scholarly journals Dechlorination of Chloral Hydrate Is Influenced by the Biofilm Adhesin Protein LapA in Pseudomonas putida LF54

2013 ◽  
Vol 79 (13) ◽  
pp. 4166-4169
Author(s):  
Wanjun Zhang ◽  
Huhe ◽  
Yuanbai Pan ◽  
Masanori Toyofuku ◽  
Nobuhiko Nomura ◽  
...  
Keyword(s):  
Pseudomonas Putida ◽  
Biofilm Formation ◽  
Chloral Hydrate ◽  
Insertion Mutagenesis ◽  
Surface Adhesion ◽  
Content Type ◽  
Adhesion Protein ◽  
Transposon Insertion ◽  
Adhesin Protein ◽  
First Time

ABSTRACTLapA is the largest surface adhesion protein ofPseudomonas putidathat initiates biofilm formation. Here, by using transposon insertion mutagenesis and a conditionallapAmutant, we demonstrate for the first time that LapA influences chloral hydrate (CH) dechlorination inP. putidaLF54.

scholarly journals Development of a Method for Markerless Gene Deletion in Pseudomonas putida

2011 ◽  
Vol 77 (15) ◽  
pp. 5549-5552 ◽  
Author(s):  
Nadja Graf ◽  
Josef Altenbuchner
Keyword(s):  
Pseudomonas Putida ◽  
Gene Deletion ◽  
Protein Gene ◽  
Surface Adhesion ◽  
Content Type ◽  
Adhesion Protein ◽  
Uracil Phosphoribosyltransferase ◽  
Flagellum Biosynthesis

ABSTRACTWe developed a negative counterselection system forPseudomonas putidabased on uracil phosphoribosyltransferase (UPRTase) and sensitivity against the antimetabolite 5-fluorouracil (5-FU). We constructed aP. putidastrain that is resistant to 5-FU and constructed vectors for the deletion of the surface adhesion protein gene, the flagellum biosynthesis operon, and two endonuclease genes. The genes were efficiently disrupted and left a markerless chromosomal in-frame deletion.

scholarly journals Genome-Wide Investigation of Biofilm Formation in Bacillus cereus

2017 ◽  
Vol 83 (13) ◽  
Author(s):  
Fang Yan ◽  
Yiyang Yu ◽  
Kevin Gozzi ◽  
Yun Chen ◽  
Jian-hua Guo ◽  
...  
Keyword(s):  
Bacillus Cereus ◽  
Biofilm Formation ◽  
Differentially Expressed ◽  
Insertion Mutagenesis ◽  
Rna Seq ◽  
Content Type ◽  
Transposon Insertion ◽  
Genome Wide ◽  
A Genome ◽  
Transcriptome Analyses

ABSTRACT Bacillus cereus is a soil-dwelling Gram-positive bacterium capable of forming structured multicellular communities, or biofilms. However, the regulatory pathways controlling biofilm formation are less well understood in B. cereus. In this work, we developed a method to study B. cereus biofilms formed at the air-liquid interface. We applied two genome-wide approaches, random transposon insertion mutagenesis to identify genes that are potentially important for biofilm formation, and transcriptome analyses by RNA sequencing (RNA-seq) to characterize genes that are differentially expressed in B. cereus when cells were grown in a biofilm-inducing medium. For the first approach, we identified 23 genes whose disruption by transposon insertion led to altered biofilm phenotypes. Based on the predicted function, they included genes involved in processes such as nucleotide biosynthesis, iron salvage, and antibiotic production, as well as genes encoding an ATP-dependent protease and transcription regulators. Transcriptome analyses identified about 500 genes that were differentially expressed in cells grown under biofilm-inducing conditions. One particular set of those genes may contribute to major metabolic shifts, leading to elevated production of small volatile molecules. Selected volatile molecules were shown to stimulate robust biofilm formation in B. cereus. Our studies represent a genome-wide investigation of B. cereus biofilm formation. IMPORTANCE In this work, we established a robust method for B. cereus biofilm studies and applied two genome-wide approaches, transposon insertion mutagenesis and transcriptome analyses by RNA-seq, to identify genes and pathways that are potentially important for biofilm formation in B. cereus. We discovered dozens of genes and two major metabolic shifts that seem to be important for biofilm formation in B. cereus. Our study represents a genome-wide investigation on B. cereus biofilm formation.

scholarly journals A Novel Gene Involved in the Survival of Streptococcus mutans under Stress Conditions

