scholarly journals Fibronectin Binding Protein BBK32 of the Lyme Disease Spirochete Promotes Bacterial Attachment to Glycosaminoglycans

2006 ◽  
Vol 74 (1) ◽  
pp. 435-441 ◽  
Author(s):  
Joshua R. Fischer ◽  
Kimberly T. LeBlanc ◽  
John M. Leong
Keyword(s):  
Lyme Disease ◽  
Mammalian Cells ◽  
Dermatan Sulfate ◽  
Shuttle Vector ◽  
Cell Types ◽  
Binding Activity ◽  
Bacterial Attachment ◽  
Mammalian Host ◽  
High Passage ◽  
Bacterial Surface

ABSTRACT Borrelia burgdorferi, the agent of Lyme disease, causes a multisystemic illness that can affect the skin, heart, joints, and nervous system and is capable of attachment to diverse cell types. Among the host components recognized by this spirochete are fibronectin and glycosaminoglycans (GAGs). Three surface-localized GAG-binding bacterial ligands, Bgp, DbpA, and DbpB, have been previously identified, but recent studies suggested that at least one additional GAG-binding ligand is expressed on the spirochetal surface when the spirochete is adapted to the mammalian host environment. BBK32 is a surface lipoprotein that is produced during infection and that has been shown to bind to fibronectin. In this study, we show that, when BBK32 was produced from a shuttle vector in an otherwise nonadherent high-passage B. burgdorferi strain, the protein localized on the bacterial surface and conferred attachment to fibronectin and to mammalian cell monolayers. In addition, the high-passage strain producing BBK32 bound to purified preparations of the GAGs dermatan sulfate and heparin, as well as to these GAGs on the surfaces of cultured mammalian cells. Recombinant BBK32 recognized purified heparin, indicating that the bacterial attachment to GAGs was due to direct binding by BBK32. This GAG-binding activity of BBK32 is apparently independent of fibronectin recognition, because exogenous heparin had no effect on BBK32-mediated bacterial binding to fibronectin.

scholarly journals Allelic Variation of the Lyme Disease Spirochete Adhesin DbpA Influences Spirochetal Binding to Decorin, Dermatan Sulfate, and Mammalian Cells

2011 ◽  
Vol 79 (9) ◽  
pp. 3501-3509 ◽  
Author(s):  
Vivian M. Benoit ◽  
Joshua R. Fischer ◽  
Yi-Pin Lin ◽  
Nikhat Parveen ◽  
John M. Leong
Keyword(s):  
Lyme Disease ◽  
Mammalian Cells ◽  
Dermatan Sulfate ◽  
Allelic Variation ◽  
Protein A ◽  
Sensu Stricto ◽  
Binding Activity ◽  
Mammalian Host ◽  
Content Type ◽  
Variable Surface

ABSTRACTAfter transmission by an infected tick, the Lyme disease spirochete,Borrelia burgdorferisensu lato, colonizes the mammalian skin and may disseminate systemically. The three major species of Lyme disease spirochete—B. burgdorferisensu stricto,B. garinii, andB. afzelii—are associated with different chronic disease manifestations. Colonization is likely promoted by the ability to bind to target tissues, and Lyme disease spirochetes utilize multiple adhesive molecules to interact with diverse mammalian components. The allelic variable surface lipoprotein decorin binding protein A (DbpA) promotes bacterial binding to the proteoglycan decorin and to the glycosaminoglycan (GAG) dermatan sulfate. To assess allelic variation of DbpA in GAG-, decorin-, and cell-binding activities, we expresseddbpAalleles derived from diverse Lyme disease spirochetes inB. burgdorferistrain B314, a noninfectious and nonadherent strain that lacksdbpA. Each DbpA allele conferred uponB. burgdorferistrain B314 the ability to bind to cultured kidney epithelial (but not glial or endothelial) cells, as well as to purified decorin and dermatan sulfate. Nevertheless, allelic variation of DbpA was associated with dramatic differences in substrate binding activity. In most cases, decorin and dermatan sulfate binding correlated well, but DbpA ofB. afzeliistrain VS461 promoted differential binding to decorin and dermatan sulfate, indicating that the two activities are separable. DbpA from a clone ofB. burgdorferistrain N40 that can cause disseminated infection in mice displayed relatively low adhesive activity, indicating that robust DbpA-mediated adhesive activity is not required for spread in the mammalian host.

