Antimicrobial Peptide Dendrimers and Quorum-Sensing Inhibitors in Formulating Next-Generation Anti-Infection Cell Therapy Dressings for Burns.

Multidrug resistance infections are the main cause of failure in the pro-regenerative cell-mediated therapy of burn wounds. The collagen-based matrices for delivery of cells could be potential substrates to support bacterial growth and subsequent lysis of the collagen leading to a cell therapy loss. In this article, we report the development of a new generation of cell therapy formulations with the capacity to resist infections through the bactericidal effect of antimicrobial peptide dendrimers and the anti-virulence effect of anti-quorum sensing MvfR (PqsR) system compounds, which are incorporated into their formulation. Anti-quorum sensing compounds limit the pathogenicity and antibiotic tolerance of pathogenic bacteria involved in the burn wound infections, by inhibiting their virulence pathways. For the first time, we report a biological cell therapy dressing incorporating live progenitor cells, antimicrobial peptide dendrimers, and anti-MvfR compounds, which exhibit bactericidal and anti-virulence properties without compromising the viability of the progenitor cells.

In Silico and in Vitro-Guided Identification of Inhibitors of Alkylquinolone-Dependent Quorum Sensing in Pseudomonas aeruginosa.

Pseudomonas aeruginosa is a major opportunistic pathogen in cystic fibrosis, wound and nosocomial infections, posing a serious burden to public health, due to its antibiotic resistance. The P. aeruginosa Pseudomonas Quinolone System (pqs) quorum sensing system, driven by the activation of the transcriptional regulator, PqsR (MvfR) by alkylquinolone (AQ) signal molecules, is a key player in the regulation of virulence and a potential target for the development of novel antibacterial agents. In this study, we performed in silico docking analysis, coupled with screening using a P. aeruginosa mCTX::PpqsA-lux chromosomal promoter fusion, to identify a series of new PqsR antagonists. The hit compounds inhibited pyocyanin and alkylquinolone signal molecule production in P. aeruginosa PAO1-L and PA14 strains. The inhibitor Ia, which showed the highest activity in PA14, reduced biofilm formation in PAO1-L and PA14, increasing their sensitivity to tobramycin. Furthermore, the hepatic and plasma stabilities for these compounds were determined in both rat and human in vitro microsomal assays, to gain a further understanding of their therapeutic potential. This work has uncovered a new class of P. aeruginosa PqsR antagonists with potential for hit to lead optimisation in the search for quorum sensing inhibitors for future anti-infective drug discovery programs.

Pharmacological Inhibition of the Pseudomonas aeruginosa MvfR Quorum-Sensing System Interferes with Biofilm Formation and Potentiates Antibiotic-Mediated Biofilm Disruption.

Pseudomonas aeruginosa biofilms contribute to its survival on biotic and abiotic surfaces and represent a major clinical threat due to their high tolerance to antibiotics. Therefore, the discovery of antibiofilm agents may hold great promise. We show that pharmacological inhibition of the P. aeruginosa quorum-sensing regulator MvfR (PqsR) using a benzamide-benzimidazole compound interferes with biofilm formation and potentiates biofilm sensitivity to antibiotics. Such a strategy could have great potential against P. aeruginosa persistence in diverse environments.

Comparative genomics reveals distinct diversification patterns among LysR-type transcriptional regulators in the ESKAPE pathogen Pseudomonas aeruginosa.

Pseudomonas aeruginosa, a harmful nosocomial pathogen associated with cystic fibrosis and burn wounds, encodes for a large number of LysR-type transcriptional regulator proteins. To understand how and why LTTR proteins evolved with such frequency and to establish whether any relationships exist within the distribution we set out to identify the patterns underpinning LTTR distribution in P. aeruginosa and to uncover cluster-based relationships within the pangenome. Comparative genomic studies revealed that in the JGI IMG database alone ~86 000 LTTRs are present across the sequenced genomes (n=699). They are widely distributed across the species, with core LTTRs present in >93 % of the genomes and accessory LTTRs present in <7 %. Analysis showed that subsets of core LTTRs can be classified as either variable (typically specific to P. aeruginosa) or conserved (and found to be distributed in other Pseudomonas species). Extending the analysis to the more extensive Pseudomonas database, PA14 rooted analysis confirmed the diversification patterns and revealed PqsR, the receptor for the Pseudomonas quinolone signal (PQS) and 2-heptyl-4-quinolone (HHQ) quorum-sensing signals, to be amongst the most variable in the dataset. Successful complementation of the PAO1 pqsR - mutant using representative variant pqsR sequences suggests a degree of structural promiscuity within the most variable of LTTRs, several of which play a prominent role in signalling and communication. These findings provide a new insight into the diversification of LTTR proteins within the P. aeruginosa species and suggests a functional significance to the cluster, conservation and distribution patterns identified.

