scholarly journals The secRNome of Listeria monocytogenes Harbors Small Noncoding RNAs That Are Potent Inducers of Beta Interferon

mBio ◽  
2019 ◽  
Vol 10 (5) ◽  
Author(s):  
Renate Frantz ◽  
Lisa Teubner ◽  
Tilman Schultze ◽  
Luigi La Pietra ◽  
Christin Müller ◽  
...  
Keyword(s):  
Listeria Monocytogenes ◽  
Bacterial Growth ◽  
Cell Envelope ◽  
Intracellular Pathogen ◽  
Type I ◽  
Type I Ifn ◽  
Intracellular Growth ◽  
Potent Inducer ◽  
Content Type ◽  
Bacterial Physiology

ABSTRACT Cellular sensing of bacterial RNA is increasingly recognized as a determinant of host-pathogen interactions. The intracellular pathogen Listeria monocytogenes induces high levels of type I interferons (alpha/beta interferons [IFN-α/β]) to create a growth-permissive microenvironment during infection. We previously demonstrated that RNAs secreted by L. monocytogenes (comprising the secRNome) are potent inducers of IFN-β. We determined the composition and diversity of the members of the secRNome and found that they are uniquely enriched for noncoding small RNAs (sRNAs). Testing of individual sRNAs for their ability to induce IFN revealed several sRNAs with this property. We examined ril32, an intracellularly expressed sRNA that is highly conserved for the species L. monocytogenes and that was the most potent inducer of IFN-β expression of all the sRNAs tested in this study, in more detail. The rli32-induced IFN-β response is RIG-I (retinoic acid inducible gene I) dependent, and cells primed with rli32 inhibit influenza virus replication. We determined the rli32 motif required for IFN induction. rli32 overproduction promotes intracellular bacterial growth, and a mutant lacking rli32 is restricted for intracellular growth in macrophages. rli32-overproducing bacteria are resistant to H2O2 and exhibit both increased catalase activity and changes in the cell envelope. Comparative transcriptome sequencing (RNA-Seq) analysis indicated that ril32 regulates expression of the lhrC locus, previously shown to be involved in cell envelope stress. Inhibition of IFN-β signaling by ruxolitinib reduced rli32-dependent intracellular bacterial growth, indicating a link between induction of the interferon system and bacterial physiology. rli32 is, to the best of our knowledge, the first secreted individual bacterial sRNA known to trigger the induction of the type I IFN response. IMPORTANCE Interferons are potent and broadly acting cytokines that stimulate cellular responses to nucleic acids of unusual structures or locations. While protective when induced following viral infections, the induction of interferons is detrimental to the host during L. monocytogenes infection. Here, we identify specific sRNAs, secreted by the bacterium, with the capacity to induce type I IFN. Further analysis of the most potent sRNA, rli32, links the ability to induce RIG-I-dependent induction of the type I IFN response to the intracellular growth properties of the bacterium. Our findings emphasize the significance of released RNA for Listeria infection and shed light on a compartmental strategy used by an intracellular pathogen to modulate host responses to its advantage.

scholarly journals c-di-AMP Accumulation Impairs Muropeptide Synthesis in Listeria monocytogenes

2020 ◽  
Vol 202 (24) ◽  
Author(s):  
Steven M. Massa ◽  
Amar Deep Sharma ◽  
Cheta Siletti ◽  
Zepeng Tu ◽  
Jared J. Godfrey ◽  
...  
Keyword(s):  
Cell Wall ◽  
Listeria Monocytogenes ◽  
Cell Envelope ◽  
Second Messenger ◽  
Fold Increase ◽  
Bacterial Cells ◽  
Content Type ◽  
Bacterial Physiology ◽  
Peptidoglycan Synthesis ◽  
Increased Sensitivity

