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Microbiological Research Sep 2021Pseudomonas fluorescens ATCC13525 is an important growth-promoting rhizobacteria (PGPR) and plant disease biocontrol bacterium. However, due to poor stress resistance,...
Pseudomonas fluorescens ATCC13525 is an important growth-promoting rhizobacteria (PGPR) and plant disease biocontrol bacterium. However, due to poor stress resistance, it is prone to be inactivated by preparation, drying and storage. In this study, we investigated the effects of different stress preadaptation methods (2.0∼3.0 wt% NaCl, 0.01∼0.20 wt% HO, and 35∼44 °C) and two stress adaptation genes (rpoS, and hfq) on the stress resistance of P. fluorescens ATCC13525 (PF-WT). After stress preadaptation with low stress of 3.0 wt% NaCl, 0.05 wt% HO, and 41 °C for 30 min, the tolerance of PF-WT toward high lethal stress environments (20.0 wt% NaCl, 1.00 wt% HO, and 47 °C) were significantly improved. Moreover, knockout of rpoS and hfq genes resulted in slower culture growth than PF-WT under the sublethal stress culture conditions (5.0 wt% NaCl, 0.08 wt% HO, and 35 °C), whereas rpoS and hfq overexpressed strains (PF-pBBR2-rpoS and PF-pBBR2-hfq) obviously grew better than the control strain PF-pBBR2. Further, we prepared biocontrol agents (BACs) of different strains after different stress preadaptation treatments. Compared to PF-WT without stress preadaptation, preadaptation by 0.05 wt% HO for 30 min resulted in 5.65 times higher survival rate, while treatment with 3.0 wt% NaCl for 30 min of PF-pBBR2-rpoS led to 5.60 times higher survival rate. This finding provides the simple and effective protection methods for P. fluorescens ATCC13525 BACs preparation by stress preadaptation and overexpression of stress adaptation genes.
Topics: Adaptation, Physiological; Bacterial Proteins; Biological Control Agents; Host Factor 1 Protein; Hydrogen Peroxide; Pseudomonas fluorescens; Sigma Factor; Stress, Physiological
PubMed: 34144508
DOI: 10.1016/j.micres.2021.126804 -
Applied and Environmental Microbiology Jul 2022Pseudomonas fluorescens 2P24 is a beneficial plant root-associated microorganism capable of suppressing several soilborne plant diseases. The capacity of P. fluorescens...
Pseudomonas fluorescens 2P24 is a beneficial plant root-associated microorganism capable of suppressing several soilborne plant diseases. The capacity of P. fluorescens to aggressively colonize the rhizosphere is an important requirement for its biocontrol trait. We previously found that the PcoI/PcoR quorum-sensing system (QS) is involved in regulating the rhizosphere colonization of P. fluorescens. Here, we revealed a sophisticated regulatory network that connects PcoR, RsaL, and MvaT proteins to fine-tune the PcoI/PcoR QS system. Our data showed that PcoR could directly bind to the promoter region of thereby inducing the PcoI/PcoR QS system, whereas RsaL binds simultaneously with PcoR to the promoter region of and represses the PcoR-dependent activation of gene. In addition, RsaL indirectly downregulates the expression of . Furthermore, we showed that disruption of enhanced the expression of , , and , whereas MvaT controls the PcoI/PcoR QS in a RsaL-independent manner. Overall, this study elucidates that PcoR, RsaL, and MvaT regulate the PcoI/PcoR QS through a multi-tiered regulatory mechanism and that PcoR is necessary in the RsaL- and MvaT-mediated repression on the expression of . The PcoI/PcoR quorum-sensing system of Pseudomonas fluorescens 2P24 is important for its effective colonization in the plant rhizosphere. Many regulatory elements appear to directly or indirectly influence the QS system. Here, we found a complex regulatory network employing transcriptional factors PcoR, RsaL, and MvaT to influence the expression of the PcoI/PcoR QS in P. fluorescens 2P24. Our results indicate that PcoR and RsaL directly bind to the promoter region of and then positively and negatively regulate the expression of , respectively. Furthermore, the H-NS family protein MvaT negatively controls the PcoI/PcoR QS in a RsaL-independent manner. Taken together, our data provide new insights into the interplays between different regulatory elements that fine-tune the QS system of P. fluorescens.
