scholarly journals Haloferax volcanii Immersed Liquid Biofilms Develop Independently of Known Biofilm Machineries and Exhibit Rapid Honeycomb Pattern Formation

mSphere ◽  
2020 ◽  
Vol 5 (6) ◽  
Author(s):  
Heather Schiller ◽  
Stefan Schulze ◽  
Zuha Mutan ◽  
Charlotte de Vaulx ◽  
Catalina Runcie ◽  
...  
Keyword(s):  
Pattern Formation ◽  
Biofilm Formation ◽  
Extracellular Polymeric Substances ◽  
Type Iv Pili ◽  
Haloferax Volcanii ◽  
Solid Surfaces ◽  
Content Type ◽  
Liquid Cultures ◽  
Type Iv ◽  
Honeycomb Pattern

ABSTRACT The ability to form biofilms is shared by many microorganisms, including archaea. Cells in a biofilm are encased in extracellular polymeric substances that typically include polysaccharides, proteins, and extracellular DNA, conferring protection while providing a structure that allows for optimal nutrient flow. In many bacteria, flagella and evolutionarily conserved type IV pili are required for the formation of biofilms on solid surfaces or floating at the air-liquid interface of liquid media. Similarly, in many archaea it has been demonstrated that type IV pili and, in a subset of these species, archaella are required for biofilm formation on solid surfaces. Additionally, in the model archaeon Haloferax volcanii, chemotaxis and AglB-dependent glycosylation play important roles in this process. H. volcanii also forms immersed biofilms in liquid cultures poured into petri dishes. This study reveals that mutants of this haloarchaeon that interfere with the biosynthesis of type IV pili or archaella, as well as a chemotaxis-targeting transposon and aglB deletion mutants, lack obvious defects in biofilms formed in liquid cultures. Strikingly, we have observed that these liquid-based biofilms are capable of rearrangement into honeycomb-like patterns that rapidly form upon removal of the petri dish lid, a phenomenon that is not dependent on changes in light or oxygen concentration but can be induced by controlled reduction of humidity. Taken together, this study demonstrates that H. volcanii requires novel, unidentified strategies for immersed liquid biofilm formation and also exhibits rapid structural rearrangements. IMPORTANCE This first molecular biological study of archaeal immersed liquid biofilms advances our basic biological understanding of the model archaeon Haloferax volcanii. Data gleaned from this study also provide an invaluable foundation for future studies to uncover components required for immersed liquid biofilms in this haloarchaeon and also potentially for liquid biofilm formation in general, which is poorly understood compared to the formation of biofilms on surfaces. Moreover, this first description of rapid honeycomb pattern formation is likely to yield novel insights into the underlying structural architecture of extracellular polymeric substances and cells within immersed liquid biofilms.

scholarly journals Haloferax volcanii immersed liquid biofilms develop independently of known biofilm machineries and exhibit rapid honeycomb pattern formation

2020 ◽  
Author(s):  
Heather Schiller ◽  
Stefan Schulze ◽  
Zuha Mutan ◽  
Charlotte de Vaulx ◽  
Catalina Runcie ◽  
...  
Keyword(s):  
Pattern Formation ◽  
Biofilm Formation ◽  
Extracellular Polymeric Substances ◽  
Type Iv Pili ◽  
Haloferax Volcanii ◽  
Solid Surfaces ◽  
Biological Study ◽  
Liquid Cultures ◽  
Type Iv ◽  
Honeycomb Pattern