ABSTRACT Cyclic di-AMP (c-di-AMP) is an essential and ubiquitous second messenger among bacteria. c-di-AMP regulates many cellular pathways through direct binding to several molecular targets in bacterial cells. c-di-AMP depletion is well known to destabilize the bacterial cell wall, resulting in increased bacteriolysis and enhanced susceptibility to cell wall targeting antibiotics. Using the human pathogen Listeria monocytogenes as a model, we found that c-di-AMP accumulation also impaired cell envelope integrity. An L. monocytogenes mutant deleted for c-di-AMP phosphodiesterases (pdeA pgpH mutant) exhibited a 4-fold increase in c-di-AMP levels and several cell wall defects. For instance, the pdeA pgpH mutant was defective for the synthesis of peptidoglycan muropeptides and was susceptible to cell wall-targeting antimicrobials. Among different muropeptide precursors, we found that the pdeA pgpH strain was particularly impaired in the synthesis of d-Ala–d-Ala, which is required to complete the pentapeptide stem associated with UDP–N-acetylmuramic acid (MurNAc). This was consistent with an increased sensitivity to d-cycloserine, which inhibits the d-alanine branch of peptidoglycan synthesis. Finally, upon examining d-Ala:d-Ala ligase (Ddl), which catalyzes the conversion of d-Ala to d-Ala–d-Ala, we found that its activity was activated by K+. Based on previous reports that c-di-AMP inhibits K+ uptake, we propose that c-di-AMP accumulation impairs peptidoglycan synthesis, partially through the deprivation of cytoplasmic K+ levels, which are required for cell wall-synthetic enzymes. IMPORTANCE The bacterial second messenger c-di-AMP is produced by a large number of bacteria and conditionally essential to many species. Conversely, c-di-AMP accumulation is also toxic to bacterial physiology and pathogenesis, but its mechanisms are largely undefined. We found that in Listeria monocytogenes, elevated c-di-AMP levels diminished muropeptide synthesis and increased susceptibility to cell wall-targeting antimicrobials. Cell wall defects might be an important mechanism for attenuated virulence in bacteria with high c-di-AMP levels.

scholarly journals Mechanism of the Intracellular Killing and Modulation of Antibiotic Susceptibility of Listeria monocytogenes in THP-1 Macrophages Activated by Gamma Interferon

1999 ◽  
Vol 43 (5) ◽  
pp. 1242-1251 ◽  
Author(s):  
Youssef Ouadrhiri ◽  
Bernard Scorneaux ◽  
Yves Sibille ◽  
Paul M. Tulkens
Keyword(s):  
Listeria Monocytogenes ◽  
Bacterial Growth ◽  
Antibiotic Activity ◽  
Gamma Interferon ◽  
Intracellular Pathogen ◽  
Host Defenses ◽  
Intracellular Killing ◽  
Cellular Accumulation ◽  
Intracellular Multiplication ◽  
Increased Activity

ABSTRACT Listeria monocytogenes, a facultative intracellular pathogen, readily enters cells and multiplies in the cytosol after escaping from phagosomal vacuoles. Macrophages exposed to gamma interferon, one of the main cellular host defenses againstListeria, become nonpermissive for bacterial growth while containing Listeria in the phagosomes. Using the human myelomonocytic cell line THP-1, we show that the combination ofl-monomethyl arginine and catalase restores bacterial growth without affecting the phagosomal containment ofListeria. A previous report (B. Scorneaux, Y. Ouadrhiri, G. Anzalone, and P. M. Tulkens, Antimicrob. Agents Chemother. 40:1225–1230, 1996) showed that intracellular Listeria was almost equally sensitive to ampicillin, azithromycin, and sparfloxacin in control cells but became insensitive to ampicillin and more sensitive to azithromycin and sparfloxacin in gamma interferon-treated cells. We show here that these modulations of antibiotic activity are largely counteracted by l-monomethyl arginine and catalase. In parallel, we show that gamma interferon enhances the cellular accumulation of azithromycin and sparfloxacin, an effect which is not reversed by addition of l-monomethyl arginine and catalase and which therefore cannot account for the increased activity of these antibiotics in gamma interferon-treated cells. We conclude that (i) the control exerted by gamma interferon on intracellular multiplication ofListeria in THP-1 macrophages is dependent on the production of nitric oxide and hydrogen peroxide; (ii) intracellularListeria may become insensitive to ampicillin in macrophages exposed to gamma interferon because the increase in reactive oxygen and nitrogen intermediates already controls bacterial growth; and (iii) azithromycin and still more sparfloxacin cooperate efficiently with gamma interferon, one of the main cellular host defenses in Listeria infection.