Skeletal muscle mitochondrial dysfunction mediated by Pseudomonas aeruginosa quorum-sensing transcription factor MvfR: reversing effects with anti-MvfR and mitochondrial-targeted compounds.

Sepsis and chronic infections with Pseudomonas aeruginosa, a leading "ESKAPE" bacterial pathogen, are associated with increased morbidity and mortality and skeletal muscle atrophy. The actions of this pathogen on skeletal muscle remain poorly understood. In skeletal muscle, mitochondria serve as a crucial energy source, which may be perturbed by infection. Here, using the well-established backburn and infection model of murine P. aeruginosa infection, we deciphered the systemic impact of the quorum-sensing transcription factor MvfR (multiple virulence factor regulator) by interrogating, 5 days post-infection, its effect on mitochondrial-related functions in the gastrocnemius skeletal muscle and the outcome of the pharmacological inhibition of MvfR function and that of the mitochondrial-targeted peptide, Szeto-Schiller 31 (SS-31). Our findings show that the MvfR perturbs adenosine triphosphate generation, oxidative phosphorylation, and antioxidant response, elevates the production of reactive oxygen species, and promotes oxidative damage of mitochondrial DNA in the gastrocnemius muscle of infected mice. These impairments in mitochondrial-related functions were corroborated by the alteration of key mitochondrial proteins involved in electron transport, mitochondrial biogenesis, dynamics and quality control, and mitochondrial uncoupling. Pharmacological inhibition of MvfR using the potent anti-MvfR lead, D88, we developed, or the mitochondrial-targeted peptide SS-31 rescued the MvfR-mediated alterations observed in mice infected with the wild-type strain PA14. Our study provides insights into the actions of MvfR in orchestrating mitochondrial dysfunction in the skeletal murine muscle, and it presents novel therapeutic approaches for optimizing clinical outcomes in affected patients. IMPORTANCE: Skeletal muscle, pivotal for many functions in the human body, including breathing and protecting internal organs, contains abundant mitochondria essential for maintaining cellular homeostasis during infection. The effect of Pseudomonas aeruginosa (PA) infections on skeletal muscle remains poorly understood. Our study delves into the role of a central quorum-sensing transcription factor, multiple virulence factor regulator (MvfR), that controls the expression of multiple acute and chronic virulence functions that contribute to the pathogenicity of PA. The significance of our study lies in the role of MvfR in the metabolic perturbances linked to mitochondrial functions in skeletal muscle and the effectiveness of the novel MvfR inhibitor and the mitochondrial-targeted peptide SS-31 in alleviating the mitochondrial disturbances caused by PA in skeletal muscle. Inhibiting MvfR or interfering with its effects can be a potential therapeutic strategy to curb PA virulence.

MvfR Controls Tolerance to Polymyxin B by Regulating rfaD in Pseudomonas aeruginosa.

Polymyxins are currently the last-resort antibiotics for the treatment of multidrug-resistant Gram-negative bacterial infections. To expand the understanding of the intrinsic resistance mechanism against polymyxins, a laboratory strain of Pseudomonas aeruginosa PAO1 was subjected to serial passage in the presence of sublethal doses of polymyxin B over a period of 30 days. By whole-genome sequencing of successively isolated polymyxin B-resistant isolates, we identified a frameshift mutation (L183fs) in the mvfR gene that further increased polymyxin resistance in the pmrB mutant background. A ΔmvfR mutation alone showed higher tolerance to polymyxin B due to altered lipopolysaccharide (LPS) on the surface of bacterial cells, which decreases its outer membrane permeability. In the ΔmvfR mutant, polymyxin B treatment caused the upregulation of rfaD, the gene involved in LPS core oligosaccharide synthesis, which is responsible for polymyxin tolerance. To the best of our knowledge, this is the first report of mvfR mutation conferring polymyxin resistance in P. aeruginosa via increased integrity of bacterial outer membrane. IMPORTANCE Antibiotic resistance imposes a considerable challenge for the treatment of P. aeruginosa infections. Polymyxins are the last-resort antibiotics for the treatment of multidrug-resistant P. aeruginosa infections. Understanding the development and mechanisms of bacterial resistance to polymyxins may provide clues for the development of new or improved therapeutic strategies effective against P. aeruginosa. In this study, using an in vitro evolution assay in combination with whole-genome sequencing, we demonstrated that MvfR controls tolerance to polymyxin B by regulating the rfaD gene in P. aeruginosa. Our results reveal a novel mechanism employed by P. aeruginosa in the defense against polymyxin antibiotics.

