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 DhhP, a Cyclic di-AMP Phosphodiesterase of Borrelia burgdorferi, Is Essential for Cell Growth and Virulence

2014 ◽  
Vol 82 (5) ◽  
pp. 1840-1849 ◽  
Author(s):  
Meiping Ye ◽  
Jun-Jie Zhang ◽  
Xin Fang ◽  
Gavin B. Lawlis ◽  
Bryan Troxell ◽  
...  
Keyword(s):  
Borrelia Burgdorferi ◽  
Wild Type Strain ◽  
Wall Structure ◽  
Mammalian Host ◽  
Dependent Manner ◽  
Gram Positive ◽  
Content Type ◽  
Gram Positive Bacteria

ABSTRACTCyclic di-AMP (c-di-AMP) is a recently discovered second messenger in bacteria. Most of work on c-di-AMP signaling has been done in Gram-positive bacteria, firmicutes, and actinobacteria, where c-di-AMP signaling pathways affect potassium transport, cell wall structure, and antibiotic resistance. Little is known about c-di-AMP signaling in other bacteria.Borrelia burgdorferi, the causative agent of Lyme disease, is a spirochete that has a Gram-negative dual membrane. In this study, we demonstrated thatB. burgdorferiBB0619, aDHH-DHHA1 domainprotein (herein designated DhhP), functions as c-di-AMP phosphodiesterase. Recombinant DhhP hydrolyzed c-di-AMP to pApA in a Mn2+- or Mg2+-dependent manner. In contrast to c-di-AMP phosphodiesterases reported thus far, DhhP appears to be essential forB. burgdorferigrowth bothin vitroand in the mammalian host. Inactivation of the chromosomaldhhPgene could be achieved only in the presence of a plasmid-encoded inducibledhhPgene. The conditionaldhhPmutant had a dramatic increase in intracellular c-di-AMP level in comparison to the isogenic wild-type strain. Unlike what has been observed in Gram-positive bacteria, elevated cellular c-di-AMP inB. burgdorferidid not result in an increased resistance to β-lactamase antibiotics, suggesting that c-di-AMP's functions in spirochetes differ from those in Gram-positive bacteria. In addition, thedhhPmutant was defective in induction of the σSfactor, RpoS, and the RpoS-dependent outer membrane virulence factor OspC, which uncovers an important role of c-di-AMP inB. burgdorferivirulence.

scholarly journals A Novel Dual-creMotif Enables Two-Way Autoregulation of CcpA inClostridium acetobutylicum

2018 ◽  
Vol 84 (8) ◽  
pp. e00114-18 ◽  
Author(s):  
Lu Zhang ◽  
Yanqiang Liu ◽  
Yunpeng Yang ◽  
Weihong Jiang ◽  
Yang Gu
Keyword(s):  
Regulatory Network ◽  
Target Genes ◽  
Protein A ◽  
Cell Physiology ◽  
Coding Region ◽  
Gram Positive ◽  
Content Type ◽  
Gram Positive Bacteria

ABSTRACTThe master regulator CcpA (catabolite control protein A) manages a large and complex regulatory network that is essential for cellular physiology and metabolism in Gram-positive bacteria. Although CcpA can affect the expression of target genes by binding to acis-acting catabolite-responsive element (cre), whether and how the expression of CcpA is regulated remain poorly explored. Here, we report a novel dual-cremotif that is employed by the CcpA inClostridium acetobutylicum, a typical solventogenicClostridiumspecies, for autoregulation. Twocresites are involved in CcpA autoregulation, and they reside in the promoter and coding regions of CcpA. In this dual-cremotif,creP, in the promoter region, positively regulatesccpAtranscription, whereascreORF, in the coding region, negatively regulates this transcription, thus enabling two-way autoregulation of CcpA. Although CcpA boundcrePmore strongly thancreORFin vitro, thein vivoassay showed thatcreORF-based repression dominates CcpA autoregulation during the entire fermentation. Finally, a synonymous mutation ofcreORFwas made within the coding region, achieving an increased intracellular CcpA expression and improved cellular performance. This study provides new insights into the regulatory role of CcpA inC. acetobutylicumand, moreover, contributes a new engineering strategy for this industrial strain.IMPORTANCECcpA is known to be a key transcription factor in Gram-positive bacteria. However, it is still unclear whether and how the intracellular CcpA level is regulated, which may be essential for maintaining normal cell physiology and metabolism. We discovered here that CcpA employs a dual-cremotif to autoregulate, enabling dynamic control of its own expression level during the entire fermentation process. This finding answers the questions above and fills a void in our understanding of the regulatory network of CcpA. Interference in CcpA autoregulation leads to improved cellular performance, providing a new useful strategy in genetic engineering ofC. acetobutylicum. Since CcpA is widespread in Gram-positive bacteria, including pathogens, this dual-cre-based CcpA autoregulation would be valuable for increasing our understanding of CcpA-based global regulation in bacteria.