scholarly journals Listeria monocytogenes σHContributes to Expression of Competence Genes and Intracellular Growth

2016 ◽  
Vol 198 (8) ◽  
pp. 1207-1217 ◽  
Author(s):  
Veronica Medrano Romero ◽  
Kazuya Morikawa
Keyword(s):  
Transcription Factor ◽  
Listeria Monocytogenes ◽  
Sigma Factor ◽  
Physiological Role ◽  
Population Level ◽  
Alternative Sigma Factor ◽  
Intracellular Growth ◽  
Content Type ◽  
Gram Positive Bacteria ◽  
Regulation Scheme

ABSTRACTThe alternative sigma factor σHhas two functions in Gram-positive bacteria: it regulates sporulation and the development of genetic competence.Listeria monocytogenesis a nonsporulating species in which competence has not yet been detected. Nevertheless, the main competence regulators and a series of orthologous genes that form the competence machinery are present in its genome; some of the competence genes play a role in optimal phagosomal escape. In this study, strains overexpressing σHand strains with a σHdeletion were used to elucidate the contribution of σHto the expression of the competence machinery genes inL. monocytogenes. Gene expression analysis showed that σHis, indeed, involved incomGandcomEregulation. Unexpectedly, we observed a unique regulation scheme in which σHand the transcription factor ComK were involved. Population-level analysis showed that even with the overexpression of both factors, only a fraction of the cells expressed the competence machinery genes. Although we could not detect competence, σHwas crucial for phagosomal escape, which implies that this alternative sigma factor has specifically evolved to regulate theL. monocytogenesintracellular life cycle.IMPORTANCEListeria monocytogenescan be an intracellular pathogen capable of causing serious infections in humans and animal species. Recently, the competence machinery genes were described as being necessary for optimal phagosomal escape, in which the transcription factor ComK plays an important role. On the other hand, our previous phylogenetic analysis suggested that the alternative sigma factor σHmight play a role in the regulation of competence genes. The present study shows that some of the competence genes belong to the σHregulon and, importantly, that σHis essential for intracellular growth, implying a unique physiological role of σHamongFirmicutes.

scholarly journals Type I Interferons Increase Host Susceptibility to Trypanosoma cruzi Infection

2011 ◽  
Vol 79 (5) ◽  
pp. 2112-2119 ◽  
Author(s):  
Anne-Danielle C. Chessler ◽  
Kacey L. Caradonna ◽  
Akram Da'dara ◽  
Barbara A. Burleigh
Keyword(s):  
Trypanosoma Cruzi ◽  
Parasite Burden ◽  
Host Susceptibility ◽  
Type I Interferons ◽  
Type I ◽  
Type I Ifn ◽  
Content Type ◽  
Type I Ifns ◽  
Ifn Γ ◽  
The Impact

ABSTRACTTrypanosoma cruzi, the protozoan parasite that causes human Chagas' disease, induces a type I interferon (IFN) (IFN-α/β) response during acute experimental infection in mice and in isolated primary cell types. To examine the potential impact of the type I IFN response in shaping outcomes in experimentalT. cruziinfection, groups of wild-type (WT) and type I IFN receptor-deficient (IFNAR−/−) 129sv/ev mice were infected with two differentT. cruzistrains under lethal and sublethal conditions and several parameters were measured during the acute stage of infection. The results demonstrate that type I IFNs are not required for early host protection againstT. cruzi. In contrast, under conditions of lethalT. cruzichallenge, WT mice succumbed to infection whereas IFNAR−/−mice were ultimately able to control parasite growth and survive.T. cruziclearance in and survival of IFNAR−/−mice were accompanied by higher levels of IFN-γ production by isolated splenocytes in response to parasite antigen. The suppression of IFN-γ in splenocytes from WT mice was independent of IL-10 levels. While the impact of type I IFNs on the production of IFN-γ and other cytokines/chemokines remains to be fully determined in the context ofT. cruziinfection, our data suggest that, under conditions of high parasite burden, type I IFNs negatively impact IFN-γ production, initiating a detrimental cycle that contributes to the ultimate failure to control infection. These findings are consistent with a growing theme in the microbial pathogenesis field in which type I IFNs can be detrimental to the host in a variety of nonviral pathogen infection models.

