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Lade, Harshad, Sung Hee Chung, Yeonhee Lee, Bajarang Vasant Kumbhar, Hwang-Soo Joo, Yun-Gon Kim, Yung-Hun Yang i Jae-Seok Kim. "Thymol Reduces agr-Mediated Virulence Factor Phenol-Soluble Modulin Production in Staphylococcus aureus". BioMed Research International 2022 (9.05.2022): 1–14. http://dx.doi.org/10.1155/2022/8221622.
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Staphylococcus aureus is a major human bacterial pathogen that carries a large number of virulence factors. Many virulence factors of S. aureus are regulated by the accessory gene regulator (agr) quorum-sensing system. Phenol-soluble modulins (PSMs) are one of the agr-mediated virulence determinants known to play a significant role in S. aureus pathogenesis. In the present study, the efficacy of thymol to inhibit PSM production including δ-toxin in S. aureus was explored. We employed liquid chromatography–mass spectrometry (LC–MS) to quantify the PSMsα1–PSMα4, PSMβ1 and PSMβ2, and δ-toxin production from culture supernatants. We found that thymol at 0.5 MIC (128 μg/mL) significantly reduced the PSMα and δ-toxin production in S. aureus WKZ-1, WKZ-2, LAC USA300, and ATCC29213. Downregulation in transcription by quantitative real-time (qRT) PCR analysis of response regulator agrA and receptor histidine kinase agrC upon 0.5 MIC thymol treatment affirmed the results of LC–MS quantification of PSMs. In silico molecular docking analysis demonstrated the binding affinity of thymol with receptors AgrA and AgrC. Transmission electron microscopy images revealed no ultrastructural alterations (cell wall and membrane) in thymol-treated WKZ-1 and WKZ-2 S. aureus strains. Here, we demonstrated that thymol reduces various PSM production in S. aureus clinical isolates and reference strains with mass spectrometry.
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Rieu, Aurélie, Stéphanie Weidmann, Dominique Garmyn, Pascal Piveteau i Jean Guzzo. "agr System of Listeria monocytogenes EGD-e: Role in Adherence and Differential Expression Pattern". Applied and Environmental Microbiology 73, nr 19 (3.08.2007): 6125–33. http://dx.doi.org/10.1128/aem.00608-07.
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ABSTRACT In this study, we investigated the agrBDCA operon in the pathogenic bacterium Listeria monocytogenes EGD-e. In-frame deletion of agrA and agrD resulted in an altered adherence and biofilm formation on abiotic surfaces, suggesting the involvement of the agr system of L. monocytogenes during the early stages of biofilm formation. Real-time PCR experiments indicated that the transcript levels of agrBDCA depended on the stage of biofilm development, since the levels were lower after the initial attachment period than during biofilm growth, whereas transcription during planktonic growth was not growth phase dependent. The mRNA quantification data also suggested that the agr system was autoregulated and pointed to a differential expression of the agr genes during sessile and planktonic growth. Although the reverse transcription-PCR experiments revealed that the four genes were transcribed as a single messenger, chemical half-life and 5′ RACE (rapid amplification of cDNA ends) experiments indicated that the full size transcript underwent cleavage followed by degradation of the agrC and agrA transcripts, which suggests a complex regulation of agr transcription.
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Srivastava, S. K., K. Rajasree, A. Fasim, G. Arakere i B. Gopal. "Influence of the AgrC-AgrA Complex on the Response Time of Staphylococcus aureus Quorum Sensing". Journal of Bacteriology 196, nr 15 (23.05.2014): 2876–88. http://dx.doi.org/10.1128/jb.01530-14.
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Zhang, Linsheng, i Guangyong Ji. "Identification of a Staphylococcal AgrB Segment(s) Responsible for Group-Specific Processing of AgrD by Gene Swapping". Journal of Bacteriology 186, nr 20 (15.10.2004): 6706–13. http://dx.doi.org/10.1128/jb.186.20.6706-6713.2004.
