scholarly journals Mammalian Cell Entry domains are required for bile resistance and virulence inSalmonella

2018 ◽  
Author(s):  
Georgia L. Isom ◽  
Jessica L. Rooke ◽  
Camila A. Antunes ◽  
Emma Sheehan ◽  
Timothy J. Wells ◽  
...  
Keyword(s):  
Mammalian Cell ◽  
Mammalian Cells ◽  
Bile Salts ◽  
Pathogenic Bacteria ◽  
Cell Entry ◽  
Direct Role ◽  
Bile Resistance ◽  
Mammalian Infection ◽  
Membrane Physiology

AbstractMCE domains were first reported inMycobacteriaas having a role inMammalianCellEntry, with subsequent studies showing their importance during infection. Here, we have examined the function of MCE proteins inSalmonellaTyphimurium during mammalian infection. We report that MCE proteins are required forSalmonellavirulence, but that this is not related to decreased adherence, entry or survival in mammalian cells. Instead, we reveal that MCE proteins are required forSalmonellabile resistance, in particular to withstand bile salts such as cholate and deoxycholate. Based on our previous work inEscherichia coli, and other studies that have reported roles for MCE proteins in membrane biogenesis, we propose thatSalmonellalacking MCE domains have a defective outer membrane that results in bile sensitivity and decreased virulencein vivo. These results suggest that MCE domains mediate fundamental aspects of bacterial membrane physiology as opposed to a proposed direct role in mammalian cell entry, explaining their conservation across both pathogenic and non-pathogenic bacteria.

scholarly journals Recruitment of Mammalian Cell Fibronectin to the Surface of Chlamydia trachomatis

2002 ◽  
Vol 70 (7) ◽  
pp. 3935-3938 ◽  
Author(s):  
Betsy J. Kleba ◽  
Erin Banta ◽  
Erika A. Lindquist ◽  
Richard S. Stephens
Keyword(s):  
Chlamydia Trachomatis ◽  
Heparan Sulfate ◽  
Host Cell ◽  
Mammalian Cell ◽  
Pathogenic Bacteria ◽  
Cell Binding ◽  
Cell Functions ◽  
Wide Range

ABSTRACT Pathogenic bacteria exploit the presence of various host cell molecules in order to colonize new tissues. Fibronectin is involved in a wide range of cell functions in vivo, and staphylococci, streptococci, and gonococci have evolved mechanisms to utilize this glycoprotein to mediate host cell binding. We show that elementary bodies (EB) from two biovars of Chlamydia trachomatis recruit fibronectin to their surfaces upon lysis of the host cell. We also demonstrate that a heparan sulfate lyase-sensitive molecule on chlamydial EB is responsible for binding at least a portion of this fibronectin.

scholarly journals Evaluation of the Probiotic Profile of the Lactobacillus Acidophilus Used in Pharmaceutical and Food Applications

2018 ◽  
Vol 40 (1) ◽  
pp. 36664
Author(s):  
Jucelaine Deon Schmitt ◽  
Luciana Oliveira de Fariña ◽  
Márcia Regina Simões ◽  
Luciana Bill Mikito Kottwitz
Keyword(s):  
Gastrointestinal Tract ◽  
Lactobacillus Acidophilus ◽  
Bile Salts ◽  
Pathogenic Bacteria ◽  
Growth Conditions ◽  
Commercial Use ◽  
Inhibitory Compounds ◽  
Food Applications ◽  
Fermented Dairy

 Lactobacillus acidophilus used in three different applications, compounding pharmacies (LA1), fermented dairy (LA2), and allopathic compoundings (LA3) were tested to evaluate the existence of significant differences between them and in different growth conditions. In the evaluation of resistance to different commercial use antibiotics, all strains were sensitive to the antibiotics ampicillin, chloramphenicol, doxycycline, and tetracycline. LA1 was considered moderately sensitive (MS) to erythromycin and LA3 was MS to clindamycin and erythromycin. LA3 was classified between MS to resistant to erythromycin. All three strains were resistant to gentamicin. When evaluating acid pH resistance, the three origins presented similar behavior, with a decrease in cell viability at pH 2, maintaining constant viability at pH 3 and 4. In the test of resistance to the gastrointestinal tract conditions and hydrophobicity, LA2 presented better results. The three strains showed production of inhibitory compounds against pathogenic bacteria and deconjugated tauroconjugated bile salts (TDCA). It was concluded that, depending on the origin, Lactobacillus acidophilus may present different behaviors that will determine its growth and, consequently, its action in vivo. Due to the practicality of access, economy, and the satisfactory results in the tests performed, LA2 can be considered the strain of choice among those studied. 

