scholarly journals Involvement of the Type IX Secretion System in Capnocytophaga ochracea Gliding Motility and Biofilm Formation

2016 ◽  
Vol 82 (6) ◽  
pp. 1756-1766 ◽  
Author(s):  
Daichi Kita ◽  
Satoshi Shibata ◽  
Yuichiro Kikuchi ◽  
Eitoyo Kokubu ◽  
Koji Nakayama ◽  
...  
Keyword(s):  
Cell Surface ◽  
Biofilm Formation ◽  
Laser Scanning ◽  
Secretion System ◽  
Gliding Motility ◽  
Confocal Laser ◽  
Bacterial Cells ◽  
Wild Type ◽  
Content Type ◽  
Type Ix Secretion System

ABSTRACTCapnocytophaga ochraceais a Gram-negative, rod-shaped bacterium that demonstrates gliding motility when cultured on solid agar surfaces.C. ochraceapossesses the ability to form biofilms; however, factors involved in biofilm formation by this bacterium are unclear. A type IX secretion system (T9SS) inFlavobacterium johnsoniaewas shown to be involved in the transport of proteins (e.g., several adhesins) to the cell surface. Genes orthologous to those encoding T9SS proteins inF. johnsoniaehave been identified in the genome ofC. ochracea; therefore, the T9SS may be involved in biofilm formation byC. ochracea. Here we constructed three ortholog-deficientC. ochraceamutants lackingsprB(which encodes a gliding motility adhesin) orgldKorsprT(which encode T9SS proteins inF. johnsoniae). Gliding motility was lost in each mutant, suggesting that, inC. ochracea, the proteins encoded bysprB,gldK, andsprTare necessary for gliding motility, and SprB is transported to the cell surface by the T9SS. For the ΔgldK, ΔsprT, and ΔsprBstrains, the amounts of crystal violet-associated biofilm, relative to wild-type values, were 49%, 34%, and 65%, respectively, at 48 h. Confocal laser scanning and scanning electron microscopy revealed that the biofilms formed by wild-typeC. ochraceawere denser and bacterial cells were closer together than in those formed by the mutant strains. Together, these results indicate that proteins exported by the T9SS are key elements of the gliding motility and biofilm formation ofC. ochracea.

scholarly journals The Carboxy-Terminal Region ofFlavobacterium johnsoniaeSprB Facilitates Its Secretion by the Type IX Secretion System and Propulsion by the Gliding Motility Machinery

2019 ◽  
Vol 201 (19) ◽  
Author(s):  
Surashree S. Kulkarni ◽  
Joseph J. Johnston ◽  
Yongtao Zhu ◽  
Zachary T. Hying ◽  
Mark J. McBride
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Cell Surface ◽  
Type A ◽  
Secretion System ◽  
Gliding Motility ◽  
Foreign Protein ◽  
Type B ◽  
Content Type ◽  
Type Ix Secretion System