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ABSTRACT The four gene products of the accessory gene regulator (agr) P2 operon of Staphylococcus aureus assemble a quorum-sensing system: AgrA and AgrC resemble a two-component signal transduction system, and AgrB and AgrD are required to produce an autoinducing peptide. Upon activation, this quorum-sensing system positively regulates the transcription of the P2 operon as well as the P3 operon, whose transcript, RNAIII, regulates the expression of virulence genes. Four groups of S. aureus have been identified based on the agr sequences and the group-specific interaction between the autoinducing peptide and AgrC. AgrB is a transmembrane protein involved in the processing of AgrD propeptide, and its interaction with AgrD is also group specific. In this study, a series of chimeric AgrBs were constructed by swapping between group I and group II AgrBs, and these mutants were used to analyze the group-specific segment(s) in AgrB that was responsible for AgrD processing. Our results revealed that the first transmembrane α-helix and the extracellular loop 1 of group I AgrB were decisive in the specific processing of group I AgrD. In contrast, two hydrophilic segments of group II AgrB played a crucial role in the group-specific processing of group II AgrD. We also found that several chimeric AgrBs were capable of processing AgrD from both groups, suggesting that all AgrB homologues may utilize the same or a similar mechanism in the processing of AgrDs.
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Somerville, Greg A., Stephen B. Beres, J. Ross Fitzgerald, Frank R. DeLeo, Robert L. Cole, Jessica S. Hoff i James M. Musser. "In Vitro Serial Passage of Staphylococcus aureus: Changes in Physiology, Virulence Factor Production, and agr Nucleotide Sequence". Journal of Bacteriology 184, nr 5 (1.03.2002): 1430–37. http://dx.doi.org/10.1128/jb.184.5.1430-1437.2002.
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ABSTRACT Recently, we observed that Staphylococcus aureus strains newly isolated from patients had twofold-higher aconitase activity than a strain passaged extensively in vitro, leading us to hypothesize that aconitase specific activity decreases over time during in vitro passage. To test this hypothesis, a strain recovered from a patient with toxic shock syndrome was serially passaged for 6 weeks, and the aconitase activity was measured. Aconitase specific activity decreased 38% (P < 0.001) by the sixth week in culture. During serial passage, S. aureus existed as a heterogeneous population with two colony types that had pronounced (wild type) or negligible zones of beta-hemolytic activity. The cell density-sensing accessory gene regulatory (agr) system regulates beta-hemolytic activity. Surprisingly, the percentage of colonies with a wild-type beta-hemolytic phenotype correlated strongly with aconitase specific activity (ρ = 0.96), suggesting a common cause of the decreased aconitase specific activity and the variation in percentage of beta-hemolytic colonies. The loss of the beta-hemolytic phenotype also coincided with the occurrence of mutations in the agrC coding region or the intergenic region between agrC and agrA in the derivative strains. Our results demonstrate that in vitro growth is sufficient to result in mutations within the agr operon. Additionally, our results demonstrate that S. aureus undergoes significant phenotypic and genotypic changes during serial passage and suggest that vigilance should be used when extrapolating data obtained from the study of high-passage strains.
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Wang, Li, Lin Qiao, Aike Li, Lixian Chen, Beibei He, Gang Liu, Weiwei Wang i Jun Fang. "Integrative Multiomics Analysis of the Heat Stress Response of Enterococcus faecium". Biomolecules 13, nr 3 (25.02.2023): 437. http://dx.doi.org/10.3390/biom13030437.
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A continuous heat-adaptation test was conducted for one Enterococcus faecium (E. faecium) strain wild-type (WT) RS047 to obtain a high-temperature-resistant strain. After domestication, the strain was screened with a significantly higher ability of heat resistance. which is named RS047-wl. Then a multi-omics analysis of transcriptomics and metabolomics was used to analyze the mechanism of the heat resistance of the mutant. A total of 98 differentially expressed genes (DEGs) and 115 differential metabolites covering multiple metabolic processes were detected in the mutant, which indicated that the tolerance of heat resistance was regulated by multiple mechanisms. The changes in AgrB, AgrC, and AgrA gene expressions were involved in quorum-sensing (QS) system pathways, which regulate biofilm formation. Second, highly soluble osmotic substances such as putrescine, spermidine, glycine betaine (GB), and trehalose-6P were accumulated for the membrane transport system. Third, organic acids metabolism and purine metabolism were down-regulated. The findings can provide target genes for subsequent genetic modification of E. faecium, and provide indications for screening heat-resistant bacteria, so as to improve the heat-resistant ability of E. faecium for production.
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Dmitrenko, Olga, Andrey Chaplin, Anna Balbutskaya, Tamara Pkhakadze i Sergey Alkhovsky. "In Silico Genome-Scale Analysis of Molecular Mechanisms Contributing to the Development of a Persistent Infection with Methicillin-Resistant Staphylococcus aureus (MRSA) ST239". International Journal of Molecular Sciences 23, nr 24 (16.12.2022): 16086. http://dx.doi.org/10.3390/ijms232416086.