2013 ◽  
Vol 80 (1) ◽  
pp. 97-103 ◽  
Author(s):  
Dan Li ◽  
Yukie Shibata ◽  
Toru Takeshita ◽  
Yoshihisa Yamashita
Keyword(s):  
Streptococcus Mutans ◽  
Streptococcus Pyogenes ◽  
Biofilm Formation ◽  
Acid Tolerance ◽  
Stress Conditions ◽  
Insertion Mutagenesis ◽  
Trna Modification ◽  
Content Type ◽  
Virulence Proteins ◽  
Random Insertion

ABSTRACTAStreptococcus mutansmutant defective in aciduricity was constructed by random-insertion mutagenesis. Sequence analysis of the mutant revealed a mutation ingidA, which is known to be involved in tRNA modification inStreptococcus pyogenes. Complementation ofgidAbyS. pyogenesgidArecovered the acid tolerance ofS. mutans. Although thegidA-inactivatedS. pyogenesmutant exhibited significantly reduced expression of multiple extracellular virulence proteins, theS. mutansmutant did not. On the other hand, thegidAmutant ofS. mutansshowed reduced ability to withstand exposure to other stress conditions (high osmotic pressure, high temperature, and bacitracin stress) besides an acidic environment. In addition, loss of GidA decreased the capacity for glucose-dependent biofilm formation by over 50%. This study revealed thatgidAplays critical roles in the survival ofS. mutansunder stress conditions, including lower pH.

PBP4 and PBP5 are involved in regulating exopolysaccharide synthesis during Escherichia coli biofilm formation

Microbiology ◽  
2021 ◽  
Vol 167 (3) ◽  
Author(s):  
Sathi Mallick ◽  
Shanti Kiran ◽  
Tapas Kumar Maiti ◽  
Anindya S. Ghosh
Keyword(s):  
Escherichia Coli ◽  
Biofilm Formation ◽  
Type Species ◽  
Pentose Phosphate ◽  
Bacterial Cells ◽  
Surface Adhesion ◽  
Content Type ◽  
Penicillin Binding Proteins ◽  
E Coli ◽  
Link Type

Escherichia coli low-molecular-mass (LMM) Penicillin-binding proteins (PBPs) help in hydrolysing the peptidoglycan fragments from their cell wall and recycling them back into the growing peptidoglycan matrix, in addition to their reported involvement in biofilm formation. Biofilms are external slime layers of extra-polymeric substances that sessile bacterial cells secrete to form a habitable niche for themselves. Here, we hypothesize the involvement of Escherichia coli LMM PBPs in regulating the nature of exopolysaccharides (EPS) prevailing in its extra-polymeric substances during biofilm formation. Therefore, this study includes the assessment of physiological characteristics of E. coli CS109 LMM PBP deletion mutants to address biofilm formation abilities, viability and surface adhesion. Finally, EPS from parent CS109 and its ΔPBP4 and ΔPBP5 mutants were purified and analysed for sugars present. Deletions of LMM PBP reduced biofilm formation, bacterial adhesion and their viability in biofilms. Deletions also diminished EPS production by ΔPBP4 and ΔPBP5 mutants, purification of which suggested an increased overall negative charge compared with their parent. Also, EPS analyses from both mutants revealed the appearance of an unusual sugar, xylose, that was absent in CS109. Accordingly, the reason for reduced biofilm formation in LMM PBP mutants may be speculated as the subsequent production of xylitol and a hindrance in the standard flow of the pentose phosphate pathway.

scholarly journals Pseudomonas aeruginosa Requires the DNA-Specific Endonuclease EndA To Degrade Extracellular Genomic DNA To Disperse from the Biofilm

2019 ◽  
Vol 201 (18) ◽  
Author(s):  
Kathryn E. Cherny ◽  
Karin Sauer
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Biofilm Formation ◽  
Genomic Dna ◽  
Early Stage ◽  
Content Type ◽  
Biofilm Dispersion ◽  
Biofilm Structure ◽  
First Time ◽  
Biofilm Matrix ◽  
Dispersed Cells

