scholarly journals Measurement of Protein Mobility in Listeria monocytogenes Reveals a Unique Tolerance to Osmotic Stress and Temperature Dependence of Diffusion

2021 ◽  
Vol 12 ◽  
Author(s):  
Buu Minh Tran ◽  
Haritha Prabha ◽  
Aditya Iyer ◽  
Conor O’Byrne ◽  
Tjakko Abee ◽  
...  
Keyword(s):  
Listeria Monocytogenes ◽  
Osmotic Stress ◽  
Turgor Pressure ◽  
Lateral Diffusion ◽  
Protein Diffusion ◽  
Media Composition ◽  
Cellular Functions ◽  
Temperature Optimum ◽  
Protein Mobility ◽  
Gram Positive Bacteria

Protein mobility in the cytoplasm is essential for cellular functions, and slow diffusion may limit the rates of biochemical reactions in the living cell. Here, we determined the apparent lateral diffusion coefficient (DL) of GFP in Listeria monocytogenes as a function of osmotic stress, temperature, and media composition. We find that DL is much less affected by hyperosmotic stress in L. monocytogenes than under similar conditions in Lactococcus lactis and Escherichia coli. We find a temperature optimum for protein diffusion in L. monocytogenes at 30°C, which deviates from predicted trends from the generalized Stokes-Einstein equation under dilute conditions and suggests that the structure of the cytoplasm and macromolecular crowding vary as a function of temperature. The turgor pressure of L. monocytogenes is comparable to other Gram-positive bacteria like Bacillus subtilis and L. lactis but higher in a knockout strain lacking the stress-inducible sigma factor SigB. We discuss these findings in the context of how L. monocytogenes survives during environmental transmission and interaction with the human host.

scholarly journals PSIX-18 Laurate and its glycerol monoester, monolaurin, as potential additives to control Listeria monocytogenes and tetracycline resistant Enterococcus faecalis in air-exposed silage

2020 ◽  
Vol 98 (Supplement_4) ◽  
pp. 414-415
Author(s):  
Yamicela Castillo-Castillo ◽  
Marina Ontiveros ◽  
Eric J Scholljegerdes ◽  
Robin Anderson ◽  
Claudio Arzola-Alvarez ◽  
...  
Keyword(s):  
Listeria Monocytogenes ◽  
Enterococcus Faecalis ◽  
Brain Heart Infusion ◽  
Bactericidal Effect ◽  
Complete Elimination ◽  
Gram Positive Bacteria ◽  
Viable Cells ◽  
Colony Forming Units ◽  
Feed Value ◽  
Risk Infection

Abstract Silages can harbor pathogenic and antimicrobial resistant microbes which risk infection of food-producing animals. Livestock producers need effective yet environmentally friendly interventions to preserve the feed value of these fermented materials. Medium chain fatty acids such as laurate and its glycerol monoester, monolaurin, are potent inhibitors of many Gram-positive bacteria and when tested at 5 mg/mL in anaerobic cultures (n = 3/treatment) inoculated with 105 colony forming units (CFU) of Listeria monocytogenes and grown at 37oC in ½ strength Brain Heart infusion broth achieved near complete elimination of viable cells after 6 h compared to a 2.2 ± 0.1 log10 CFU/mL increase observed in controls. Culture of a tetracycline-resistant Enterococcus faecalis with 5 mg laurate/mL likewise achieved near complete elimination of viable cells (5 log10 CFU/mL) by 6 h incubation. The bactericidal effect of 5 mg monolaurin was less against E. faecalis, achieving a decrease of 1.8 ± 0.2 log10 CFU/mL and not decreased further after 24 h. When tested against air-exposed silage, pH 7.53 (4 g), mixed with 4 mL water, 5 mg laurate or monolaurin decreased viability of experimentally-inoculated L. monocytogenes (105 CFU/g silage) more (P < 0.05) than untreated controls after 24 h aerobic incubation (22oC), with viable counts being decreased 6.3 ± 0.1, 5.9 ± 0.8 and 4.5 ± 0.1 log10 CFU/g, respectively. In contrast, viable recovery of the experimentally-inoculated (105 CFU/g) tetracycline-resistant E. faecalis was reduced more (P < 0.05) than controls (decreased 0.7 ± 0.1 log10 CFU/g) after 6 h incubation when similarly tested with laurate and monolaurin (1.7 ± 0.5 and 3.0 ± 0.9 log10 CFU/g, respectively) but counts after 24 h were similar, decreasing on average 2.0 ± 0.5 log10 CFU/g). Results indicate laurate and monolaurin may be useful in killing L. monocytogenes and tetracycline-resistant E. faecalis during silage feed-out.