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The increasing frequency of isolation of methicillin-resistant Staphylococcus aureus (MRSA) limits the chances for the effective antibacterial therapy of staphylococcal diseases and results in the development of persistent infection such as bacteremia and osteomyelitis. The aim of this study was to identify features of the MRSAST239 0943-1505-2016 (SA943) genome that contribute to the formation of both acute and chronic musculoskeletal infections. The analysis was performed using comparative genomics data of the dominant epidemic S. aureus lineages, namely ST1, ST8, ST30, ST36, and ST239. The SA943 genome encodes proteins that provide resistance to the host’s immune system, suppress immunological memory, and form biofilms. The molecular mechanisms of adaptation responsible for the development of persistent infection were as follows: amino acid substitution in PBP2 and PBP2a, providing resistance to ceftaroline; loss of a large part of prophage DNA and restoration of the nucleotide sequence of beta-hemolysin, that greatly facilitates the escape of phagocytosed bacteria from the phagosome and formation of biofilms; dysfunction of the AgrA system due to the presence of psm-mec and several amino acid substitutions in the AgrC; partial deletion of the nucleotide sequence in genomic island vSAβ resulting in the loss of two proteases of Spl—operon; and deletion of SD repeats in the SdrE amino acid sequence.
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Steiner, Elisabeth, Jamie Scott, Nigel P. Minton i Klaus Winzer. "AnagrQuorum Sensing System That Regulates Granulose Formation and Sporulation in Clostridium acetobutylicum". Applied and Environmental Microbiology 78, nr 4 (16.12.2011): 1113–22. http://dx.doi.org/10.1128/aem.06376-11.
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ABSTRACTThe Gram-positive, anaerobic, endospore-forming bacteriumClostridium acetobutylicumhas considerable biotechnological potential due to its ability to produce solvents as fermentation products, in particular the biofuel butanol. Its genome contains a putativeagrlocus,agrBDCA, known in staphylococci to constitute a cyclic peptide-based quorum sensing system. In staphylococci,agrBDis required for the generation of a peptide signal that, upon extracellular accumulation, is sensed by anagrCA-encoded two-component system. Using ClosTron technology,agrB,agrC, andagrAmutants ofC. acetobutylicumATCC 824 were generated and phenotypically characterized. Mutants and wild type displayed similar growth kinetics and no apparent differences in solvent formation under the conditions tested. However, the number of heat-resistant endospores formed by the mutants in liquid culture was reduced by about one order of magnitude. On agar-solidified medium, spore formation was more strongly affected, particularly inagrAandagrCmutants. Similarly, accumulation of the starch-like storage compound granulose was almost undetectable in colonies ofagrB,agrA, andagrCmutants. Importantly, these defects could be genetically complemented, demonstrating that they were directly linked toagrinactivation. A diffusible factor produced byagrBD-expressing strains was found to restore granulose and spore formation in theagrBmutant. Furthermore, a synthetic cyclic peptide, designed on the basis of theC. acetobutylicumAgrD sequence, was also capable of complementing the defects of theagrBmutant when added exogenously to the culture. Together, these findings support the hypothesis thatagr-dependent quorum sensing is involved in the regulation of sporulation and granulose formation inC. acetobutylicum.
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Sung, Julia M. L., Peter D. Chantler i David H. Lloyd. "Accessory Gene Regulator Locus of Staphylococcus intermedius". Infection and Immunity 74, nr 5 (maj 2006): 2947–56. http://dx.doi.org/10.1128/iai.74.5.2947-2956.2006.
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ABSTRACT The accessory gene regulator (agr) locus, a candidate system for the regulation of the production of virulence factors in Staphylococcus intermedius, has been characterized. Using PCR-based genome walking, we have obtained the first complete sequence (3,436 bp) of the accessory gene regulator (agr) gene in this organism. Sequence analysis of the agr gene has identified five open reading frames (ORFs), agrB, agrD, agrC, agrA, and hld. The translated ORF contained amino acid motifs characteristic of the response regulator and histidine protein kinase signal transducer of the classic two-component regulatory system. Sequencing of the agrD PCR products amplified from DNA from 20 different isolates has facilitated detection of genetic variation in the putative autoinducing peptide (AIP) within the agr gene of S. intermedius, revealing the presence of at least three agr specificity groups within this species. Classification of the agr gene from S. intermedius was supported by phylogenetic analysis. Real-time PCR also revealed that the effector molecule of the agr system, RNAIII, was regulated in an autocrine manner in S. intermedius and demonstrated positive correlation with the temporal gene expression patterns of luk and entC. Transcription of RNAIII was also dependent on self secreted cues. Cyclic self and nonself peptides were synthesized on the basis of the novel AIPs produced by S. intermedius, which lack the cysteine necessary to form the thiolactone ring in analogous peptides from Staphylococcus aureus and Staphylococcus epidermidis. Experiments with these synthetic cyclic peptides indicated that self peptides led to up-regulation of RNAIII—findings in support of the assumption that activation of the agr gene is initiated by growth- and species-specific factors generated during bacterial growth.