ABSTRACT The dispersion of biofilms is an active process resulting in the release of planktonic cells from the biofilm structure. While much is known about the process of dispersion cue perception and the subsequent modulation of the c-di-GMP pool, little is known about subsequent events resulting in the release of cells from the biofilm. Given that dispersion coincides with void formation and an overall erosion of the biofilm structure, we asked whether dispersion involves degradation of the biofilm matrix. Here, we focused on extracellular genomic DNA (eDNA) due to its almost universal presence in the matrix of biofilm-forming species. We identified two probable nucleases, endA and eddB, and eddA encoding a phosphatase that were significantly increased in transcript abundance in dispersed cells. However, only inactivation of endA but not eddA or eddB impaired dispersion by Pseudomonas aeruginosa biofilms in response to glutamate and nitric oxide (NO). Heterologously produced EndA was found to be secreted and active in degrading genomic DNA. While endA inactivation had little effect on biofilm formation and the presence of eDNA in biofilms, eDNA degradation upon induction of dispersion was impaired. In contrast, induction of endA expression coincided with eDNA degradation and resulted in biofilm dispersion. Thus, released cells demonstrated a hyperattaching phenotype but remained as resistant to tobramycin as biofilm cells from which they egress, indicating EndA-dispersed cells adopted some but not all of the phenotypes associated with dispersed cells. Our findings indicate for the first time a role of DNase EndA in dispersion and suggest weakening of the biofilm matrix is a requisite for biofilm dispersion. IMPORTANCE The finding that exposure to DNase I impairs biofilm formation or leads to the dispersal of early stage biofilms has led to the realization of extracellular genomic DNA (eDNA) as a structural component of the biofilm matrix. However, little is known about the contribution of intrinsic DNases to the weakening of the biofilm matrix and dispersion of established biofilms. Here, we demonstrate for the first time that nucleases are induced in dispersed Pseudomonas aeruginosa cells and are essential to the dispersion response and that degradation of matrix eDNA by endogenously produced/secreted EndA is required for P. aeruginosa biofilm dispersion. Our findings suggest that dispersing cells mediate their active release from the biofilm matrix via the induction of nucleases.

scholarly journals Targeting Fibronectin To Disrupt In Vivo Candida albicans Biofilms

2016 ◽  
Vol 60 (5) ◽  
pp. 3152-3155 ◽  
Author(s):  
Jeniel E. Nett ◽  
Jonathan Cabezas-Olcoz ◽  
Karen Marchillo ◽  
Deane F. Mosher ◽  
David R. Andes
Keyword(s):  
Candida Albicans ◽  
Biofilm Formation ◽  
Drug Targets ◽  
Venous Catheter ◽  
Surface Adhesion ◽  
Inhibitory Protein ◽  
Content Type ◽  
Novel Target

ABSTRACTNew drug targets are of great interest for the treatment of fungal biofilms, which are routinely resistant to antifungal therapies. We theorized that the interaction ofCandida albicanswith matricellular host proteins would provide a novel target. Here, we show that an inhibitory protein (FUD) targetingCandida-fibronectin interactions disrupts biofilm formationin vitroandin vivoin a rat venous catheter model. The peptide appears to act by blocking the surface adhesion ofCandida, halting biofilm formation.

scholarly journals Self-Regulation and Interplay of Rsm Family Proteins Modulate the Lifestyle of Pseudomonas putida

2016 ◽  
Vol 82 (18) ◽  
pp. 5673-5686 ◽  
Author(s):  
Óscar Huertas-Rosales ◽  
María Isabel Ramos-González ◽  
Manuel Espinosa-Urgel
Keyword(s):  
Pseudomonas Putida ◽  
Biofilm Formation ◽  
Expression Patterns ◽  
Self Regulation ◽  
Bacterial Attachment ◽  
Triple Mutant ◽  
Content Type ◽  
Global Regulators ◽  
Glass Surfaces ◽  
Plastic Surfaces

ABSTRACTIn the plant-beneficial bacteriumPseudomonas putidaKT2440, three genes have been identified that encode posttranscriptional regulators of the CsrA/RsmA family. Their regulatory roles in the motile and sessile lifestyles ofP. putidahave been investigated by generating single-, double-, and triple-null mutants and by overexpressing each protein (RsmA, RsmE, and RsmI) in different genetic backgrounds. Thersmtriple mutant shows reduced swimming and swarming motilities and increased biofilm formation, whereas overexpression of RsmE or RsmI results in reduced bacterial attachment. However, biofilms formed on glass surfaces by the triple mutant are more labile than those of the wild-type strain and are easily detached from the surface, a phenomenon that is not observed on plastic surfaces. Analysis of the expression of adhesins and exopolysaccharides in the different genetic backgrounds suggests that the biofilm phenotypes are due to alterations in the composition of the extracellular matrix and in the timing of synthesis of its elements. We have also studied the expression patterns of Rsm proteins and obtained data that indicate the existence of autoregulation mechanisms.IMPORTANCEProteins of the CsrA/RsmA family function as global regulators in different bacteria. More than one of these proteins is present in certain species. In this study, all of the RsmA homologs inP. putidaare characterized and globally taken into account to investigate their roles in controlling bacterial lifestyles and the regulatory interactions among them. The results offer new perspectives on how biofilm formation is modulated in this environmentally relevant bacterium.

