. Thus, this single transcriptional regulator, which responds to changes in available carbon sources, can regulate multiple critical behaviors for causing disease.

Gene expression profiles based flux balance model to predict the carbon source for Bacillus subtilis

2019 ◽  
Author(s):  
Kulwadee Thanamit ◽  
Franziska Hoerhold ◽  
Marcus Oswald ◽  
Rainer Koenig
Keyword(s):  
Gene Expression ◽  
Carbon Source ◽  
Drug Targets ◽  
Expression Profiles ◽  
Carbon Sources ◽  
Gene Expression Profiles ◽  
Fast Method ◽  
Flux Balance ◽  
Metabolic Fluxes

ABSTRACTFinding drug targets for antimicrobial treatment is a central focus in biomedical research. To discover new drug targets, we developed a method to identify which nutrients are essential for microorganisms. Using 13C labeled metabolites to infer metabolic fluxes is the most informative way to infer metabolic fluxes to date. However, the data can get difficult to acquire in complicated environments, for example, if the pathogen homes in host cells. Although data from gene expression profiling is less informative compared to metabolic tracer derived data, its generation is less laborious, and may still provide the relevant information. Besides this, metabolic fluxes have been successfully predicted by flux balance analysis (FBA). We developed an FBA based approach using the stoichiometric knowledge of the metabolic reactions of a cell combining them with expression profiles of the coding genes. We aimed to identify essential drug targets for specific nutritional uptakes of microorganisms. As a case study, we predicted each single carbon source out of a pool of eight different carbon sources for B. subtilis based on gene expression profiles. The models were in good agreement to models basing on 13C metabolic flux data of the same conditions. We could well predict every carbon source. Later, we applied successfully the model to unseen data from a study in which the carbon source was shifted from glucose to malate and vice versa. Technically, we present a new and fast method to reduce thermodynamically infeasible loops, which is a necessary preprocessing step for such model-building algorithms.SIGNIFICANCEIdentifying metabolic fluxes using 13C labeled tracers is the most informative way to gain insight into metabolic fluxes. However, obtaining the data can be laborious and challenging in a complex environment. Though transcriptional data is an indirect mean to estimate the fluxes, it can help to identify this. Here, we developed a new method employing constraint-based modeling to predict metabolic fluxes embedding gene expression profiles in a linear regression model. As a case study, we used the data from Bacillus subtilis grown under different carbon sources. We could well predict the correct carbon source. Additionally, we established a novel and fast method to remove thermodynamically infeasible loops.

scholarly journals De Novo Design of Antimicrobial Peptides With a Special Charge Pattern and Their Application in Combating Plant Pathogens

2021 ◽  
Vol 12 ◽  
Author(s):  
Eric H. -L. Chen ◽  
Cheng-Wei Weng ◽  
Yi-Min Li ◽  
Ming-Chin Wu ◽  
Chien-Chih Yang ◽  
...  
Keyword(s):  
Antimicrobial Activity ◽  
Antimicrobial Peptides ◽  
Plant Pathogens ◽  
De Novo ◽  
Plant Protection ◽  
Acyl Chain ◽  
Fatty Acyl ◽  
Plant Diseases ◽  
Basic Residue ◽  
Wide Range

Plant diseases are important issues in agriculture, and the development of effective and environment-friendly means of disease control is crucial and highly desired. Antimicrobial peptides (AMPs) are known as potential alternatives to chemical pesticides because of their potent broad-spectrum antimicrobial activity and because they have no risk, or have only a low risk, of developing chemical-resistant pathogens. In this study, we designed a series of amphipathic helical peptides with different spatial distributions of positive charges and found that the peptides that had a special sequence pattern “BBHBBHHBBH” (“B” for basic residue and “H” for hydrophobic residue) displayed excellent bactericidal and fungicidal activities in a wide range of economically important plant pathogens. The peptides with higher helical propensity had lower antimicrobial activity. When we modified the peptides with a long acyl chain at their N-terminus, their plant protection effect improved. Our application of the fatty acyl-modified peptides on the leaves of tomato and Arabidopsis plants lessened the infection caused by Pectobacterium carotovorum subsp. carotovorum and Botrytis cinerea. Our study provides important insights on the development of more potent novel AMPs for plant protection.

Cellular and environmental factors influencing biofilm formation and colonization of plant tissue by a beneficial strain of bacteria, Pseudomonas donghuensis P482.