The structural mechanics of cell division in Trypanosoma brucei

2008 ◽  
Vol 36 (3) ◽  
pp. 421-424 ◽  
Author(s):  
Sue Vaughan ◽  
Keith Gull
Keyword(s):  
Cell Division ◽  
Trypanosoma Brucei ◽  
Mammalian Cells ◽  
Reverse Genetics ◽  
Protozoan Parasite ◽  
Cell Types ◽  
Single Copy ◽  
Structural Mechanics ◽  
Sub Saharan Africa ◽  
Mammalian Host

Undoubtedly, there are fundamental processes driving the structural mechanics of cell division in eukaryotic organisms that have been conserved throughout evolution and are being revealed by studies on organisms such as yeast and mammalian cells. Precision of structural mechanics of cytokinesis is however probably no better illustrated than in the protozoa. A dramatic example of this is the protozoan parasite Trypanosoma brucei, a unicellular flagellated parasite that causes a devastating disease (African sleeping sickness) across Sub-Saharan Africa in both man and animals. As trypanosomes migrate between and within a mammalian host and the tsetse vector, there are periods of cell proliferation and cell differentiation involving at least five morphologically distinct cell types. Much of the existing cytoskeleton remains intact during these processes, necessitating a very precise temporal and spatial duplication and segregation of the many single-copy organelles. This structural precision is aiding progress in understanding these processes as we apply the excellent reverse genetics and post-genomic technologies available in this system. Here we outline our current understanding of some of the structural aspects of cell division in this fascinating organism.

scholarly journals Peptidomimetic Polyurethanes Disrupt Surface Established Bacterial Biofilms and Prevent Biofilm Formation

2021 ◽  
Author(s):  
Apoorva Vishwakarma ◽  
Francis Dang ◽  
Allison Ferrell ◽  
Hazel A. Barton ◽  
Abraham Joy
Keyword(s):  
Biofilm Formation ◽  
Mammalian Cells ◽  
Bacterial Infections ◽  
Polymyxin B ◽  
Bacterial Attachment ◽  
Water Soluble ◽  
Chronic Infections ◽  
Bacterial Surface ◽  
Host Immune Responses ◽  
Conventional Antibiotics

Over 80% of all chronic bacterial infections in humans are associated with biofilms, which are surface-associated bacterial communities encased within a secreted exopolysaccharide matrix that can provide resistance to environmental and chemical insults. Biofilm formation triggers broad adaptive changes in the bacteria, allowing them to be almost a thousand-fold more resistant to conventional antibiotic treatments and host immune responses. The failure of antibiotics to eliminate biofilms leads to persistent chronic infections and can promote the development of antibiotic-resistant strains. Therefore, there is an urgent need to develop agents that effectively prevent biofilm formation and eradicate established biofilms. Herein, we present water-soluble synthetic peptidomimetic polyurethanes that can disrupt surface established biofilms of <i>Pseudomonas aeruginosa, Staphylococcus aureus, </i>and <i>Escherichia coli</i>, all of which show tolerance to the conventional antibiotics polymyxin B and ciprofloxacin. Furthermore, these polyurethanes prevent bacterial attachment and stimulate bacterial surface motility to inhibit biofilm formation of both Gram-positive and Gram-negative bacteria at sub-inhibitory concentrations, without being toxic to mammalian cells. Our results show that these polyurethanes show promise as a platform for the development of therapeutics that target biofilms and modulate surface interactions of bacteria for the treatment of chronic biofilm-associated infections and as antibiofilm agents.

scholarly journals Peptidomimetic Polyurethanes Disrupt Surface Established Bacterial Biofilms and Prevent Biofilm Formation

2021 ◽  
Author(s):  
Apoorva Vishwakarma ◽  
Francis Dang ◽  
Allison Ferrell ◽  
Hazel A. Barton ◽  
Abraham Joy
Keyword(s):  
Biofilm Formation ◽  
Mammalian Cells ◽  
Bacterial Infections ◽  
Polymyxin B ◽  
Bacterial Attachment ◽  
Water Soluble ◽  
Chronic Infections ◽  
Bacterial Surface ◽  
Host Immune Responses ◽  
Conventional Antibiotics