scholarly journals PapRIV, a BV-2 microglial cell activating quorum sensing peptide

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Yorick Janssens ◽  
Nathan Debunne ◽  
Anton De Spiegeleer ◽  
Evelien Wynendaele ◽  
Marta Planas ◽  
...  
Keyword(s):  
Quorum Sensing ◽  
Cell Model ◽  
Dependent Manner ◽  
Communication Signals ◽  
Gram Positive Bacteria ◽  
A Cell ◽  
Gastro Intestinal Tract

AbstractQuorum sensing peptides (QSPs) are bacterial peptides produced by Gram-positive bacteria to communicate with their peers in a cell-density dependent manner. These peptides do not only act as interbacterial communication signals, but can also have effects on the host. Compelling evidence demonstrates the presence of a gut-brain axis and more specifically, the role of the gut microbiota in microglial functioning. The aim of this study is to investigate microglial activating properties of a selected QSP (PapRIV) which is produced by Bacillus cereus species. PapRIV showed in vitro activating properties of BV-2 microglia cells and was able to cross the in vitro Caco-2 cell model and reach the brain. In vivo peptide presence was also demonstrated in mouse plasma. The peptide caused induction of IL-6, TNFα and ROS expression and increased the fraction of ameboid BV-2 microglia cells in an NF-κB dependent manner. Different metabolites were identified in serum, of which the main metabolite still remained active. PapRIV is thus able to cross the gastro-intestinal tract and the blood–brain barrier and shows in vitro activating properties in BV-2 microglia cells, hereby indicating a potential role of this quorum sensing peptide in gut-brain interaction.

scholarly journals In Vitro and In Vivo Antibacterial Activities of DW-224a, a New Fluoronaphthyridone

2006 ◽  
Vol 50 (6) ◽  
pp. 2261-2264 ◽  
Author(s):  
Hee-Soo Park ◽  
Hyun-Joo Kim ◽  
Min-Jung Seol ◽  
Dong-Rack Choi ◽  
Eung-Chil Choi ◽  
...  
Keyword(s):  
Antibacterial Activities ◽  
The Other ◽  
Vitro Activity ◽  
Gram Negative Bacteria ◽  
In Vitro Activity ◽  
Gram Positive ◽  
Gram Negative ◽  
Gram Positive Bacteria

ABSTRACT DW-224a showed the most potent in vitro activity among the quinolone compounds tested against clinical isolates of gram-positive bacteria. Against gram-negative bacteria, DW-224a was slightly less active than the other fluoroquinolones. The in vivo activities of DW-224a against gram-positive bacteria were more potent than those of other quinolones.

scholarly journals In Vitro and In Vivo Activities of DA-7867, a New Oxazolidinone, against Aerobic Gram-Positive Bacteria

2005 ◽  
Vol 49 (6) ◽  
pp. 2498-2500 ◽  
Author(s):  
Eun Jeong Yoon ◽  
Yeong Woo Jo ◽  
Sung Hak Choi ◽  
Tae Ho Lee ◽  
Jae Keol Rhee ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Streptococcus Pneumoniae ◽  
Gram Positive ◽  
Methicillin Resistant ◽  
Gram Positive Bacteria ◽  
Vancomycin Resistant Enterococci ◽  
Infection Models ◽  
Vancomycin Resistant

ABSTRACT In vitro and in vivo activities of DA-7867 were assessed against methicillin-resistant Staphylococcus aureus, vancomycin-resistant enterococci, and penicillin-resistant Streptococcus pneumoniae. All isolates were inhibited by DA-7867 at ≤0.78 μg/ml, a four-times-lower concentration than that of inhibition by linezolid. For murine infection models, DA-7867 also exhibited greater efficacy than linezolid against all isolates tested.

scholarly journals Restoration of Bioactive Lantibiotic Suicin from a RemnantlanLocus of Pathogenic Streptococcus suis Serotype 2

2013 ◽  
Vol 80 (3) ◽  
pp. 1062-1071 ◽  
Author(s):  
Jian Wang ◽  
Yong Gao ◽  
Kunling Teng ◽  
Jie Zhang ◽  
Shutao Sun ◽  
...  
Keyword(s):  
Gene Promoter ◽  
Streptococcus Suis ◽  
Gene Clusters ◽  
Disulfide Bridge ◽  
Gram Positive ◽  
Content Type ◽  
Gram Positive Bacteria ◽  
System A ◽  
Bactericidal Activities