ABSTRACTFlavobacterium johnsoniaeSprB moves rapidly along the cell surface, resulting in gliding motility. SprB secretion requires the type IX secretion system (T9SS). Proteins secreted by the T9SS typically have conserved C-terminal domains (CTDs) belonging to the type A CTD or type B CTD family. Attachment of 70- to 100-amino-acid type A CTDs to a foreign protein allows its secretion. Type B CTDs are common but have received little attention. Secretion of the foreign protein superfolder green fluorescent protein (sfGFP) fused to regions spanning the SprB type B CTD (sfGFP-CTDSprB) was analyzed. CTDs of 218 amino acids or longer resulted in secretion of sfGFP, whereas a 149-amino-acid region did not. Some sfGFP was secreted in soluble form, whereas the rest was attached on the cell surface. Surface-attached sfGFP was rapidly propelled along the cell, suggesting productive interaction with the motility machinery. This did not result in rapid cell movement, which apparently requires additional regions of SprB. Secretion of sfGFP-CTDSprBrequired coexpression withsprF, which lies downstream ofsprB. SprF is similar in sequence toPorphyromonas gingivalisPorP. MostF. johnsoniaegenes encoding proteins with type B CTDs lie immediately upstream ofporP/sprF-like genes. sfGFP was fused to the type B CTD from one such protein (Fjoh_3952). This resulted in secretion of sfGFP only when it was coexpressed with its cognate PorP/SprF-like protein. These results highlight the need for extended regions of type B CTDs and for coexpression with the appropriate PorP/SprF-like protein for efficient secretion and cell surface localization of cargo proteins.IMPORTANCETheF. johnsoniaegliding motility adhesin SprB is delivered to the cell surface by the type IX secretion system (T9SS) and is rapidly propelled along the cell by the motility machinery. How this 6,497-amino-acid protein interacts with the secretion and motility machines is not known. Fusion of the C-terminal 218 amino acids of SprB to a foreign cargo protein resulted in its secretion, attachment to the cell surface, and rapid movement by the motility machinery. Efficient secretion of SprB required coexpression with the outer membrane protein SprF. Secreted proteins that have sequence similarity to SprB in their C-terminal regions are common in the phylumBacteroidetesand may have roles in adhesion, motility, and virulence.

scholarly journals Flavobacterium johnsoniae PorV Is Required for Secretion of a Subset of Proteins Targeted to the Type IX Secretion System

2014 ◽  
Vol 197 (1) ◽  
pp. 147-158 ◽  
Author(s):  
Sampada S. Kharade ◽  
Mark J. McBride
Keyword(s):  
Cell Surface ◽  
Secretion System ◽  
Gliding Motility ◽  
Content Type ◽  
Flavobacterium Johnsoniae ◽  
Protein Secretion System ◽  
Surface Motility ◽  
Type Ix Secretion System ◽  
Carboxy Terminal ◽  
Novel Protein

Flavobacterium johnsoniaeexhibits gliding motility and digests many polysaccharides, including chitin. A novel protein secretion system, the type IX secretion system (T9SS), is required for gliding and chitin utilization. The T9SS secretes the cell surface motility adhesins SprB and RemA and the chitinase ChiA. Proteins involved in secretion by the T9SS include GldK, GldL, GldM, GldN, SprA, SprE, and SprT.Porphyromonas gingivalishas orthologs for each of these that are required for secretion of gingipain protease virulence factors by its T9SS.P. gingivalisporUandporVhave also been linked to T9SS-mediated secretion, andF. johnsoniaehas orthologs of these. Mutations inF. johnsoniaeporUandporVwere constructed to determine if they function in secretion. Cells of aporVdeletion mutant were deficient in chitin utilization and failed to secrete ChiA. They were also deficient in secretion of the motility adhesin RemA but retained the ability to secrete SprB. SprB is involved in gliding motility and is needed for formation of spreading colonies on agar, and theporVmutant exhibited gliding motility and formed spreading colonies. However, theporVmutant was partially deficient in attachment to glass, apparently because of the absence of RemA and other adhesins on the cell surface. TheporVmutant also appeared to be deficient in secretion of numerous other proteins that have carboxy-terminal domains associated with targeting to the T9SS. PorU was not required for secretion of ChiA, RemA, or SprB, indicating that it does not play an essential role in theF. johnsoniaeT9SS.

scholarly journals The Type 9 Secretion System Is Required for Flavobacterium johnsoniae Biofilm Formation

2021 ◽  
Vol 12 ◽  
Author(s):  
Todd J. Eckroat ◽  
Camillus Greguske ◽  
David W. Hunnicutt
Keyword(s):  
Biofilm Formation ◽  
Deletion Mutant ◽  
Secretion System ◽  
Gliding Motility ◽  
Microplate Assay ◽  
Partial Reduction ◽  
Wild Type ◽  
Flavobacterium Johnsoniae ◽  
Biofilm Production ◽  
Type Ix Secretion System