Interconnections of Pseudomonas aeruginosa Quorum-Sensing Systems in Intestinal Permeability and Inflammation.

Quorum sensing (QS) is a highly conserved microbial communication mechanism based on the production and sensing of secreted signaling molecules. The recalcitrant pathogen Pseudomonas aeruginosa is a problematic nosocomial pathogen with complex interconnected QS systems controlling multiple virulence functions. The relevance of QS in P. aeruginosa pathogenesis is well established; however, the regulatory interrelationships of the three major QS systems, LasR/LasI, MvfR (PqsR)/PqsABCD, and RhlR/RhlI, have been studied primarily in vitro. It is, therefore, unclear how these relationships translate to the host environment during infection. Here, we use a collection of P. aeruginosa QS mutants of the three major QS systems to assess the interconnections and contributions in intestinal inflammation and barrier function in vivo. This work reveals that MvfR, not LasR or RhlR, promotes intestinal inflammation during infection. In contrast, we find that P. aeruginosa-driven murine intestinal permeability is controlled by an interconnected QS network involving all three regulators, with MvfR situated upstream of LasR and RhlR. This study demonstrates the importance of understanding the interrelationships of the QS systems during infection and provides critical insights for developing successful antivirulence strategies. Moreover, this work provides a framework to interrogate QS systems in physiologically relevant settings. IMPORTANCE Pseudomonas aeruginosa is a common multidrug-resistant bacterial pathogen that seriously threatens critically ill and immunocompromised patients. Intestinal colonization by this pathogen is associated with elevated mortality rates. Disrupting bacterial communication is a desirable anti-infective approach since these systems coordinate multiple acute and chronic virulence functions in P. aeruginosa. Here, we investigate the role of each of the three major communication systems in the host intestinal functions. This work reveals that P. aeruginosa influences intestinal inflammation and permeability through distinct mechanisms.

Quorum-Sensing Signaling Molecule 2-Aminoacetophenone Mediates the Persistence of Pseudomonas aeruginosa in Macrophages by Interference with Autophagy through Epigenetic Regulation of Lipid Biosynthesis.

Macrophages are crucial components of the host's defense against pathogens. Recent studies indicate that macrophage functions are influenced by lipid metabolism. However, knowledge of how bacterial pathogens exploit macrophage lipid metabolism for their benefit remains rudimentary. We have shown that the Pseudomonas aeruginosa MvfR-regulated quorum-sensing (QS) signaling molecule 2-aminoacetophenone (2-AA) mediates epigenetic and metabolic changes associated with this pathogen's persistence in vivo. We provide evidence that 2-AA counteracts the ability of macrophages to clear the intracellular P. aeruginosa, leading to persistence. The intracellular action of 2-AA in macrophages is linked to reduced autophagic functions and the impaired expression of a central lipogenic gene, stearoyl-CoA desaturase 1 (Scd1), which catalyzes the biosynthesis of monounsaturated fatty acids. 2-AA also reduces the expression of the autophagic genes Unc-51-like autophagy activating kinase 1 (ULK1) and Beclin1 and the levels of the autophagosomal membrane protein microtubule-associated protein 1, light chain 3 isoform B (LC3B) and p62. Reduced autophagy is accompanied by the reduced expression of the lipogenic gene Scd1, preventing bacterial clearance. Adding the SCD1 substrates palmitoyl-CoA and stearoyl-CoA increases P. aeruginosa clearance by macrophages. The impact of 2-AA on lipogenic gene expression and autophagic machinery is histone deacetylase 1 (HDAC1) mediated, implicating the HDAC1 epigenetic marks at the promoter sites of Scd1 and Beclin1 genes. This work provides novel insights into the complex metabolic alterations and epigenetic regulation promoted by QS and uncovers additional 2-AA actions supporting P. aeruginosa sustainment in macrophages. These findings may aid in designing host-directed therapeutics and protective interventions against P. aeruginosa persistence. IMPORTANCE This work sheds new light on how P. aeruginosa limits bacterial clearance in macrophages through 2-aminoacetophenone (2-AA), a secreted signaling molecule by this pathogen that is regulated by the quorum-sensing transcription factor MvfR. The action of 2-AA on the lipid biosynthesis gene Scd1 and the autophagic genes ULK1 and Beclin1 appears to secure the reduced intracellular clearance of P. aeruginosa by macrophages. In support of the 2-AA effect on lipid biosynthesis, the ability of macrophages to reduce the intracellular P. aeruginosa burden is reinstated following the supplementation of palmitoyl-CoA and stearoyl-CoA. The 2-AA-mediated reduction of Scd1 and Beclin1 expression is linked to chromatin modifications, implicating the enzyme histone deacetylase 1 (HDAC1), thus opening new avenues for future strategies against this pathogen's persistence. Overall, the knowledge obtained from this work provides for developing new therapeutics against P. aeruginosa.

