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5-Fluorouracil blocks quorum-sensing of biofilm-embedded methicillin-resistant Staphylococcus aureus in mice.

ABSTRACT: Antibiotic-resistant pathogens often escape antimicrobial treatment by forming protective biofilms in response to quorum-sensing communication via diffusible autoinducers. Biofilm formation by the nosocomial pathogen methicillin-resistant Staphylococcus aureus (MRSA) is triggered by the quorum-sensor autoinducer-2 (AI-2), whose biosynthesis is mediated by methylthioadenosine/S-adenosylhomocysteine nucleosidase (MTAN) and S-ribosylhomocysteine lyase (LuxS). Here, we present a high-throughput screening platform for small-molecular inhibitors of either enzyme. This platform employs a cell-based assay to report non-toxic, bioavailable and cell-penetrating inhibitors of AI-2 production, utilizing engineered human cells programmed to constitutively secrete AI-2 by tapping into the endogenous methylation cycle via ectopic expression of codon-optimized MTAN and LuxS. Screening of a library of over 5000 commercial compounds yielded 66 hits, including the FDA-licensed cytostatic anti-cancer drug 5-fluorouracil (5-FU). Secondary screening and validation studies showed that 5-FU is a potent quorum-quencher, inhibiting AI-2 production and release by MRSA, Staphylococcus epidermidis, Escherichia coli and Vibrio harveyi. 5-FU efficiently reduced adherence and blocked biofilm formation of MRSA in vitro at an order-of-magnitude-lower concentration than that clinically relevant for anti-cancer therapy. Furthermore, 5-FU reestablished antibiotic susceptibility and enabled daptomycin-mediated prevention and clearance of MRSA infection in a mouse model of human implant-associated infection.

SUBMITTER: Sedlmayer F

PROVIDER: S-EPMC8287944 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Novel Peptide from Commensal Staphylococcus simulans Blocks Methicillin-Resistant Staphylococcus aureus Quorum Sensing and Protects Host Skin from Damage.

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Control of <i>Staphylococcus aureus</i> Quorum Sensing by a Membrane-Embedded Peptidase.

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| S-EPMC6479040 | biostudies-literature

Nuclease modulates biofilm formation in community-associated methicillin-resistant Staphylococcus aureus.

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| S-EPMC3214024 | biostudies-literature

Salicylic acid stabilizes Staphylococcus aureus biofilm by impairing the agr quorum-sensing system.

Project description:Salicylic acid (SAL) has recently been shown to induce biofilm formation in Staphylococcus aureus and to affect the expression of virulence factors. This study was aimed to investigate the effect of SAL on the regulatory agr system and its impact on S. aureus biofilm formation. The agr quorum-sensing system, which is a central regulator in S. aureus pathogenicity, plays a pivotal role in the dispersal of S. aureus mature biofilms and contributes to the creation of new colonization sites. Here, we demonstrate that SAL impairs biofilm dispersal by interfering with agr expression. As revealed by our work, protease and surfactant molecule production is diminished, and bacterial cell autolysis is also negatively affected by SAL. Furthermore, as a consequence of SAL treatment, the S. aureus biofilm matrix revealed the lack of extracellular DNA. In silico docking and simulation of molecular dynamics provided evidence for a potential interaction of AgrA and SAL, resulting in reduced activity of the agr system. In conclusion, SAL stabilized the mature S. aureus biofilms, which may prevent bacterial cell dissemination. However, it may foster the establishment of infections locally and consequently increase bacterial persistence leading to therapeutic failure.

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Genomic analyses of two novel biofilm-degrading methicillin-resistant Staphylococcus aureus phages.