AbstractThe ability to form biofilms is shared by many microorganisms, including archaea. Cells in a biofilm are encased in extracellular polymeric substances that typically include polysaccharides, proteins, and extracellular DNA, conferring protection while providing a structure that allows for optimal nutrient flow. In many bacteria, flagella and evolutionarily conserved type IV pili are required for the formation of biofilms on solid surfaces or floating at the air-liquid interface of liquid media. Similarly, in many archaea it has been demonstrated that type IV pili and, in a subset of these species, archaella are required for biofilm formation on solid surfaces. Additionally, in the model archaeon Haloferax volcanii, chemotaxis and AglB-dependent glycosylation play important roles in this process. H. volcanii also forms immersed biofilms in liquid cultures poured into Petri dishes. This study reveals that mutants of this haloarchaeon that interfere with the biosynthesis of type IV pili or archaella, as well as a chemotaxis-targeting transposon and aglB-deletion mutants, lack obvious defects in biofilms formed in liquid cultures. Strikingly, we have observed that these liquid-based biofilms are capable of rearrangement into honeycomb-like patterns that rapidly form upon removal of the Petri dish lid, a phenomenon that is not dependent on changes in light or oxygen concentration but can be induced by controlled reduction of humidity. Taken together, this study demonstrates that H. volcanii requires novel, as-yet-unidentified strategies for immersed liquid biofilm formation and also exhibits rapid structural rearrangements.ImportanceThis first molecular biological study of archaeal immersed liquid biofilms advances our basic-biological understanding of the model archaeon Haloferax volcanii. Data gleaned from this study also provide an invaluable foundation for future studies to uncover components required for immersed liquid biofilms in this haloarchaeon and also potentially for liquid biofilm formation in general, which is poorly understood compared to the formation of biofilms on surfaces. Moreover, this first description of rapid honeycomb pattern formation is likely to yield novel insights into the underlying structural architecture of extracellular polymeric substances and cells within immersed liquid biofilms.

scholarly journals Characterization of Zoospore Type IV Pili inActinoplanes missouriensis

2019 ◽  
Vol 201 (14) ◽  
Author(s):  
Tomohiro Kimura ◽  
Takeaki Tezuka ◽  
Daisuke Nakane ◽  
Takayuki Nishizaka ◽  
Shin-Ichi Aizawa ◽  
...  
Keyword(s):  
Vegetative Growth ◽  
Physiological State ◽  
Type Iv Pili ◽  
Solid Surfaces ◽  
Cellular Biology ◽  
Content Type ◽  
Type Iv ◽  
Transmission Electron ◽  
Rare Actinomycetes ◽  
Transcriptional Units

ABSTRACTThe rare actinomyceteActinoplanes missouriensisproduces terminal sporangia containing a few hundred flagellated spores. After release from the sporangia, the spores swim rapidly in aquatic environments as zoospores. The zoospores stop swimming and begin to germinate in niches for vegetative growth. Here, we report the characterization and functional analysis of zoospore type IV pili inA. missouriensis. The pilus gene (pil) cluster, consisting of three apparently σFliA-dependent transcriptional units, is activated during sporangium formation similarly to the flagellar gene cluster, indicating that the zoospore has not only flagella but also pili. With a new method in which zoospores were fixed with glutaraldehyde to prevent pilus retraction, zoospore pili were observed relatively easily using transmission electron microscopy, showing 6 ± 3 pili per zoospore (n = 37 piliated zoospores) and a length of 0.62 ± 0.35 μm (n = 206), via observation offliC-deleted, nonflagellated zoospores. No pili were observed in the zoospores of a prepilin-encodingpilAdeletion (ΔpilA) mutant. In addition, the deletion ofpilT, which encodes an ATPase predicted to be involved in pilus retraction, substantially reduced the frequency of pilus retraction. Several adhesion experiments using wild-type and ΔpilAzoospores indicated that the zoospore pili are required for the sufficient adhesion of zoospores to hydrophobic solid surfaces. Many zoospore-forming rare actinomycetes conserve thepilcluster, which indicates that the zoospore pili yield an evolutionary benefit in the adhesion of zoospores to hydrophobic materials as footholds for germination in their mycelial growth.IMPORTANCEBacterial zoospores are interesting cells in that their physiological state changes dynamically: they are dormant in sporangia, show temporary mobility after awakening, and finally stop swimming to germinate in niches for vegetative growth. However, the cellular biology of a zoospore remains largely unknown. This study describes unprecedented zoospore type IV pili in the rare actinomyceteActinoplanes missouriensis. Similar to the case for the usual bacterial type IV pili, zoospore pili appeared to be retractable. Our findings that the zoospore pili have a functional role in the adhesion of zoospores to hydrophobic solid surfaces and that the zoospores use both pili and flagella properly according to their different purposes provide an important insight into the cellular biology of the zoospore.

