Genome rearrangements induce biofilm formation inEscherichia coliC – an old model organism with a new application in biofilm research

AbstractEscherichia coliC forms more robust biofilms than the other laboratory strains. Biofilm formation and cell aggregation under a high shear force depends on temperature and salt concentrations. It is the last of fiveE. colistrains (C, K12, B, W, Crooks) designated as safe for laboratory purposes whose genome has not been sequenced. Here we present the complete genomic sequence of this strain in which we utilized both long-read PacBio-based sequencing and high resolution optical mapping to confirm a large inversion in comparison to the other laboratory strains. Notably, DNA sequence comparison revealed the absence of several genes thought to be involved in biofilm formation, including antigen 43,waaSBOJYZULfor LPS synthesis, andcpsBfor curli synthesis. The first main difference we identified that likely affects biofilm formation is the presence of an IS3-like insertion sequence in front of the carbon storage regulatorcsrAgene. This insertion is located 86 bp upstream of thecsrAstart codon inside the −35 region of P4 promoter and blocks the transcription from the sigma32and sigma70promoters P1-P3 located further upstream. The second is the presence of an IS5/IS1182 in front of thecsgDgene, which may drive its overexpression in biofilm. And finally,E. coliC encodes an additional sigma70subunit overexpressed in biofilm and driven by the same IS3-like insertion sequence. Promoter analyses using GFP gene fusions and total expression profiles using RNA-seq analyses comparing planktonic and biofilm envirovars provided insights into understanding this regulatory pathway inE. coli.IMPORTANCEBiofilms are crucial for bacterial survival, adaptation, and dissemination in natural, industrial, and medical environments. Most laboratory strains ofE. coligrown for decadesin vitrohave evolved and lost their ability to form biofilm, while environmental isolates that can cause infections and diseases are not safe to work with. Here, we show that the historic laboratory strain ofE. coliC produces a robust biofilm and can be used as a model organism for multicellular bacterial research. Furthermore, we ascertained the full genomic sequence as well as gene expression profiles of both the biofilm and planktonic envirovars of this classic strain, which provide for a base level of characterization and make it useful for many biofilm-based applications.

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Genome rearrangements induce biofilm formation in Escherichia coli C – an old model organism with a new application in biofilm research

BMC Genomics ◽

10.1186/s12864-019-6165-4 ◽

2019 ◽

Vol 20 (1) ◽

Cited By ~ 2

Author(s):

Jarosław E. Król ◽

Donald C. Hall ◽

Sergey Balashov ◽

Steven Pastor ◽

Justin Sibert ◽

...

Keyword(s):

Escherichia Coli ◽

Biofilm Formation ◽

Insertion Sequence ◽

Genomic Sequence ◽

Model Organism ◽

Start Codon ◽

Bacterial Survival ◽

E Coli ◽

Full Genomic Sequence

Abstract Background Escherichia coli C forms more robust biofilms than other laboratory strains. Biofilm formation and cell aggregation under a high shear force depend on temperature and salt concentrations. It is the last of five E. coli strains (C, K12, B, W, Crooks) designated as safe for laboratory purposes whose genome has not been sequenced. Results Here we present the complete genomic sequence of this strain in which we utilized both long-read PacBio-based sequencing and high resolution optical mapping to confirm a large inversion in comparison to the other laboratory strains. Notably, DNA sequence comparison revealed the absence of several genes thought to be involved in biofilm formation, including antigen 43, waaSBOJYZUL for lipopolysaccharide (LPS) synthesis, and cpsB for curli synthesis. The first main difference we identified that likely affects biofilm formation is the presence of an IS3-like insertion sequence in front of the carbon storage regulator csrA gene. This insertion is located 86 bp upstream of the csrA start codon inside the − 35 region of P4 promoter and blocks the transcription from the sigma32 and sigma70 promoters P1-P3 located further upstream. The second is the presence of an IS5/IS1182 in front of the csgD gene. And finally, E. coli C encodes an additional sigma70 subunit driven by the same IS3-like insertion sequence. Promoter analyses using GFP gene fusions provided insights into understanding this regulatory pathway in E. coli. Conclusions Biofilms are crucial for bacterial survival, adaptation, and dissemination in natural, industrial, and medical environments. Most laboratory strains of E. coli grown for decades in vitro have evolved and lost their ability to form biofilm, while environmental isolates that can cause infections and diseases are not safe to work with. Here, we show that the historic laboratory strain of E. coli C produces a robust biofilm and can be used as a model organism for multicellular bacterial research. Furthermore, we ascertained the full genomic sequence of this classic strain, which provides for a base level of characterization and makes it useful for many biofilm-based applications.