scholarly journals Elucidation of Gram-Positive Bacterial Iron(III) Reduction for Kaolinite Clay Refinement

Molecules ◽  
2021 ◽  
Vol 26 (11) ◽  
pp. 3084
Author(s):  
Hao Jing ◽  
Zhao Liu ◽  
Seng How Kuan ◽  
Sylvia Chieng ◽  
Chun Loong Ho
Keyword(s):  
Iron Reduction ◽  
Growth Conditions ◽  
Cellular Interaction ◽  
Kaolin Clay ◽  
Media Composition ◽  
Gram Positive ◽  
Kaolinite Clay ◽  
Gram Positive Bacteria ◽  
Rich Media ◽  
The Media

Recently, microbial-based iron reduction has been considered as a viable alternative to typical chemical-based treatments. The iron reduction is an important process in kaolin refining, where iron-bearing impurities in kaolin clay affects the whiteness, refractory properties, and its commercial value. In recent years, Gram-negative bacteria has been in the center stage of iron reduction research, whereas little is known about the potential use of Gram-positive bacteria to refine kaolin clay. In this study, we investigated the ferric reducing capabilities of five microbes by manipulating the microbial growth conditions. Out of the five, we discovered that Bacillus cereus and Staphylococcus aureus outperformed the other microbes under nitrogen-rich media. Through the biochemical changes and the microbial behavior, we mapped the hypothetical pathway leading to the iron reduction cellular properties, and found that the iron reduction properties of these Gram-positive bacteria rely heavily on the media composition. The media composition results in increased basification of the media that is a prerequisite for the cellular reduction of ferric ions. Further, these changes impact the formation of biofilm, suggesting that the cellular interaction for the iron(III)oxide reduction is not solely reliant on the formation of biofilms. This article reveals the potential development of Gram-positive microbes in facilitating the microbial-based removal of metal contaminants from clays or ores. Further studies to elucidate the corresponding pathways would be crucial for the further development of the field.

scholarly journals Antimicrobial Nodule-Specific Cysteine-Rich Peptides Induce Membrane Depolarization-Associated Changes in the Transcriptome of Sinorhizobium meliloti

2013 ◽  
Vol 79 (21) ◽  
pp. 6737-6746 ◽  
Author(s):  
Hilda Tiricz ◽  
Attila Szűcs ◽  
Attila Farkas ◽  
Bernadett Pap ◽  
Rui M. Lima ◽  
...  
Keyword(s):  
Stress Responses ◽  
Sinorhizobium Meliloti ◽  
Antimicrobial Agents ◽  
Cellular Functions ◽  
Gram Positive ◽  
Content Type ◽  
Gram Positive Bacteria ◽  
Peptide Family ◽  
Wide Range ◽  
Natural Target

ABSTRACTLeguminous plants establish symbiosis with nitrogen-fixing alpha- and betaproteobacteria, collectively called rhizobia, which provide combined nitrogen to support plant growth. Members of the inverted repeat-lacking clade of legumes impose terminal differentiation on their endosymbiotic bacterium partners with the help of the nodule-specific cysteine-rich (NCR) peptide family composed of close to 600 members. Among the few tested NCR peptides, cationic ones had antirhizobial activity measured by reduction or elimination of the CFU and uptake of the membrane-impermeable dye propidium iodide. Here, the antimicrobial spectrum of two of these peptides, NCR247 and NCR335, was investigated, and their effect on the transcriptome of the natural targetSinorhizobium melilotiwas characterized. Both peptides were able to kill quickly a wide range of Gram-negative and Gram-positive bacteria; however, their spectra were only partially overlapping, and differences were found also in their efficacy on given strains, indicating that the actions of NCR247 and NCR335 might be similar though not identical. Treatment ofS. meliloticultures with either peptide resulted in a quick downregulation of genes involved in basic cellular functions, such as transcription-translation and energy production, as well as upregulation of genes involved in stress and oxidative stress responses and membrane transport. Similar changes provoked mainly in Gram-positive bacteria by antimicrobial agents were coupled with the destruction of membrane potential, indicating that it might also be a common step in the bactericidal actions of NCR247 and NCR335.

scholarly journals Visualizing the dynamics of exported bacterial proteins with the chemogenetic fluorescent reporter FAST