scholarly journals Imipenem Treatment Induces Expression of Important Genes and Phenotypes in a Resistant Acinetobacter baumannii Isolate

2015 ◽  
Vol 60 (3) ◽  
pp. 1370-1376 ◽  
Author(s):  
Ghulam Nasser Dhabaan ◽  
Sazaly AbuBakar ◽  
Gustavo Maia Cerqueira ◽  
Mohammed Al-Haroni ◽  
Sui Ping Pang ◽  
...  
Keyword(s):  
Acinetobacter Baumannii ◽  
Biofilm Formation ◽  
Clinical Success ◽  
Multidrug Resistant ◽  
Type Iv Pili ◽  
Control Measures ◽  
Content Type ◽  
Type Iv ◽  
Abiotic Surfaces ◽  
Colonization Factors

Acinetobacter baumanniihas emerged as a notorious multidrug-resistant pathogen, and development of novel control measures is of the utmost importance. Understanding the factors that play a role in drug resistance may contribute to the identification of novel therapeutic targets. Pili are essential forA. baumanniiadherence to and biofilm formation on abiotic surfaces as well as virulence. In the present study, we found that biofilm formation was significantly induced in an imipenem-resistant (Impr) strain treated with a subinhibitory concentration of antibiotic compared to that in an untreated control and an imipenem-susceptible (Imps) isolate. Using microarray and quantitative PCR analyses, we observed that several genes responsible for the synthesis of type IV pili were significantly upregulated in the Imprbut not in the Impsisolate. Notably, this finding is corroborated by an increase in the motility of the Imprstrain. Our results suggest that the ability to overproduce colonization factors in response to imipenem treatment confers biological advantage toA. baumanniiand may contribute to clinical success.

scholarly journals Regulation of Type IV Pili Contributes to Surface Behaviors of Historical and Epidemic Strains of Clostridium difficile

2015 ◽  
Vol 198 (3) ◽  
pp. 565-577 ◽  
Author(s):  
Erin B. Purcell ◽  
Robert W. McKee ◽  
Eric Bordeleau ◽  
Vincent Burrus ◽  
Rita Tamayo
Keyword(s):  
Clostridium Difficile ◽  
Biofilm Formation ◽  
The United States ◽  
Type Iv Pili ◽  
Flagellar Motility ◽  
Content Type ◽  
Type Iv ◽  
Intestinal Pathogen ◽  
Surface Motility ◽  
Pilin Gene

ABSTRACTThe intestinal pathogenClostridium difficileis an urgent public health threat that causes antibiotic-associated diarrhea and is a leading cause of fatal nosocomial infections in the United States.C. difficilerates of recurrence and mortality have increased in recent years due to the emergence of so-called “hypervirulent” epidemic strains. A great deal of the basic biology ofC. difficilehas not been characterized. Recent findings that flagellar motility, toxin synthesis, and type IV pilus (TFP) formation are regulated by cyclic diguanylate (c-di-GMP) reveal the importance of this second messenger forC. difficilegene regulation. However, the function(s) of TFP inC. difficileremains largely unknown. Here, we examine TFP-dependent phenotypes and the role of c-di-GMP in controlling TFP production in the historical 630 and epidemic R20291 strains ofC. difficile. We demonstrate that TFP contribute toC. difficilebiofilm formation in both strains, but with a more prominent role in R20291. Moreover, we report that R20291 is capable of TFP-dependent surface motility, which has not previously been described inC. difficile. The expression and regulation of thepilA1pilin gene differs between R20291 and 630, which may underlie the observed differences in TFP-mediated phenotypes. The differences inpilA1expression are attributable to greater promoter-driven transcription in R20291. In addition, R20291, but not 630, upregulates c-di-GMP levels during surface-associated growth, suggesting that the bacterium senses its substratum. The differential regulation of surface behaviors in historical and epidemicC. difficilestrains may contribute to the different infection outcomes presented by these strains.IMPORTANCEHowClostridium difficileestablishes and maintains colonization of the host bowel is poorly understood. Surface behaviors ofC. difficileare likely relevant during infection, representing possible interactions between the bacterium and the intestinal environment. Pili mediate bacterial interactions with various surfaces and contribute to the virulence of many pathogens. We report that type IV pili (TFP) contribute to biofilm formation byC. difficile. TFP are also required for surface motility, which has not previously been demonstrated forC. difficile. Furthermore, an epidemic-associatedC. difficilestrain showed higher pilin gene expression and greater dependence on TFP for biofilm production and surface motility. Differences in TFP regulation and their effects on surface behaviors may contribute to increased virulence in recent epidemic strains.