A double-label confocal imaging method for detecting the internalization of bacteria by non-phagocytic mammalian cells

1994 ◽  
Vol 52 ◽  
pp. 216-217
Author(s):  
M. H. Chestnut ◽  
L. L. Odioso ◽  
T. E. Otte ◽  
B.C. Hulette
Keyword(s):  
Plasma Membrane ◽  
Mammalian Cell ◽  
Mammalian Cells ◽  
Pathogenic Bacteria ◽  
Confocal Imaging ◽  
Imaging Method ◽  
Thin Sections ◽  
Double Label ◽  
Human Gingival Epithelial Cells ◽  
Immunofluorescence Labeling

Internalization by non-phagocytic mammalian cells is a key part of the life cycle of several important pathogenic bacteria. Mechanistic studies of this event often include transmission electron micrographs intended to demonstrate the presence of bacteria within the mammalian cell, but these data are often uncompelling. It is very difficult to be certain, based on the appearance and number of phospholipid membranes surrounding the bacterial cell, if it is enclosed in a vesicle, is surrounded directly by mammalian cell cytoplasm, or is merely contained within an invagination of the plasma membrane. Serial thin sections could provide the needed clarification, but are almost never presented, presumably because of the effort involved. Here we offer an alternative method, combining immunofluorescence labeling of the bacterium and fluorescence staining of the mammalian cell plasma membrane with a lipophilic dye, to visualize the internalization of bacteria by mammalian cells.Freshly isolated human gingival epithelial cells were cultured in keratinocyle serum-free medium (KSFM). Second passage cells were seeded on coverslips and placed in 24-well plates.

Novel Bioreactor-System for In Situ-Cultivation of Artificial Tissue

2016 ◽  
Vol 879 ◽  
pp. 1002-1007
Author(s):  
Jordanka Kostova ◽  
Sylvio Schneider ◽  
Sabine Sauer ◽  
Andrea Böhme ◽  
Mauro Casalboni ◽  
...  
Keyword(s):  
Mammalian Cell ◽  
Mammalian Cells ◽  
Ph Value ◽  
Artificial Organs ◽  
Parallel Operation ◽  
Single Chamber ◽  
Mammalian Cell Lines ◽  
Micro Pump ◽  
Bioreactor System

A bioreactor is a device simulating physiological environments for different biotechnological applications. In highly promising research fields like tissue engineering micro-sized bioreactors were utilized successfully promoting mammalian cells to grow and build 3D cell structures similar to in vivo environments. For any practical application and even for improved R&D it is necessary to generate and maintain a physiological environment over the whole cultivation period (hours, days or weeks, in case of artificial organs even up to months). Depending on the field of application physiological environments can comprise different parameters. In case of mammalian cell lines these parameters require a complex supply and monitoring system. Thus, we developed a semi-automated bioreactor-system for long-term cultivation of different mammalian cell types imitating physiological conditions. The system included detection and control of the following parameters: temperature, pH-value, gas concentration and the continuous supply with nutrients. A micro fluidic network was established enabling a high through-put cultivating system as bioreactor-system. The bioreactor-system consists of several micro-sized chambers in a microliter scale (the related article discussing the micro-sized chambers “Miniaturized Flow-Through Bioreactor for Processing and Testing in Pharmacology” by Boehme et al is published within this issue). The chambers were placed in a polymeric slide each with an individual medium supply and disposal. Every single chamber thus was connected to an individual syringe-based micro-pump setup and supplied by nutrients solution with a velocity of 100μl/h. The pH-value was observed optically and controlled via CO2 supply. All gas interchanges into every single chamber were realized via semi permeable membranes. The required temperature was adjusted via an appropriate custom-fit heating system utilizing MOSFETs allocated on an aluminum board along the slides. Two slides each were housed in a PMMA case. This bioreactor-system is a first prototype for larger systems aiming for the parallel operation of up to 100 micro-sized reaction chambers.

scholarly journals The inlAB locus mediates the entry of Listeria monocytogenes into hepatocytes in vivo.