ABSTRACTLantibiotics are ribosomally synthesized, posttranslationally modified antimicrobial peptides. Their biosynthesis genes are usually organized in gene clusters, which are mainly found in Gram-positive bacteria, including pathogenic streptococci. Three highly virulentStreptococcus suisserotype 2 strains (98HAH33, 05ZYH33, and SC84) have been shown to contain an 89K pathogenicity island. Here, on these islands, we unveiled and reannotated a putative lantibiotic locus designatedsuiwhich contains a virulence-associated two-component regulator,suiK-suiR. In silicoanalysis revealed that the putative lantibiotic modification genesuiMwas interrupted by a 7.9-kb integron and that other biosynthesis-related genes contained various frameshift mutations. By reconstituting the intactsuiMinEscherichia colitogether with a semi-in vitrobiosynthesis system, a putative lantibiotic named suicin was produced with bactericidal activities against a variety of Gram-positive strains, including pathogenic streptococci and vancomycin-resistant enterococci. Ring topology dissection indicated that the 34-amino-acid lantibiotic contained two methyllanthionine residues and one disulfide bridge, which render suicin in an N-terminal linear and C-terminal globular shape. To confirm the function ofsuiK-suiR, SuiR was overexpressed and purified.In vitroanalysis showed that SuiR could specifically bind to thesuiAgene promoter. Its coexpression withsuiKcould activatesuiAgene promoter inLactococcus lactisNZ9000. Conclusively, we obtained a novel lantibiotic suicin by restoring its production from the remnantsuilocus and demonstrated that virulence-associated SuiK-SuiR regulates its production.

PTS 50: Past, Present and Future, or Diauxie Revisited

2015 ◽  
Vol 25 (2-3) ◽  
pp. 79-93 ◽  
Author(s):  
Joseph W. Lengeler
Keyword(s):  
Catabolite Repression ◽  
Biological Significance ◽  
Cellular Growth ◽  
Gram Positive ◽  
Gram Negative ◽  
Inducer Exclusion ◽  
Gram Positive Bacteria ◽  
Cellular Control

<b><i>Past:</i></b> The title ‘PTS 50 or The PTS after 50 years' relies on the first description in 1964 of the phosphoenolpyruvate-dependent carbohydrate:phosphotransferase system (PTS) by Kundig, Gosh and Roseman [Proc Natl Acad Sci USA 1964;52:1067-1074]. The system comprised proteins named Enzyme I, HPr and Enzymes II, as part of a novel PTS for carbohydrates in Gram-negative and Gram-positive bacteria, whose ‘biological significance remained unclear'. In contrast, studies which would eventually lead to the discovery of the central role of the PTS in bacterial metabolism had been published since before 1942. They are primarily linked to names like Epps and Gale, J. Monod, Cohn and Horibata, and B. Magasanik, and to phenomena like ‘glucose effects', ‘diauxie', ‘catabolite repression' and carbohydrate transport. <b><i>Present:</i></b> The pioneering work from Roseman's group initiated a flood of publications. The extraordinary progress from 1964 to this day in the qualitative and in vitro description of the genes and enzymes of the PTS, and of its multiple roles in global cellular control through ‘inducer exclusion', gene induction and ‘catabolite repression', in cellular growth, in cell differentiation and in chemotaxis, as well as the differences of its functions between Gram-positive and Gram-negative bacteria, was one theme of the meeting and will not be treated in detail here. <b><i>Future:</i></b> At the 1988 Paris meeting entitled ‘The PTS after 25 years', Saul Roseman predicted that ‘we must describe these interactions [of the PTS components] in a quantitative way [under] in vivo conditions'. I will present some results obtained by our group during recent years on the old phenomenon of diauxie by means of very fast and quantitative tests, measured in vivo, and obtained from cultures of isogenic mutant strains growing under chemostat conditions. The results begin to hint at the problems relating to future PTS research, but also to the ‘true science' of Roseman.

scholarly journals Activity of the Pore-Forming Virulence Factor Listeriolysin O Is Reversibly Inhibited by Naturally Occurring S-Glutathionylation

2017 ◽  
Vol 85 (4) ◽  
Author(s):  
Jonathan L. Portman ◽  
Qiongying Huang ◽  
Michelle L. Reniere ◽  
Anthony T. Iavarone ◽  
Daniel A. Portnoy
Keyword(s):  
Protein Function ◽  
Inhibitory Effect ◽  
Cysteine Residue ◽  
Infection Model ◽  
Listeriolysin O ◽  
Content Type ◽  
Pore Forming Proteins