Flavobacterium johnsoniae forms biofilms in low nutrient conditions. Protein secretion and cell motility may have roles in biofilm formation. The F. johnsoniae type IX secretion system (T9SS) is important for both secretion and motility. To determine the roles of each process in biofilm formation, mutants defective in secretion, in motility, or in both processes were tested for their effects on biofilm production using a crystal violet microplate assay. All mutants that lacked both motility and T9SS-mediated secretion failed to produce biofilms. A porV deletion mutant, which was severely defective for secretion, but was competent for motility, also produced negligible biofilm. In contrast, mutants that retained secretion but had defects in gliding formed biofilms. An sprB mutant that is severely but incompletely defective in gliding motility but retains a fully functional T9SS was similar to the wild type in biofilm formation. Mutants with truncations of the gldJ gene that compromise motility but not secretion showed partial reduction in biofilm formation compared to wild type. Unlike the sprB mutant, these gldJ truncation mutants were essentially nonmotile. The results show that a functional T9SS is required for biofilm formation. Gliding motility, while not required for biofilm formation, also appears to contribute to formation of a robust biofilm.

scholarly journals Calcite Biomineralization by Bacterial Isolates from the Recently Discovered Pristine Karstic Herrenberg Cave

2011 ◽  
Vol 78 (4) ◽  
pp. 1157-1167 ◽  
Author(s):  
Anna Rusznyák ◽  
Denise M. Akob ◽  
Sándor Nietzsche ◽  
Karin Eusterhues ◽  
Kai Uwe Totsche ◽  
...  
Keyword(s):  
Laser Scanning ◽  
Extracellular Polymeric Substances ◽  
Large Fraction ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Rrna Gene ◽  
Seepage Water ◽  
Bacterial Cells ◽  
Content Type ◽  
Rich Media

ABSTRACTKarstic caves represent one of the most important subterranean carbon storages on Earth and provide windows into the subsurface. The recent discovery of the Herrenberg Cave, Germany, gave us the opportunity to investigate the diversity and potential role of bacteria in carbonate mineral formation. Calcite was the only mineral observed by Raman spectroscopy to precipitate as stalactites from seepage water. Bacterial cells were found on the surface and interior of stalactites by confocal laser scanning microscopy. Proteobacteria dominated the microbial communities inhabiting stalactites, representing more than 70% of total 16S rRNA gene clones. Proteobacteria formed 22 to 34% of the detected communities in fluvial sediments, and a large fraction of these bacteria were also metabolically active. A total of 9 isolates, belonging to the generaArthrobacter,Flavobacterium,Pseudomonas,Rhodococcus,Serratia, andStenotrophomonas, grew on alkaline carbonate-precipitating medium. Two cultures with the most intense precipitate formation,Arthrobacter sulfonivoransandRhodococcus globerulus, grew as aggregates, produced extracellular polymeric substances (EPS), and formed mixtures of calcite, vaterite, and monohydrocalcite.R. globerulusformed idiomorphous crystals with rhombohedral morphology, whereasA. sulfonivoransformed xenomorphous globular crystals, evidence for taxon-specific crystal morphologies. The results of this study highlighted the importance of combining various techniques in order to understand the geomicrobiology of karstic caves, but further studies are needed to determine whether the mineralogical biosignatures found in nutrient-rich media can also be found in oligotrophic caves.

scholarly journals Bioinformatic and Molecular Analysis of Inverse Autotransporters from Escherichia coli

mSphere ◽  
2019 ◽  
Vol 4 (4) ◽  
Author(s):  
Kelvin G. K. Goh ◽  
Danilo G. Moriel ◽  
Steven J. Hancock ◽  
Minh-Duy Phan ◽  
Mark A. Schembri
Keyword(s):  
Escherichia Coli ◽  
Cell Surface ◽  
Biofilm Formation ◽  
Secretion System ◽  
Content Type ◽  
E Coli ◽  
Wide Range ◽  
Type V Secretion ◽  
K 12 ◽  
Type V

