scholarly journals Detoxification of methylglyoxal by the glyoxalase system is required for glutathione availability and virulence activation in Listeria monocytogenes

2021 ◽  
Vol 17 (8) ◽  
pp. e1009819
Author(s):  
Andrea Anaya-Sanchez ◽  
Ying Feng ◽  
John C. Berude ◽  
Daniel A. Portnoy
Keyword(s):  
Listeria Monocytogenes ◽  
Transcriptional Activation ◽  
Protein Glycation ◽  
Host Cells ◽  
Intracellular Pathogen ◽  
Cell Monolayers ◽  
Food Borne ◽  
Virulence Regulator ◽  
Allosteric Activator

Listeria monocytogenes is a Gram-positive, food-borne pathogen that lives a biphasic lifestyle, cycling between the environment and as a facultative intracellular pathogen of mammals. Upon entry into host cells, L. monocytogenes upregulates expression of glutathione synthase (GshF) and its product, glutathione (GSH), which is an allosteric activator of the master virulence regulator PrfA. Although gshF mutants are highly attenuated for virulence in mice and form very small plaques in host cell monolayers, these virulence defects can be fully rescued by mutations that lock PrfA in its active conformation, referred to as PrfA*. While PrfA activation can be recapitulated in vitro by the addition of reducing agents, the precise biological cue(s) experienced by L. monocytogenes that lead to PrfA activation are not known. Here we performed a genetic screen to identify additional small-plaque mutants that were rescued by PrfA* and identified gloA, which encodes glyoxalase A, a component of a GSH-dependent methylglyoxal (MG) detoxification system. MG is a toxic byproduct of metabolism produced by both the host and pathogen, which if accumulated, causes DNA damage and protein glycation. As a facultative intracellular pathogen, L. monocytogenes must protect itself from MG produced by its own metabolic processes and that of its host. We report that gloA mutants grow normally in broth, are sensitive to exogenous MG and severely attenuated upon IV infection in mice, but are fully rescued for virulence in a PrfA* background. We demonstrate that transcriptional activation of gshF increased upon MG challenge in vitro, and while this resulted in higher levels of GSH for wild-type L. monocytogenes, the glyoxalase mutants had decreased levels of GSH, presumably due to the accumulation of the GSH-MG hemithioacetal adduct. These data suggest that MG acts as a host cue that leads to GSH production and activation of PrfA.

scholarly journals The Virulence Regulator PrfA Promotes Biofilm Formation by Listeria monocytogenes

2010 ◽  
Vol 192 (15) ◽  
pp. 3969-3976 ◽  
Author(s):  
Katherine P. Lemon ◽  
Nancy E. Freitag ◽  
Roberto Kolter
Keyword(s):  
Listeria Monocytogenes ◽  
Biofilm Formation ◽  
Virulence Genes ◽  
Constitutive Expression ◽  
Host Cells ◽  
Intracellular Pathogen ◽  
Initial Surface ◽  
Protein Variant ◽  
Biofilm Development ◽  
Virulence Regulator

ABSTRACT Listeria monocytogenes is a food-borne facultative intracellular pathogen. It is widespread in the environment and has several distinct life-styles. The key transcriptional activator PrfA positively regulates L. monocytogenes virulence genes to mediate the transition from extracellular, flagellum-propelled cell to intracellular pathogen. Here we report the first evidence that PrfA also has a significant positive impact on extracellular biofilm formation. Mutants lacking prfA were defective in surface-adhered biofilm formation. The ΔprfA mutant exhibited wild-type flagellar motility, and its biofilm defect occurred after initial surface adhesion. We also observed that mutations that led to the constitutive expression of PrfA-dependent virulence genes had a minimal impact on biofilm formation. Furthermore, biofilm development was enhanced in a mutant encoding a PrfA protein variant unable to fully transition from the extracellular form to the virulent, intracellular activity conformation. These results indicate that PrfA positively regulates biofilm formation and suggest that PrfA has a global role in modulating the life-style of L. monocytogenes. The requirement of PrfA for optimal biofilm formation may provide selective pressure to maintain this critical virulence regulator when L. monocytogenes is outside host cells in the environment.

