scholarly journals ThemsaABCROperon Regulates Resistance in Vancomycin-Intermediate Staphylococcus aureus Strains

2014 ◽  
Vol 58 (11) ◽  
pp. 6685-6695 ◽  
Author(s):  
Dhritiman Samanta ◽  
Mohamed O. Elasri
Keyword(s):  
Public Health ◽  
Staphylococcus Aureus ◽  
Cell Wall ◽  
Binding Capacity ◽  
Difficult Problem ◽  
Cell Wall Synthesis ◽  
Wild Type ◽  
Content Type ◽  
Type Strains

ABSTRACTVancomycin-intermediateStaphylococcus aureus(VISA) strains present an increasingly difficult problem in terms of public health. However, the molecular mechanism for this resistance is not yet understood. In this study, we define the role of themsaABCRoperon in vancomycin resistance in three clinical VISA strains, i.e., Mu50, HIP6297, and LIM2. Deletion of themsaABCRoperon resulted in significant decreases in the vancomycin MIC (from 6.25 to 1.56 μg/ml) and significant reductions of cell wall thickness in strains Mu50 and HIP6297. Growth of the mutants in medium containing vancomycin at concentrations greater than 2 μg/ml resulted in decreases in the growth rate, compared with the wild-type strains. Mutation of themsaABCRoperon also reduced the binding capacity for vancomycin. We conclude that themsaABCRoperon contributes to resistance to vancomycin and cell wall synthesis inS. aureus.

scholarly journals Causal Role of Single Nucleotide Polymorphisms within themprFGene of Staphylococcus aureus in Daptomycin Resistance

2013 ◽  
Vol 57 (11) ◽  
pp. 5658-5664 ◽  
Author(s):  
Soo-Jin Yang ◽  
Nagendra N. Mishra ◽  
Aileen Rubio ◽  
Arnold S. Bayer
Keyword(s):  
Staphylococcus Aureus ◽  
Single Nucleotide Polymorphisms ◽  
Point Mutations ◽  
Nucleotide Polymorphisms ◽  
Wild Type ◽  
Single Nucleotide ◽  
Content Type ◽  
Reading Frame ◽  
A Charge

ABSTRACTSingle nucleotide polymorphisms (SNPs) within themprFopen reading frame (ORF) have been commonly observed in daptomycin-resistant (DAPr)Staphylococcus aureusstrains. Such SNPs are usually associated with a gain-in-function phenotype, in terms of either increased synthesis or enhanced translocation (flipping) of lysyl-phosphatidylglycerol (L-PG). However, it is unclear if suchmprFSNPs are causal in DAPrstrains or are merely a biomarker for this phenotype. In this study, we used an isogenic set ofS. aureusstrains: (i) Newman, (ii) its isogenic ΔmprFmutant, and (iii) several intransplasmid complementation constructs, expressing either a wild-type or point-mutated form of themprFORF cloned from two isogenic DAP-susceptible (DAPs)-DAPrstrain pairs (616-701 and MRSA11/11-REF2145). Complementation of the ΔmprFstrain with singly point-mutatedmprFgenes (mprFS295LormprFT345A) revealed that (i) individual and distinct point mutations within themprFORF can recapitulate phenotypes observed in donor strains (i.e., changes in DAP MICs, positive surface charge, and cell membrane phospholipid profiles) and (ii) these gain-in-function SNPs (i.e., enhanced L-PG synthesis) likely promote reduced DAP binding toS. aureusby a charge repulsion mechanism. Thus, for these two DAPrstrains, the definedmprFSNPs appear to be causally related to this phenotype.

scholarly journals Role of the msaABCR Operon in Cell Wall Biosynthesis, Autolysis, Integrity, and Antibiotic Resistance in Staphylococcus aureus

2019 ◽  
Vol 63 (10) ◽  
Author(s):  
Bibek G C ◽  
Gyan S. Sahukhal ◽  
Mohamed O. Elasri
Keyword(s):  
Staphylococcus Aureus ◽  
Cell Wall ◽  
Antibiotic Resistance ◽  
Teichoic Acid ◽  
Cross Linking ◽  
Wall Defect ◽  
Content Type ◽  
Mrsa Strains ◽  
Mutant Cells