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Qin, Xiang, Kavindra V. Singh, George M. Weinstock i Barbara E. Murray. "Effects of Enterococcus faecalis fsrGenes on Production of Gelatinase and a Serine Protease and Virulence". Infection and Immunity 68, nr 5 (1.05.2000): 2579–86. http://dx.doi.org/10.1128/iai.68.5.2579-2586.2000.
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ABSTRACT Three agr-like genes (fsrA,fsrB, and fsrC, for Enterococcus faecalis regulator) were found upstream of the previously reported gelatinase gene (gelE) and a downstream putative serine protease gene (sprE; accession number Z12296 ) ofEnterococcus faecalis OG1RF. The deduced amino acid sequence of fsrA shows 26% identity and 38% similarity toStaphylococcus aureus AgrA (the response regulator of the accessory gene regulator system in the agr locus), FsrB shows 23% identity and 41% similarity to S. aureus AgrB, and FsrC shows 23% identity and 36% similarity to S. aureus AgrC (the sensor transducer of Agr system). Northern blot analysis suggested that gelE and sprE are cotranscribed and that fsrB and fsrC are also cotranscribed in OG1RF. Northern blot analysis of fsrA,fsrB, fsrC, gelE, andsprE insertion mutants showed that fsrB,fsrC, gelE, and sprE are not expressed in fsrA, fsrB, and fsrCmutants, while insertion in an open reading frame further upstream offsrA did not effect the expression of these genes, suggesting that agr-like genes may be autoregulated and that they regulate gelE and sprE expression, as further confirmed by complementation of fsr gene mutations with a 6-kb fragment which contains all three fsr genes in the shuttle vector, pAT18. Testing of 95 other isolates of E. faecalis showed that 62% produced gelatinase (Gel+), while 91% (including all Gel+ strains) hybridized to agelE probe; 71% (including all Gel+ strains) hybridized to an fsr probe, corroborating the conclusion that both gelE and fsr are necessary for gelatinase production. Testing of fsrA, fsrB, and sprE mutants in a mouse peritonitis model showed that sprE and agr-like gene mutants resulted in highly significantly prolonged survival compared to the parent strain OG1RF, a finding similar to what we had previously shown for agelE mutant. These results suggest that sprEand agr-like genes contribute to the virulence of E. faecalis OG1RF in this model.
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Spiegel, Christopher, Stephan Josef Maria Steixner i Débora C. Coraça-Huber. "Antibiofilm Activity of Omega-3 Fatty Acids and Its Influence on the Expression of Biofilm Formation Genes on Staphylococcus aureus". Antibiotics 11, nr 7 (11.07.2022): 932. http://dx.doi.org/10.3390/antibiotics11070932.
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Background: Currently, 1–2% of all prosthetic joint surgeries are followed by an infection. These infections cause approximately 4% of deaths in the first year after surgery, while the 5-year mortality rate is up to 21%. Prosthetic joint infections are mainly caused by Staphylococcus aureus or Staphylococcus epidermis strains. Both species share the capability of biofilm formation and methicillin resistance. The formation of biofilm helps bacterial cells to withstand critical environmental conditions. Due to their tolerance against antibacterial substances, biofilms are a significant problem in modern medicine. Alternatives for the use of methicillin as a therapeutic are not yet widespread. The use of omega-3 fatty acids, such as docosahexaenoic acid, may help against prosthetic joint infections and lower mortality rates. The aim of this study is to evaluate if docosahexaenoic acid offers a safe anti-biofilm activity against Staphylococcus aureus and MRSA without enhancing icaADBC-dependent biofilm formation or additional stress responses, therefore enhancing antibiotic tolerance and resistance. Methods: In this study, we examined the gene expression of biofilm-associated genes and regulators. We performed RT-qPCR after RNA extraction of Staphylococcus aureus ATCC 29213 and one clinical MRSA strain. We compared gene expression of icaADBC, SarA, SigB, and agrAC under the influence of 1.25 mg /L and 0.625 mg/L of docosahexaenoic acid to their controls. Results: We found a higher expression of regulatory genes such as SarA, SigB, agrA, and agrC at 1.25 mg/L of docosahexaenoic acid in ATCC 29213 and a lower increase in gene expression levels in clinical MRSA isolates. icaADBC was not affected in both strains at both concentration levels by docosahexaenoic acid. Conclusions: Docosahexaenoic acid does not enhance icaADBC-dependent biofilm formation while still reducing bacterial CFU in biofilms. Docosahexaenoic acid can be considered an option as a therapeutic substance against biofilm formation and may be a good alternative in reducing the risk of MRSA formation.