A Pseudomonas aeruginosa PQS quorum-sensing system inhibitor with anti-staphylococcal activity sensitizes polymicrobial biofilms to tobramycin.

As single- and mixed-species biofilms, Staphylococcus aureus and Pseudomonas aeruginosa cause difficult-to-eradicate chronic infections. In P. aeruginosa, pseudomonas quinolone (PQS)-dependent quorum sensing regulates virulence and biofilm development that can be attenuated via antagonists targeting the transcriptional regulator PqsR (MvfR). Here, we exploited a quinazolinone (QZN) library including PqsR agonists and antagonists for their activity against S. aureus alone, when co-cultured with P. aeruginosa, and in combination with the aminoglycoside tobramycin. The PqsR inhibitor, QZN 34 killed planktonic Gram-positives but not Gram-negatives. QZN 34 prevented S. aureus biofilm formation, severely damaged established S. aureus biofilms, and perturbed P. aeruginosa biofilm development. Although P. aeruginosa protected S. aureus from tobramycin in mixed biofilms, the combination of aminoglycoside antibiotic with QZN 34 eradicated the mixed-species biofilm. The mechanism of action of QZN 34 toward Gram-positive bacteria is shown to involve membrane perturbation and dissipation of transmembrane potential.

An Immunochemical Approach to Detect the Quorum Sensing-Regulated Virulence Factor 2-Heptyl-4-Quinoline N-Oxide (HQNO) Produced by Pseudomonas aeruginosa Clinical Isolates.

Understanding quorum sensing (QS) and its role in the development of pathogenesis may provide new avenues for diagnosing, surveillance, and treatment of infectious diseases. For this purpose, the availability of reliable and efficient analytical diagnostic tools suitable to specifically detect and quantify these essential QS small molecules and QS regulated virulence factors is crucial. Here, we reported the development and evaluation of antibodies and an enzyme-linked immunosorbent assay (ELISA) for HQNO (2-heptyl-4-quinoline N-oxide), a QS product of the PqsR system, which has been found to act as a major virulence factor that interferes with the growth of other microorganisms. Despite the nonimmunogenic character of HQNO, the antibodies produced showed high avidity and the microplate-based ELISA developed could detect HQNO in the low nM range. Hence, a limit of detection (LOD) of 0.60 ± 0.13 nM had been reached in Müeller Hinton (MH) broth, which was below previously reported levels using sophisticated equipment based on liquid chromatography coupled to mass spectrometry. The HQNO profile of release of different Pseudomonas aeruginosa clinical isolates analyzed using this ELISA showed significant differences depending on whether the clinical isolates belonged to patients with acute or chronic infections. These data point to the possibility of using HQNO as a specific biomarker to diagnose P. aeruginosa infections and for patient surveillance. Considering the role of HQNO in inhibiting the growth of coinfecting bacteria, the present ELISA will allow the investigation of these complex bacterial interactions underlying infections. IMPORTANCE Bacteria use quorum sensing (QS) as a communication mechanism that releases small signaling molecules which allow synchronizing a series of activities involved in the pathogenesis, such as the biosynthesis of virulence factors or the regulation of growth of other bacterial species. HQNO is a metabolite of the Pseudomonas aeruginosa-specific QS signaling molecule PQS (Pseudomonas quinolone signal). In this work, the development of highly specific antibodies and an immunochemical diagnostic technology (ELISA) for the detection and quantification of HQNO was reported. The ELISA allowed profiling of the release of HQNO by clinical bacterial isolates, showing its potential value for diagnosing and surveillance of P. aeruginosa infections. Moreover, the antibodies and the ELISA reported here may contribute to the knowledge of other underlying conditions related to the pathology, such as the role of the interactions with other bacteria of a particular microbiota environment.