Topics: Bacterial Proteins; Gene Expression Regulation, Bacterial; Pseudomonas aeruginosa; Pseudomonas fluorescens; Quorum Sensing; Transcription Factors
PubMed: 35695573
DOI: 10.1128/aem.00625-22 -
Frontiers in Microbiology 2023CFBP2392 has been recognized as a potential biocontrol agent due to its ability to suppress damping-off and root rot disease. This isolate has antibacterial activity...
CFBP2392 has been recognized as a potential biocontrol agent due to its ability to suppress damping-off and root rot disease. This isolate has antibacterial activity as many other strains from the complex. In this work, the antibacterial and antifungal activity of the strain were explored. Dual culture assays evidenced the antifungal activity of the strain against different phytopathogens: sp., , , and . Purification of an antifungal fraction was performed by preparative HPLC from the chemical extraction of growth media. The fraction showed altered growth and ultrastructure. Transmission electron microscopy revealed the purified compound hypertrophied mitochondria, membranous vesicles, and a higher number of vacuoles in cytoplasm. In addition, co-cultivation of CFBP2392 with resulted in an enlarged and deformed cell wall. To gain genomic insights on this inhibition, the complete genome of CFBP2392 was obtained with Oxford Nanopore technology. Different biosynthetic gene clusters (BGCs) involved in specialized metabolites production including a lokisin-like and a koreenceine-like cluster were identified. In accordance with the putative BGCs identified, sequence phylogeny analysis of the MacB transporter in the lokisin-like cluster further supports the similarity with other transporters from the amphisin family. Our results give insights into the cellular effects of the purified microbial metabolite in ultrastructure and provide a genomic background to further explore the specialized metabolite potential.
PubMed: 38033591
DOI: 10.3389/fmicb.2023.1286926 -
Molecules (Basel, Switzerland) Oct 2021A biofungicide is a natural product that can be derived from various sources such as, among others, microorganisms, higher plants, animal products, phytochemicals,...
A biofungicide is a natural product that can be derived from various sources such as, among others, microorganisms, higher plants, animal products, phytochemicals, semiochemicals, and antagonist microorganisms. One of the most important approaches for the production of biofungicides is the combination of biocontrol agents. This study showed the inhibition growth of and treated with cell-free extracts of . Using thin-layer chromatography and plate assays it was also demonstrated that the cell-free extracts of contained siderophores and derivates of 4-diacetylphloroglucinol and phenazine. Moreover, the combination of cell-free extracts of and chitosan [50-1.5% (/)] had a synergistic effect since they notably inhibited the mycelial growth of and . Various morphological alterations to the mycelia and conidia of the treated fungi as a result of this combination were also observed. The present study could be a starting point to control other fungal phytopathogens using different cell-free extracts and chitosan as biocontrol agents.
Topics: Anti-Infective Agents; Cell Extracts; Chitosan; Fungi; Microbial Sensitivity Tests; Plant Diseases; Pseudomonas fluorescens
PubMed: 34770767
DOI: 10.3390/molecules26216359 -
ELife Nov 2019Microscopic water films allow bacteria to survive the seemingly dry surface of plant leaves.
Microscopic water films allow bacteria to survive the seemingly dry surface of plant leaves.
Topics: Plant Leaves; Pseudomonas fluorescens
PubMed: 31674912
DOI: 10.7554/eLife.52123 -
Frontiers in Microbiology 2022A Gram-negative bacteria () was exposed to different concentrations (0, 20, and 40 mg/L) of dimethyl phthalate (DMP) for 8 h, and then Fourier transform infrared...
A Gram-negative bacteria () was exposed to different concentrations (0, 20, and 40 mg/L) of dimethyl phthalate (DMP) for 8 h, and then Fourier transform infrared spectroscopy (FTIR) analysis, lipopolysaccharide content detection, analysis of fatty acids, calcein release test, proteomics, non-targeted metabolomics, and enzyme activity assays were used to evaluate the toxicological effect of DMP on . The results showed that DMP exposure caused an increase in the unsaturated fatty acid/saturated fatty acid (UFA/SFA) ratio and in the release of lipopolysaccharides (LPSs) from the cell outer membrane (OM) of . Moreover, DMP regulated the abundances of phosphatidyl ethanolamine (PE) and phosphatidyl glycerol (PG) of and induced dye leakage from an artificial membrane. Additionally, excessive reactive oxygen species (ROS), malondialdehyde (MDA), and changes in antioxidant enzymes (i.e., catalase [CAT] and superoxide dismutase [SOD]) activities, as well as the inhibition of Ca-Mg-ATPase and Na/K-ATPase activities in , which were induced by the DMP. In summary, DMP could disrupt the lipid asymmetry of the outer membrane, increase the fluidity of the cell membrane, and destroy the integrity of the cell membrane of through lipid peroxidation, oxidative stress, and ion imbalance.