Over 80% of all chronic bacterial infections in humans are associated with biofilms, which are surface-associated bacterial communities encased within a secreted exopolysaccharide matrix that can provide resistance to environmental and chemical insults. Biofilm formation triggers broad adaptive changes in the bacteria, allowing them to be almost a thousand-fold more resistant to conventional antibiotic treatments and host immune responses. The failure of antibiotics to eliminate biofilms leads to persistent chronic infections and can promote the development of antibiotic-resistant strains. Therefore, there is an urgent need to develop agents that effectively prevent biofilm formation and eradicate established biofilms. Herein, we present water-soluble synthetic peptidomimetic polyurethanes that can disrupt surface established biofilms of <i>Pseudomonas aeruginosa, Staphylococcus aureus, </i>and <i>Escherichia coli</i>, all of which show tolerance to the conventional antibiotics polymyxin B and ciprofloxacin. Furthermore, these polyurethanes prevent bacterial attachment and stimulate bacterial surface motility to inhibit biofilm formation of both Gram-positive and Gram-negative bacteria at sub-inhibitory concentrations, without being toxic to mammalian cells. Our results show that these polyurethanes show promise as a platform for the development of therapeutics that target biofilms and modulate surface interactions of bacteria for the treatment of chronic biofilm-associated infections and as antibiofilm agents.

scholarly journals Strain Variation in Glycosaminoglycan Recognition Influences Cell-Type-Specific Binding by Lyme Disease Spirochetes

1999 ◽  
Vol 67 (4) ◽  
pp. 1743-1749 ◽  
Author(s):  
Nikhat Parveen ◽  
Douglas Robbins ◽  
John M. Leong
Keyword(s):  
Lyme Disease ◽  
Heparan Sulfate ◽  
Dermatan Sulfate ◽  
Specific Binding ◽  
C6 Glioma Cells ◽  
Cell Type ◽  
Cell Binding ◽  
High Passage ◽  
Binding Preference ◽  
Cell Type Specific

ABSTRACT Lyme disease, a chronic multisystemic disorder that can affect the skin, heart, joints, and nervous system is caused by Borrelia burgdorferi sensu lato. Lyme disease spirochetes were previously shown to bind glycosaminoglycans (GAGs). In the current study, the GAG-binding properties of eight Lyme disease strains were determined. Binding by two high-passage HB19 derivatives to Vero cells could not be inhibited by enzymatic removal of GAGs or by the addition of exogenous GAG. The other six strains, which included a different high-passage HB19 derivative (HB19 clone 1), were shown to recognize both heparan sulfate and dermatan sulfate in cell-binding assays, but the relative efficiency of binding to these two GAGs varied among the strains. Strains N40, CA20-2A, and PBi bound predominantly to heparan sulfate, PBo bound both heparan sulfate and dermatan sulfate roughly equally, and VS461 and HB19 clone 1 recognized primarily dermatan sulfate. Cell binding by strain HB19 clone 1 was inhibited better by exogenous dermatan sulfate than by heparin, whereas heparin was the better inhibitor of binding by strain N40. The GAG-binding preference of a Lyme disease strain was reflected in its cell-type-specific binding. Strains that recognized predominantly heparan sulfate bound efficiently to both C6 glioma cells and EA-Hy926 cells, whereas strains that recognized predominantly dermatan sulfate bound well only to the glial cells. The effect of lyase treatment of these cells on bacterial binding was consistent with the model that cell-type-specific binding was a reflection of the GAG-binding preference. We conclude that the GAG-binding preference varies with the strain of Lyme disease spirochete and that this variation influences cell-type-specific binding in vitro.

scholarly journals A novel cell permeability assay for macromolecules

2020 ◽  
Author(s):  
Yensi Alejandra Flores Bueso ◽  
Sidney Peter Walker ◽  
Jennifer Quinn ◽  
Mark Tangney
Keyword(s):  
Mammalian Cells ◽  
Cell Types ◽  
Binding Activity ◽  
Cell Permeability ◽  
Bacterial Cells ◽  
Bacterial Membranes ◽  
Specific Permeability ◽  
Biotin Binding ◽  
Permeability Assay ◽  
Formalin Fixed