Project description:BackgroundMethicillin-resistant Staphylococcus aureus (MRSA) biofilm producers represent an important etiological agent of many chronic human infections. Antibiotics and host immune responses are largely ineffective against bacteria within biofilms. Alternative actions and novel antimicrobials should be considered. In this context, the use of phages to destroy MRSA biofilms presents an innovative alternative mechanism.ResultsTwenty-five MRSA biofilm producers were used as substrates to isolate MRSA-specific phages. Despite the difficulties in obtaining an isolate of this phage, two phages (UPMK_1 and UPMK_2) were isolated. Both phages varied in their ability to produce halos around their plaques, host infectivity, one-step growth curves, and electron microscopy features. Furthermore, both phages demonstrated antagonistic infectivity on planktonic cultures. This was validated in an in vitro static biofilm assay (in microtiter-plates), followed by the visualization of the biofilm architecture in situ via confocal laser scanning microscopy before and after phage infection, and further supported by phages genome analysis. The UPMK_1 genome comprised 152,788 bp coding for 155 putative open reading frames (ORFs), and its genome characteristics were between the Myoviridae and Siphoviridae family, though the morphological features confined it more to the Siphoviridae family. The UPMK_2 has 40,955 bp with 62 putative ORFs; morphologically, it presented the features of the Podoviridae though its genome did not show similarity with any of the S. aureus in the Podoviridae family. Both phages possess lytic enzymes that were associated with a high ability to degrade biofilms as shown in the microtiter plate and CLSM analyses.ConclusionsThe present work addressed the possibility of using phages as potential biocontrol agents for biofilm-producing MRSA.

| S-EPMC6540549 | biostudies-literature

Quorum sensing in Staphylococcus aureus biofilms.

Project description: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.

| S-EPMC355980 | biostudies-literature

Targeted and untargeted analysis of secondary metabolites to monitor growth and quorum sensing inhibition for methicillin-resistant Staphylococcus aureus (MRSA).

Project description:Drug resistant infections are an increasing problem world-wide, responsible for an estimated 700,000 annual mortalities. The use of antibiotics to treat such infections has resulted in the development of resistant bacterial pathogens such as methicillin-resistant Staphylococcus aureus (MRSA). One potential alternative strategy for treating drug resistant bacterial infections is to inhibit the production of toxins, thereby making the bacteria less harmful to the host, a so called "anti-virulence" approach. In MRSA, the agr quorum sensing system is one of the major regulators of toxin production, and quorum sensing inhibitors that target this system are a promising anti-virulence strategy. With this study, we developed a method that enables the activity of quorum sensing inhibitors to be measured using ultra-performance liquid chromatography coupled to mass spectrometry (UPLC-MS). This method is an improvement over existing methods because it can be employed to distinguish antimicrobial activity from quorum sensing inhibition activity based on the UPLC-MS data. This is possible by simultaneously tracking production of metabolites regulated by the agr quorum sensing system (AIP-I and formylated δ-toxin) and a metabolite that appears not to be agr regulated under the conditions of this study (aureusimine B). The newly developed method provides more nuanced indication of how metabolite production changes over time and in response to quorum sensing or growth inhibition than is possible with commonly employed spectroscopic methods.

| S-EPMC8485213 | biostudies-literature

Staphylococcus aureus autoinducer-2 quorum sensing decreases biofilm formation in an icaR-dependent manner.

Project description:BackgroundStaphylococcus aureus is an important pathogen that causes biofilm-associated infection in humans. Autoinducer 2 (AI-2), a quorum-sensing (QS) signal for interspecies communication, has a wide range of regulatory functions in both Gram-positive and Gram-negative bacteria, but its exact role in biofilm formation in S. aureus remains unclear.ResultsHere we demonstrate that mutation of the AI-2 synthase gene luxS in S. aureus RN6390B results in increased biofilm formation compared with the wild-type (WT) strain under static, flowing and anaerobic conditions and in a mouse model. Addition of the chemically synthesized AI-2 precursor in the luxS mutation strain (ΔluxS) restored the WT phenotype. Real-time RT-PCR analysis showed that AI-2 activated the transcription of icaR, a repressor of the ica operon, and subsequently a decreased level of icaA transcription, which was presumably the main reason why luxS mutation influences biofilm formation. Furthermore, we compared the roles of the agr-mediated QS system and the LuxS/AI-2 QS system in the regulation of biofilm formation using the ΔluxS strain, RN6911 and the Δagr ΔluxS strain. Our data indicate a cumulative effect of the two QS systems on the regulation of biofilm formation in S. aureus.ConclusionThese findings demonstrate that AI-2 can decrease biofilm formation in S. aureus via an icaR-activation pathway. This study may provide clues for therapy in S. aureus biofilm-associated infection.

| S-EPMC3539994 | biostudies-literature

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