scholarly journals Effect of Gaseous Ozone on Listeria monocytogenes Planktonic Cells and Biofilm: An In Vitro Study

Foods ◽  
2021 ◽  
Vol 10 (7) ◽  
pp. 1484
Author(s):  
Felice Panebianco ◽  
Selene Rubiola ◽  
Francesco Chiesa ◽  
Tiziana Civera ◽  
Pierluigi Aldo Di Ciccio
Keyword(s):  
Listeria Monocytogenes ◽  
In Vitro Study ◽  
Planktonic Cells ◽  
Free Living ◽  
Biofilm Inhibition ◽  
Additional Tool ◽  
Complete Inactivation ◽  
Gaseous Ozone ◽  
Food Borne

Among food-borne pathogens, Listeria monocytogenes continues to pose concerns to food business operators due to its capacity to form biofilm in processing environments. Ozone may be an eco-friendly technology to control microbial contaminations, but data concerning its effect on Listeria monocytogenes biofilm are still limited. In this study, the effect of gaseous ozone at 50 ppm on planktonic cells and biofilm of reference and food-related Listeria monocytogenes strains was evaluated. Ozone caused a reduction in microbial loads of 3.7 ± 0.4 and 3.9 ± 0.4 Log10 CFU/mL after 10 and 30 min, respectively. A complete inactivation of planktonic cells after 6 h of treatment was observed. Biofilm inhibition and eradication treatments (50 ppm, 6 h) resulted in a significant decrease of the biofilm biomass for 59% of the strains tested, whilst a slight dampening of live cell loads in the biofilm state was observed. In conclusion, gaseous ozone is not sufficient to completely counteract Listeria monocytogenes biofilm, but it may be useful as an additional tool to contrast Listeria monocytogenes free-living cells and to improve the existing sanitization procedures in food processing environments.

scholarly journals Use of multi-locus sequencing typing as identification method for the food-borne pathogen Listeria monocytogenes: a review

2015 ◽  
Vol 5 (1) ◽  
Author(s):  
Sonia Lamon ◽  
Domenico Meloni ◽  
Simonetta Gianna Consolati ◽  
Anna Mureddu ◽  
Rina Mazzette
Keyword(s):  
Listeria Monocytogenes ◽  
Detection Methods ◽  
Causative Organism ◽  
Intracellular Pathogen ◽  
Bacterial Strains ◽  
New Approach ◽  
Food Borne ◽  
Global Epidemiology ◽  
Selection Of

<em>Listeria monocytogenes</em> is an ubiquitous, intracellular pathogen which has been implicated within the past decade as the causative organism in several outbreaks of foodborne diseases. In this review, a new approach to molecular typing primarily designed for global epidemiology has been described: multi-<em>locus</em> sequencing typing (MLST). This approach is novel, in that it uses data that allow the unambiguous characterization of bacterial strains via the Internet. Our aim is to present the currently available selection of references on <em>L. monocytogenes</em> MLST detection methods and to discuss its use as <em>gold</em> <em>standard</em> to <em>L. monocytogenes</em> subtyping method.

Metal oxide nanoparticle-functionalized sebacic acid-grafted PHEAM nanocarriers for enriched activity of metronidazole against food borne bacteria: in vitro and in vivo study

2018 ◽  
Vol 42 (22) ◽  
pp. 18437-18447 ◽  
Author(s):  
Murugesan Gowri ◽  
Kannan Suganya ◽  
Nachimuthu Latha ◽  
Marudhamuthu Murugan ◽  
Mukhtar Ahmed ◽  
...  
Keyword(s):  
Listeria Monocytogenes ◽  
Metal Oxide ◽  
Sebacic Acid ◽  
In Vivo Study ◽  
Metal Oxide Nanoparticle ◽  
Food Borne ◽  
Oxide Nanoparticle