ABSTRACT Proteins secreted by the type V secretion system possess multiple functions, including the capacity to mediate adhesion, aggregation, and biolfilm formation. The type V secretion system can be divided into five subclasses, one of which is the type Ve system. Proteins of the type Ve secretion system are also referred to as inverse autotransporters (IATs). In this study, we performed an in silico analysis of 126 completely sequenced Escherichia coli genomes available in the NCBI database and identified several distinct IAT-encoding gene families whose distribution varied throughout the E. coli phylogeny. The genes included three characterized IATs (intimin, fdeC, and yeeJ) and four uncharacterized IATs (here named iatA, iatB, iatC, and iatD). The four iat genes were cloned from the completely sequenced environmental E. coli strain SMS-3-5 and characterized. Three of these IAT proteins (IatB, IatC, and IatD) were expressed at the cell surface and possessed the capacity to mediate biofilm formation in a recombinant E. coli K-12 strain. Further analysis of the iatB gene, which showed a unique association with extraintestinal E. coli strains, suggested that its regulation is controlled by the LeuO global regulator. Overall, this study provides new data describing the prevalence, sequence variation, domain structure, function, and regulation of IATs found in E. coli. IMPORTANCE Escherichia coli is one of the most prevalent facultative anaerobes of the human gut. E. coli normally exists as a harmless commensal but can also cause disease following the acquisition of genes that enhance its pathogenicity. Adhesion is an important first step in colonization of the host and is mediated by an array of cell surface components. In E. coli, these include a family of adhesins secreted by the type V secretion system. Here, we identified and characterized new proteins from an emerging subclass of the type V secretion system known as the inverse autotransporters (IATs). We found that IAT-encoding genes are present in a wide range of strains and showed that three novel IATs were localized on the E. coli cell surface and mediated biofilm formation. Overall, this study provides new insight into the prevalence, function, and regulation of IATs in E. coli.

scholarly journals Type IX Secretion System Cargo Proteins Are Glycosylated at the C Terminus with a Novel Linking Sugar of the Wbp/Vim Pathway

mBio ◽  
2020 ◽  
Vol 11 (5) ◽  
Author(s):  
Paul D. Veith ◽  
Mikio Shoji ◽  
Richard A. J. O’Hair ◽  
Michael G. Leeming ◽  
Shuai Nie ◽  
...  
Keyword(s):  
Cell Surface ◽  
Porphyromonas Gingivalis ◽  
Biosynthetic Pathway ◽  
Secretion System ◽  
Tannerella Forsythia ◽  
Diverse Range ◽  
Content Type ◽  
C Terminus ◽  
Cargo Proteins ◽  
Type Ix Secretion System

ABSTRACT Porphyromonas gingivalis and Tannerella forsythia use the type IX secretion system to secrete cargo proteins to the cell surface where they are anchored via glycolipids. In P. gingivalis, the glycolipid is anionic lipopolysaccharide (A-LPS), of partially known structure. Modified cargo proteins were deglycosylated using trifluoromethanesulfonic acid and digested with trypsin or proteinase K. The residual modifications were then extensively analyzed by tandem mass spectrometry. The C terminus of each cargo protein was amide-bonded to a linking sugar whose structure was deduced to be 2-N-seryl, 3-N-acetylglucuronamide in P. gingivalis and 2-N-glycyl, 3-N-acetylmannuronic acid in T. forsythia. The structures indicated the involvement of the Wbp pathway to produce 2,3-di-N-acetylglucuronic acid and a WbpS amidotransferase to produce the uronamide form of this sugar in P. gingivalis. The wbpS gene was identified as PGN_1234 as its deletion resulted in the inability to produce the uronamide. In addition, the P. gingivalis vimA mutant which lacks A-LPS was successfully complemented by the T. forsythia vimA gene; however, the linking sugar was altered to include glycine rather than serine. After removal of the acetyl group at C-2 by the putative deacetylase, VimE, VimA presumably transfers the amino acid to complete the biosynthesis. The data explain all the enzyme activities required for the biosynthesis of the linking sugar accounting for six A-LPS-specific genes. The linking sugar is therefore the key compound that enables the attachment of cargo proteins in P. gingivalis and T. forsythia. We propose to designate this novel linking sugar biosynthetic pathway the Wbp/Vim pathway. IMPORTANCE Porphyromonas gingivalis and Tannerella forsythia, two pathogens associated with severe gum disease, use the type IX secretion system (T9SS) to secrete and attach toxic arrays of virulence factor proteins to their cell surfaces. The proteins are tethered to the outer membrane via glycolipid anchors that have remained unidentified for more than 2 decades. In this study, the first sugar molecules (linking sugars) in these anchors are identified and found to be novel compounds. The novel biosynthetic pathway of these linking sugars is also elucidated. A diverse range of bacteria that do not have the T9SS were found to have the genes for this pathway, suggesting that they may synthesize similar linking sugars for utilization in different systems. Since the cell surface attachment of virulence factors is essential for virulence, these findings reveal new targets for the development of novel therapies.

