Quorum Sensing Signal Molecules Research Articles

Study on the inhibition activity and mechanism of Tanreqing against Klebsiella pneumoniae biofilm formation in vitro and in vivo.

To evaluate the antibacterial effect of Tanreqing (TRQ) against K. pneumoniae and its inhibition activity on bacterial biofilm formation in vitro and in vivo, and to explore the mechanism of the inhibitory effects of TRQ on K. pneumoniae biofilm formation. An in vitro biofilm model of K. pneumoniae was established, and the impact of TRQ on biofilm formation was evaluated using crystal violet staining and scanning electron microscopy (SEM). Furthermore, the clearance effect of TRQ against K. pneumoniae in the biofilm was assessed using the viable plate counting method; q-RT PCR was used to evaluate the inhibitory effect of different concentrations of TRQ on the expression of biofilm-related genes in Klebsiella pneumoniae; The activity of quorum sensing signal molecule AI-2 was detected by Vibrio harveyi bioluminescence assay; Meanwhile, a guinea pig lung infection model of Klebsiella pneumoniae was constructed, and after treated with drugs, pathological analysis of lung tissue and determination of bacterial load in lung tissue were performed. The treatment groups included TRQ group, imipenem(IPM) group, TRQ+IPM group, and sterile saline group as the control. The formation of K. pneumoniae biofilm was significantly inhibited by TRQ in vitro experiments. Furthermore, when combined with IPM, the clearance of K. pneumoniae in the biofilm was notably increased compared to the TRQ group and IPM group alone. q-RT PCR analysis revealed that TRQ down-regulated the expression of genes related to biofilm formation in K. pneumoniae, specifically luxS, wbbm, wzm, and lsrK, and also inhibited the activity of AI-2 molecules in the bacterium. In vivo experiments demonstrated that TRQ effectively treated guinea pig lung infections, resulting in reduced lung inflammation. Additionally, when combined with IPM, there was a significant reduction in the bacterial load in lung tissue. TRQ as a potential therapeutic agent plays a great role in the treatment of K. pneumoniae infections, particularly in combination with conventional antibiotics. And TRQ can enhanced the clearance effect on the bacterium by inhibiting the K. pneumoniae biofilm formation, which provided experimental evidence in support of clinical treatment of TRQ against K. pneumoniae infections.

A novel photoelectrochemical self-screening aptamer biosensor based on CAU-17-derived Bi2WO6/Bi2S3 for rapid detection of quorum sensing signal molecules

Quorum sensing signal molecule is an important biomarker released by some microorganisms, which can regulate the adhesion and aggregation of marine microorganisms on the surface of engineering facilities. Thus, it is significant to exploit a convenient method that can effectively monitor the formation and development of marine biofouling. In this work, an advanced photoelectrochemical (PEC) aptamer biosensing platform was established and firstly applied for the rapid and ultrasensitive determination of N-(3-Oxodecanoyl)-l-homoserine lactone (3-O-C10-HL) released from marine fouling microorganism Ponticoccus sp. PD-2. The visible-light-driven Bi2WO6/Bi2S3 heterojunction derived from metal-organic frameworks (MOFs) CAU-17 and self-screened aptamer were employed as the photoactive materials and bioidentification elements, respectively. Appropriate amount of MoS2 quantum dots (QDs) conjugated with single-stranded DNA were introduced by hybridization to enhance the photocurrent response of the PEC biosensor. The self-screening aptamer can specifically recognize 3-O-C10-HL, accompanied by increasing the steric hindrance and forcing MoS2 QDs to leave the electrode surface, resulting in an obvious reduction of photocurrent and achieving a dual-inhibition signal amplification effect. Under the optimized conditions, the photocurrent response of PEC aptasensor was linear with 3-O-C10-HL concentration from 1 nM to 10 μM, and the detection limit was as low as 0.26 nM. The detection strategy also showed a high reproducibility, superior specificity and good stability. This work not only provides a simple, rapid and ultrasensitive PEC aptamer biosensing strategy for monitoring quorum sensing signal molecules in marine biofouling, but also broadens the application of MOFs-based heterojunctions in PEC sensors.