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Nickel Promotes Biofilm Formation by Escherichia coli K-12 Strains That Produce Curli

Applied and Environmental Microbiology ◽

10.1128/aem.02171-08 ◽

2009 ◽

Vol 75 (6) ◽

pp. 1723-1733 ◽

Cited By ~ 40

Author(s):

Claire Perrin ◽

Romain Briandet ◽

Gregory Jubelin ◽

Philippe Lejeune ◽

Marie-Andrée Mandrand-Berthelot ◽

...

Keyword(s):

Escherichia Coli ◽

Biofilm Formation ◽

Molecular Mechanisms ◽

Selective Advantage ◽

Bacterial Survival ◽

Toxic Compounds ◽

E Coli ◽

Transcriptional Effect ◽

K 12 ◽

Encoding Genes

ABSTRACT The survival of bacteria exposed to toxic compounds is a multifactorial phenomenon, involving well-known molecular mechanisms of resistance but also less-well-understood mechanisms of tolerance that need to be clarified. In particular, the contribution of biofilm formation to survival in the presence of toxic compounds, such as nickel, was investigated in this study. We found that a subinhibitory concentration of nickel leads Escherichia coli bacteria to change their lifestyle, developing biofilm structures rather than growing as free-floating cells. Interestingly, whereas nickel and magnesium both alter the global cell surface charge, only nickel promotes biofilm formation in our system. Genetic evidence indicates that biofilm formation induced by nickel is mediated by the transcriptional induction of the adhesive curli-encoding genes. Biofilm formation induced by nickel does not rely on efflux mechanisms using the RcnA pump, as these require a higher concentration of nickel to be activated. Our results demonstrate that the nickel-induced biofilm formation in E. coli is an adaptational process, occurring through a transcriptional effect on genes coding for adherence structures. The biofilm lifestyle is obviously a selective advantage in the presence of nickel, but the means by which it improves bacterial survival needs to be investigated.

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Antibacterial and antibiofilm effects of garlic (Allium sativum), ginger (Zingiber officinale) and mint (Mentha piperta) on Escherichia coli biofilms

Food Science and Applied Biotechnology ◽

10.30721/fsab2021.v4.i2.146 ◽

2021 ◽

Vol 4 (2) ◽

pp. 166

Author(s):

Ndaindila Haindongo ◽

Amara Anyogu ◽

Osmond Ekwebelem ◽

Christian Anumudu ◽

Helen Onyeaka

Keyword(s):

Escherichia Coli ◽

Biofilm Formation ◽

Food Industry ◽

Allium Sativum ◽

Zingiber Officinale ◽

Bacterial Survival ◽

Planktonic Cells ◽

E Coli ◽

Foodborne Outbreaks ◽

Potential Use

Biofilms are a significant concern in the food industry because of their potential to enhance bacterial survival and cause foodborne outbreaks. Escherichia coli (E. coli) is among the leading pathogens responsible for foodborne outbreaks and this can be attributed to its ability to form biofilms in food containers and food preparatory surfaces. The purpose of this study was to investigate the antibacterial and antibiofilm properties of garlic, ginger and mint and their potential to inhibit E.coli and biofilm formation. Disc diffusion assays and 96-well plate crystal violet-based methods were used to achieve these objectives. The plant extracts were diluted from 1 mg/ml to 0.1 mg/ml and incubated 25°C and 37°C to investigate the antimicrobial and antibiofilm effects on E. coli. The findings of this study showed that low temperatures induced the formation of E. coli biofilms and all tested extracts contain a broad spectrum of antibacterial and antibiofilm properties. This study provided new insights on the combined antimicrobial and antibiofilm properties of garlic, ginger and mint against planktonic cells and biofilms of E. coli MG 1655 and highlight the potential use of these extracts in the food industry to prevent biofilm formation by E. coli.