scholarly journals Significance of a Posttranslational Modification of the PilA Protein of Geobacter sulfurreducens for Surface Attachment, Biofilm Formation, and Growth on Insoluble Extracellular Electron Acceptors

2017 ◽  
Vol 199 (8) ◽  
Author(s):  
Lubna V. Richter ◽  
Ashley E. Franks ◽  
Robert M. Weis ◽  
Steven J. Sandler
Keyword(s):  
Electron Transfer ◽  
Amino Acid ◽  
Posttranslational Modification ◽  
Biofilm Formation ◽  
Electron Acceptors ◽  
Type Iv Pili ◽  
Geobacter Sulfurreducens ◽  
Surface Attachment ◽  
Content Type ◽  
Type Iv

ABSTRACT Geobacter sulfurreducens, an anaerobic metal-reducing bacterium, possesses type IV pili. These pili are intrinsic structural elements in biofilm formation and, together with a number of c-type cytochromes, are thought to serve as conductive nanowires enabling long-range electron transfer (ET) to metal oxides and graphite anodes. Here, we report that a posttranslational modification of a nonconserved amino acid residue within the PilA protein, the structural subunit of the type IV pili, is crucial for growth on insoluble extracellular electron acceptors. Matrix-assisted laser desorption ionization (MALDI) mass spectrometry of the secreted PilA protein revealed a posttranslational modification of tyrosine-32 with a moiety of a mass consistent with a glycerophosphate group. Mutating this tyrosine into a phenylalanine inhibited cell growth with Fe(III) oxides as the sole electron acceptor. In addition, this amino acid substitution severely diminished biofilm formation on graphite surfaces and impaired current output in microbial fuel cells. These results demonstrate that the capability to attach to insoluble electron acceptors plays a crucial role for the cells' ability to utilize them. The work suggests that glycerophosphate modification of Y32 is a key factor contributing to the surface charge of type IV pili, influencing the adhesion of Geobacter to specific surfaces. IMPORTANCE Type IV pili are bacterial appendages that function in cell adhesion, virulence, twitching motility, and long-range electron transfer (ET) from bacterial cells to insoluble extracellular electron acceptors. The mechanism and role of type IV pili for ET in Geobacter sulfurreducens is still a subject of research. In this study, we identified a posttranslational modification of the major G. sulfurreducens type IV pilin, suggested to be a glycerophosphate moiety. We show that a mutant in which the glycerophosphate-modified tyrosine-32 is replaced with a phenylalanine has reduced abilities for ET and biofilm formation compared with those of the wild type. The results show the importance of the glycerophosphate-modified tyrosine for surface attachment and electron transfer in electrode- or Fe(III)-respiring G. sulfurreducens cells.

scholarly journals Role of Cyclic Di-GMP and Exopolysaccharide in Type IV Pilus Dynamics