Potential of the quorum-quenching and plant-growth promoting halotolerant Bacillus toyonensis AA1EC1 as biocontrol agent.

The use of fertilizers and pesticides to control plant diseases is widespread in intensive farming causing adverse effects together with the development of antimicrobial resistance pathogens. As the virulence of many Gram-negative phytopathogens is controlled by N-acyl-homoserine lactones (AHLs), the enzymatic disruption of this type of quorum-sensing (QS) signal molecules, mechanism known as quorum quenching (QQ), has been proposed as a promising alternative antivirulence therapy. In this study, a novel strain of Bacillus toyonensis isolated from the halophyte plant Arthrocaulon sp. exhibited numerous traits associated with plant growth promotion (PGP) and degraded a broad range of AHLs. Three lactonases and an acylase enzymes were identified in the bacterial genome and verified invitro. The AHL-degrading activity of strain AA1EC1 significantly attenuated the virulence of relevant phytopathogens causing reduction of soft rot symptoms on potato and carrots. Invivo assays showed that strain AA1EC1 significantly increased plant length, stem width, root and aerial dry weights and total weight of tomato and protected plants against Pseudomonas syringae pv. tomato. To our knowledge, this is the first report to demonstrate PGP and QQ activities in the species B. toyonensis that make this strain as a promising phytostimulant and biocontrol agent.

A novel thermostable YtnP lactonase from Stenotrophomonas maltophilia inhibits Pseudomonas aeruginosa virulence in vitro and in vivo

Infections caused by multidrug-resistant pathogens are one of the biggest challenges facing the healthcare system today. Quorum quenching (QQ) enzymes have the potential to be used as innovative enzyme-based antivirulence therapeutics to combat infections caused by multidrug-resistant pathogens. The main objective of this research was to describe the novel YtnP lactonase derived from the clinical isolate Stenotrophomonas maltophilia and to investigate its antivirulence potential against multidrug-resistant Pseudomonas aeruginosa MMA83. YtnP lactonase, the QQ enzyme, belongs to the family of metallo-β-lactamases. The recombinant enzyme has several advantageous biotechnological properties, such as high thermostability, activity in a wide pH range, and no cytotoxic effect. High-performance liquid chromatography analysis revealed the activity of recombinant YtnP lactonase toward a wide range of N-acyl-homoserine lactones (AHLs), quorum sensing signaling molecules, with a higher preference for long-chain AHLs. Recombinant YtnP lactonase was shown to inhibit P. aeruginosa MMA83 biofilm formation, induce biofilm decomposition, and reduce extracellular virulence factors production. Moreover, the lifespan of MMA83-infected Caenorhabditis elegans was prolonged with YtnP lactonase treatment. YtnP lactonase showed synergistic inhibitory activity in combination with gentamicin and acted additively with meropenem against MMA83. The described properties make YtnP lactonase a promising therapeutic candidate for the development of next-generation antivirulence agents.

Pretreatment by a novel photo-electro reactor to control organic and biofouling during reverse osmosis filtration of reclaimed water

This study reports the performance of a novel photo-electro filtration (PEF) reactor consisting of an IrO2/RuO2 coated anodic filter and UV lamp to pretreat biologically treated wastewater effluent prior to reverse osmosis (RO) filtration. Synthetic model solutions and biologically treated wastewater effluent were used for systematic evaluation of the PEF reactor performance in terms of organic degradation, bacterial inactivation, biofilm formation reduction, and ultimately RO fouling mitigation. Experiments with model solutions containing proteins (Bovine Serum Albumin) and bacteria (Pseudomonas aeruginosa) show that UV irradiation and electrochemical treatment with increasing voltage input resulted in superior bacterial inactivation, biofilm formation reduction, and effective organic degradation. When the PEF reactor was used to pretreat biologically treated wastewater effluent, electrochemical treatment at 3 V alone and the combination of electrochemical treatment at 3 V and UV could achieve near complete bacterial inactivation and good fouling reduction during subsequent RO operation. However, the combination of electrochemical treatment at 3 V and UV was superior over electrochemical treatment at 3 V alone in several aspects including fouling reduction and reduced risk of membrane degradation due to chlorine residue. The underlying mechanisms responsible for effective PEF pretreatment performance were elucidated by monitoring the fate of free chlorine, hydroxyl radicals (indirectly monitored by measuring concentration of the quenching chemical N-n-dimethyl-p-nitrosoaniline), H2O2, quorum sensing signal molecules, and organic fractions during UV irradiation and electrochemical treatment. Effective bio and organic fouling mitigation by PEF pretreatment was attributed to the enhanced bacterial inactivation and degradation of organic compounds (primarily protein-like and humic-like substances) by hydroxyl radicals (∙OH) produced from photo-electrochemical treatment.