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Phosphodiesterase activity of NbdA from Pseudomonas aeruginosa

10.5194/biofilms9-89 ◽

2020 ◽

Author(s):

Anna Scherhag ◽

Martina Rüger ◽

Katrin Gerbracht ◽

Jaqueline Rehner ◽

Susanne Zehner ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Biofilm Formation ◽

Model Organism ◽

Functional Complementation ◽

Full Length ◽

Protein Activity ◽

E Coli ◽

Diguanylate Cyclases ◽

Ggdef Domain

The molecule c-di-GMP is a bacterial second messenger that controls various processes such as motility or biofilm formation in bacteria [1]. To synthesize and degrade c-di-GMP, enzymes called diguanylate cyclases (DGC) containing a GGDEF-domain and phosphodiesterases (PDE) containing an EAL-domain or HD-GYP-domain are important [1, 2]. Pseudomonas aeruginosa, a model organism for biofilm formation and dispersion, encodes for 18 GGDEF, 5 EAL, 16 GGDEF / EAL, and 3 HD-GYP-domain-containing proteins [3]. One of the GGDEF / EAL-containing proteins is NbdA. This protein also harbors an N-terminal membrane anchored MHYT-domain, that is predicted to be a sensor for NO, CO or O2 [4]. In this work, recombinant and affinity purified NbdA was tested for its PDE activity. Three different methods were used to measure the PDE activity of NbdA: a bis-pNPP-assay in which the conversion of the pseudosubstrate bis-pNPP into p-nitrophenol was detected spectroscopically, an HPLC-analysis of an enzymatic assay with the native substrate c-di-GMP, and a MANT-c-di-GMP-assay in which a fluorescently labeled form of the presumed substrate c-di-GMP was utilized. To establish these methods, the two known phosphodiesterases, PdeH from Escherichia coli [5] and RocR from P. aeruginosa [6], were also produced and tested. Subsequently, three variants of NbdA were investigated: the full-length version and two truncated versions of the protein. Activity was further assessed using functional complementation of an E. coli phosphodiesterase deficient strain with full-length and truncated NbdA variants confirming PDE activity in vivo. &#160; &#160; [1] Hengge, R. (2009) Nature Rev. Microbiol. 7: 263-273. [2] R&#246;mling, U., Gomelsky, M., Galperin, M.Y. (2005). Mol. Microbiol. 57: 629&#8211;639. [3] Valentini, M., Filloux, A. (2016). J. Biol. Chem. 291: 12547&#8211;12555. [4] Galperin, M.Y., Gaidenko, T.A., Mulkidjanian, A.Y., Nakano, M., und Price, C.W. (2001). FEMS Microbiol. Lett. 205, 17&#8211;23. [5] Pesavento, C., Becker, G., Sommerfeldt, N., Possling, A., Tschowri, N., Mehlis, A., Hengge, R. (2008). Genes Dev. 22: 2434&#8211;2446. [6] Chen et al. (2012) Chen, M.W., Kotaka, M., Vonrhein, C., Bricogne, G., Rao, F., Chuah, M.L.C., Svergun, D., Schneider, G., Liang, Z.-X., Lescar, J.&#160; (2012). Signaling. J. Bacteriol. 194: 4837&#8211;4846 &#160;

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Interactions Screenings Unearth Potential New Divisome Components in the Chlamydia-Related Bacterium, Waddlia chondrophila

Microorganisms ◽

10.3390/microorganisms7120617 ◽

2019 ◽

Vol 7 (12) ◽

pp. 617

Author(s):