scholarly journals Two Zinc Uptake Systems Contribute to the Full Virulence of Listeria monocytogenes during GrowthIn VitroandIn Vivo

2011 ◽  
Vol 80 (1) ◽  
pp. 14-21 ◽  
Author(s):  
David Corbett ◽  
Jiahui Wang ◽  
Stephanie Schuler ◽  
Gloria Lopez-Castejon ◽  
Sarah Glenn ◽  
...  
Keyword(s):  
Listeria Monocytogenes ◽  
Zinc Uptake ◽  
Detectable Effect ◽  
Intracellular Growth ◽  
Content Type ◽  
Zinc Sensing ◽  
Identification And Characterization ◽  
Cell Deletion

ABSTRACTWe report here the identification and characterization of two zinc uptake systems, ZurAM and ZinABC, in the intracellular pathogenListeria monocytogenes. Transcription of both operons was zinc responsive and regulated by the zinc-sensing repressor Zur. Deletion of eitherzurAMorzinAhad no detectable effect on growth in defined media, but a doublezurAM zinAmutant was unable to grow in the absence of zinc supplementation. Deletion ofzinAhad no detectable effect on intracellular growth in HeLa epithelial cells. In contrast, growth of thezurAMmutant was significantly impaired in these cells, indicating the importance of the ZurAM system during intracellular growth. Notably, the deletion of bothzinAandzurAMseverely attenuated intracellular growth, with the double mutant being defective in actin-based motility and unable to spread from cell to cell. Deletion of eitherzurAMorzinAhad a significant effect on virulence in an oral mouse model, indicating that both zinc uptake systems are importantin vivoand establishing the importance of zinc acquisition during infection byL. monocytogenes. The presence of two zinc uptake systems may offer a mechanism by whichL. monocytogenescan respond to zinc deficiency within a variety of environments and during different stages of infection, with each system making distinct contributions under different stress conditions.

scholarly journals Hofbauer Cells Spread Listeria monocytogenes among Placental Cells and Undergo Pro-Inflammatory Reprogramming while Retaining Production of Tolerogenic Factors

mBio ◽  
2021 ◽  
Author(s):  
Siavash Azari ◽  
Lauren J. Johnson ◽  
Amy Webb ◽  
Sophia M. Kozlowski ◽  
Xiaoli Zhang ◽  
...  
Keyword(s):  
Listeria Monocytogenes ◽  
Pregnancy Complications ◽  
Intracellular Pathogen ◽  
Content Type ◽  
Placental Cells ◽  
Hofbauer Cells ◽  
Anti Inflammatory

Infection of the placental/fetal unit by the facultative intracellular pathogen Listeria monocytogenes results in severe pregnancy complications. Hofbauer cells (HBCs) are fetal macrophages that play homeostatic anti-inflammatory functions in healthy placentas.

scholarly journals Investigating the Roles of Listeria monocytogenes Peroxidases in Growth and Virulence

2021 ◽  
Author(s):  
Monica R. Cesinger ◽  
Nicole H. Schwardt ◽  
Cortney R. Halsey ◽  
Maureen K. Thomason ◽  
Michelle L. Reniere
Keyword(s):  
Reactive Oxygen Species ◽  
Immune System ◽  
Listeria Monocytogenes ◽  
Causative Agent ◽  
Foodborne Illness ◽  
Intracellular Pathogen ◽  
Host Immune System ◽  
Oxygen Species ◽  
Aerobic Growth ◽  
Content Type