Food borne infection is a serious complication caused by Listeria monocytogenes (L. monocytogenes), a dangerous bacteria.

scholarly journals Rapid modulation of electrolyte transport in Caco-2 cell monolayers by enteropathogenic Escherichia coli (EPEC) infection

Gut ◽  
1998 ◽  
Vol 42 (2) ◽  
pp. 200-207 ◽  
Author(s):  
G K Collington ◽  
I W Booth ◽  
S Knutton
Keyword(s):  
Escherichia Coli ◽  
Signal Transduction ◽  
Short Circuit ◽  
Electrolyte Transport ◽  
Host Cells ◽  
Ussing Chambers ◽  
Cell Monolayers ◽  
Initial Adhesion ◽  
Rapid Modulation

Background and aims—The pathophysiology of enteropathogenic Escherichia coli (EPEC) diarrhoea remains uncertain. EPEC adhere to enterocytes and transduce signals which produce a characteristic “attaching and effacing” (A/E) lesion in the brush border membrane. The present in vitro study was designed to determine whether signal transduction by EPEC also influences electrolyte transport.Methods—Caco-2 cell monolayers were rapidly infected with wild type EPEC strain E2348/69, or the signal transduction-defective mutant 14.2.1(1), and mounted in Ussing chambers.Results—Strain E2348/69 stimulated a rapid but transient increase in short circuit current (Isc) which coincided with A/E lesion formation; this Isc response was absent on infection with strain 14.2.1(1). While the initial rise inIsc induced by E2348/69 was partially (∼35%) dependent on chloride, the remainder possibly represents an influx of sodium and amino acid(s) across the apical membrane.Conclusions—The study directly shows that, after initial adhesion, EPEC induce major alterations in host cell electrolyte transport. The observed Isc responses indicate a rapid modulation of electrolyte transport in Caco-2 cells by EPEC, including stimulation of chloride secretion, for which signal transduction to host cells is a prerequisite.

scholarly journals Expression of Truncated Internalin A Is Involved in Impaired Internalization of Some Listeria monocytogenes Isolates Carried Asymptomatically by Humans

2003 ◽  
Vol 71 (3) ◽  
pp. 1217-1224 ◽  
Author(s):  
Maïwenn Olier ◽  
Fabrice Pierre ◽  
Sandrine Rousseaux ◽  
Jean-Paul Lemaître ◽  
André Rousset ◽  
...  
Keyword(s):  
Listeria Monocytogenes ◽  
Surface Protein ◽  
Human Infection ◽  
Host Cells ◽  
Genetic Lineage ◽  
Chick Embryos ◽  
Asymptomatic Carriage ◽  
High Degree ◽  
Internalin A

ABSTRACT Fourteen human carriage Listeria monocytogenes isolates were compared to sporadic and epidemic-associated human strains in order to ascertain the pathogenic behavior of these unrecognized asymptomatic strains. Experimental infection of 14-day-old chick embryos revealed that the majority of the carriage strains were attenuated for virulence. Of the 10 attenuated carriage strains, 5 were affected in their invasion capacities in vitro. Western blot analysis with antibody directed against InlA, the surface protein implicated in the internalization in host cells, allowed correlation between the ability of the carriage strains to enter Caco-2 cells and InlA expression. Indeed, these five carriage strains produced truncated forms of InlA. Four of the five truncated forms of InlA had an apparent molecular mass of 47 kDa. In order to assess the existence of a genetic lineage, partial sequences of inlA gene of these four strains were compared and revealed that they had a high degree of sequence conservation at the gene (99.86%) and amino acid (100%) levels. Comparison of their nucleotide sequences with that of the corresponding segment of inlA from EGD-e and Scott A strains, taken as epidemic references, showed more divergence. Taken together, these observations suggest the presence of specific traits that characterize L. monocytogenes strains isolated during asymptomatic carriage. Some of these traits could provide some explanations about the determinants that make them unable to cause systemic human infection.