scholarly journals Hypothetical Protein Avin_16040 as the S-Layer Protein of Azotobacter vinelandii and Its Involvement in Plant Root Surface Attachment

2015 ◽  
Vol 81 (21) ◽  
pp. 7484-7495 ◽  
Author(s):  
Pauline Woan Ying Liew ◽  
Bor Chyan Jong ◽  
Nazalan Najimudin
Keyword(s):  
Cell Surface ◽  
Deletion Mutant ◽  
Azotobacter Vinelandii ◽  
Plant Root ◽  
Hypothetical Protein ◽  
Surface Hydrophobicity ◽  
Root Surface ◽  
Bacterial Cells ◽  
Wild Type ◽  
Content Type

ABSTRACTA proteomic analysis of a soil-dwelling, plant growth-promotingAzotobacter vinelandiistrain showed the presence of a protein encoded by the hypotheticalAvin_16040gene when the bacterial cells were attached to theOryza sativaroot surface. AnAvin_16040deletion mutant demonstrated reduced cellular adherence to the root surface, surface hydrophobicity, and biofilm formation compared to those of the wild type. By atomic force microscopy (AFM) analysis of the cell surface topography, the deletion mutant displayed a cell surface architectural pattern that was different from that of the wild type.Escherichia colitransformed with the wild-typeAvin_16040gene displayed on its cell surface organized motifs which looked like the S-layer monomers ofA. vinelandii. The recombinantE. colialso demonstrated enhanced adhesion to the root surface.

scholarly journals The Type IX Secretion System Is Required for Virulence of the Fish Pathogen Flavobacterium columnare

2017 ◽  
Vol 83 (23) ◽  
Author(s):  
Nan Li ◽  
Yongtao Zhu ◽  
Benjamin R. LaFrentz ◽  
Jason P. Evenhuis ◽  
David W. Hunnicutt ◽  
...  
Keyword(s):  
Secretion System ◽  
Gliding Motility ◽  
Control Measures ◽  
Flavobacterium Columnare ◽  
Wild Type ◽  
Secretion Systems ◽  
Content Type ◽  
Freshwater Aquaculture ◽  
Columnaris Disease ◽  
Spent Media