ABSTRACT Staphylococcus aureus is an important human pathogen in both community and health care settings. One of the challenges with S. aureus as a pathogen is its acquisition of antibiotic resistance. Previously, we showed that deletion of the msaABCR operon reduces cell wall thickness, resulting in decreased resistance to vancomycin in vancomycin-intermediate S. aureus (VISA). In this study, we investigated the nature of the cell wall defect in the msaABCR operon mutant in the Mu50 (VISA) and USA300 LAC methicillin-resistant Staphylococcus aureus (MRSA) strains. Results showed that msaABCR mutant cells had decreased cross-linking in both strains. This defect is typically due to increased murein hydrolase activity and/or nonspecific processing of murein hydrolases mediated by increased protease activity in mutant cells. The defect was enhanced by a decrease in teichoic acid content in the msaABCR mutant. Therefore, we propose that deletion of the msaABCR operon results in decreased peptidoglycan cross-linking, leading to increased susceptibility toward cell wall-targeting antibiotics, such as β-lactams and vancomycin. Moreover, we also observed significantly downregulated transcription of early cell wall-synthesizing genes, supporting the finding that msaABCR mutant cells have decreased peptidoglycan synthesis. More specifically, the msaABCR mutant in the USA300 LAC strain (MRSA) showed significantly reduced expression of the murA gene, whereas the msaABCR mutant in the Mu50 strain (VISA) showed significantly reduced expression of glmU, murA, and murD. Thus, we conclude that the msaABCR operon controls the balance between cell wall synthesis and cell wall hydrolysis, which is required for maintaining a robust cell wall and acquiring resistance to cell wall-targeting antibiotics, such as vancomycin and the β-lactams.

scholarly journals LcpB Is a Pyrophosphatase Responsible for Wall Teichoic Acid Synthesis and Virulence in Staphylococcus aureus Clinical Isolate ST59

2021 ◽  
Vol 12 ◽  
Author(s):  
Ting Pan ◽  
Jing Guan ◽  
Yujie Li ◽  
Baolin Sun
Keyword(s):  
Staphylococcus Aureus ◽  
Cell Wall ◽  
Molecular Mechanisms ◽  
Teichoic Acid ◽  
Acid Synthesis ◽  
Cell Wall Synthesis ◽  
Wall Teichoic Acid ◽  
Independent Manner ◽  
Methicillin Resistant

The community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) causes severe pandemics primarily consisting of skin and soft tissue infections. However, the underlying pathomechanisms of the bacterium are yet to fully understood. The present study identifies LcpB protein, which belongs to the LytR-A-Psr (LCP) family, is crucial for cell wall synthesis and virulence in S. aureus. The findings revealed that LcpB is a pyrophosphatase responsible for wall teichoic acid synthesis. The results also showed that LcpB regulates enzyme activity through specific key arginine sites in its LCP domain. Furthermore, knockout of lcpB in the CA-MRSA isolate ST59 resulted in enhanced hemolytic activity, enlarged of abscesses, and increased leukocyte infiltration. Meanwhile, we also found that LcpB regulates virulence in agr-independent manner and the key sites for pyrophosphatase of LcpB play critical roles in regulating the virulence. In addition, the results showed that the role of LcpB was different between methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-sensitive Staphylococcus aureus (MSSA). This study therefore highlights the dual role of LcpB in cell wall synthesis and regulation of virulence. These insights on the underlying molecular mechanisms can thus guide the development of novel anti-infective strategies.

scholarly journals Role of VraSR in Antibiotic Resistance and Antibiotic-Induced Stress Response in Staphylococcus aureus

2006 ◽  
Vol 50 (10) ◽  
pp. 3424-3434 ◽  
Author(s):  
S. Gardete ◽  
S. W. Wu ◽  
S. Gill ◽  
A. Tomasz
Keyword(s):  
Staphylococcus Aureus ◽  
Cell Wall ◽  
Stress Response ◽  
Critical Role ◽  
Regulatory System ◽  
Experimental System ◽  
Cell Wall Synthesis ◽  
Beta Lactam ◽  
Active Inhibitor

ABSTRACT Exposure of Staphylococcus aureus to cell wall inhibitors induces massive overexpression of a number of genes, provided that the VraSR two-component sensory regulatory system is intact. Inactivation of vraS blocks this transcriptional response and also causes a drastic reduction in the levels of resistance to beta-lactam antibiotics and vancomycin. We used an experimental system in which the essential cell wall synthesis gene of S. aureus, pbpB, was put under the control of an isopropyl-β-d-thiogalactopyranoside-inducible promoter in order to induce reversible perturbations in cell wall synthesis without the use of any cell wall-active inhibitor. Changes in the level of transcription of pbpB were rapidly followed by parallel changes in the vraSR signal, and the abundance of the pbpB transcript was precisely mirrored by the abundance of the transcripts of vraSR and some additional genes that belong to the VraSR regulon. Beta-lactam resistance in S. aureus appears to involve a complex stress response in which VraSR performs the critical role of a sentinel system capable of sensing the perturbation of cell wall synthesis and allowing mobilization of genes that are essential for the generation of a highly resistant phenotype. One of the sites in cell wall synthesis “sensed” by the VraSR system appears to be a step catalyzed by PBP 2.