1996 ◽  
Vol 183 (2) ◽  
pp. 359-369 ◽  
Author(s):  
J L Gaillard ◽  
F Jaubert ◽  
P Berche
Keyword(s):  
Listeria Monocytogenes ◽  
Mammalian Cells ◽  
Defense Mechanisms ◽  
Inflammatory Cells ◽  
Cell Infection ◽  
Chromosomal Locus ◽  
Direct Role ◽  
Innate Defense ◽  
A Cell

The intracellular parasite Listeria monocytogenes is able to induce its internalization by cultured mammalian cells that are not normally phagocytic. This process requires the expression of the chromosomal locus inlAB. We studied the virulence of an inlAB mutant and of its parent in murine listeriosis. Irrespective of the route of inoculation, the inlAB mutant was severely attenuated for growth in the liver. The livers of mice inoculated with the inlAB mutant displayed much smaller infectious foci than the parent as early as 24 h after infection. Electron microscopy showed that these foci consisted of a few inflammatory cells, with few bacteria; bacteria were rarely found within hepatocytes. In contrast, foci in livers of mice inoculated with the parent consisted of islets of heavily infected hepatocytes that were infiltrated by numerous neutrophils; bacteria seemed intact within hepatocytes and damaged within neutrophils. A direct role of inlAB for the entry of L. monocytogenes into hepatocytes was confirmed in a cell infection system using the murine embryonic hepatocyte cell line TIB73. The inlAB mutant was approximately 20-fold less invasive in trans. The "invasion locus" inlAB contributes to protect L. monocytogenes from the host's innate defense mechanisms by promoting its entry into hepatocytes.

scholarly journals CodY-Mediated c-di-GMP-Dependent Inhibition of Mammalian Cell Invasion in Listeria monocytogenes

2017 ◽  
Vol 200 (5) ◽  
Author(s):  
Ahmed M. Elbakush ◽  
Kurt W. Miller ◽  
Mark Gomelsky
Keyword(s):  
Transcription Factor ◽  
Listeria Monocytogenes ◽  
Mammalian Cell ◽  
Cell Invasion ◽  
Mammalian Cells ◽  
Pathogenic Bacteria ◽  
Second Messenger ◽  
Content Type ◽  
Branched Chain

ABSTRACT Elevated levels of the second messenger c-di-GMP suppress virulence in diverse pathogenic bacteria, yet mechanisms are poorly characterized. In the foodborne pathogen Listeria monocytogenes , high c-di-GMP levels inhibit mammalian cell invasion. Here, we show that invasion is impaired because of the decreased expression levels of internalin genes whose products are involved in invasion. We further show that at high c-di-GMP levels, the expression of the entire virulence regulon is suppressed, and so is the expression of the prfA gene encoding the master activator of the virulence regulon. Analysis of mechanisms controlling prfA expression pointed to the transcription factor CodY as a c-di-GMP-sensitive component. In high-c-di-GMP strains, codY gene expression is decreased, apparently due to the lower activity of CodY, which functions as an activator of codY transcription. We found that listerial CodY does not bind c-di-GMP in vitro and therefore investigated whether c-di-GMP levels affect two known cofactors of listerial CodY, branched-chain amino acids and GTP. Our manipulation of branched-chain amino acid levels did not perturb the c-di-GMP effect; however, our replacement of listerial CodY with the streptococcal CodY homolog, whose activity is GTP independent, abolished the c-di-GMP effect. The results of this study suggest that elevated c-di-GMP levels decrease the activity of the coordinator of metabolism and virulence, CodY, possibly via lower GTP levels, and that decreased CodY activity suppresses L. monocytogenes virulence by the decreased expression of the PrfA virulence regulon. IMPORTANCE Listeria monocytogenes is a pathogen causing listeriosis, a disease responsible for the highest mortality rate among foodborne diseases. Understanding how the virulence of this pathogen is regulated is important for developing treatments to decrease the frequency of listerial infections in susceptible populations. In this study, we describe the mechanism through which elevated levels of the second messenger c-di-GMP inhibit listerial invasion in mammalian cells. Inhibition is caused by the decreased activity of the transcription factor CodY that coordinates metabolism and virulence.

Injection Molding of ProNectin®F Dispersed in Polystyrene for The Fabrication of Plastic Ware Activated Towards Attachment of Mammalian Cells

1993 ◽  
Vol 330 ◽  
Author(s):  
Erwin R. Stedronsky ◽  
Joseph Cappello ◽  
Samuel David ◽  
David M. Donofrio ◽  
Tina Mcarthur ◽  
...  
Keyword(s):  
Injection Molding ◽  
Mammalian Cell ◽  
Mammalian Cells ◽  
De Novo ◽  
Cell Attachment ◽  
Biologically Active ◽  
Polymeric Surfactant ◽  
New Paradigm

ABSTRACTProNectin®F is a recombinant engineered protein polymer of de novo design which incorporates the RGD epitope recognized by mammalian cell integrins. It is biologically active as a cell attachment protein, manifests properties of a planar polymeric surfactant, and is extremely resistant to thermal degradation. ProNectin®F was dispersed onto polystyrene powder, fabricated into plastic ware through injection molding, and the plastic ware was shown to have cell attachment activity. This technology represents a new paradigm for the production of plastic ware useful for mammalian cell culture under serum free conditions; and more generally, for the production of molded devices for use in contact with cells in vitro or in vivo.