Deciphering the role of hydraulic retention time (HRT) in integrated fixed-film activated sludge (IFAS) system under aniline stress: Effects on microbial succession and communication

Integrated fixed-film activated sludge (IFAS) system offered an energy-efficient option for nitrogen removal from aniline wastewater. The regulation of hydraulic retention time (HRT) has been reported to affect the fate of functional microbial communities with different generation cycles, but the role of HRT in IFAS under aniline stress has not been sufficiently revealed. Herein, it was observed that shortening the HRT altered the mechanisms underlying the adaptation of a continuous flow micro-aerated IFAS system to aniline stress, thereby negligibly impacting the removal of aniline. The total nitrogen removal rate in Y2 (HRT = 0.5 day) was 40.28 % higher than Y1 (1 day), and 23.7 % higher than Y3 (0.25 day). This could be attributed to Y2 having an ample supply of electron donors for denitrification compared to Y1, as well as showing weaker stress to inhibit nitrification compared to Y3. Furthermore, with the progressive intensification of stress induced by HRT reduction, microorganisms gradually enhanced their adaptation to aniline. This led to an increased migration of microorganisms towards the inner layers of the biofilm, resulting in enhanced functional complementarity between the sludge and biofilm in Y2. Also, there was a distinct development of autotrophic denitrification, with arenimonas predominant in Y1, and Thauera predominant in Y2 and Y3. Moreover, Y2 possessed a high abundance of functional genes related to the production and reception of quorum sensing signal molecules, which may enhance the electron transport-dependent nitrogen metabolism reaction.

Constructions of quorum sensing signaling network for activated sludge microbial community.

In wastewater treatment systems, the interactions among various microbes based on chemical signals, namely quorum sensing (QS), play critical roles in influencing microbial structure and function. However, it is challenging to understand the QS-controlled behaviors and the underlying mechanisms in complex microbial communities. In this study, we constructed a QS signaling network, providing insights into the intra- and interspecies interactions of activated sludge microbial communities based on diverse QS signal molecules. Our research underscores the role of diffusible signal factors in both intra- and interspecies communication among activated sludge microorganisms, and signal molecules commonly considered to mediate intraspecies communication may also participate in interspecies communication. QS signaling molecules play an important role as communal resources among the entire microbial group. The communication network within the microbial community is highly redundant, significantly contributing to the stability of natural microbial systems. This work contributes to the establishment of QS signaling network for activated sludge microbial communities, which may complement metabolic exchanges in explaining activated sludge microbial community structure and may help with a variety of future applications, such as making the dynamics and resilience of highly complex ecosystems more predictable.

Facet-dependent haloperoxidase-like activities of CeO2 nanoparticles contribute to their excellent biofilm formation suppression abilities

Biofilms adhering to different surfaces have significant negative impacts in various fields. Cerium oxide nanoparticles can serve as mimics of haloperoxidase for biological biofilm inhibition applications. The regulation of the exposed facet of CeO2 nanoparticles influences their efficiency in various catalytic processes. However, there is still a lack of systematic studies on the facet-dependent haloperoxidase-like activity of CeO2. In the present study, the facet-dependent haloperoxidase activities and antibiofilm performance of CeO2 nanoparticles were elucidated through experiment analysis and density function theory calculation. The as-prepared CeO2 nanoparticles inhibited bacterial survival and catalyzed the oxidative bromination of quorum sensing signaling molecules, achieving biofilm inhibition performance. The antibacterial and biofilm formation suppression abilities were consistent with their haloperoxidase activities. The {111}- and {110}-facet CeO2 nanopolyhedra, as well as the {110}- and {100}-facet CeO2 nanorods, which had higher haloperoxidase activity showed better antibiofilm performance than the {100}-facet CeO2 cubes. The present findings provide a comprehensive understanding of the facet-dependent haloperoxidase-like activity of CeO2. Furthermore, engineering CeO2 morphologies with different crystal facets may represent a novel method for significantly adjusting their haloperoxidase-like activity.