2020 ◽  
Author(s):  
Magdalena Rajewska ◽  
Marta Matuszewska ◽  
Sylwia Jafra
Keyword(s):  
Antimicrobial Activity ◽  
Environmental Factors ◽  
Biofilm Formation ◽  
Genetic Background ◽  
Plant Pathogens ◽  
Knock Out ◽  
Factors Influencing ◽  
Fungal Plant Pathogens ◽  
Artificial Surfaces ◽  
Strain Type

<p>The ability to colonize different environmental niches by bacteria is most often determined by the ability to form biofilms - complex, multicellular communities. This, in turn, depends on both cellular and extracellular factors such as genetic background of the strain, type of surface (biotic or abiotic) to which bacteria attach, availability of nutrients, temperature, <em>etc</em>. <em>Pseudomonas donghuensis</em> P482 strain is a little-known isolate from tomato rhizosphere, exhibiting antimicrobial activity towards bacterial and fungal plant pathogens. Studies have shown that it efficiently colonizes plant rhizosphere and forms biofilm on artificial surfaces. Which genetic or environmental factors underlie the mechanism of biofilm formation were yet to be elucidated. The presented research aimed at identifying those factors. Basing on the analysis of genome, knock-out mutants of the P482 strain were constructed in the genes potentially involved in biofilm formation and further analyzed for motility, colony morphology, attachment to artificial surfaces in different culture conditions, and colonization of maize and tomato rhizosphere.</p>

scholarly journals Reference Gene Selection for Gene Expression Analyses in Mouse Models of Acute Lung Injury

2021 ◽  
Vol 22 (15) ◽  
pp. 7853
Author(s):  
Athanassios Fragoulis ◽  
Kristina Biller ◽  
Stephanie Fragoulis ◽  
Dennis Lex ◽  
Stefan Uhlig ◽  
...  
Keyword(s):  
Gene Expression ◽  
Acute Lung Injury ◽  
Lung Injury ◽  
Reference Gene ◽  
Reference Genes ◽  
Gene Selection ◽  
Ex Vivo ◽  
Qrt Pcr ◽  
The Impact

qRT-PCR still remains the most widely used method for quantifying gene expression levels, although newer technologies such as next generation sequencing are becoming increasingly popular. A critical, yet often underappreciated, problem when analysing qRT-PCR data is the selection of suitable reference genes. This problem is compounded in situations where up to 25% of all genes may change (e.g., due to leukocyte invasion), as is typically the case in ARDS. Here, we examined 11 widely used reference genes for their suitability in commonly used models of acute lung injury (ALI): ventilator-induced lung injury (VILI), in vivo and ex vivo, lipopolysaccharide plus mechanical ventilation (MV), and hydrochloric acid plus MV. The stability of reference gene expression was determined using the NormFinder, BestKeeper, and geNorm algorithms. We then proceeded with the geNorm results because this is the only algorithm that provides the number of reference genes required to achieve normalisation. We chose interleukin-6 (Il‑6) and C-X-C motif ligand 1 (Cxcl-1) as the genes of interest to analyse and demonstrate the impact of inappropriate normalisation. Reference gene stability differed between the ALI models and even within the subgroup of VILI models, no common reference gene index (RGI) could be determined. NormFinder, BestKeeper, and geNorm produced slightly different, but comparable results. Inappropriate normalisation of Il-6 and Cxcl1 gene expression resulted in significant misinterpretation in all four ALI settings. In conclusion, choosing an inappropriate normalisation strategy can introduce different kinds of bias such as gain or loss as well as under- or overestimation of effects, affecting the interpretation of gene expression data.

scholarly journals The Cyclic AMP Receptor Protein Regulates Quorum Sensing and Global Gene Expression in Yersinia pestis During Planktonic Growth and Growth in Biofilms

2019 ◽  
Author(s):  
Jeremy T. Ritzert ◽  
George Minasov ◽  
Ryan Embry ◽  
Matthew J. Schipma ◽  
Karla J. F. Satchell
Keyword(s):  
Gene Expression ◽  
Quorum Sensing ◽  
Carbon Source ◽  
Yersinia Pestis ◽  
Bacterial Species ◽  
Carbon Sources ◽  
Transcriptional Regulator ◽  
Adenosine Monophosphate ◽  
Receptor Protein ◽  
Bacterial Physiology