2020 ◽  
Author(s):  
Yankel Chekli ◽  
Caroline Peron-Cane ◽  
Dario Dell’Arciprete ◽  
Jean-François Allemand ◽  
Chenge Li ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Listeria Monocytogenes ◽  
Cell Surface ◽  
Bacterial Cell ◽  
Surface Proteins ◽  
Cellular Functions ◽  
Reporter Protein ◽  
Cell Surface Proteins ◽  
Fluorescent Reporter ◽  
Bacterial Proteins

AbstractBacterial proteins exported to the cell surface play key cellular functions. However, despite the interest to study the localization of surface proteins such as adhesins, transporters or hydrolases, monitoring their dynamics in live imaging remains challenging, due to the limited availability of fluorescent probes remaining functional after secretion. In this work, we used the Escherichia coli intimin and the Listeria monocytogenes InlB invasin as surface exposed scaffolds fused with the recently developed chemogenetic fluorescent reporter protein FAST. Using both membrane permeant (HBR-3,5DM) and non-permeant (HBRAA-3E) fluorogens that fluoresce upon binding to FAST, we demonstrated that fully functional FAST can be exposed at the cell surface and specifically tagged on the external side of the bacterial envelop in both diderm and monoderm bacteria. Our work opens new avenues to study of the organization and dynamics of the bacterial cell surface proteins.

scholarly journals Protein Diffusion in the Periplasm of E. coli under Osmotic Stress

2011 ◽  
Vol 100 (1) ◽  
pp. 22-31 ◽  
Author(s):  
Kem A. Sochacki ◽  
Irina A. Shkel ◽  
M. Thomas Record ◽  
James C. Weisshaar
Keyword(s):  
Osmotic Stress ◽  
Protein Diffusion ◽  
E Coli

scholarly journals Methyltransferase DnmA is responsible for genome-wide N6-methyladenosine modifications at non-palindromic recognition sites in Bacillus subtilis

2020 ◽  
Vol 48 (10) ◽  
pp. 5332-5348
Author(s):  
Taylor M Nye ◽  
Lieke A van Gijtenbeek ◽  
Amanda G Stevens ◽  
Jeremy W Schroeder ◽  
Justin R Randall ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Bacillus Subtilis ◽  
Consensus Sequence ◽  
Dna Uptake ◽  
Cellular Functions ◽  
Promoter Regions ◽  
Gram Positive Bacteria ◽  
Genome Wide ◽  
Domains Of Life ◽  
Reporter Assays

Abstract The genomes of organisms from all three domains of life harbor endogenous base modifications in the form of DNA methylation. In bacterial genomes, methylation occurs on adenosine and cytidine residues to include N6-methyladenine (m6A), 5-methylcytosine (m5C), and N4-methylcytosine (m4C). Bacterial DNA methylation has been well characterized in the context of restriction-modification (RM) systems, where methylation regulates DNA incision by the cognate restriction endonuclease. Relative to RM systems less is known about how m6A contributes to the epigenetic regulation of cellular functions in Gram-positive bacteria. Here, we characterize site-specific m6A modifications in the non-palindromic sequence GACGmAG within the genomes of Bacillus subtilis strains. We demonstrate that the yeeA gene is a methyltransferase responsible for the presence of m6A modifications. We show that methylation from YeeA does not function to limit DNA uptake during natural transformation. Instead, we identify a subset of promoters that contain the methylation consensus sequence and show that loss of methylation within promoter regions causes a decrease in reporter expression. Further, we identify a transcriptional repressor that preferentially binds an unmethylated promoter used in the reporter assays. With these results we suggest that m6A modifications in B. subtilis function to promote gene expression.

scholarly journals Mutations of the Listeria monocytogenes PeptidoglycanN-Deacetylase andO-Acetylase Result in Enhanced Lysozyme Sensitivity, Bacteriolysis, and Hyperinduction of Innate Immune Pathways

2011 ◽  
Vol 79 (9) ◽  
pp. 3596-3606 ◽  
Author(s):  
Chris S. Rae ◽  
Aimee Geissler ◽  
Paul C. Adamson ◽  
Daniel A. Portnoy
Keyword(s):  
Listeria Monocytogenes ◽  
Transcriptional Responses ◽  
Gram Positive ◽  
Content Type ◽  
Growth Defects ◽  
Gram Positive Bacteria ◽  
Host Cell Death