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O’Keeffe, Triona, Colin Hill i R. Paul Ross. "Characterization and Heterologous Expression of the Genes Encoding Enterocin A Production, Immunity, and Regulation inEnterococcus faecium DPC1146". Applied and Environmental Microbiology 65, nr 4 (1.04.1999): 1506–15. http://dx.doi.org/10.1128/aem.65.4.1506-1515.1999.
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ABSTRACT Enterocin A is a small, heat-stable, antilisterial bacteriocin produced by Enterococcus faecium DPC1146. The sequence of a 10,879-bp chromosomal region containing at least 12 open reading frames (ORFs), 7 of which are predicted to play a role in enterocin biosynthesis, is presented. The genes entA,entI, and entF encode the enterocin A prepeptide, the putative immunity protein, and the induction factor prepeptide, respectively. The deduced proteins EntK and EntR resemble the histidine kinase and response regulator proteins of two-component signal transducing systems of the AgrC-AgrA type. The predicted proteins EntT and EntD are homologous to ABC (ATP-binding cassette) transporters and accessory factors, respectively, of several other bacteriocin systems and to proteins implicated in the signal-sequence-independent export of Escherichia colihemolysin A. Immediately downstream of the entT andentD genes are two ORFs, the product of one of which, ORF4, is very similar to the product of the yteI gene ofBacillus subtilis and to E. coli protease IV, a signal peptide peptidase known to be involved in outer membrane lipoprotein export. Another potential bacteriocin is encoded in the opposite direction to the other genes in the enterocin cluster. This putative bacteriocin-like peptide is similar to LafX, one of the components of the lactacin F complex. A deletion which included one of two direct repeats upstream of the entA gene abolished enterocin A activity, immunity, and ability to induce bacteriocin production. Transposon insertion upstream of the entF gene also had the same effect, but this mutant could be complemented by exogenously supplied induction factor. The putative EntI peptide was shown to be involved in the immunity to enterocin A. Cloning of a 10.5-kb amplicon comprising all predicted ORFs and regulatory regions resulted in heterologous production of enterocin A and induction factor in Enterococcus faecalis, while a four-gene construct (entAITD) under the control of a constitutive promoter resulted in heterologous enterocin A production in both E. faecalis and Lactococcus lactis.
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Sloan, Tim J., Ewan Murray, Maho Yokoyama, Ruth C. Massey, Weng C. Chan, Boyan B. Bonev i Paul Williams. "Timing Is Everything: Impact of Naturally Occurring Staphylococcus aureus AgrC Cytoplasmic Domain Adaptive Mutations on Autoinduction". Journal of Bacteriology 201, nr 20 (29.07.2019). http://dx.doi.org/10.1128/jb.00409-19.
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ABSTRACT Mutations in the polymorphic Staphylococcus aureus agr locus responsible for quorum sensing (QS)-dependent virulence gene regulation occur frequently during host adaptation. In two genomically closely related S. aureus clinical isolates exhibiting marked differences in Panton-Valentine leukocidin production, a mutation conferring an N267I substitution was identified in the cytoplasmic domain of the QS sensor kinase, AgrC. This natural mutation delayed the onset and accumulation of autoinducing peptide (AIP) and showed reduced responsiveness to exogenous AIPs. Other S. aureus strains harboring naturally occurring AgrC cytoplasmic domain mutations were identified, including T247I, I311T, A343T, L245S, and F264C. These mutations were associated with reduced cytotoxicity, delayed/reduced AIP production, and impaired sensitivity to exogenous AIP. Molecular dynamics simulations were used to model the AgrC cytoplasmic domain conformational changes arising. Although mutations were localized in different parts of the C-terminal domain, their impact on molecular structure was manifested by twisting of the leading helical hairpin α1-α2, accompanied by repositioning of the H-box and G-box, along with closure of the flexible loop connecting the two and occlusion of the ATP-binding site. Such conformational rearrangements of key functional subdomains in these mutants highlight the cooperative response of molecular structure involving dimerization and ATP binding and phosphorylation, as well as the binding site for the downstream response element AgrA. These appear to increase the threshold for agr activation via AIP-dependent autoinduction, thus reducing virulence and maintaining S. aureus in an agr-downregulated “colonization” mode. IMPORTANCE Virulence factor expression in Staphylococcus aureus is regulated via autoinducing peptide (AIP)-dependent activation of the sensor kinase AgrC, which forms an integral part of the agr quorum sensing system. In response to bound AIP, the cytoplasmic domain of AgrC (AgrC-cyt) undergoes conformational changes resulting in dimerization, autophosphorylation, and phosphotransfer to the response regulator AgrA. Naturally occurring mutations in AgrC-cyt are consistent with repositioning of key functional domains, impairing dimerization and restricting access to the ATP-binding pocket. Strains harboring specific AgrC-cyt mutations exhibit reduced AIP autoinduction efficiency and a timing-dependent attenuation of cytotoxicity which may confer a survival advantage during established infection by promoting colonization while restricting unnecessary overproduction of exotoxins.