scholarly journals Improving promiscuous mammalian cell entry by the baculovirus Autographa californica multiple nuclear polyhedrosis virus

2012 ◽  
Vol 33 (1) ◽  
Author(s):  
Neil M. J. O’Flynn ◽  
Avnish Patel ◽  
Jan Kadlec ◽  
Ian M. Jones
Keyword(s):  
Mammalian Cell ◽  
Mammalian Cells ◽  
Nuclear Polyhedrosis Virus ◽  
Low Ph ◽  
Autographa Californica ◽  
Cell Entry ◽  
Surface Glycoprotein ◽  
Mammalian Cell Lines ◽  
Polyhedrosis Virus ◽  
Nuclear Polyhedrosis

The insect baculovirus AcMNPV (Autographa californica multiple nuclear polyhedrosis virus) enters many mammalian cell lines, prompting its application as a general eukaryotic gene delivery agent, but the basis of entry is poorly understood. For adherent mammalian cells, we show that entry is favoured by low pH and by increasing the available cell-surface area through a transient release from the substratum. Low pH also stimulated baculovirus entry into mammalian cells grown in suspension which, optimally, could reach 90% of the transduced population. The basic loop, residues 268–281, of the viral surface glycoprotein gp64 was required for entry and a tetra mutant with increasing basicity increased entry into a range of mammalian cells. The same mutant failed to plaque in Sf9 cells, instead showing individual cell entry and minimal cell-to-cell spread, consistent with an altered fusion phenotype. Viruses grown in different insect cells showed different mammalian cell entry efficiencies, suggesting that additional factors also govern entry.

scholarly journals Differential Effects of Paromomycin on Ribosomes ofLeishmania mexicanaand Mammalian Cells

2010 ◽  
Vol 55 (1) ◽  
pp. 86-93 ◽  
Author(s):  
Marisa M. Fernández ◽  
Emilio L. Malchiodi ◽  
Israel D. Algranati
Keyword(s):  
Protein Synthesis ◽  
Mammalian Cell ◽  
Mammalian Cells ◽  
Aminoglycoside Antibiotic ◽  
Surface Plasmon Resonance Analysis ◽  
Differential Effects ◽  
Resonance Analysis ◽  
Neomycin B

ABSTRACTParomomycin, an aminoglycoside antibiotic having low mammalian cell toxicity, is one of the drugs currently used in the chemotherapy of cutaneous and visceral leishmaniasis. In order to understand the mode of action of this antibiotic at the molecular level, we have investigated the effects of paromomycin on protein synthesis inLeishmaniaand its mammalian hosts. We were able to demonstrate thatin vivoprotein synthesis in the promastigote stage of the parasite and its proliferation rate are markedly inhibited by paromomycin while being only slightly affected by other aminoglycoside antibiotics, such as streptomycin and neomycin B. Furthermore, bothin vitropolypeptide synthesis induced by poly(U) as mRNA and accuracy of translation are significantly decreased by paromomycin in cell-free systems containing ribosomal particles ofLeishmaniapromastigotes. Conversely, when ribosomes from mammalian cells are used instead of the protozoan particles, polyphenylalanine synthesis is only barely reduced by the antibiotic and the translation misreading remains almost unaltered. Surface plasmon resonance analysis of the interaction between paromomycin and protozoan or mammalian cell ribosomal RNAs shows a strong binding of antibiotic to the parasite ribosomal decoding site and practically no interaction with the mammalian cell counterpart. Our results indicating differential effects of paromomycin on the translation processes of theLeishmaniaparasite and its mammalian hosts can explain the therapeutic efficiency of this antibiotic as an antileishmaniasis agent.

Targeting intracellular bacteria with an extended cationic amphiphilic polyproline helix

2015 ◽  
Vol 13 (21) ◽  
pp. 5930-5936 ◽  
Author(s):  
Manish Nepal ◽  
Shankar Thangamani ◽  
Mohamed N. Seleem ◽  
Jean Chmielewski
Keyword(s):  
Antibacterial Activity ◽  
Mammalian Cell ◽  
Mammalian Cells ◽  
Pathogenic Bacteria ◽  
Intracellular Bacteria ◽  
Cell Penetration

Eradicating pathogenic bacteria that reside within mammalian cells is currently quite difficult. Herein we describe an agent with the dual properties of efficient mammalian cell penetration and potent antibacterial activity. Significantly, these activities can be combined to target pathogenic bacteria within macrophages.

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