ABSTRACTCyclic adenosine monophosphate (cAMP) receptor protein (Crp) is an important transcriptional regulator of Yersinia pestis. Expression of crp increases during pneumonic plague as the pathogen depletes glucose and forms large biofilms within lungs. To better understand control of Y. pestis Crp, we determined a 1.8 Å crystal structure of the protein-cAMP complex. We found that compared to Escherichia coli Crp, C helix amino acid substitutions in Y. pestis Crp did not impact cAMP dependency of Crp to bind DNA promoters. To investigate Y. pestis Crp-regulated genes during plague pneumonia, we performed RNA-sequencing on both wild-type and Δcrp mutant bacteria growing in planktonic and biofilm states in minimal media with glucose or glycerol. Y. pestis Crp is found to dramatically alter expression of hundreds of genes dependent upon carbon source and growth state. Gel shift assays confirmed direct regulation of the malT and ptsG promoters and Crp was then linked to Y. pestis growth on maltose as a sole carbon source. Iron-regulation genes ybtA and fyuA were found to be indirectly regulated by Crp. A new connection between carbon source and quorum sensing was revealed as Crp was found to regulate production of acyl-homoserine lactones (AHLs) through direct and indirect regulation of genes for AHL synthetases and receptors. AHLs were subsequently identified in the lungs of Y. pestis infected mice when crp expression is highest in Y. pestis biofilms. Thus, in addition to well-studied pla, other Crp-regulated genes likely have important functions during plague infection.IMPORTANCEBacterial pathogens have evolved extensive signaling pathways to translate environmental signals into changes in gene expression. While Crp has long been appreciated for its role in regulating metabolism of carbon sources in many bacterial species, transcriptional profiling has revealed that this protein regulates many other aspects of bacterial physiology. The plague pathogen, Y. pestis, requires this global regulator to survive in blood, skin, and lungs. During disease progression, this organism adapts to changes within these niches. In addition to regulating genes for metabolism of non-glucose sugars, we find the Crp regulates genes for virulence, metal acquisition and quorum sensing by direct or indirect mechanisms. Thus, this single transcriptional regulator, that responds to changes in available carbon sources, can regulate multiple critical behaviors for causing disease.

Production, partial purification, and antimicrobial activity of a novel antibiotic produced by a Bacillus subtilis isolate from Ulmus americana

1988 ◽  
Vol 66 (11) ◽  
pp. 2338-2346 ◽  
Author(s):  
L. R. Schreiber ◽  
Garold F. Gregory ◽  
C. R. Krause ◽  
J. M. Ichida
Keyword(s):  
Antimicrobial Activity ◽  
Bacillus Subtilis ◽  
High Performance ◽  
Plant Pathogens ◽  
Antibiotic Activity ◽  
Dutch Elm Disease ◽  
Absolute Methanol ◽  
Partial Purification ◽  
Ulmus Americana ◽  
Fungal Plant Pathogens

Antibiotic substances produced by a Bacillus subtilis isolate from the xylem of an American elm (Ulmus americana L.) inhibited several fungal plant pathogens including the Dutch elm disease pathogen, Ceratocystis ulrni. Bioassays indicated the bacterium produced a diffusible, methanol-soluble antibiotic on potato dextrose agar at 30 °C. Cellulose powder column chromatography separated the antibiotic, which was further purified on a Sephadex LH-20 column, yielding two peaks of antibiotic activity, BS1 and BS2. These were further purified by paper chromatography. High-performance liquid chromatography, using C-18 reverse phase chromatography under isocratic conditions, resulted in five 280-nm absorbing peaks. Only one peak contained antibiotic activity. Upon ultrafiltration, the antibiotics passed through a 500 mol. wt. filter. The antibiotic was soluble in water, absolute methanol, ethanol, and chloroform but not in hexane or petroleum ether. Antimicrobial activity differed from that of other B. subtilis antibiotics including iturin A, bacillomycin L, mycosubtilin, fengycin, and bacilysin. Ceratocystis ulmi exposed to BS1 produced short, twisted, and swollen hyphae with irregularly thickened cell walls.

scholarly journals Intact double stranded RNA is mobile and triggers RNAi against viral and fungal plant pathogens

2021 ◽  
Author(s):  
Christopher A Brosnan ◽  
Anne Sawyer ◽  
Filipe Fenselau Felippes ◽  
Bernard J Carroll ◽  
Peter M Waterhouse ◽  
...  
Keyword(s):  
Topical Application ◽  
Plant Pathogens ◽  
Plant Protection ◽  
Critical Role ◽  
Crop Protection ◽  
Double Stranded Rna ◽  
Ability To Act ◽  
Fungal Plant Pathogens ◽  
Transgenic Breeding ◽  
Protection Strategies

Topical application of double-stranded RNA (dsRNA) as RNA interference(RNAi) based biopesticides represents a sustainable alternative to traditional transgenic, breeding-based or chemical crop protection strategies. A key feature of RNAi is its ability to act non-cell autonomously, a process that plays a critical role in plant protection. However, the uptake of dsRNA upon topical application, and its ability to move and act non-cell autonomously remains debated and largely unexplored. Here we show that when applied to a leaf, unprocessed full-length dsRNA enters the vasculature and rapidly moves to multiple distal below ground, vegetative and reproductive tissue types in several model plant and crop hosts. Intact unprocessed dsRNA was detected in the apoplast of leaves, roots and flowers after leaf application and maintained in subsequent new growth. Furthermore, we show mobile dsRNA is functional against root infecting fungal and foliar viral pathogens. Our demonstration of the uptake and maintained movement of intact and functional dsRNA stands to add significant benefit to the emerging field of RNAi-based plant protection.

Sign in / Sign up

Export Citation Format

Share Document