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Williams, Paul, Phil Hill, Boyan Bonev i Weng C. Chan. "Quorum-sensing, intra- and inter-species competition in the staphylococci". Microbiology 169, nr 8 (3.08.2023). http://dx.doi.org/10.1099/mic.0.001381.
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In Gram-positive bacteria such as Staphylococcus aureus and the coagulase-negative staphylococci (CoNS), the accessory gene regulator (agr) is a highly conserved but polymorphic quorum-sensing system involved in colonization, virulence and biofilm development. Signalling via agr depends on the interaction of an autoinducing peptide (AIP) with AgrC, a transmembrane sensor kinase that, once phosphorylated activates the response regulator AgrA. This in turn autoinduces AIP biosynthesis and drives target gene expression directly via AgrA or via the post-transcriptional regulator, RNAIII. In this review we describe the molecular mechanisms underlying the agr-mediated generation of, and response to, AIPs and the molecular basis of AIP-dependent activation and inhibition of AgrC. How the environment impacts on agr functionality is considered and the consequences of agr dysfunction for infection explored. We also discuss the concept of AIP-driven competitive interference between S. aureus and the CoNS and its anti-infective potential.
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Cosgriff, Chance J., Chelsea R. White, Wei Ping Teoh, James P. Grayczyk i Francis Alonzo. "Control ofStaphylococcus aureusQuorum Sensing by a Membrane-Embedded Peptidase". Infection and Immunity 87, nr 5 (4.03.2019). http://dx.doi.org/10.1128/iai.00019-19.
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ABSTRACTGram-positive bacteria process and release small peptides, or pheromones, that act as signals for the induction of adaptive traits, including those involved in pathogenesis. One class of small signaling pheromones is the cyclic autoinducing peptides (AIPs), which regulate expression of genes that orchestrate virulence and persistence in a range of microbes, including staphylococci, listeriae, clostridia, and enterococci. In a genetic screen forStaphylococcus aureussecreted virulence factors, we identified anS. aureusmutant containing an insertion in the geneSAUSA300_1984(mroQ), which encodes a putative membrane-embedded metalloprotease. A ΔmroQmutant exhibited impaired induction of Toll-like receptor 2-dependent inflammatory responses from macrophages but elicited greater production of the inflammatory cytokine interleukin-1β and was attenuated in a murine skin and soft tissue infection model. The ΔmroQmutant phenocopies anS. aureusmutant containing a deletion of the accessory gene regulatory system (Agr), wherein both strains have significantly reduced production of secreted toxins and virulence factors but increased surface protein A abundance. The Agr system controls virulence factor gene expression inS. aureusby sensing the accumulation of AIP via the histidine kinase AgrC and the response regulator AgrA. We provide evidence to suggest that MroQ acts within the Agr pathway to facilitate the optimal processing or export of AIP for signal amplification through AgrC/A and induction of virulence factor gene expression. Mutation of MroQ active-site residues significantly reduces AIP signaling and attenuates virulence. Altogether, this work identifies a new component of the Agr quorum-sensing circuit that is critical for the production ofS. aureusvirulence factors.
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Liu, Yunge, Lina Wu, Jina Han, Pengcheng Dong, Xin Luo, Yimin Zhang i Lixian Zhu. "Inhibition of Biofilm Formation and Related Gene Expression of Listeria monocytogenes in Response to Four Natural Antimicrobial Compounds and Sodium Hypochlorite". Frontiers in Microbiology 11 (14.01.2021). http://dx.doi.org/10.3389/fmicb.2020.617473.
