scholarly journals Prevalence of Agr phase variants in Staphylococcus aureus

2020 ◽  
Vol 2 (7A) ◽  
Author(s):  
Vishal Gor ◽  
Mitsuaki Hoshi ◽  
Aya Takemura ◽  
Masato Higashide ◽  
Veronica Romero ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Quorum Sensing ◽  
Virulence Factors ◽  
Human Pathogen ◽  
Accessory Gene ◽  
Accessory Gene Regulator ◽  
Inversion Event ◽  
First Time ◽  
Phase Variants ◽  
Phenotypic Reversion

Staphylococcus aureus is an important human pathogen whose success is largely attributed to its vast arsenal of virulence factors that facilitate its invasion into, and survival within, the human host. The expression of these virulence factors is controlled by the quorum sensing Accessory Gene Regulator (Agr) system. However, a large proportion of clinical S. aureus isolates are consistently found to have a mutationally inactivated Agr system. These mutants have a survival advantage in the host but are considered irreversible mutants. Here we show, for the first time, that a fraction of Agr-negative mutants can revert their Agr activity. By serially passaging Agr negative strains and screening for phenotypic reversion of haemolysis and subsequent sequencing, we identified two mutational events responsible for reversion: a genetic duplication plus inversion event and a poly(A) tract alteration. Additionally, we demonstrate that one clinical Agr-negative MRSA isolate could reproducibly generate Agr-revertant colonies with a poly(A) tract genetic mechanism. We also show that these revertants activate their Agr system upon phagocytosis. To assess the significance of our findings we screened a series of primary clinical isolates, which had undergone minimal handling post-isolation, and successfully identified a fraction which were Agr phase variants. Taken together, we propose a model where some Agr-negative S. aureus strains are phase variants who can revert their Agr activity and may act as a cryptic insurance strategy against host-mediated stress.

scholarly journals Finding of Agr Phase Variants in Staphylococcus aureus

mBio ◽  
2019 ◽  
Vol 10 (4) ◽  
Author(s):  
Vishal Gor ◽  
Aya J. Takemura ◽  
Masami Nishitani ◽  
Masato Higashide ◽  
Veronica Medrano Romero ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Virulence Factors ◽  
Phase Variation ◽  
Accessory Gene ◽  
Content Type ◽  
Accessory Gene Regulator ◽  
Immune Mediated ◽  
A Minor ◽  
First Time ◽  
Phase Variants

ABSTRACT Staphylococcus aureus is an important human pathogen whose success is largely attributed to its vast arsenal of virulence factors that facilitate its invasion into, and survival within, the human host. The expression of these virulence factors is controlled by the quorum sensing accessory gene regulator (Agr) system. However, a large proportion of clinical S. aureus isolates are consistently found to have a mutationally inactivated Agr system. These mutants have a survival advantage in the host but are considered irreversible mutants. Here we show, for the first time, that a fraction of Agr-negative mutants can revert their Agr activity. By serially passaging Agr-negative strains and screening for phenotypic reversion of hemolysis and subsequent sequencing, we identified two mutational events responsible for reversion: a genetic duplication plus inversion event and a poly(A) tract alteration. Additionally, we demonstrate that one clinical Agr-negative methicillin-resistant S. aureus (MRSA) isolate could reproducibly generate Agr-revertant colonies with a poly(A) tract genetic mechanism. We also show that these revertants activate their Agr system upon phagocytosis. We propose a model in which a minor fraction of Agr-negative S. aureus strains are phase variants that can revert their Agr activity and may act as a cryptic insurance strategy against host-mediated stress. IMPORTANCE Staphylococcus aureus is responsible for a broad range of infections. This pathogen has a vast arsenal of virulence factors at its disposal, but avirulent strains are frequently isolated as the cause of clinical infections. These isolates have a mutated agr locus and have been believed to have no evolutionary future. Here we show that a fraction of Agr-negative strains can repair their mutated agr locus with mechanisms resembling phase variation. The agr revertants sustain an Agr OFF state as long as they exist as a minority but can activate their Agr system upon phagocytosis. These revertant cells might function as a cryptic insurance strategy to survive immune-mediated host stress that arises during infection.

What role does the quorum-sensing accessory gene regulator system play during Staphylococcus aureus bacteremia?