PQS and pyochelin in Pseudomonas aeruginosa share inner membrane transporters to mediate iron uptake.

Bacteria absorb different forms of iron through various channels to meet their needs. Our previous studies have shown that TseF, a type VI secretion system effector for Fe uptake, facilitates the delivery of outer membrane vesicle-associated Pseudomonas quinolone signal (PQS)-Fe3+ to bacterial cells by a process involving the Fe(III) pyochelin receptor FptA and the porin OprF. However, the form in which the PQS-Fe3+ complex enters the periplasm and how it is moved into the cytoplasm remain unclear. Here, we first demonstrate that the PQS-Fe3+ complex enters the cell directly through FptA or OprF. Next, we show that inner membrane transporters such as FptX, PchHI, and FepBCDG are not only necessary for Pseudomonas aeruginosa to absorb PQS-Fe3+ and pyochelin (PCH)-Fe3+ but are also necessary for the virulence of P. aeruginosa toward Galleria mellonella larvae. Furthermore, we suggest that the function of PQS-Fe3+ (but not PQS)-mediated quorum-sensing regulation is dependent on FptX, PchHI, and FepBCDG. Additionally, the findings indicate that unlike FptX, neither FepBCDG nor PchHI play roles in the autoregulatory loop involving PchR, but further deletion of fepBCDG and pchHI can reverse the inactive PchR phenotype caused by fptX deletion and reactivate the expression of the PCH pathway genes under iron-limited conditions. Finally, this work identifies the interaction between FptX, PchHI, and FepBCDG, indicating that a larger complex could be formed to mediate the uptake of PQS-Fe3+ and PCH-Fe3+. These results pave the way for a better understanding of the PQS and PCH iron absorption pathways and provide future directions for research on tackling P. aeruginosa infections.IMPORTANCEPseudomonas aeruginosa has evolved a number of strategies to acquire the iron it needs from its host, with the most common being the synthesis, secretion, and uptake of siderophores such as pyoverdine, pyochelin, and the quorum-sensing signaling molecule Pseudomonas quinolone signal (PQS). However, despite intensive studies of the siderophore uptake pathways of P. aeruginosa, our understanding of how siderophores transport iron across the inner membrane into the cytoplasm is still incomplete. Herein, we reveal that PQS and pyochelin in P. aeruginosa share inner membrane transporters such as FptX, PchHI, and FepBCDG to mediate iron uptake. Meanwhile, PQS and pyochelin-mediated signaling operate to a large extent via these inner membrane transporters. Our study revealed the existence of shared uptake pathways between PQS and pyochelin, which could lead us to reexamine the role of these two molecules in the iron uptake and virulence of P. aeruginosa.

Carrier with cyclodextrin and quorum sensing synergy: An efficient method for selective enrichment of anammox bacteria

High-efficiency enrichment of functional bacteria is one of the biggest technical barriers to the application of anaerobic ammonium oxidation (anammox). This study combined biofilm carriers and quorum sensing to achieve rapid biomass accumulation. A biofilm carrier with biomass adsorption and quorum sensing regulation functions was developed for the rapid start-up of the reactor and enrichment of anammox bacteria (AnAOB). The nitrogen removal rates of the three reactors were further tested, and the carrier mechanism was explored. The start-up time of the reactor with the modified carriers was 28 days, which was ∼44 % shorter than that of the reactor without carriers. After 90 days of operation, the absolute abundance of total AnAOB was 3.2 times that in the control group. The modified carrier promoted the rapid adhesion of biomass through the hydrophobic cavity of cyclodextrin and induced rapid proliferation, enhancement of activity, and extracellular protein secretion of AnAOB through quorum sensing signaling molecules. Finally, a special hydrophobic biofilm with granular sludge characteristics was formed on the modified carrier through the self-assembly of hydrophobic biomass particles. Overall, we propose a novel approach to enrich anammox bacteria involving a combination of quorum sensing and biofilm carriers.