scholarly journals An Intracellular Peptidyl-Prolyl cis/trans Isomerase Is Required for Folding and Activity of the Staphylococcus aureus Secreted Virulence Factor Nuclease

2016 ◽  
Vol 199 (1) ◽  
Author(s):  
Richard E. Wiemels ◽  
Stephanie M. Cech ◽  
Nikki M. Meyer ◽  
Caleb A. Burke ◽  
Andy Weiss ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Cell Wall ◽  
Virulence Factors ◽  
Virulence Factor ◽  
Active Form ◽  
Ppiase Activity ◽  
Content Type ◽  
Associated Proteins

ABSTRACT Staphylococcus aureus is an important human pathogen that relies on a large repertoire of secreted and cell wall-associated proteins for pathogenesis. Consequently, the ability of the organism to cause disease is absolutely dependent on its ability to synthesize and successfully secrete these proteins. In this study, we investigate the role of peptidyl-prolyl cis/trans isomerases (PPIases) on the activity of the S. aureus secreted virulence factor nuclease (Nuc). We identify a staphylococcal cyclophilin-type PPIase (PpiB) that is required for optimal activity of Nuc. Disruption of ppiB results in decreased nuclease activity in culture supernatants; however, the levels of Nuc protein are not altered, suggesting that the decrease in activity results from misfolding of Nuc in the absence of PpiB. We go on to demonstrate that PpiB exhibits PPIase activity in vitro, is localized to the bacterial cytosol, and directly interacts with Nuc in vitro to accelerate the rate of Nuc refolding. Finally, we demonstrate an additional role for PpiB in S. aureus hemolysis and demonstrate that the S. aureus parvulin-type PPIase PrsA also plays a role in the activity of secreted virulence factors. The deletion of prsA leads to a decrease in secreted protease and phospholipase activity, similar to that observed in other Gram-positive pathogens. Together, these results demonstrate, for the first time to our knowledge, that PPIases play an important role in the secretion of virulence factors in S. aureus. IMPORTANCE Staphylococcus aureus is a highly dangerous bacterial pathogen capable of causing a variety of infections throughout the human body. The ability of S. aureus to cause disease is largely due to an extensive repertoire of secreted and cell wall-associated proteins, including adhesins, toxins, exoenzymes, and superantigens. These virulence factors, once produced, are typically transported across the cell membrane by the secretory (Sec) system in a denatured state. Consequently, once outside the cell, they must refold into their active form. This step often requires the assistance of bacterial folding proteins, such as PPIases. In this work, we investigate the role of PPIases in S. aureus and uncover a cyclophilin-type enzyme that assists in the folding/refolding of staphylococcal nuclease.

scholarly journals The Peptidoglycan Hydrolase of Staphylococcus aureus Bacteriophage ϕ11 Plays a Structural Role in the Viral Particle

2013 ◽  
Vol 79 (19) ◽  
pp. 6187-6190 ◽  
Author(s):  
Lorena Rodríguez-Rubio ◽  
Nuria Quiles-Puchalt ◽  
Beatriz Martínez ◽  
Ana Rodríguez ◽  
José R. Penadés ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Burst Size ◽  
Wild Type ◽  
Content Type ◽  
Wild Type Phenotype ◽  
Viral Particles ◽  
Peptidoglycan Hydrolases ◽  
Host Lysis ◽  
Phage Growth

ABSTRACTThe role of virion-associated peptidoglycan hydrolases (VAPGHs) in the phage infection cycle is not clear. gp49, the VAPGH fromStaphylococcus aureusphage ϕ11, is not essential for phage growth but stabilizes the viral particles. ϕ11Δ49 phages showed a reduced burst size and delayed host lysis. Complementation of gp49 with HydH5 from bacteriophage vB_SauS-phiIPLA88 restored the wild-type phenotype.

scholarly journals Epigallocatechin Gallate Induces Upregulation of the Two-Component VraSR System by Evoking a Cell Wall Stress Response in Staphylococcus aureus