Hit Identification of New Potent PqsR Antagonists as Inhibitors of Quorum Sensing in Planktonic and Biofilm Grown Pseudomonas aeruginosa.

Current treatments for Pseudomonas aeruginosa infections are becoming less effective because of the increasing rates of multi-antibiotic resistance. Pharmacological targeting of virulence through inhibition of quorum sensing (QS) dependent virulence gene regulation has considerable therapeutic potential. In P. aeruginosa, the pqs QS system regulates the production of multiple virulence factors as well as biofilm maturation and is a promising approach for developing antimicrobial adjuvants for combatting drug resistance. In this work, we report the hit optimisation for a series of potent novel inhibitors of PqsR, a key regulator of the pqs system, bearing a 2-((5-methyl-5H-[1,2,4]triazino[5,6-b]indol-3-yl)thio) acetamide scaffold. The initial hit compound 7 (PAO1-L IC50 0.98 ± 0.02 µM, PA14 inactive at 10 µM) was obtained through a virtual screening campaign performed on the PqsR ligand binding domain using the University of Nottingham Managed Chemical Compound Collection. Hit optimisation gave compounds with enhanced potency against strains PAO1-L and PA14, evaluated using P. aeruginosa pqs-based QS bioreporter assays. Compound 40 (PAO1-L IC50 0.25 ± 0.12 µM, PA14 IC50 0.34 ± 0.03 µM) is one of the most potent PqsR antagonists reported showing significant inhibition of P. aeruginosa pyocyanin production and pqs system signaling in both planktonic cultures and biofilms. The co-crystal structure of 40 with the PqsR ligand binding domain revealed the specific binding interactions occurring between inhibitor and this key regulatory protein.

The Anti-activator QslA Negatively Regulates Phenazine-1-Carboxylic Acid Biosynthesis by Interacting With the Quorum Sensing Regulator MvfR in the Rhizobacterium Pseudomonas aeruginosa Strain PA1201.

Two almost identical gene clusters (phz1 and phz2) are responsible for phenazine-1-carboxylic acid (PCA) production in Pseudomonas aeruginosa (P. aeruginosa) strain MSH (derived from strain PA1201). Here, we showed that the anti-activator QslA negatively regulated PCA biosynthesis and phz1 expression in strain PA1201 but had little effect on phz2 expression. This downregulation was mediated by a 56-bp region within the 5'-untranslated region (5'-UTR) of the phz1 promoter and was independent of LasR and RsaL signaling. QslA also negatively regulated Pseudomonas quinolone signal (PQS) production. Indeed, QslA controlled the PQS threshold concentration needed for PQS-dependent PCA biosynthesis. The quorum sensing regulator MvfR was required for the QslA-dependent inhibition of PCA production. We identified a direct protein-protein interaction between QslA and MvfR. The ligand-binding domain of MvfR (residues 123-306) was involved in this interaction. Our results suggested that MvfR bound directly to the promoter of the phz1 cluster. QslA interaction with MvfR prevented the binding of MvfR to the phz1 promoter regions. Thus, this study depicted a new mechanism by which QslA controls PCA and PQS biosynthesis in P. aeruginosa.

Molecular Insights into Function and Competitive Inhibition of Pseudomonas aeruginosa Multiple Virulence Factor Regulator.