2014 ◽  
Vol 22 (12) ◽  
pp. 676-685 ◽  
Author(s):  
Kimberley L. Painter ◽  
Aishwarya Krishna ◽  
Sivaramesh Wigneshweraraj ◽  
Andrew M. Edwards
Keyword(s):  
Staphylococcus Aureus ◽  
Quorum Sensing ◽  
Accessory Gene ◽  
Accessory Gene Regulator ◽  
Staphylococcus Aureus Bacteremia

scholarly journals A Novel Aza-Derivative Inhibits agr Quorum Sensing Signaling and Synergizes Methicillin-Resistant Staphylococcus aureus to Clindamycin

2021 ◽  
Vol 12 ◽  
Author(s):  
Giulia Bernabè ◽  
Matteo Dal Pra ◽  
Vittoria Ronca ◽  
Anthony Pauletto ◽  
Giovanni Marzaro ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Quorum Sensing ◽  
Virulence Factors ◽  
Response Regulator ◽  
Accessory Gene ◽  
Mobility Shift ◽  
Signaling Mechanism ◽  
Methicillin Resistant ◽  
Qrt Pcr ◽  
Mobility Shift Assay

Increasing antibiotic resistance and diminishing pharmaceutical industry investments have increased the need for molecules that can treat infections caused by dangerous pathogens such as methicillin-resistant Staphylococcus aureus (MRSA). Quorum Sensing (QS) is a signaling mechanism that regulates bacterial virulence in pathogens. A report demonstrating that the anti-inflammatory drug Diflunisal reduces MRSA virulence factors’ expression prompted us to design, synthesize and test 16 aza-analogs as inhibitors of S. aureus virulence factors controlled by the accessory gene regulator (agr) QS system. At first, we evaluated by qRT-PCR the activity of compounds on rnaIII expression, a QS related gene. Azan-7 was the most active molecule tested and it did not show cytotoxic activity in human cell lines. Moreover, we demonstrated that it did not affect bacterial proliferation. Regulation of MRSA virulence genes by Azan-7 was investigated using qRT-PCR and RNAseq. Azan-7 significantly reduced hla, psmα, hysA, agrA, cap1A, and cap1C gene expression. In silico docking demonstrated that Azan-7 binds the response regulator AgrA. This data was confirmed by electrophoretic mobility shift assay (EMSA) reporting that Azan-7 binding to AgrA protein strongly reduced the AgrA-DNA complex formation at the P3 promoter region involved in the regulation of rnaIII transcription. Azan-7 inhibited MRSA-mediated haemolysis, reduced survival of the pathogen at low pH levels, and increased macrophage killing. In addition, Azan-7 enhanced MRSA susceptibility to clindamycin both in planktonic growth and biofilm. Azan-7 did not induce resistance over 10 days in culture. It was equally active against all the AgrA MRSA subtypes encountered among clinical isolates, but it was not active against Staphylococcus epidermidis, although the AgrA proteins show an approximate 80% homology. These results demonstrate that Azan-7 inhibits the expression of MRSA virulence factors by interfering in the QS and synergizes MRSA biofilm with clindamycin, indicating the compound as a promising candidate for the treatment of MRSA infections.

scholarly journals Generation of Virulence Factor Variants in Staphylococcus aureus Biofilms

2007 ◽  
Vol 189 (22) ◽  
pp. 7961-7967 ◽  
Author(s):  
Jeremy M. Yarwood ◽  
Kara M. Paquette ◽  
Ilya B. Tikh ◽  
Esther M. Volper ◽  
E. Peter Greenberg
Keyword(s):  
Staphylococcus Aureus ◽  
Virulence Factors ◽  
Hemolytic Activity ◽  
Bacterial Species ◽  
Phase Variation ◽  
Accessory Gene ◽  
Altered Expression ◽  
Sensing System ◽  
Accessory Gene Regulator ◽  
Different Strains

ABSTRACT Several serious diseases are caused by biofilm-associated Staphylococcus aureus. Colonial variants occur in biofilms of other bacterial species, and S. aureus variants are frequently isolated from biofilm-associated infections. Thus, we studied the generation of variants with altered expression of virulence factors in S. aureus biofilms. We observed that the number of variants found in biofilms, as measured by hemolytic activity, varied for different strains. Further study of hemolytic activity and signaling by the accessory gene regulator (Agr) quorum-sensing system in one S. aureus strain revealed three primary biofilm subpopulations: nonhemolytic (Agr deficient), hemolytic (Agr positive), and hyperhemolytic (also Agr positive). The nonhemolytic variant became the numerically dominant subpopulation in the biofilm. The nonhemolytic variant phenotype was stable and heritable, indicating a genetic perturbation, whereas the hyperhemolytic phenotype was unstable, suggesting a phase variation. Transcription profiling revealed that expression of the agr locus and many extracellular virulence factors was repressed in the nonhemolytic variant. Expression of the agr-activating gene, sarU, was also repressed in the nonhemolytic variant, suggesting one potential regulatory pathway responsible for the Agr-deficient phenotype. We suggest that the development of these variants in biofilms may have important clinical implications.