ABSTRACT Flavobacterium columnare, a member of the phylum Bacteroidetes, causes columnaris disease in wild and aquaculture-reared freshwater fish. The mechanisms responsible for columnaris disease are not known. Many members of the phylum Bacteroidetes use type IX secretion systems (T9SSs) to secrete enzymes, adhesins, and proteins involved in gliding motility. The F. columnare genome has all of the genes needed to encode a T9SS. gldN, which encodes a core component of the T9SS, was deleted in wild-type strains of F. columnare. The F. columnare ΔgldN mutants were deficient in the secretion of several extracellular proteins and lacked gliding motility. The ΔgldN mutants exhibited reduced virulence in zebrafish, channel catfish, and rainbow trout, and complementation restored virulence. PorV is required for the secretion of a subset of proteins targeted to the T9SS. An F. columnare ΔporV mutant retained gliding motility but exhibited reduced virulence. Cell-free spent media from exponentially growing cultures of wild-type and complemented strains caused rapid mortality, but spent media from ΔgldN and ΔporV mutants did not, suggesting that soluble toxins are secreted by the T9SS. IMPORTANCE Columnaris disease, caused by F. columnare, is a major problem for freshwater aquaculture. Little is known regarding the virulence factors produced by F. columnare, and control measures are limited. Analysis of targeted gene deletion mutants revealed the importance of the type IX protein secretion system (T9SS) and of secreted toxins in F. columnare virulence. T9SSs are common in members of the phylum Bacteroidetes and likely contribute to the virulence of other animal and human pathogens.

scholarly journals High Adhesion and Increased Cell Death Contribute to Strong Biofilm Formation in Klebsiella pneumoniae

Pathogens ◽  
2019 ◽  
Vol 8 (4) ◽  
pp. 277
Author(s):  
Siddhi Desai ◽  
Kinjal Sanghrajka ◽  
Devarshi Gajjar
Keyword(s):  
Cell Death ◽  
Klebsiella Pneumoniae ◽  
Biofilm Formation ◽  
Laser Scanning ◽  
Laser Scanning Microscopy ◽  
Extracellular Dna ◽  
Confocal Laser ◽  
Bacterial Cells ◽  
High Adhesion ◽  
Biofilm Matrix

Klebsiella pneumoniae (Kp), is a frequent cause of hospital and community-acquired infections and WHO had declared it as a “priority pathogen”. Biofilm is a major virulence factor of Kp and yet the mechanism of strong biofilm formation in Kp is unclear. A key objective of the present study is to investigate the differences between strong and weak biofilms formed by clinical isolates of Kp on various catheters and in different media conditions and to identify constituents contributing to strong biofilm formation. Quantification of matrix components (extracellular DNA (eDNA), protein, exopolysaccharides (EPS), and bacterial cells), confocal laser scanning microscopy (CLSM), field emission gun scanning electron microscopy (FEG-SEM) and flow-cytometry analysis were performed to compare strong and weak biofilm matrix. Our results suggest increased biofilm formation on latex catheters compared to silicone and silicone-coated latex catheters. Higher amounts of eDNA, protein, EPS, and dead cells were observed in the strong biofilm of Kp. High adhesion capacity and cell death seem to play a major role in formation of strong Kp biofilms. The enhanced eDNA, EPS, and protein in the biofilm matrix appear as a consequence of increased cell death.

scholarly journals The Type IX Secretion System: Advances in Structure, Function and Organisation

2020 ◽  
Vol 8 (8) ◽  
pp. 1173 ◽  
Author(s):  
Dhana G. Gorasia ◽  
Paul D. Veith ◽  
Eric C. Reynolds
Keyword(s):  
Cell Surface ◽  
Structure Function ◽  
Secretion System ◽  
Gliding Motility ◽  
Flavobacterium Johnsoniae ◽  
Bacteroidetes Phylum ◽  
Transcription Regulators ◽  
Type Ix Secretion System ◽  
Insight Into ◽  
Made In

The type IX secretion system (T9SS) is specific to the Bacteroidetes phylum. Porphyromonas gingivalis, a keystone pathogen for periodontitis, utilises the T9SS to transport many proteins—including its gingipain virulence factors—across the outer membrane and attach them to the cell surface. Additionally, the T9SS is also required for gliding motility in motile organisms, such as Flavobacterium johnsoniae. At least nineteen proteins have been identified as components of the T9SS, including the three transcription regulators, PorX, PorY and SigP. Although the components are known, the overall organisation and the molecular mechanism of how the T9SS operates is largely unknown. This review focusses on the recent advances made in the structure, function, and organisation of the T9SS machinery to provide further insight into this highly novel secretion system.

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