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The aim of this study was to assess the efficacy of four natural antimicrobial compounds (cinnamaldehyde, eugenol, resveratrol and thymoquinone) plus a control chemical disinfectant (sodium hypochlorite) in inhibiting biofilm formation by Listeria monocytogenes CMCC54004 (Lm 54004) at a minimum inhibitory concentration (MIC) and sub-MICs. Crystal violet staining assay and microscopic examination were employed to investigate anti-biofilm effects of the evaluated compounds, and a real-time PCR assay was used to investigate the expression of critical genes by Lm 54004 biofilm. The results showed that five antimicrobial compounds inhibited Lm 54004 biofilm formation in a dose dependent way. Specifically, cinnamaldehyde and resveratrol showed better anti-biofilm effects at 1/4 × MIC, while sodium hypochlorite exhibited the lowest inhibitory rates. A swimming assay confirmed that natural compounds at sub-MICs suppressed Lm 54004 motility to a low degree. Supporting these findings, expression analysis showed that all four natural compounds at 1/4 × MIC significantly down-regulated quorum sensing genes (agrA, agrC, and agrD) rather than suppressing the motility- and flagella-associated genes (degU, motB, and flaA). This study revealed that sub-MICs of natural antimicrobial compounds reduced biofilm formation by suppressing the quorum sensing system rather than by inhibiting flagella formation.
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Saadati, Fatemeh, Shahab Shahryari, Naeema Mohseni Sani, Davoud Farajzadeh, Hossein Shahbani Zahiri, Hojatollah Vali i Kambiz Akbari Noghabi. "Effect of MA01 rhamnolipid on cell viability and expression of quorum-sensing (QS) genes involved in biofilm formation by methicillin-resistant Staphylococcus aureus". Scientific Reports 12, nr 1 (1.09.2022). http://dx.doi.org/10.1038/s41598-022-19103-w.
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AbstractA group of biosurfactants, called rhamnolipids, have been shown to have antibacterial and antibiofilm activity against multidrug-resistant bacteria. Here, we examined the effect of rhamnolipid biosurfactants extracted from Pseudomonas aeruginosa MA01 on cell growth/viability, biofilm formation, and membrane permeability of methicillin-resistant Staphylococcus aureus (MRSA) ATCC6538 bacterial cells. The results obtained from flow cytometry analysis showed that by increasing the concentration of rhamnolipid from 30 to 120 mg/mL, the cell viability decreased by about 70%, and the cell membrane permeability increased by approximately 20%. In fact, increasing rhamnolipid concentration was directly related to cell membrane permeability and inversely related to cell survival. Microtiter plate biofilm assay and laser scanning confocal microscopy analysis revealed that rhamnolipid, at a concentration of 60 mg/mL, exerts a reducing effect on the biofilm formation of Staphylococcus aureus. Real-time PCR analysis for monitoring the relative changes in the expression of agrA, agrC, icaA, and icaD genes involved in biofilm formation and related to the quorum-sensing pathway after treatment with rhamnolipid indicated a reduced expression level of these genes, as well as sortase A gene. The results of the present study deepen our knowledge regarding the use of microbial natural products as promising candidates for therapeutic applications.
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Huber, Charlotte, Ivonne Stamm, Wilma Ziebuhr, Gabriella Marincola, Markus Bischoff, Birgit Strommenger, Greta Jaschkowitz i in. "Silence as a way of niche adaptation: mecC-MRSA with variations in the accessory gene regulator (agr) functionality express kaleidoscopic phenotypes". Scientific Reports 10, nr 1 (8.09.2020). http://dx.doi.org/10.1038/s41598-020-71640-4.
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Abstract Functionality of the accessory gene regulator (agr) quorum sensing system is an important factor promoting either acute or chronic infections by the notorious opportunistic human and veterinary pathogen Staphylococcus aureus. Spontaneous alterations of the agr system are known to frequently occur in human healthcare-associated S. aureus lineages. However, data on agr integrity and function are sparse regarding other major clonal lineages. Here we report on the agr system functionality and activity level in mecC-carrying methicillin resistant S. aureus (MRSA) of various animal origins (n = 33) obtained in Europe as well as in closely related human isolates (n = 12). Whole genome analysis assigned all isolates to four clonal complexes (CC) with distinct agr types (CC599 agr I, CC49 agr II, CC130 agr III and CC1943 agr IV). Agr functionality was assessed by a combination of phenotypic assays and proteome analysis. In each CC, isolates with varying agr activity levels were detected, including the presence of completely non-functional variants. Genomic comparison of the agr I–IV encoding regions associated these phenotypic differences with variations in the agrA and agrC genes. The genomic changes were detected independently in divergent lineages, suggesting that agr variation might foster viability and adaptation of emerging MRSA lineages to distinct ecological niches.