ABSTRACTListeria monocytogenesis a Gram-positive intracellular pathogen that is naturally resistant to lysozyme. Recently, it was shown that peptidoglycan modification by N-deacetylation or O-acetylation confers resistance to lysozyme in various Gram-positive bacteria, includingL. monocytogenes.L. monocytogenespeptidoglycan is deacetylated by the action ofN-acetylglucosamine deacetylase (Pgd) and acetylated byO-acetylmuramic acid transferase (Oat). We characterized Pgd−, Oat−, and double mutants to determine the specific role ofL. monocytogenespeptidoglycan acetylation in conferring lysozyme sensitivity during infection of macrophages and mice. Pgd−and Pgd−Oat−double mutants were attenuated approximately 2 and 3.5 logs, respectively,in vivo. In bone-marrow derived macrophages, the mutants demonstrated intracellular growth defects and increased induction of cytokine transcriptional responses that emanated from a phagosome and the cytosol. Lysozyme-sensitive mutants underwent bacteriolysis in the macrophage cytosol, resulting in AIM2-dependent pyroptosis. Each of thein vitrophenotypes was rescued upon infection of LysM−macrophages. The addition of extracellular lysozyme to LysM−macrophages restored cytokine induction, host cell death, andL. monocytogenesgrowth inhibition. This surprising observation suggests that extracellular lysozyme can access the macrophage cytosol and act on intracellular lysozyme-sensitive bacteria.

scholarly journals Biofilm Formation among Clinical and Food Isolates ofListeria monocytogenes

2013 ◽  
Vol 2013 ◽  
pp. 1-6 ◽  
Author(s):  
Joana Barbosa ◽  
Sandra Borges ◽  
Ruth Camilo ◽  
Rui Magalhães ◽  
Vânia Ferreira ◽  
...  
Keyword(s):  
Tissue Culture ◽  
Listeria Monocytogenes ◽  
Osmotic Stress ◽  
Environmental Conditions ◽  
Biofilm Formation ◽  
Stress Conditions ◽  
Acidic Stress ◽  
Biofilm Production ◽  
Oflisteria Monocytogenes ◽  
Clinical Cases

Objective. A total of 725Listeria monocytogenesisolates, 607 from various foods and 118 from clinical cases of listeriosis, were investigated concerning their ability to form biofilms, at 4°C during 5 days and at 37°C during 24 h.Methods. Biofilm production was carried out on polystyrene tissue culture plates. FiveL. monocytogenesisolates were tested for biofilm formation after being exposed to acidic and osmotic stress conditions.Results. Significant differences (P<0.01) between clinical and food isolates were observed. At 37°C for 24 h, most food isolates were classified as weak or moderate biofilm formers whereas all the clinical isolates were biofilm producers, although the majority were weak. At 4°C during 5 days, 65 and 59% isolates, from food and clinical cases, respectively, were classified as weak. After both sublethal stresses, at 37°C just one of the five isolates tested was shown to be more sensitive to subsequent acidic exposure. However, at 4°C both stresses did not confer either sensitivity or resistance.Conclusions. Significant differences between isolates origin, temperature, and sublethal acidic stress were observed concerning the ability to form biofilms. Strain, origin, and environmental conditions can determine the level of biofilm production byL. monocytogenesisolates.

scholarly journals Lateral diffusion in nuclear membranes.

1985 ◽  
Vol 100 (5) ◽  
pp. 1408-1414 ◽  
Author(s):  
M Schindler ◽  
J F Holland ◽  
M Hogan
Keyword(s):  
Diffusion Coefficient ◽  
Outer Membrane ◽  
Wheat Germ Agglutinin ◽  
Wheat Germ ◽  
Lipid Analysis ◽  
Membrane Integrity ◽  
Lateral Diffusion ◽  
Protein Diffusion ◽  
Inner Membrane ◽  
Lectin Receptors

Chemical modification of rat liver nuclei with citraconic anhydride selectively removed outer nuclear membrane. This conclusion was based on (a) transmission electron microscopy, (b) lipid analysis, (c) lamin B as an inner membrane-associated marker, and (d) the demonstration of phospholipid lateral mobility on outer membrane-depleted nuclei as a criteria for inner membrane integrity. Addition of urea or N-ethylmaleimide resulted in the additional disruption of inner membrane. Fluorescence photobleaching was used to determine the long range (greater than 4 microns) lateral transport of lectin receptors and a phospholipid analog in both membranes. The diffusion coefficient for wheat germ agglutinin on whole nuclei was 3.9 X 10(-10) cm2/s whereas the diffusion coefficient for wheat germ agglutinin in outer membrane-depleted nuclei was less than or equal to 10(-12) cm2/s. Phospholipid mobilities were the same in whole and outer membrane-depleted nuclei (3.8 X 10(-9) cm2/s). The protein diffusion differences observed between whole and outer membrane-depleted nuclei may be interpreted in the context of two functionally different membrane systems that compose the double bilayer of the nucleus.

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