Cross-talk between QseBC and PmrAB two-component systems is crucial for regulation of motility and colistin resistance in Enteropathogenic Escherichia coli.

The quorum sensing two-component system (TCS) QseBC has been linked to virulence, motility and metabolism regulation in multiple Gram-negative pathogens, including Enterohaemorrhagic Escherichia coli (EHEC), Uropathogenic E. coli (UPEC) and Salmonella enterica. In EHEC, the sensor histidine kinase (HK) QseC detects the quorum sensing signalling molecule AI-3 and also acts as an adrenergic sensor binding host epinephrine and norepinephrine. Downstream changes in gene expression are mediated by phosphorylation of its cognate response regulator (RR) QseB, and 'cross-talks' with non-cognate regulators KdpE and QseF to activate motility and virulence. In UPEC, cross-talk between QseBC and TCS PmrAB is crucial in the regulation and phosphorylation of QseB RR that acts as a repressor of multiple pathways, including motility. Here, we investigated QseBC regulation of motility in the atypical Enteropathogenic E. coli (EPEC) strain O125ac:H6, causative agent of persistent diarrhoea in children, and its possible cross-talk with the KdpDE and PmrAB TCS. We showed that in EPEC QseB acts as a repressor of genes involved in motility, virulence and stress response, and in absence of QseC HK, QseB is likely activated by the non-cognate PmrB HK, similarly to UPEC. We show that in absence of QseC, phosphorylated QseB activates its own expression, and is responsible for the low motility phenotypes seen in a QseC deletion mutant. Furthermore, we showed that KdpD HK regulates motility in an independent manner to QseBC and through a third unidentified party different to its own response regulator KdpE. We showed that PmrAB has a role in iron adaptation independent to QseBC. Finally, we showed that QseB is the responsible for activation of colistin and polymyxin B resistance genes while PmrA RR acts by preventing QseB activation of these resistance genes.

Bacterial biofilm formation and anti-biofilm strategies

Bacteria are ubiquitous prokaryotes. They are involved in biofilm formation and also have the ability to produce anti-biofilm products for biofilm mitigation. This special issue entitled: “Biofilms- community structure, applications and mitigation” of the journal Research in Microbiology was designed to discuss the flexibility of bacterial biofilms and their products under particular circumstances. Given that quorum sensing (QS) controls biofilm growth in some situations, especially for pathogenic bacteria antibiotic evading strategies. In Gram-negative bacteria, N-acyl homoserine lactones are the major quorum sensing signaling molecules. Another approach to prevent bacterial biofilm formation may be to inhibit the QS-regulated activities using quorum quenching (QQ). In this context, QS inhibitors and QS enzymes are important because they, respectively, interfere with signal creation, perception, or degradation and chemical modification. There have been numerous reports of QQ enzymes from bacteria. Treatment failure and recurrent staphylococcal infections are also brought on by biofilm development, which boosts an organism's ability to withstand antibiotics and is thought to be a virulence factor in patients. However, polyphenol quercetin antibiofilm activity is naturally available against drug-resistant Staphylococcus aureus.

The Construction of the Self-Induced Sal System and Its Application in Salicylic Acid Production

The design and construction of more complex and delicate genetic control circuits suffer from poor orthogonality in quorum sensing (QS) systems. The Sal system, which relies on salicylic acid as a signaling molecule, is an artificially engineered regulatory system with a structure that differs significantly from that of natural QS signaling molecules. Salicylic acid is an important drug precursor, mainly used in the production of drugs such as aspirin and anti-HIV drugs. However, there have been no reports on the construction of a self-induced Sal system in single cells. In this study, a high-copy plasmid backbone was used to construct the regulatory proteins and a self-induced promoter of salicylic acid in E. coli by adjusting the precise regulation of key gene expression; the sensitivity and induction range of this system were improved. Subsequently, the exogenous gene pchBA was introduced in E. coli to extend the shikimate pathway and synthesize salicylic acid, resulting in the construction of the first complete self-induced Sal system. Finally, the self-induced Sal System was combined with artificial trans-encoded sRNAs (atsRNAs) to repress the growth-essential gene ppc and accumulate the precursor substance PEP, thereby increasing the titer of salicylic acid by 151%. This construction of a self-induced artificial system introduces a new tool for selecting communication tools and induction systems in synthetic biology and metabolic engineering, but also demonstrates a self-inducible pathway design strategy for salicylic acid biosynthesis.