Listeria monocytogenes is a facultative intracellular pathogen and the causative agent of the foodborne illness listeriosis. L. monocytogenes must contend with reactive oxygen species generated extracellularly during aerobic growth and intracellularly by the host immune system. However, the mechanisms by which L. monocytogenes defends against peroxide toxicity have not yet been defined.

scholarly journals Identification of Conserved and Species-Specific Functions of the Listeria monocytogenes PrsA2 Secretion Chaperone

2015 ◽  
Vol 83 (10) ◽  
pp. 4028-4041 ◽  
Author(s):  
Laty A. Cahoon ◽  
Nancy E. Freitag
Keyword(s):  
Listeria Monocytogenes ◽  
Protein Translocation ◽  
Functional Complementation ◽  
Broth Culture ◽  
Cell Physiology ◽  
Gram Positive ◽  
Content Type ◽  
Bacterial Physiology ◽  
Gram Positive Bacteria ◽  
Species Specific

The Gram-positive bacteriumListeria monocytogenesis a facultative intracellular pathogen that relies on the regulated secretion and activity of a variety of proteins that sustain life within diverse environments. PrsA2 has recently been identified as a secreted peptidyl-prolylcis/transisomerase and chaperone that is dispensable for bacterial growth in broth culture but essential forL. monocytogenesvirulence. Following host infection, PrsA2 contributes to the proper folding and activity of secreted proteins that are required for bacterial replication within the host cytosol and for bacterial spread to adjacent cells. PrsA2 is one member of a family of Gram-positive secretion chaperones that appear to play important roles in bacterial physiology; however, it is not known how these proteins recognize their substrate proteins or the degree to which their function is conserved across diverse Gram-positive species. We therefore examined PrsA proteins encoded by a variety of Gram-positive bacteria for functional complementation ofL. monocytogenesmutants lackingprsA2. PrsA homologues encoded byBacillus subtilis,Streptococcus pyogenes,Streptococcus pneumoniae,Streptococcus mutans,Staphylococcus aureus, andLactococcus lactiswere examined for functional complementation of a variety ofL. monocytogenesPrsA2-associated phenotypes central toL. monocytogenespathogenesis and bacterial cell physiology. Our results indicate that while selected aspects of PrsA2 function are broadly conserved among diverse Gram-positive bacteria, PrsA2 exhibits unique specificity forL. monocytogenestarget proteins required for pathogenesis. TheL. monocytogenesPrsA2 chaperone thus appears evolutionarily optimized for virulence factor secretion within the host cell cytosol while still maintaining aspects of activity relevant to more general features of Gram-positive protein translocation.

scholarly journals Listeria Phage and Phage Tail Induction Triggered by Components of Bacterial Growth Media (Phosphate, LiCl, Nalidixic Acid, and Acriflavine)

2015 ◽  
Vol 81 (6) ◽  
pp. 2117-2124 ◽  
Author(s):  
Jean-Paul Lemaître ◽  
Amandine Duroux ◽  
Romain Pimpie ◽  
Jean-Marie Duez ◽  
Marie-Louise Milat
Keyword(s):  
Listeria Monocytogenes ◽  
Nalidixic Acid ◽  
Bacterial Growth ◽  
False Negative ◽  
Brain Heart Infusion ◽  
False Negative Result ◽  
Growth Media ◽  
Content Type ◽  
Phage Tail ◽  
Initial Ph

ABSTRACTThe detection ofListeria monocytogenesfrom food is currently carried out using a double enrichment. For the ISO methodology, this double enrichment is performed using half-Fraser and Fraser broths, in which the overgrowth ofL. innocuacan occur in samples where both species are present. In this study, we analyzed the induction of phages and phage tails ofListeriaspp. in these media and in two brain heart infusion (BHI) broths (BHIM [bioMérieux] and BHIK [Biokar]) to identify putative effectors. It appears that Na2HPO4at concentrations ranging from 1 to 40 g/liter with an initial pH of 7.5 can induce phage or phage tail production ofListeriaspp., especially with 10 g/liter of Na2HPO4and a pH of 7.5, conditions present in half-Fraser and Fraser broths. Exposure to LiCl in BHIM (18 to 21 g/liter) can also induce phage and phage tail release, but in half-Fraser and Fraser broths, the concentration of LiCl is much lower (3 g/liter). Low phage titers were induced by acriflavine and/or nalidixic acid. We also show that the production of phages and phage tails can occur in half-Fraser and Fraser broths. This study points out that induction of phages and phage tails could be triggered by compounds present in enrichment media. This could lead to a false-negative result for the detection ofL. monocytogenesin food products.