Firuza Bayramova ◽

Nicolas Jacquier ◽

Gilbert Greub

Keyword(s):

Iron Homeostasis ◽

Expression Profiles ◽

Model Organism ◽

E Coli ◽

Obligate Intracellular ◽

Gene And Protein Expression ◽

Related Bacterium ◽

Obligate Intracellular Bacteria ◽

Protein Expression Profiles ◽

Two Hybrid

Chlamydiales order members are obligate intracellular bacteria, dividing by binary fission. However, Chlamydiales lack the otherwise conserved homologue of the bacterial division organizer FtsZ and certain division protein homologues. FtsZ might be functionally replaced in Chlamydiales by the actin homologue MreB. RodZ, the membrane anchor of MreB, localizes early at the division septum. In order to better characterize the organization of the chlamydial divisome, we performed co-immunoprecipitations and yeast-two hybrid assays to study the interactome of RodZ, using Waddlia chondrophila, a potentially pathogenic Chlamydia-related bacterium, as a model organism. Three potential interactors were further investigated: SecA, FtsH, and SufD. The gene and protein expression profiles of these three genes were measured and are comparable with recently described division proteins. Moreover, SecA, FtsH, and SufD all showed a peripheral localization, consistent with putative inner membrane localization and interaction with RodZ. Notably, heterologous overexpression of the abovementioned proteins could not complement E. coli mutants, indicating that these proteins might play different functions in these two bacteria or that important regulators are not conserved. Altogether, this study brings new insights to the composition of the chlamydial divisome and points to links between protein secretion, degradation, iron homeostasis, and chlamydial division.

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Global Gene Expression Profiling of Asymptomatic Bacteriuria Escherichia coli during Biofilm Growth in Human Urine

Infection and Immunity ◽

10.1128/iai.01748-06 ◽

2006 ◽

Vol 75 (2) ◽

pp. 966-976 ◽

Cited By ~ 57

Author(s):

Viktoria Hancock ◽

Per Klemm

Keyword(s):

Escherichia Coli ◽

Urinary Tract ◽

Expression Profile ◽

Human Urine ◽

Biofilm Formation ◽

Expression Profiles ◽

Asymptomatic Bacteriuria ◽

Knockout Mutant ◽

Biofilm Growth ◽

E Coli

ABSTRACT Urinary tract infection (UTI) is an important health problem worldwide, with many millions of cases each year, and Escherichia coli is the most common organism causing UTI in humans. Also, E. coli is responsible for most infections in patients with chronic indwelling bladder catheter. The two asymptomatic bacteriuria (ABU) E. coli strains 83972 and VR50 are significantly better biofilm formers in their natural growth medium, human urine, than the two uropathogenic E. coli isolates CFT073 and 536. We used DNA microarrays to monitor the expression profile during biofilm growth in urine of the two ABU strains 83972 and VR50. Significant differences in expression levels were seen between the biofilm expression profiles of the two strains with the corresponding planktonic expression profiles in morpholinepropanesulfonic acid minimal laboratory medium and human urine; 417 and 355 genes were up- and down-regulated, respectively, during biofilm growth in urine of 83972 and VR50. Many genes involved in transcription and stress were up-regulated in biofilm-grown cells. The role in biofilm formation of four of the up-regulated genes, i.e., yceP, yqgA, ygiD, and aaeX, was investigated by creating single-knockout mutant versions of 83972 and VR50; all mutants showed reduced biofilm formation in urine by 18 to 43% compared with the wild type (P < 0.05). Also, the expression profile of strain 83972 in the human urinary tract partially overlaps with the biofilm expression profile.

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Biofilm Formation on Biotic and Abiotic Surfaces in the Presence of Antimicrobials by Escherichia coli Isolates from Cases of Bovine Mastitis

Applied and Environmental Microbiology ◽

10.1128/aem.01953-14 ◽

2014 ◽

Vol 80 (19) ◽

pp. 6136-6145 ◽

Cited By ~ 20

Author(s):

Vitor O. Silva ◽

Larissa O. Soares ◽

Abelardo Silva Júnior ◽

Hilário C. Mantovani ◽

Yung-Fu Chang ◽

...