2012 ◽  
Vol 78 (22) ◽  
pp. 7954-7959 ◽  
Author(s):  
Oren Levinger ◽  
Tamar Bikels-Goshen ◽  
Elad Landau ◽  
Merav Fichman ◽  
Roni Shapira
Keyword(s):  
Staphylococcus Aureus ◽  
Cell Wall ◽  
Epigallocatechin Gallate ◽  
Wall Stress ◽  
Null Mutant ◽  
Wild Type ◽  
Content Type ◽  
Cell Wall Stress ◽  
Two Component ◽  
A Cell

ABSTRACTWe previously found that a short exposure ofStaphylococcus aureusto subinhibitory (SI) doses of epigallocatechin gallate (EGCG) results in increased cell wall thickness, adaptation, and enhanced tolerance to cell-wall-targeted antibiotics. In this study, the response to EGCG ofsigBandvraSRtranscription factor mutants was characterized. We show that in contrast to the results observed for wild-type (WT) strains, anS. aureus315vraSRnull mutant exposed to SI doses of EGCG did not exhibit increased tolerance to EGCG and oxacillin. A diminished increase in tolerance to ampicillin (from 16-fold to 4-fold) and no change in the magnitude of resistance to vancomycin were observed. Preexposure to EGCG enhanced the tolerance of wild-type andsigBnull mutant cells to lysostaphin, but this enhancement was much weaker in thevraSRnull mutant. Marked upregulation (about 60-fold) ofvraRand upregulation of the peptidoglycan biosynthesis-associated genesmurA,murF, andpbp2(2-, 5-, and 6-fold, respectively) in response to SI doses of EGCG were determined by quantitative reverse transcription-PCR (qRT-PCR). EGCG also induced the promoter ofsas016(encoding a cell wall stress protein of unknown function which is not induced invraSRnull mutants) in a concentration-dependent manner, showing kinetics comparable to those of cell-wall-targeting antibiotics. Taken together, our results suggest that the two-component VraSR system is involved in modulating the cell response to SI doses of EGCG.

scholarly journals σBInhibits Poly-N-Acetylglucosamine Exopolysaccharide Synthesis and Biofilm Formation inStaphylococcus aureus

2019 ◽  
Vol 201 (11) ◽  
Author(s):  
Jaione Valle ◽  
Maite Echeverz ◽  
Iñigo Lasa
Keyword(s):  
Staphylococcus Aureus ◽  
Biofilm Formation ◽  
Flow Conditions ◽  
Content Type ◽  
Environmental Signals ◽  
General Stress Response ◽  
Biofilm Development ◽  
Type Strains ◽  
Complex Regulatory Network

ABSTRACTStaphylococcus aureusclinical strains are able to produce at least two distinct types of biofilm matrixes: biofilm matrixes made of the polysaccharide intercellular adhesin (PIA) or poly-N-acetylglucosamine (PNAG), whose synthesis is mediated by theicaADBClocus, and biofilm matrixes built of proteins (polysaccharide independent). σBis a conserved alternative sigma factor that regulates the expression of more than 100 genes in response to changes in environmental conditions. While numerous studies agree that σBis required for polysaccharide-independent biofilms, controversy persists over the role of σBin the regulation of PIA/PNAG-dependent biofilm development. Here, we show that genetically unrelatedS. aureusσB-deficient strains produced stronger biofilms under both static and flow conditions and accumulated higher levels of PIA/PNAG exopolysaccharide than their corresponding wild-type strains. The increased accumulation of PIA/PNAG in the σBmutants correlated with a greater accumulation of the IcaC protein showed that it was not due to adjustments inicaADBCoperon transcription and/oricaADBCmRNA stability. Overall, our results reveal that in the presence of active σB, the turnover of Ica proteins is accelerated, reducing the synthesis of PIA/PNAG exopolysaccharide and consequently the PIA/PNAG-dependent biofilm formation capacity.IMPORTANCEDue to its multifaceted lifestyle,Staphylococcus aureusneeds a complex regulatory network to connect environmental signals with cellular physiology. One particular transcription factor, named σB(SigB), is involved in the general stress response and the expression of virulence factors. For many years, great confusion has existed about the role of σBin the regulation of the biofilm lifestyle inS. aureus. Our study demonstrated that σBis not necessary for exopolysaccharide-dependent biofilms and, even more, thatS. aureusproduces stronger biofilms in the absence of σB. The increased accumulation of exopolysaccharide correlates with higher stability of the proteins responsible for its synthesis. The present findings reveal an additional regulatory layer to control biofilm exopolysaccharide synthesis under stress conditions.