scholarly journals prfA-Like Transcription Factor Genelmo0753Contributes to l-Rhamnose Utilization in Listeria monocytogenes Strains Associated with Human Food-Borne Infections

2013 ◽  
Vol 79 (18) ◽  
pp. 5584-5592 ◽  
Author(s):  
Joelle K. Salazar ◽  
Zhuchun Wu ◽  
P. David McMullen ◽  
Qin Luo ◽  
Nancy E. Freitag ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Listeria Monocytogenes ◽  
Bacterial Pathogen ◽  
Phenol Red ◽  
Flagellar Motility ◽  
Content Type ◽  
Genetic Lineages ◽  
Functional Roles ◽  
Food Borne ◽  
Virulence Regulator

ABSTRACTListeria monocytogenesis a food-borne bacterial pathogen and the causative agent of human and animal listeriosis. Among the three major genetic lineages ofL. monocytogenes(i.e., LI, LII, and LIII), LI and LII are predominantly associated with food-borne listeriosis outbreaks, whereas LIII is rarely implicated in human infections. In a previous study, we identified a Crp/Fnr family transcription factor gene,lmo0753, that was highly specific to outbreak-associated LI and LII but absent from LIII. Lmo0753 shares two conserved functional domains, including a DNA binding domain, with the well-characterized master virulence regulator PrfA inL. monocytogenes. In this study, we constructedlmo0753deletion and complementation mutants in two fully sequencedL. monocytogenesLII strains, 10403S and EGDe, and compared the flagellar motility, phospholipase C production, hemolysis, and intracellular growth of the mutants and their respective wild types. Our results suggested thatlmo0753plays a role in hemolytic activity in both EGDe and 10403S. More interestingly, we found that deletion oflmo0753led to the loss ofl-rhamnose utilization in EGDe, but not in 10403S. RNA-seq analysis of EGDe Δ0753incubated in phenol red medium containingl-rhamnose as the sole carbon source revealed that 126 (4.5%) and 546 (19.5%) out of 2,798 genes in the EGDe genome were up- and downregulated more than 2-fold, respectively, compared to the wild-type strain. Genes related to biotin biosynthesis, general stress response, and rhamnose metabolism were shown to be differentially regulated. Findings from this study collectively suggested varied functional roles oflmo0753in different LIIL. monocytogenesstrain backgrounds associated with human listeriosis outbreaks.

scholarly journals Some Listeria monocytogenes Outbreak Strains Demonstrate Significantly Reduced Invasion, inlA Transcript Levels, and Swarming Motility In Vitro

2009 ◽  
Vol 75 (17) ◽  
pp. 5647-5658 ◽  
Author(s):  
A. J. Roberts ◽  
S. K. Williams ◽  
M. Wiedmann ◽  
K. K. Nightingale
Keyword(s):  
Listeria Monocytogenes ◽  
Molecular Subtype ◽  
Phylogenetic Analyses ◽  
Data Sets ◽  
Intestinal Epithelial ◽  
Swarming Motility ◽  
Transcript Levels ◽  
Hepatic Cells ◽  
Food Borne

ABSTRACT Listeria monocytogenes can cause a severe invasive food-borne disease known as listeriosis, and large outbreaks of this disease occur occasionally. Based on molecular-subtype data, epidemic clone (EC) strains have been defined, including ECI and ECIa, which have caused listeriosis outbreaks on different continents. While a number of molecular-subtyping studies of outbreak strains have been reported, few comprehensive data sets of virulence-associated characteristics of these strains are available. We assembled a set of human clinical isolates from 15 outbreaks that occurred worldwide between 1975 and 2002. Initial characterization of these strains showed significant variation in the ability to invade human Caco-2 intestinal epithelial cells and HepG2 hepatic cells; four strains showed consistently reduced invasion in both cell lines. DNA sequencing of inlA, which encodes a protein required for efficient Caco-2 and HepG2 invasion, showed that none of the invasion-attenuated strains contained known virulence-attenuating mutations in inlA. Phylogenetic analyses of inlA sequences revealed a well-supported clade containing a fully invasive ECI strain and three invasion-attenuated ECI strains, along with a fully invasive ECIa strain and an invasion-attenuated ECIa strain. Of the four invasion-attenuated strains, one strain showed both reduced inlA transcript levels and impaired swarming, one strain showed reduced inlA transcript levels, and two strains showed reduced swarming. Overall, our data show that (i) L. monocytogenes strains from outbreaks vary significantly in invasion efficiency and (ii) different mechanisms may contribute to reduced invasion efficiency. Association between EC strains and listeriosis outbreaks may involve characteristics other than virulence phenotypes, including survival and growth in food-associated environments.