PubMed: 36071970
DOI: 10.3389/fmicb.2022.949590 -
Applied and Environmental Microbiology Sep 2021Within soil, bacteria are found in multispecies communities, where interactions can lead to emergent community properties. Studying bacteria in a social context is...
Exploration of Social Spreading Reveals That This Behavior Is Prevalent among and Pseudomonas fluorescens Isolates and That There Are Variations in the Induction of the Phenotype.
Within soil, bacteria are found in multispecies communities, where interactions can lead to emergent community properties. Studying bacteria in a social context is critical for investigating community-level functions. We previously showed that cocultured Pseudomonas fluorescens Pf0-1 and sp. V48 engage in interspecies social spreading (ISS) on a hard agar surface, a behavior which required close contact and depended on the nutritional environment. Here, we investigate whether social spreading is widespread among P. fluorescens and isolates and whether the requirements for interaction vary. We find that this phenotype is not restricted to the interaction between P. fluorescens Pf0-1 and sp. V48 but is a prevalent behavior found in one clade in the P. fluorescens group and two clades in the genus. We show that the interaction with certain isolates occurred without close contact, indicating induction of spreading by a putative diffusible signal. As with ISS by Pf0-1+V48, the motility of interacting pairs is influenced by the environment, with no spreading behaviors (or induction of motility) observed under high nutrient conditions. While Pf0-1+V48 require low nutrient but high NaCl conditions, in the broader range of interacting pairs, the high salt influence was variable. The prevalence of motility phenotypes observed here and found within the literature indicates that community-induced locomotion in general, and social spreading in particular, is likely important within the environment. It is crucial that we continue to study microbial interactions and their emergent properties to gain a fuller understanding of the functions of microbial communities. Interspecies social spreading (ISS) is an emergent behavior observed when Pseudomonas fluorescens Pf0-1 and sp. V48 interact, during which both species move together across a surface. Importantly, this environment does not permit the movement of either individual species. This group behavior suggests that communities of microbes can function in ways not predictable by knowledge of the individual members. Here, we have asked whether ISS is widespread and thus potentially of importance in soil microbial communities. The significance of this research is the demonstration that surface spreading behaviors are not unique to the Pf0-1-V48 interaction but rather is a more widespread phenomenon observed among members of distinct clades of both P. fluorescens and isolates. Furthermore, we identify differences in mechanisms of signaling and nutritional requirements for ISS. Emergent traits resulting from bacterial interactions are widespread, and their characterization is necessary for a complete understanding of microbial community function.
Topics: Microbial Interactions; Pedobacter; Phenotype; Phylogeny; Pseudomonas fluorescens; RNA, Ribosomal, 16S; Soil Microbiology
PubMed: 34288708
DOI: 10.1128/AEM.01344-21 -
Journal of Bacteriology Oct 2022Cells in microbial communities on surfaces live and divide in close proximity, which greatly enhances the potential for social interactions. Spatiogenetic structures are...
Cells in microbial communities on surfaces live and divide in close proximity, which greatly enhances the potential for social interactions. Spatiogenetic structures are manifested through competitive and cooperative interactions among the same and different genotypes within a shared space, and extracellular secretions appear to function dynamically at the forefront. A previous experimental evolution study utilizing Pseudomonas fluorescens Pf0-1 colonies demonstrated that diverse mutations in the gene were repeatedly and exclusively selected through the formation of a dominant spatial structure. RsmE's primary molecular function is translation repression, and its homologs regulate various social and virulence phenotypes. Pseudomonas spp. possess multiple paralogs of Rsm proteins, and RsmA, RsmE, and RsmI are the most prevalent. Here, we demonstrate that the production of a mucoid polymer and a biosurfactant are exclusively regulated through RsmE, contradicting the generalized notion of functional redundancy among the Rsm paralogs. Furthermore, we identified the biosurfactant as the cyclic lipopeptide gacamide A. Competition and microscopy analyses showed that the mucoid polymer is solely responsible for creating a space of low cellular density, which is shared exclusively by the same genotype. Gacamide A and other RsmE-regulated products appear to establish a physical boundary that prevents the encroachment of the competing genotype into the newly created space. Although cyclic lipopeptides and other biosurfactants are best known for their antimicrobial properties and reducing surface tension to promote the spreading of cells on various surfaces, they also appear to help define spatial structure formation within a dense community. In densely populated colonies of the bacterium Pseudomonas fluorescens Pf0-1, diverse mutations in the gene are naturally selected by solving the problem of overcrowding. Here, we show that RsmE-regulated secretions function together to create and protect space of low cell density. A biosurfactant generally promotes the spreading of bacterial cells on abiotic surfaces; however, it appears to function atypically within a crowded population by physically defining genotypic boundaries. Another significant finding is that these secretions are not regulated by RsmE's paralogs that share high sequence similarity. The experimental pipeline described in this study is highly tractable and should facilitate future studies to explore additional RsmE-regulated products and address why RsmE is functionally unique from its paralogs.