ABSTRACT Cholesterol-dependent cytolysins (CDCs) represent a family of homologous pore-forming proteins secreted by many Gram-positive bacterial pathogens. CDCs mediate membrane binding partly through a conserved C-terminal undecapeptide, which contains a single cysteine residue. While mutational changes to other residues in the undecapeptide typically have severe effects, mutation of the cysteine residue to alanine has minor effects on overall protein function. Thus, the role of this highly conserved reactive cysteine residue remains largely unknown. We report here that the CDC listeriolysin O (LLO), secreted by the facultative intracellular pathogen Listeria monocytogenes, was posttranslationally modified by S-glutathionylation at this conserved cysteine residue and that either endogenously synthesized or exogenously added glutathione was sufficient to form this modification. When recapitulated with purified protein in vitro, this modification completely ablated the activity of LLO, and this inhibitory effect was fully reversible by treatment with reducing agents. A cysteine-to-alanine mutation in LLO rendered the protein completely resistant to inactivation by S-glutathionylation, and a mutant expressing this mutation retained full hemolytic activity. A mutant strain of L. monocytogenes expressing the cysteine-to-alanine variant of LLO was able to infect and replicate within bone marrow-derived macrophages indistinguishably from the wild type in vitro, yet it was attenuated 4- to 6-fold in a competitive murine infection model in vivo. This study suggests that S-glutathionylation may represent a mechanism by which CDC-family proteins are posttranslationally modified and regulated and help explain an evolutionary pressure to retain the highly conserved undecapeptide cysteine.

Zinc oxide microrods and nanorods: different antibacterial activity and their mode of action against Gram-positive bacteria

RSC Advances ◽  
2014 ◽  
Vol 4 (99) ◽  
pp. 56031-56040 ◽  
Author(s):  
Ilaria Rago ◽  
Chandrakanth Reddy Chandraiahgari ◽  
Maria P. Bracciale ◽  
Giovanni De Bellis ◽  
Elena Zanni ◽  
...  
Keyword(s):  
Zinc Oxide ◽  
Antibacterial Activity ◽  
Mode Of Action ◽  
Gram Positive ◽  
Gram Positive Bacteria

ZnO micro and nanorods, produced through simple and inexpensive techniques, resulted to be strong antimicrobials against Gram-positive bacteria, in vitro as well as in vivo, by altering cell outer structures like membrane and exopolysaccharides.

scholarly journals A75 ROLE OF LRRK2 IN INFLAMMATORY BOWEL DISEASE

2018 ◽  
Vol 1 (suppl_2) ◽  
pp. 118-118
Author(s):  
J Rocha ◽  
C Sun ◽  
M Glogauer ◽  
D Philpott
Keyword(s):  
Listeria Monocytogenes ◽  
Myeloid Cell ◽  
Mucosal Barrier ◽  
Ros Generation ◽  
Transwell Assay ◽  
Bacteria Growth ◽  
Tlr4 Ligand

Abstract Background Variants of the leucine-rich repeat kinase 2 (LRRK2) are associated with an increased susceptibility to Parkinson disease but also Crohn’s disease (CD). Aims The present research is designed to develop a comprehensive understanding of the role of LRRK2 in immune system modulation, and how dysfunction of this pathway may lead to the development of CD. Methods WT and LRRK2-deficient neutrophil were infected with Gram-positive Bacteria (Listeria monocytogenes-LM) in a gentamicin protection assays and colony-forming unit assessment will determine the competence of LRRK2 deficient cells for bacterial phagocytosis as well as killing capacity). To examine how LRRK2 is involved in the generation of ROS during the respiratory burst, we will first examine if neutrophil from LRRK2-KO mice have altered ROS generation upon infection with LM and addition of PMA. We evaluate in vitro the ability of neutrophils from LRRK2-KO versus WT mice to transmigrate in vitro in a transwell assay using fMLP as a chemattractant. Also, we investigate the peritoneal cells (by FACS analysis) after injection of different microbial stimuli including FK105 (NOD1 ligand), MDP (NOD2 ligand) and LPS (TLR4 ligand) and anti-cd3 model of ielitis. Results We found that LRRK2 KO mice have a defect in migration of neutrophils to the peritoneal cavity after injection of different microbial stimuli including FK10565 (NOD1 ligand), MDP (NOD2 ligand) and LPS (TLR4 ligand). Neutrophils from LRRK2 mice were compromised in their ability to transmigrate in vitro in a transwell assay using fMLP as a chemoattractant. Chemotaxis was also compromised. In parallel, we designed experiments to examine reactive oxygen species (ROS) produced in response to infection of myeloid cells with bacteria. Neutrophils from LRRK2 KO mice infected with Listeria monocytogenes were less able to restrict bacteria growth compared to WT cells. Consistent with these findings, cells from LRRK2 KO mice produced lower levels of ROS following bacterial infection. In order to determine whether myeloid cell migration is compromised in vivo during inflammation, we performed experiments in WT and KO mice looking at different models of ileitis/colitis. Conclusions With this work we will further characterize the role of LRRK2 in intestinal homeostasis and mucosal barrier maintenance, including how its deficiency may predispose an individual to developing CD. Funding Agencies CAG, CIHR

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