2017 ◽  
Vol 199 (8) ◽  
Author(s):  
Jan Ribbe ◽  
Amy E. Baker ◽  
Sebastian Euler ◽  
George A. O'Toole ◽  
Berenike Maier
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Biofilm Formation ◽  
Molecular Motor ◽  
Type Iv Pili ◽  
Force Generation ◽  
Surface Attachment ◽  
Content Type ◽  
Type Iv ◽  
Critical Components ◽  
Planktonic State

ABSTRACT For Pseudomonas aeruginosa, levels of cyclic di-GMP (c-di-GMP) govern the transition from the planktonic state to biofilm formation. Type IV pili (T4P) are crucial determinants of biofilm structure and dynamics, but it is unknown how levels of c-di-GMP affect pilus dynamics. Here, we scrutinized how c-di-GMP affects molecular motor properties and adhesive behavior of T4P. By means of retraction, T4P generated forces of ∼30 pN. Deletion mutants in the proteins with known roles in biofilm formation, swarming motility, and exopolysaccharide (EPS) production (specifically, the diguanylate cyclases sadC and roeA or the c-di-GMP phosphodiesterase bifA) showed only modest effects on velocity or force of T4P retraction. At high levels of c-di-GMP, the production of exopolysaccharides, particularly of Pel, is upregulated. We found that Pel production strongly enhances T4P-mediated surface adhesion of P. aeruginosa, suggesting that T4P-matrix interactions may be involved in biofilm formation by P. aeruginosa. Finally, our data support the previously proposed model of slingshot-like “twitching” motility of P. aeruginosa. IMPORTANCE Type IV pili (T4P) play various important roles in the transition of bacteria from the planktonic state to the biofilm state, including surface attachment and surface sensing. Here, we investigate adhesion, dynamics, and force generation of T4P after bacteria engage a surface. Our studies showed that two critical components of biofilm formation by Pseudomonas aeruginosa, T4P and exopolysaccharides, contribute to enhanced T4P-mediated force generation by attached bacteria. These data indicate a crucial role for the coordinated impact of multiple biofilm-promoting factors during the early stages of attachment to a surface. Our data are also consistent with a previous model explaining why pilus-mediated motility in P. aeruginosa results in characteristic “twitching” behavior.

scholarly journals Biofilm formation by Pseudomonas aeruginosa wild type, flagella and type IV pili mutants

2003 ◽  
Vol 48 (6) ◽  
pp. 1511-1524 ◽  
Author(s):  
Mikkel Klausen ◽  
Arne Heydorn ◽  
Paula Ragas ◽  
Lotte Lambertsen ◽  
Anders Aaes-Jørgensen ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Biofilm Formation ◽  
Type Iv Pili ◽  
Wild Type ◽  
Type Iv

scholarly journals Axenic Biofilm Formation and Aggregation bySynechocystissp. Strain PCC 6803 Are Induced by Changes in Nutrient Concentration and Require Cell Surface Structures

2019 ◽  
Vol 85 (7) ◽  
Author(s):  
Rey Allen ◽  
Bruce E. Rittmann ◽  
Roy Curtiss
Keyword(s):  
Biofilm Formation ◽  
Molecular Mechanisms ◽  
Type Iv Pili ◽  
Biofuel Production ◽  
Oxygenic Photosynthesis ◽  
Phototrophic Biofilm ◽  
Bg11 Medium ◽  
Phototrophic Biofilms ◽  
Type Iv ◽  
Cell Surface Structures