scholarly journals Virulence Reversion in Staphylococcus aureus

Proceedings ◽  
2021 ◽  
Vol 66 (1) ◽  
pp. 24
Author(s):  
Vishal Gor ◽  
Mitsuaki Hoshi ◽  
Aya J. Takemura ◽  
Masato Higashide ◽  
Veronica Medrano Romero ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Regulatory System ◽  
Opportunistic Pathogen ◽  
Accessory Gene ◽  
Genetic Changes ◽  
Accessory Gene Regulator ◽  
Heavy Burden ◽  
The Common ◽  
Phase Variants ◽  
Inactivating Mutations

Staphylococcus aureus is a Gram-positive opportunistic pathogen that imposes a heavy burden on society. What sets this pathogen apart is the sheer spectrum of infections it can cause, which range from benign skin and soft tissue infections to lethal endocarditis and bacteraemia. The ability of S. aureus to cause this gamut of infections is conferred by its arsenal of virulence factors that are under the control of the Accessory Gene Regulator (Agr) system. However, a large proportion of clinical isolates have inactivating mutations in this important regulatory system. We previously showed that, contrary to the common dogma, not all these mutations are evolutionary ‘dead-ends’ and a fraction are phase variants which can revert to an Agr active state. Here we report that some Agr deficient isolates can revert a haemolytic phenotype without repairing their Agr system. We collected a series of 30 Agr negative primary patient samples in order to assess the significance of our previous findings on the existence of Agr phase variants. We used primary samples to avoid strains that had undergone multiple clonal expansions before being tested for reversibility. We assessed Agr reversibility by serially passaging strains and screening for phenotypic reversion of haemolysis. We show that two strains reverted haemolysis and one reverted alpha haemolysin activity without any genetic changes in agr (and hla for the alpha revertant). These results add further complexity to the phenomenon of Agr shutdown observed in the clinical setting and corroborate recent findings of compensatory mutations arising in Agr deficient clinical strains.

scholarly journals Control of Staphylococcus aureus quorum sensing by a membrane-embedded peptidase

2019 ◽  
Author(s):  
Chance J. Cosgriff ◽  
Chelsea R. White ◽  
Wei Ping Teoh ◽  
James P. Grayczyk ◽  
Francis Alonzo
Keyword(s):  
Gene Expression ◽  
Staphylococcus Aureus ◽  
Quorum Sensing ◽  
Virulence Factors ◽  
Virulence Factor ◽  
Inflammatory Responses ◽  
Infection Model ◽  
Accessory Gene ◽  
Factor Gene ◽  
Virulence Factor Gene

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 auto-inducing peptides (AIPs), which regulate expression of genes that orchestrate virulence and persistence in a range of microbes including Staphylococci, Listeria, Clostridia, and Enterococci. In a genetic screen for Staphylococcus aureus secreted virulence factors, we identified a S. aureus mutant containing an insertion in gene SAUSA300_1984 (mroQ), which encodes a putative membrane-embedded metalloprotease. A ΔmroQ mutant exhibits impaired induction of TLR2-dependent inflammatory responses from macrophages, but elicits greater production of the inflammatory cytokine IL-1β and is attenuated in a murine skin and soft tissue infection model. The ΔmroQ mutant phenocopies a S. aureus mutant 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 in S. aureus through sensing accumulation of AIP via the histidine kinase AgrC and response regulator AgrA. We provide evidence to suggest that MroQ acts within the Agr pathway to facilitate 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 of S. aureus virulence factors.

scholarly journals Quorum Sensing in Staphylococcus aureus Biofilms

2004 ◽  
Vol 186 (6) ◽  
pp. 1838-1850 ◽  
Author(s):  
Jeremy M. Yarwood ◽  
Douglas J. Bartels ◽  
Esther M. Volper ◽  
E. Peter Greenberg
Keyword(s):  
Staphylococcus Aureus ◽  
Quorum Sensing ◽  
Biofilm Formation ◽  
Time Lapse ◽  
Confocal Scanning Laser Microscopy ◽  
Accessory Gene ◽  
Fluorescent Reporter ◽  
Accessory Gene Regulator ◽  
Confocal Scanning Laser ◽  
Biofilm Development