Keyword(s):

Escherichia Coli ◽

Biofilm Formation ◽

Expression Profiles ◽

Bovine Mastitis ◽

Pcr Analysis ◽

Content Type ◽

E Coli ◽

Biofilm Production ◽

Abiotic Surfaces

ABSTRACTEscherichia coliis a highly adaptive microorganism, and its ability to form biofilms under certain conditions can be critical for antimicrobial resistance. The adhesion of fourE. coliisolates from bovine mastitis to bovine mammary alveolar (MAC-T) cells, biofilm production on a polystyrene surface, and the expression profiles of the genesfliC,csgA,fimA, andluxSin the presence of enrofloxacin, gentamicin, co-trimoxazole, and ampicillin at half of the MIC were investigated. Increased adhesion ofE. coliisolates in the presence of antimicrobials was not observed; however, increased internalization of some isolates was observed by confocal microscopy. All of the antimicrobials induced the formation of biofilms by at least one isolate, whereas enrofloxacin and co-trimoxazole decreased biofilm formation by at least one isolate. Quantitative PCR analysis revealed that all four genes were differentially expressed when bacteria were exposed to subinhibitory concentrations of antimicrobials, with expression altered on the order of 1.5- to 22-fold. However, it was not possible to associate gene expression with induction or reduction of biofilm formation in the presence of the antimicrobials. Taken together, the results demonstrate that antimicrobials could induce biofilm formation by some isolates, in addition to inducing MAC-T cell invasion, a situation that might occurin vivo, potentially resulting in a bacterial reservoir in the udder, which might explain some cases of persistent mastitis in herds.

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Regulation of Resistance to Copper in Xanthomonas axonopodis pv. vesicatoria

Applied and Environmental Microbiology ◽

10.1128/aem.71.2.782-789.2005 ◽

2005 ◽

Vol 71 (2) ◽

pp. 782-789 ◽

Cited By ~ 35

Author(s):

Andreas E. Voloudakis ◽

Therese M. Reignier ◽

Donald A. Cooksey

Keyword(s):

Pseudomonas Syringae ◽

Transcriptional Activation ◽

Genomic Sequence ◽

Direct Interaction ◽

Protein Product ◽

Inducible Expression ◽

Copper Resistance ◽

Start Codon ◽

Xanthomonas Axonopodis ◽

E Coli

ABSTRACT Copper-resistant strains of Xanthomonas axonopodis pv. vesicatoria were previously shown to carry plasmid-borne copper resistance genes related to the cop and pco operons of Pseudomonas syringae and Escherichia coli, respectively. However, instead of the two-component (copRS and pcoRS) systems determining copper-inducible expression of the operons in P. syringae and E. coli, a novel open reading frame, copL, was found to be required for copper-inducible expression of the downstream multicopper oxidase copA in X. axonopodis. copL encodes a predicted protein product of 122 amino acids that is rich in histidine and cysteine residues, suggesting a possible direct interaction with copper. Deletions or frameshift mutations within copL, as well as an amino acid substitution generated at the putative start codon of copL, caused a loss of copper-inducible transcriptional activation of copA. A nonpolar insertion of a kanamycin resistance gene in copL resulted in copper sensitivity in the wild-type strain. However, repeated attempts to complement copL mutations in trans failed. Analysis of the genomic sequence databases shows that there are copL homologs upstream of copAB genes in X. axonopodis pv. citri, X. campestris pv. campestris, and Xylella fastidiosa. The cloned promoter area upstream of copA in X. axonopodis pv. vesicatoria did not function in Pseudomonas syringae or in E. coli, nor did the P. syringae cop promoter function in Xanthomonas. However, a transcriptional fusion of the Xanthomonas cop promoter with the Pseudomonas copABCDRS was able to confer resistance to copper in Xanthomonas, showing divergence in the mechanisms of regulation of the resistance to copper in phytopathogenic bacteria.