scholarly journals Characterization and Two-Dimensional Crystallization of Membrane Component AlkB of the Medium-Chain Alkane Hydroxylase System from Pseudomonas putida GPo1

2012 ◽  
Vol 78 (22) ◽  
pp. 7946-7953 ◽  
Author(s):  
Hernan Alonso ◽  
Anna Roujeinikova
Keyword(s):  
Pseudomonas Putida ◽  
Active Form ◽  
Control Sample ◽  
Membrane Component ◽  
Alkane Hydroxylase ◽  
Oligomeric State ◽  
Content Type ◽  
Neopentyl Glycol ◽  
Wide Range ◽  
First Time

ABSTRACTThe alkane hydroxylase system ofPseudomonas putidaGPo1 allows it to use alkanes as the sole source of carbon and energy. Bacterial alkane hydroxylases have tremendous potential as biocatalysts for the stereo- and regioselective transformation of a wide range of chemically inert unreactive alkanes into valuable reactive chemical precursors. We have produced and characterized the first 2-dimensional crystals of the integral membrane component of theP. putidaalkane hydroxylase system, the nonheme di-iron alkane monooxygenase AlkB. Our analysis reveals for the first time that AlkB reconstituted into a lipid bilayer forms trimers. Addition of detergents that do not disrupt the AlkB oligomeric state (decyl maltose neopentyl glycol [DMNG], lauryl maltose neopentyl glycol [LMNG], and octaethylene glycol monododecyl ether [C12E8]) preserved its activity at a level close to that of the detergent-free control sample. In contrast, the monomeric form of AlkB produced by purification inn-decyl-β-d-maltopyranoside (DM),n-dodecyl-β-d-maltopyranoside (DDM), octyl glucose neopentyl glycol (OGNG), andn-dodecyl-N,N-dimethylamine-N-oxide (LDAO) was largely inactive. This is the first indication that the physiologically active form of membrane-embedded AlkB may be a multimer. We present for the first time experimental evidence that 1-octyne acts as a mechanism-based inhibitor of AlkB. Therefore, despite the lack of any significant full-length sequence similarity with members of other monooxygenase classes that catalyze the terminal oxidation of alkanes, AlkB is likely to share a similar catalytic mechanism.

scholarly journals Membrane Vesicle Formation as a Multiple-Stress Response Mechanism Enhances Pseudomonas putida DOT-T1E Cell Surface Hydrophobicity and Biofilm Formation

2012 ◽  
Vol 78 (17) ◽  
pp. 6217-6224 ◽  
Author(s):  
Thomas Baumgarten ◽  
Stefanie Sperling ◽  
Jana Seifert ◽  
Martin von Bergen ◽  
Frank Steiniger ◽  
...  
Keyword(s):  
Stress Response ◽  
Cell Surface ◽  
Pseudomonas Putida ◽  
Biofilm Formation ◽  
Membrane Vesicle ◽  
Cell Surface Hydrophobicity ◽  
Surface Hydrophobicity ◽  
Outer Membrane Vesicles ◽  
Stress Factors ◽  
Content Type

ABSTRACTAmong the adaptive responses of bacteria to rapid changes in environmental conditions, those of the cell envelope are known to be the most crucial. Therefore, several mechanisms with which bacteria change their cell surface and membranes in the presence of different environmental stresses have been elucidated. Among these mechanisms, the release of outer membrane vesicles (MV) in Gram-negative bacteria has attracted particular research interest because of its involvement in pathogenic processes, such as that ofPseudomonas aeruginosabiofilm formation in cystic fibrosis lungs. In this study, we investigated the role of MV formation as an adaptive response ofPseudomonas putidaDOT-T1E to several environmental stress factors and correlated it to the formation of biofilms. In the presence of toxic concentrations of long-chain alcohols, under osmotic stress caused by NaCl, in the presence of EDTA, and after heat shock, cells of this strain released MV within 10 min in the presence of a stressor. The MV formed showed similar size and charge properties, as well as comparable compositions of proteins and fatty acids. MV release caused a significant increase in cell surface hydrophobicity, and an enhanced tendency to form biofilms was demonstrated in this study. Therefore, the release of MV as a stress response could be put in a physiological context.

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