Abstract Background: Many cell permeabilisation methods to mediate internalisation of various molecules to mammalian or bacterial cells have been developed. However, no size-specific permeability assay suitable for both cell types exists. Results: We report the use of intrinsically biotinylated cell components as the target for reporter molecules for assessing permeabilisation. Due to its well-described biotin binding activity, we developed an assay using Streptavidin (SAv) as a molecular weight marker for assessing eukaryotic and prokaryotic cell internalisation, using flow cytometry as a readout. This concept was tested here as part of the development of host DNA depletion strategies for microbiome analysis of formalin-fixed (FF) samples. Host depletion (HD) strategies require differential cell permeabilisation, where mammalian cells but not bacterial cells are permeabilised, and are subsequently treated with a nuclease. Here, the internalisation of a SAv-conjugate was used as a reference for nucleases of similar dimensions. With this assay, it was possible to demonstrate that formalin fixation does not generate pores which allow the introduction of 60 KDa molecules in mammalian or bacterial membranes/envelopes. Among surfactants tested, Saponin derived from Quillaja bark showed the best selectivity for mammalian cell permeabilisation, which, when coupled with Benzonase nuclease, provided the best results for host DNA depletion, representing a new HD strategy for formalin fixed samples.Conclusion: The assay presented provides researchers with a sensitive and accessible tool for discerning membrane/cell envelop permeability for different size macromolecules.

scholarly journals Identification and Functional Assessment of the First Placental Adhesin of Treponema pallidum That May Play Critical Role in Congenital Syphilis

2020 ◽  
Vol 11 ◽  
Author(s):  
Shekerah Primus ◽  
Sandra C. Rocha ◽  
Lorenzo Giacani ◽  
Nikhat Parveen
Keyword(s):  
Cell Lines ◽  
Dermatan Sulfate ◽  
Congenital Syphilis ◽  
Treponema Pallidum ◽  
Bacterial Attachment ◽  
Host Cells ◽  
Mammalian Host ◽  
Bewo Cell ◽  
Specific Cell ◽  
Mammalian Cell Lines

Syphilis is a global, re-emerging sexually transmitted infection and congenital syphilis remains a major cause of adverse pregnancy outcomes due to bacterial infection in developing nations with a high rate of fetus loss. The molecular mechanisms involved in pathogenesis of the causative agent, Treponema pallidum subsp. pallidum remain poorly understood due to the difficulties of working with this pathogen, including the inability to grow it in pure culture. To reduce the spread of syphilis, we must first increase our knowledge of the virulence factors of T. pallidum and their contribution to syphilis manifestations. Tp0954 was predicted to be a surface lipoprotein of T. pallidum. Therefore, we experimentally demonstrated that Tp0954 is indeed a surface protein and further investigated its role in mediating bacterial attachment to various mammalian host cells. We found that expression of Tp0954 in a poorly adherent, but physiologically related derivative strain of the Lyme disease causing spirochete Borrelia burgdorferi B314 strain promotes its binding to epithelial as well as non-epithelial cells including glioma and placental cell lines. We also found that Tp0954 expression facilitates binding of this strain to purified dermatan sulfate and heparin, and also that bacterial binding to mammalian cell lines is mediated by the presence of heparan sulfate and dermatan sulfate in the extracellular matrix of the specific cell lines. These results suggest that Tp0954 may be involved not only in initiating T. pallidum infection by colonizing skin epithelium, but it may also contribute to disseminated infection and colonization of distal tissues. Significantly, we found that Tp0954 promotes binding to the human placental choriocarcinoma BeWo cell line, which is of trophoblastic endocrine cell type, as well as human placental tissue sections, suggesting its role in placental colonization and possible contribution to transplacental transmission of T. pallidum. Altogether, these novel findings offer an important step toward unraveling syphilis pathogenesis, including placental colonization and T. pallidum vertical transmission from mother to fetus during pregnancy.

scholarly journals A novel cell permeability assay for macromolecules

2020 ◽  
Vol 21 (1) ◽  
Author(s):  
Yensi Flores Bueso ◽  
Sidney Walker ◽  
Jennifer Quinn ◽  
Mark Tangney
Keyword(s):  
Mammalian Cells ◽  
Cell Types ◽  
Binding Activity ◽  
Cell Permeability ◽  
Bacterial Cells ◽  
Bacterial Membranes ◽  
Specific Permeability ◽  
Biotin Binding ◽  
Permeability Assay ◽  
Formalin Fixed