scholarly journals Roles of Lytic Transglycosylases in Biofilm Formation and β-Lactam Resistance in Methicillin-Resistant Staphylococcus aureus

2019 ◽  
Vol 63 (12) ◽  
Author(s):  
Anne-Aurelie Lopes ◽  
Yutaka Yoshii ◽  
Satomi Yamada ◽  
Mari Nagakura ◽  
Yuki Kinjo ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Staphylococcus Epidermidis ◽  
Biofilm Formation ◽  
Wild Type ◽  
Potential Therapeutic Target ◽  
Methicillin Resistant ◽  
Content Type ◽  
Lytic Transglycosylases ◽  
Community Outbreaks ◽  
Type Strains

ABSTRACT Staphylococcus aureus is responsible for numerous community outbreaks and is one of the most frequent causes of nosocomial infections with significant morbidity and mortality. While the function of lytic transglycosylases (LTs) in relation to cell division, biofilm formation, and antibiotic resistance has been determined for several bacteria, their role in S. aureus remains largely unknown. The only known LTs in S. aureus are immunodominant staphylococcal antigen A (IsaA) and Staphylococcus epidermidis D protein (SceD). Our study demonstrates that, in strains of methicillin-resistant S. aureus (MRSA), IsaA and SceD contribute differently to biofilm formation and β-lactam resistance. Deletion of isaA, but not sceD, led to decreased biofilm formation. Additionally, in isaA-deleted strains, β-lactam resistance was significantly decreased compared with that of wild-type strains. Plasmid-based expression of mecA, a major determinant of β-lactam resistance in MRSA, in an isaA-deleted strain did not restore β-lactam resistance, demonstrating that the β-lactam susceptibility phenotype is exhibited by the isaA mutant regardless of the production level of PBP2a. Overall, our results suggest that IsaA is a potential therapeutic target for MRSA infections.

scholarly journals Putative Role of the Aldo-Keto Reductase from Trypanosoma cruzi in Benznidazole Metabolism

2016 ◽  
Vol 60 (5) ◽  
pp. 2664-2670 ◽  
Author(s):  
Patricia Andrea Garavaglia ◽  
Marc Laverrière ◽  
Joaquín J. B. Cannata ◽  
Gabriela Andrea García
Keyword(s):  
Trypanosoma Cruzi ◽  
Inhibitory Concentration ◽  
Reductase Activity ◽  
Differential Susceptibility ◽  
Type I ◽  
Wild Type ◽  
Content Type ◽  
Type Strains ◽  
Dependent Type

ABSTRACTBenznidazole (Bz), the drug used for treatment of Chagas' disease (caused by the protozoanTrypanosoma cruzi), is activated by a parasitic NADH-dependent type I nitroreductase (NTR I). However, several studies have shown that other enzymes are involved. The aim of this study was to evaluate whether the aldo-keto reductase fromT. cruzi(TcAKR), a NADPH-dependent oxido-reductase previously described by our group, uses Bz as the substrate. We demonstrated that both recombinant and nativeTcAKR enzymes reduce Bz by using NADPH, but not NADH, as a cofactor.TcAKR-overexpressing epimastigotes showed higher NADPH-dependent Bz reductase activity and a 50% inhibitory concentration (IC50) value for Bz 1.8-fold higher than that of the controls, suggesting thatTcAKR is involved in Bz detoxification instead of activation. To understand the role ofTcAKR in Bz metabolism, we studiedTcAKR expression and NADPH/NADH-dependent Bz reductase activities in twoT. cruzistrains with differential susceptibility to Bz: CL Brener and Nicaragua. Taking into account the results obtained withTcAKR-overexpressing epimastigotes, we expected the more resistant strain, Nicaragua, to have higherTcAKR levels than CL Brener. However, the results were the opposite. CL Brener showed 2-fold higherTcAKR expression and 5.7-fold higher NADPH-Bz reduction than the Nicaragua strain. In addition, NADH-dependent Bz reductase activity, characteristic of NTR I, was also higher in CL Brener than in Nicaragua. We conclude that althoughTcAKR uses Bz as the substrate,TcAKR activity is not a determinant of Bz resistance in wild-type strains and may be overcome by other enzymes involved in Bz activation, such as NADPH- and NADH-dependent reductases.

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