ABSTRACT Several serious diseases are caused by biofilm-associated Staphylococcus aureus, infections in which the accessory gene regulator (agr) quorum-sensing system is thought to play an important role. We studied the contribution of agr to biofilm development, and we examined agr-dependent transcription in biofilms. Under some conditions, disruption of agr expression had no discernible influence on biofilm formation, while under others it either inhibited or enhanced biofilm formation. Under those conditions where agr expression enhanced biofilm formation, biofilms of an agr signaling mutant were particularly sensitive to rifampin but not to oxacillin. Time lapse confocal scanning laser microscopy showed that, similar to the expression of an agr-independent fluorescent reporter, biofilm expression of an agr-dependent reporter was in patches within cell clusters and oscillated with time. In some cases, loss of fluorescence appeared to coincide with detachment of cells from the biofilm. Our studies indicate that the role of agr expression in biofilm development and behavior depends on environmental conditions. We also suggest that detachment of cells expressing agr from biofilms may have important clinical implications.

scholarly journals Quorum Sensing and Toxin Production in Staphylococcus aureus Osteomyelitis: Pathogenesis and Paradox

Toxins ◽  
2020 ◽  
Vol 12 (8) ◽  
pp. 516
Author(s):  
Casey E. Butrico ◽  
James E. Cassat
Keyword(s):  
Staphylococcus Aureus ◽  
Quorum Sensing ◽  
Toxin Production ◽  
High Morbidity ◽  
Invasive Infection ◽  
Accessory Gene ◽  
Accessory Gene Regulator ◽  
Production Factors ◽  
Regulatory Systems ◽  
Skeletal Homeostasis

Staphylococcus aureus is a Gram-positive pathogen capable of infecting nearly every vertebrate organ. Among these tissues, invasive infection of bone (osteomyelitis) is particularly common and induces high morbidity. Treatment of osteomyelitis is notoriously difficult and often requires debridement of diseased bone in conjunction with prolonged antibiotic treatment to resolve infection. During osteomyelitis, S. aureus forms characteristic multicellular microcolonies in distinct niches within bone. Virulence and metabolic responses within these multicellular microcolonies are coordinated, in part, by quorum sensing via the accessory gene regulator (agr) locus, which allows staphylococcal populations to produce toxins and adapt in response to bacterial density. During osteomyelitis, the Agr system significantly contributes to dysregulation of skeletal homeostasis and disease severity but may also paradoxically inhibit persistence in the host. Moreover, the Agr system is subject to complex crosstalk with other S. aureus regulatory systems, including SaeRS and SrrAB, which can significantly impact the progression of osteomyelitis. The objective of this review is to highlight Agr regulation, its implications on toxin production, factors that affect Agr activation, and the potential paradoxical influences of Agr regulation on disease progression during osteomyelitis.

scholarly journals Identification and characterization of msa (SA1233), a gene involved in expression of SarA and several virulence factors in Staphylococcus aureus

Microbiology ◽  
2006 ◽  
Vol 152 (9) ◽  
pp. 2559-2572 ◽  
Author(s):  
Karthik Sambanthamoorthy ◽  
Mark S. Smeltzer ◽  
Mohamed O. Elasri
Keyword(s):  
Staphylococcus Aureus ◽  
Virulence Factors ◽  
Protein A ◽  
Laboratory Strain ◽  
Reporter System ◽  
Accessory Gene ◽  
Accessory Gene Regulator ◽  
Genes Encoding ◽  
Membrane Spanning ◽  
The Impact

The staphylococcal accessory regulator (sarA) plays a central role in the regulation of virulence in Staphylococcus aureus. To date, studies involving sarA have focused on its activity as a global regulator that modulates transcription of a wide variety of genes (>100) and its role in virulence. However, there is also evidence to suggest the existence of accessory elements that modulate SarA production and/or function. A reporter system was developed to identify such elements, and a new gene, msa (SA1233), mutation of which results in reduced expression of SarA, was identified and characterized. Additionally, it was shown that mutation of msa resulted in altered transcription of the accessory gene regulator (agr) and the genes encoding several virulence factors including alpha toxin (hla) and protein A (spa). However, the impact of mutating msa was different in the laboratory strain RN6390 and the clinical isolate UAMS-1. For instance, mutation of msa caused a decrease in spa and hla transcription in RN6390 but had a different effect in UAMS-1. The strain-dependent effects of the msa mutation were similar to those observed previously, which suggests that msa may modulate the production of specific virulence factors through its impact on sarA. Interestingly, sequence analysis of Msa suggests that it is a putative membrane protein with three membrane-spanning regions, indicating that Msa might interact with the environment. The findings show that msa is involved in the expression of SarA and several virulence factors.

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