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Valliammai, Alaguvel, Sivasamy Sethupathy, Arumugam Priya, Anthonymuthu Selvaraj, James Prabhanand Bhaskar, Venkateswaran Krishnan i Shunmugiah Karutha Pandian. "5-Dodecanolide interferes with biofilm formation and reduces the virulence of Methicillin-resistant Staphylococcus aureus (MRSA) through up regulation of agr system". Scientific Reports 9, nr 1 (24.09.2019). http://dx.doi.org/10.1038/s41598-019-50207-y.
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Abstract Methicillin resistant Staphylococcus aureus (MRSA) is a predominant human pathogen with high morbidity that is listed in the WHO high priority pathogen list. Being a primary cause of persistent human infections, biofilm forming ability of S. aureus plays a pivotal role in the development of antibiotic resistance. Hence, targeting biofilm is an alternative strategy to fight bacterial infections. The present study for the first time demonstrates the non-antibacterial biofilm inhibitory efficacy of 5-Dodecanolide (DD) against ATCC strain and clinical isolates of S. aureus. In addition, DD is able to inhibit adherence of MRSA on human plasma coated Titanium surface. Further, treatment with DD significantly reduced the eDNA synthesis, autoaggregation, staphyloxanthin biosynthesis and ring biofilm formation. Reduction in staphyloxanthin in turn increased the susceptibility of MRSA to healthy human blood and H2O2 exposure. Quantitative PCR analysis revealed the induced expression of agrA and agrC upon DD treatment. This resulted down regulation of genes involved in biofilm formation such as fnbA and fnbB and up regulation of RNAIII, hld, psmα and genes involved in biofilm matrix degradation such as aur and nuc. Inefficacy of DD on the biofilm formation of agr mutant further validated the agr mediated antibiofilm potential of DD. Notably, DD was efficient in reducing the in vivo colonization of MRSA in Caenorhabditis elegans. Results of gene expression studies and physiological assays unveiled the agr mediated antibiofilm efficacy of DD.
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Wang, Hongyan, Yiyi Shi, Junwen Chen, Yu Wang, Zhanwen Wang, Zhijian Yu, Jinxin Zheng i Yongpeng Shang. "The antiviral drug efavirenz reduces biofilm formation and hemolysis by Staphylococcus aureus". Journal of Medical Microbiology 70, nr 10 (20.10.2021). http://dx.doi.org/10.1099/jmm.0.001433.
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Introduction. Biofilm formation and hemolysis are closely related to the pathogenicity of Staphylococcus aureus . Hypothesis/Gap Statement. Strategies that reduce the mortality of S. aureus infections may involve novel antimicrobials and/or drugs that decrease S. aureus virulence, such as biofilm formation. The antiviral drug efavirenz is a non-nucleoside reverse transcriptase inhibitor, which also has shown antibacterial effect on Bacillus subtilis and Escherichia coli . Its effect on pathogen virulence has not yet been explored. Aim. This study investigates the antimicrobial and anti-virulence effect of efavirenz on S. aureus . Methodology. Biofilm biomasses were detected by crystal violet staining. Hemolysis activities of S. aureus were determined by rabbit erythrocytes lysis assay. RNA levels of transcriptional regulatory genes, biofilm-related genes, and virulence-related genes of S. aureus were determined by RT-qPCR. Results. Efavirenz showed an inhibitory effect on the growth of S. aureus , Enterococcus faecalis and Streptococcus agalactiae at 50 µM. Efavirenz significantly inhibited biofilm formation of both methicillin-sensitive S. aureus (MSSA) and methicillin-resistant S. aureus (MRSA) at 25 µM, but did not affect the growth of planktonic S. aureus cells. Moreover, hemolysis by S. aureus was inhibited by efavirenz at 25 µM. The expression levels of RNA transcriptional regulatory genes (agrA, agrC, sigB, saeR and saeS), biofilm-related genes (cidA, clfA, clfB, fnbA, fnbB), and virulence-related genes (hla, hld, staphopain B, alpha-3 PSM, beta PSM, delta PSM) of S. aureus decreased significantly at 25 µM efavirenz. Conclusion. Efavirenz inhibits S. aureus biofilm formation and virulence in vitro.