scholarly journals Cyclic di-AMP Is Critical for Listeria monocytogenes Growth, Cell Wall Homeostasis, and Establishment of Infection

mBio ◽  
2013 ◽  
Vol 4 (3) ◽  
Author(s):  
Chelsea E. Witte ◽  
Aaron T. Whiteley ◽  
Thomas P. Burke ◽  
John-Demian Sauer ◽  
Daniel A. Portnoy ◽  
...  
Keyword(s):  
Cell Wall ◽  
Listeria Monocytogenes ◽  
Bacterial Growth ◽  
Genetic Screen ◽  
Content Type ◽  
Forward Genetic Screen ◽  
Innate Immune Signaling ◽  
Food Borne Illness ◽  
Multidrug Resistance Transporters

ABSTRACT Listeria monocytogenes infection leads to robust induction of an innate immune signaling pathway referred to as the cytosolic surveillance pathway (CSP), characterized by expression of beta interferon (IFN-β) and coregulated genes. We previously identified the IFN-β stimulatory ligand as secreted cyclic di-AMP. Synthesis of c-di-AMP in L. monocytogenes is catalyzed by the diadenylate cyclase DacA, and multidrug resistance transporters are necessary for secretion. To identify additional bacterial factors involved in L. monocytogenes detection by the CSP, we performed a forward genetic screen for mutants that induced altered levels of IFN-β. One mutant that stimulated elevated levels of IFN-β harbored a transposon insertion in the gene lmo0052. Lmo0052, renamed here PdeA, has homology to a cyclic di-AMP phosphodiesterase, GdpP (formerly YybT), of Bacillus subtilis and is able to degrade c-di-AMP to the linear dinucleotide pApA. Reduction of c-di-AMP levels by conditional depletion of the di-adenylate cyclase DacA or overexpression of PdeA led to marked decreases in growth rates, both in vitro and in macrophages. Additionally, mutants with altered levels of c-di-AMP had different susceptibilities to peptidoglycan-targeting antibiotics, suggesting that the molecule may be involved in regulating cell wall homeostasis. During intracellular infection, increases in c-di-AMP production led to hyperactivation of the CSP. Conditional depletion of dacA also led to increased IFN-β expression and a concomitant increase in host cell pyroptosis, a result of increased bacteriolysis and subsequent bacterial DNA release. These data suggest that c-di-AMP coordinates bacterial growth, cell wall stability, and responses to stress and plays a crucial role in the establishment of bacterial infection. IMPORTANCE Listeria monocytogenes is a Gram-positive intracellular pathogen and the causative agent of the food-borne illness listeriosis. Upon infection, L. monocytogenes stimulates expression of IFN-β and coregulated genes dependent upon host detection of a secreted bacterial signaling nucleotide, c-di-AMP. Using a forward genetic screen for mutants that induced high levels of host IFN-β expression, we identified a c-di-AMP phosphodiesterase, PdeA, that degrades c-di-AMP. Here we characterize L. monocytogenes mutants that express enhanced or diminished levels of c-di-AMP. Decreased c-di-AMP levels by conditional depletion of the diadenylate cyclase (DacA) or overexpression of PdeA attenuated bacterial growth and led to bacteriolysis, suggesting that its production is essential for viability and may regulate cell wall metabolism. Mutants lacking PdeA had a distinct transcriptional profile, which may provide insight into additional roles for the molecule. This work demonstrates that c-di-AMP is a critical signaling molecule required for bacterial replication, cell wall stability, and pathogenicity.

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