Topics: Pseudomonas fluorescens; Gene Expression Regulation, Bacterial; Bacterial Proteins; Pseudomonas; Peptides, Cyclic; Lipopeptides; Polymers
PubMed: 36165622
DOI: 10.1128/jb.00285-22 -
Microorganisms Mar 2023Biofilms are often tolerant towards routine cleaning and disinfection processes. As they can grow on fabrics in household or healthcare settings, resulting in odors and...
Biofilms are often tolerant towards routine cleaning and disinfection processes. As they can grow on fabrics in household or healthcare settings, resulting in odors and serious health problems, it is necessary to contain biofilms through eradication strategies. The current study proposes a novel test model for the growth and removal of biofilms on textiles with and the opportunistic nosocomial pathogen as model organisms. To assess the biofilm removal on fabrics, (1) a detergent-based, (2) enzyme-based, and (3) combined formulation of both detergent and enzymes (F1/2) were applied. Biofilms were analyzed microscopically (FE-SEM, SEM, 3D laser scanning- and epifluorescence microscopy), via a quartz crystal microbalance with mass dissipation monitoring (QCM-D) as well as plate counting of colonies. This study indicated that spp. form robust biofilms on woven cellulose that can be efficiently removed via F1/2, proven by a significant reduction ( < 0.001) of viable bacteria in biofilms. Moreover, microscopic analysis indicated a disruption and almost complete removal of the biofilms after F1/2 treatment. QCM-D measurements further confirmed a maximal mass dissipation change after applying F1/2. The combination strategy applying both enzymes and detergent is a promising antibiofilm approach to remove bacteria from fabrics.
PubMed: 37110314
DOI: 10.3390/microorganisms11040892 -
Frontiers in Microbiology 2023Bacterial communication is a fundamental process used to synchronize gene expression and collective behavior among the bacterial population. The most studied bacterial...
Bacterial communication is a fundamental process used to synchronize gene expression and collective behavior among the bacterial population. The most studied bacterial communication system is quorum sensing, a cell density system, in which the concentration of inductors increases to a threshold level allowing detection by specific receptors. As a result, bacteria can change their behavior in a coordinated way. While in quorum sensing based on the synthesis of -acyl homoserine lactone molecules is well studied, volatile organic compounds, although considered to be communication signals in the rhizosphere, are understudied. The MFE01 strain has a very active type six secretion system that can kill some competitive bacteria. Furthermore, MFE01 emits numerous volatile organic compounds, including 1-undecene, which contributes to the aerial inhibition of growth. Finally, MFE01 appears to be deprived of -acyl homoserine lactone synthase. The main objective of this study was to explore the role of 1-undecene in the communication of MFE01. We constructed a mutant affected in gene encoding the enzyme responsible for 1-undecene synthesis to provide further insight into the role of 1-undecene in MFE01. First, we studied the impacts of this mutation both on volatile organic compounds emission, using headspace solid-phase microextraction combined with gas chromatography-mass spectrometry and on long-range inhibition. Then, we analyzed influence of 1-undecene on MFE01 coordinated phenotypes, including type six secretion system activity and biofilm formation. Next, to test the ability of MFE01 to synthesize -acyl homoserine lactones in our conditions, we investigated the presence of corresponding genes across the MFE01 genome and we exposed its biofilms to an acyl homoserine lactone-degrading enzyme. Finally, we examined the effects of 1-undecene emission on MFE01 biofilm maturation and aerial communication using an original experimental set-up. This study demonstrated that the Δ mutant is impaired in biofilm maturation. An exposure of the Δ mutant to the volatile compounds emitted by MFE01 during the biofilm development restored the biofilm maturation process. These findings indicate that MFE01 uses 1-undecene emission for aerial communication, reporting for the first time this volatile organic compound as bacterial intraspecific communication signal.
PubMed: 37908545
DOI: 10.3389/fmicb.2023.1264801