ABSTRACTPhototrophic biofilms are key to nutrient cycling in natural environments and bioremediation technologies, but few studies describe biofilm formation by pure (axenic) cultures of a phototrophic microbe. The cyanobacteriumSynechocystissp. strain PCC 6803 (hereSynechocystis) is a model microorganism for the study of oxygenic photosynthesis and biofuel production. We report here that wild-type (WT)Synechocystiscaused extensive biofilm formation in a 2,000-liter outdoor nonaxenic photobioreactor under conditions attributed to nutrient limitation. We developed a biofilm assay and found that axenicSynechocystisforms biofilms of cells and extracellular material but only when cells are induced by an environmental signal, such as a reduction in the concentration of growth medium BG11. Mutants lacking cell surface structures, namely type IV pili and the S-layer, do not form biofilms. To further characterize the molecular mechanisms of cell-cell binding bySynechocystis, we also developed a rapid (8-h) axenic aggregation assay. Mutants lacking type IV pili were unable to aggregate, but mutants lacking a homolog to Wza, a protein required for type 1 exopolysaccharide export inEscherichia coli, had a superbinding phenotype. In WT cultures, 1.2× BG11 medium induced aggregation to the same degree as 0.8× BG11 medium. Overall, our data support that Wza-dependent exopolysaccharide is essential to maintain stable, uniform suspensions of WTSynechocystiscells in unmodified growth medium and that this mechanism is counteracted in a pilus-dependent manner under altered BG11 concentrations.IMPORTANCEMicrobes can exist as suspensions of individual cells in liquids and also commonly form multicellular communities attached to surfaces. Surface-attached communities, called biofilms, can confer antibiotic resistance to pathogenic bacteria during infections and establish food webs for global nutrient cycling in the environment. Phototrophic biofilm formation is one of the earliest phenotypes visible in the fossil record, dating back over 3 billion years. Despite the importance and ubiquity of phototrophic biofilms, most of what we know about the molecular mechanisms, genetic regulation, and environmental signals of biofilm formation comes from studies of heterotrophic bacteria. We aim to help bridge this knowledge gap by developing new assays forSynechocystis, a phototrophic cyanobacterium used to study oxygenic photosynthesis and biofuel production. With the aid of these new assays, we contribute to the development ofSynechocystisas a model organism for the study of axenic phototrophic biofilm formation.

scholarly journals Molecular and Physical Factors That Influence Attachment of Vibrio vulnificus to Chitin

2015 ◽  
Vol 81 (18) ◽  
pp. 6158-6165 ◽  
Author(s):  
Tiffany C. Williams ◽  
Mesrop Ayrapetyan ◽  
James D. Oliver
Keyword(s):  
Quorum Sensing ◽  
Vibrio Vulnificus ◽  
The United States ◽  
Type Iv Pili ◽  
Negative Regulator ◽  
Physical Factors ◽  
Wound Infections ◽  
Environmental Persistence ◽  
Content Type ◽  
Type Iv

ABSTRACTThe human pathogenVibrio vulnificusis the leading cause of seafood-related deaths in the United States. Strains are genotyped on the basis of alleles that correlate with isolation source, with clinical (C)-genotype strains being more often implicated in disease and environmental (E)-genotype strains being more frequently isolated from oysters and estuarine waters. Previously, we have shown that the ecologically distinct C- and E-genotype strains ofV. vulnificusdisplay different degrees of chitin attachment, with C-genotype strains exhibiting reduced attachment relative to their E-genotype strain counterparts. We identified type IV pili to be part of the molecular basis for this observed genotypic variance, as E-genotype strains exhibit higher levels of expression of these genes than C-genotype strains. Here, we used a C-genotype quorum-sensing (QS) mutant to demonstrate that quorum sensing is a negative regulator of type IV pilus expression, which results in decreased chitin attachment. Furthermore, calcium depletion reduced E-genotype strain attachment to chitin, which suggests that calcium is necessary for proper functioning of the type IV pili in E-genotype strains. We also found that starvation or dormancy can alter the efficiency of chitin attachment, which has significant implications for the environmental persistence ofV. vulnificus. With the increasing incidence of wound infections caused byV. vulnificus, we investigated a subset of E-genotype strains isolated from human wound infections and discovered that they attached to chitin in a manner more similar to that of C-genotype strains. This study enhances our understanding of the molecular and physical factors that mediate chitin attachment inV. vulnificus, providing insight into the mechanisms that facilitate the persistence of this pathogen in its native environment.

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