Abstract Background Many cell permeabilisation methods to mediate internalisation of various molecules to mammalian or bacterial cells have been developed. However, no size-specific permeability assay suitable for both cell types exists. Results We report the use of intrinsically biotinylated cell components as the target for reporter molecules for assessing permeabilisation. Due to its well-described biotin binding activity, we developed an assay using Streptavidin (SAv) as a molecular weight marker for assessing eukaryotic and prokaryotic cell internalisation, using flow cytometry as a readout. This concept was tested here as part of the development of host DNA depletion strategies for microbiome analysis of formalin-fixed (FF) samples. Host depletion (HD) strategies require differential cell permeabilisation, where mammalian cells but not bacterial cells are permeabilised, and are subsequently treated with a nuclease. Here, the internalisation of a SAv-conjugate was used as a reference for nucleases of similar dimensions. With this assay, it was possible to demonstrate that formalin fixation does not generate pores which allow the introduction of 60 KDa molecules in mammalian or bacterial membranes/envelopes. Among surfactants tested, Saponin derived from Quillaja bark showed the best selectivity for mammalian cell permeabilisation, which, when coupled with Benzonase nuclease, provided the best results for host DNA depletion, representing a new HD strategy for formalin fixed samples. Conclusion The assay presented provides researchers with a sensitive and accessible tool for discerning membrane/cell envelop permeability for different size macromolecules.

scholarly journals Review onTrypanosoma cruzi: Host Cell Interaction

2010 ◽  
Vol 2010 ◽  
pp. 1-18 ◽  
Author(s):  
Wanderley de Souza ◽  
Tecia Maria Ulisses de Carvalho ◽  
Emile Santos Barrias
Keyword(s):  
Trypanosoma Cruzi ◽  
Host Cell ◽  
Mammalian Cells ◽  
Cell Types ◽  
Cell Interaction ◽  
Host Cells ◽  
Mammalian Host ◽  
Blood Sucking ◽  
Number Of Individuals ◽  
Host Cell Interaction

Trypanosoma cruzi, the causative agent of Chagas' disease, which affects a large number of individuals in Central and South America, is transmitted to vertebrate hosts by blood-sucking insects. This protozoan is an obligate intracellular parasite. The infective forms of the parasite are metacyclic and bloodstream trypomastigote and amastigote. Metacyclic trypomastigotes are released with the feces of the insect while amastigotes and bloodstream trypomastigotes are released from the infected host cells of the vertebrate host after a complex intracellular life cycle. The recognition between parasite and mammalian host cell involves numerous molecules present in both cell types. Here, we present a brief review of the interaction betweenTrypanosoma cruziand its host cells, mainly emphasizing the mechanisms and molecules that participate in theT. cruziinvasion process of the mammalian cells.

scholarly journals Leptospira interrogans Binds to Human Cell Surface Receptors Including Proteoglycans

2009 ◽  
Vol 77 (12) ◽  
pp. 5528-5536 ◽  
Author(s):  
Deborah D. Breiner ◽  
Mark Fahey ◽  
Ryan Salvador ◽  
Jana Novakova ◽  
Jenifer Coburn
Keyword(s):  
Mammalian Cells ◽  
Public Health Problem ◽  
Binding Activity ◽  
Bacterial Attachment ◽  
Leptospira Interrogans ◽  
Global Public Health ◽  
Host Animal ◽  
Surface Receptors ◽  
Causative Agents ◽  
New Hosts

ABSTRACT Leptospirosis is a global public health problem, primarily in the tropical developing world. The pathogenic mechanisms of the causative agents, several members of the genus Leptospira, have been underinvestigated. The exception to this trend has been the demonstration of the binding of pathogenic leptospires to the extracellular matrix (ECM) and its components. In this work, interactions of Leptospira interrogans bacteria with mammalian cells, rather than the ECM, were examined. The bacteria bound more efficiently to the cells than to the ECM, and a portion of this cell-binding activity was attributable to attachment to glycosaminoglycan (GAG) chains of proteoglycans (PGs). Chondroitin sulfate B PGs appeared to be the primary targets of L. interrogans attachment, while heparan sulfate PGs were much less important. Inhibition of GAG/PG-mediated attachment resulted in partial inhibition of bacterial attachment, suggesting that additional receptors for L. interrogans await identification. GAG binding may participate in the pathogenesis of leptospirosis within the host animal. In addition, because GAGs are expressed on the luminal aspects of epithelial cells in the proximal tubules of the kidneys, this activity may play a role in targeting the bacteria to this critical site. Because GAGs are shed in the urine, GAG binding may also be important for transmission to new hosts through the environment.

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