scholarly journals Identification of the agr Locus of Listeria monocytogenes: Role in Bacterial Virulence

2003 ◽  
Vol 71 (8) ◽  
pp. 4463-4471 ◽  
Author(s):  
Nicolas Autret ◽  
Catherine Raynaud ◽  
Iharilalao Dubail ◽  
Patrick Berche ◽  
Alain Charbit
Keyword(s):  
Listeria Monocytogenes ◽  
Response Regulator ◽  
Lethal Dose ◽  
Fold Increase ◽  
Bacterial Virulence ◽  
Intravenous Route ◽  
Two Component System ◽  
Food Borne ◽  
Severe Infections

ABSTRACT Listeria monocytogenes is a gram-positive facultative intracellular food-borne pathogen that can cause severe infections in humans and animals. We have recently adapted signature-tagged transposon mutagenesis (STM) to identify genes involved in the virulence of L. monocytogenes. A new round of STM allowed us to identify a new locus encoding a protein homologous to AgrA, the well-studied response regulator of Staphylococcus aureus and part of a two-component system involved in bacterial virulence. The production of several secreted proteins was modified in the agrA mutant of L. monocytogenes grown in broth, indicating that the agr locus influenced protein secretion. Inactivation of agrA did not affect the ability of the pathogen to invade and multiply in cells in vitro. However, the virulence of the agrA mutant was attenuated in the mouse (a 10-fold increase in the 50% lethal dose by the intravenous route), demonstrating for the first time a role for the agr locus in the virulence of L. monocytogenes.

scholarly journals Active Escape of Orientia tsutsugamushi from Cellular Autophagy

2012 ◽  
Vol 81 (2) ◽  
pp. 552-559 ◽  
Author(s):  
Youngho Ko ◽  
Ji-Hye Choi ◽  
Na-Young Ha ◽  
Ik-Sang Kim ◽  
Nam-Hyuk Cho ◽  
...  
Keyword(s):  
Defense Mechanism ◽  
Host Cells ◽  
Intracellular Pathogen ◽  
Orientia Tsutsugamushi ◽  
Content Type ◽  
Cellular Autophagy ◽  
A Cell ◽  
Infected Cells ◽  
Obligate Intracellular Pathogen

ABSTRACTOrientia tsutsugamushi, the causative agent of scrub typhus, is an obligate intracellular pathogen. After entry into host cells, the bacterium rapidly escapes from the endosomal pathway and replicates in the cytosol of eukaryotic host cells. Here we show thatO. tsutsugamushiinfection efficiently promotes cellular autophagy, a cell-autonomous defense mechanism of innate immunity. However, most of the internalized bacteria barely colocalized with the induced autophagosomes, even when stimulated with rapamycin, a chemical inducer of autophagy. Treatment of infected cells with tetracycline suppressed bacterial evasion from autophagy and facilitatedO. tsutsugamushitargeting to autophagosomes, suggesting that the intracellular pathogen may be equipped with a bacterial factor or factors that block autophagic recognition. Finally, we also found that chemical modulators of cellular autophagy or genetic knockout of theatg3gene does not significantly affect the intracellular growth ofO. tsutsugamushiin vitro. These results suggest thatO. tsutsugamushihas evolved to block autophagic microbicidal defense by evading autophagic recognition even though it activates the autophagy pathway during the early phase of infection.

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