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Effects of Essential Oils from Cymbopogon spp. and Cinnamomum verum on Biofilm and Virulence Properties of Escherichia coli O157:H7

Antibiotics ◽

10.3390/antibiotics10020113 ◽

2021 ◽

Vol 10 (2) ◽

pp. 113 ◽

Cited By ~ 1

Author(s):

Raffaella Scotti ◽

Annarita Stringaro ◽

Laura Nicolini ◽

Miriam Zanellato ◽

Priscilla Boccia ◽

...

Keyword(s):

Escherichia Coli ◽

Essential Oils ◽

Biofilm Formation ◽

Expression Profiles ◽

Enterohemorrhagic Escherichia Coli ◽

Zinc Homeostasis ◽

Transcriptional Analysis ◽

Escherichia Coli O157 ◽

Biofilm Inhibition ◽

E Coli

Every year, the pharmaceutical and food industries produce over 1000 tons of essential oils (EOs) exploitable in different fields as the development of eco-friendly and safe antimicrobial inhibitors. In this work we investigated the potential of some EOs, namely Cinnamomum verum, Cymbopogon martini, Cymbopogoncitratus and Cymbopogon flexuosus, on the growth, biofilm formation and gene expression in four strains of enterohemorrhagic Escherichia coli O157:H7. All EOs were analyzed by gas chromatography-mass spectrometry (GC-MS). The antimicrobial activity was performed by using dilutions of EOs ranging from 0.001 to 1.2% (v/v). Subinhibitory doses were used for biofilm inhibition assay. The expression profiles were obtained by RT-PCR. E. coli O157:H7 virulence was evaluated in vivo in the nematode Caenorhabditis elegans. All EOs showed minimal inhibitory concentrations (MICs) ranging from 0.0075 to 0.3% (v/v). Cinnamomum verum bark EO had the best activity (MIC of 0.0075% (v/v) in all strains) while the C. verum leaf EO had an intermediate efficacy with MIC of 0.175% (v/v) in almost all strains. The Cymbopogon spp. showed the more variable MICs (ranging from 0.075 to 0.3% (v/v)) depending on the strain used. Transcriptional analysis showed that C. martini EO repressed several genes involved in biofilm formation, virulence, zinc homeostasis and encoding some membrane proteins. All EOs affected zinc homeostasis, reducing ykgM and zinT expression, and reduced the ability of E. coli O157:H7 to infect the nematode C. elegans. In conclusion, we demonstrated that these EOs, affecting E. coli O157:H7 infectivity, have a great potential to be used against infections caused by microorganisms.

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Three-Dimensional Reconstruction of Two Forms of the Chaperonin GRO EL

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100180045 ◽

1990 ◽

Vol 48 (1) ◽

pp. 258-259

Author(s):

J.L. Carrascosa ◽

G. Abella ◽

S. Marco ◽

M. Muyal ◽

J.M. Carazo

Keyword(s):

Three Dimensional ◽

The Other ◽

Two Dimensional ◽

Series Method ◽

Three Dimensional Reconstruction ◽

Structural Components ◽

E Coli ◽

Dimensional Reconstruction ◽

Morphogenetic Factors ◽

Tilt Series

Chaperonins are a class of proteins characterized by their role as morphogenetic factors. They trantsiently interact with the structural components of certain biological aggregates (viruses, enzymes etc), promoting their correct folding, assembly and, eventually transport. The groEL factor from E. coli is a conspicuous member of the chaperonins, as it promotes the assembly and morphogenesis of bacterial oligomers and/viral structures.We have studied groEL-like factors from two different bacteria:E. coli and B.subtilis. These factors share common morphological features , showing two different views: one is 6-fold, while the other shows 7 morphological units. There is also a correlation between the presence of a dominant 6-fold view and the fact of both bacteria been grown at low temperature (32°C), while the 7-fold is the main view at higher temperatures (42°C). As the two-dimensional projections of groEL were difficult to interprete, we studied their three-dimensional reconstruction by the random conical tilt series method from negatively stained particles.

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