New approaches to antimicrobial drug discovery are urgently needed to combat intractable infections caused by multidrug-resistant (MDR) bacteria. Multiple virulence factor regulator (MvfR or PqsR), a Pseudomonas aeruginosa quorum sensing transcription factor, regulates functions important in both acute and persistent infections. Recently identified non-ligand-based benzamine-benzimidazole (BB) inhibitors of MvfR suppress both acute and persistent P. aeruginosa infections in mice without perturbing bacterial growth. Here, we elucidate the crystal structure of the MvfR ligand binding domain (LBD) in complex with one potent BB inhibitor, M64. Structural analysis indicated that M64 binds, like native ligands, to the MvfR hydrophobic cavity. A hydrogen bond and pi interaction were found to be important for MvfR-M64 affinity. Surface plasmon resonance analysis demonstrated that M64 is a competitive inhibitor of MvfR. Moreover, a protein engineering approach revealed that Gln194 and Tyr258 are critical for the interaction between MvfR and M64. Random mutagenesis of the full-length MvfR protein identified a single-amino-acid substitution, I68F, at a DNA binding linker domain that confers M64 insensitivity. In the presence of M64, I68F but not the wild-type (WT) MvfR protein retained DNA binding ability. Our findings strongly suggest that M64 promotes conformational change at the DNA binding domain of MvfR and that the I68F mutation may compensate for this change, indicating allosteric inhibition. This work provides critical new insights into the molecular mechanism of MvfR function and inhibition that could aid in the optimization of anti-MvfR compounds and improve our understanding of MvfR regulation.IMPORTANCEPseudomonas aeruginosa is an opportunistic Gram-negative pathogen that causes serious acute, persistent, and relapsing infections. New approaches to antimicrobial drug discovery are urgently needed to combat intractable infections caused by this pathogen. The Pseudomonas aeruginosa quorum sensing transcription factor MvfR regulates functions important in both acute and persistent infections. We used recently identified inhibitors of MvfR to perform structural studies and reveal important insights that would benefit the optimization of anti-MvfR compounds. Altogether, the results reported here provide critical detailed mechanistic insights into the function of MvfR domains that may benefit the optimization of the chemical, pharmacological, and safety properties of MvfR antagonist series.

In-depth Profiling of MvfR-Regulated Small Molecules in Pseudomonas aeruginosa after Quorum Sensing Inhibitor Treatment.

Pseudomonas aeruginosa is a Gram-negative bacterium, which causes opportunistic infections in immuno-compromised individuals. Due to its multiple resistances toward antibiotics, the development of new drugs is required. Interfering with Quorum Sensing (QS), a cell-to-cell communication system, has shown to be highly efficient in reducing P. aeruginosa pathogenicity. One of its QS systems employs Pseudomonas Quinolone Signal (PQS) and 4-hydroxy-2-heptylquinoline (HHQ) as signal molecules. Both activate the transcriptional regulator MvfR (Multiple Virulence Factor Regulator), also called PqsR, driving the production of QS molecules as well as toxins and biofilm formation. The aim of this work was to elucidate the effects of QS inhibitors (QSIs), such as MvfR antagonists and PqsBC inhibitors, on the biosynthesis of the MvfR-regulated small molecules 2'-aminoacetophenone (2-AA), dihydroxyquinoline (DHQ), HHQ, PQS, and 4-hydroxy-2-heptylquinoline-N-oxide (HQNO). The employed synthetic MvfR antagonist fully inhibited pqs small molecule formation showing expected sigmoidal dose-response curves for 2-AA, HQNO, HHQ and PQS. Surprisingly, DHQ levels were enhanced at lower antagonist concentrations followed by a full suppression at higher QSI amounts. This particular bi-phasic profile hinted at the accumulation of a biosynthetic intermediate resulting in the observed overproduction of the shunt product DHQ. Additionally, investigations on PqsBC inhibitors showed a reduction of MvfR natural ligands, while increased 2-AA, DHQ and HQNO levels compared to the untreated cells were detected. Moreover, PqsBC inhibitors did not show any significant effect in PA14 pqsC mutant demonstrating their target selectivity. As 2-AA is important for antibacterial tolerance, the QSIs were evaluated in their capability to attenuate persistence. Indeed, persister cells were reduced along with 2-AA inhibition resulting from MvfR antagonism, but not from PqsBC inhibition. In conclusion, antagonizing MvfR using a dosage capable of fully suppressing this QS system will lead to a favorable therapeutic outcome as DHQ overproduction is avoided and bacterial persistence is reduced.

Honey's Ability to Counter Bacterial Infections Arises from Both Bactericidal Compounds and QS Inhibition.

The ability of honey to kill bacterial pathogens in vitro and quickly clear even chronic or drug-resistant infections has been demonstrated by several studies. Most current research is focused on identifying the bactericidal compounds in honey, but the action of the compounds discovered is not sufficient to explain honey's activity. By diluting honey to sub-inhibitory levels, we were able to study its impact on bacterial coordinated behavior, and discovered that honey inhibits bacterial quorum sensing (QS). Experiments to characterize and quantify honey's effect on the QS networks of Pseudomonas aeruginosa revealed that low concentrations of honey inhibited the expression of MvfR, las, and rhl regulons, including the associated virulence factors. This research also establishes that inhibition of QS is associated with honey's sugar content. Therefore, honey combats infections by two independent mechanisms acting in tandem: bactericidal components, which actively kill cells, and disruption of QS, which weakens bacterial coordination and virulence.