Exploration of Chemical Diversity in Intercellular Quorum Sensing Signalling Systems in Prokaryotes.

Quorum sensing (QS) serves as a vital means of intercellular signalling in a variety of prokaryotes, which enables single cells to act in multicellular configurations. The potential to control community-wide responses has also sparked numerous recent biotechnological innovations. However, our capacity to utilize intercellular communication is hindered due to a scarcity of complementary signalling systems and a restricted comprehension of interconnections between these systems caused by variations in their dynamic range. In this study, we utilize uniform manifold approximation and projection and extended-connectivity fingerprints to explore the available chemical space of QS signalling molecules. We investigate and experimentally characterize a set of closely related QS signalling ligands, consisting of N-acyl homoserine lactones and the aryl homoserine lactone p-coumaroyl, as well as a set of more widely diverging QS ligands, consisting of photopyrones, dialkylresorcinols, 3,5-dimethylpyrazin-2-ol and autoinducer-2, and define their performance. We report on a set of six signal- and promoter-orthogonal intercellular QS signalling systems, significantly expanding the toolkit for engineering community-wide behaviour. Furthermore, we demonstrate that ligand diversity can serve as a statistically significant tool to predict much more complicated ligand-receptor interactions. This approach highlights the potential of dimensionality reduction to explore chemical diversity in microbial dynamics.

Study of the biofilm mechanism of C4-HSL and C6-HSL in the degradation of quinoline.

Quinoline is a hard-to-degrade organic compound widely found in coal chemical wastewater, that seriously affects the ecological environment and human health. A number of biochemical methods are already available for quinoline degradation, but the use of microbial community sensing for quinoline degradation has not been studied in depth. Therefore, this paper focuses on the enhanced mechanism of quorum-sensing signaling molecules in the biofilm formation process during quinoline degradation by functional strains of bacteria. In this paper, the effects of the signal molecules C4-HSL and C6-HSL on the adhesion ability, colony diameter, biofilm formation ability and biofilm morphology of functional strains of quinoline degrading bacteria (Ochrobactrum sp., LC-1) were investigated, and the results showed that both signal molecules promoted the biofilm formation process during the degradation of quinoline by exhibiting an efficient biofortification effect. Both signal molecules could enhance the colony diameter of strain LC-1, where C4-HSL could enhance the biomass of strain LC-1 and stimulate the secretion of extracellular polysaccharides; and C6-HSL could induce the enhancement of adhesion performance and the secretion of extracellular proteins from strain LC-1; both molecules together enhanced the biofilm formation process of strain LC-1. This study has practical application in the degradation of quinoline in coal chemical wastewater.

Quorum sensing N-acyl homoserine lactones-SdiA enhances the biofilm formation of E. coli by regulating sRNA CsrB expression

As an important virulence phenotype of Escherichia coli, the regulation mechanism of biofilm by non-coding RNA and quorum sensing system has not been clarified. Here, by transcriptome sequencing and RT-PCR analysis, we found CsrB, a non-coding RNA of the carbon storage regulation system, was positively regulated by the LuxR protein SdiA. Furthermore, β-galactosidase reporter assays showed that SdiA enhanced promoter transcriptional activity of csrB. The consistent dynamic expression levels of SdiA and CsrB during Escherichia coli growth were also detected. Moreover, curli assays and biofilm assays showed sdiA deficiency in Escherichia coli SM10λπ or BW25113 led to a decreased formation of biofilm, and was significantly restored by over-expression of CsrB. Interestingly, the regulations of SdiA on CsrB in biofilm formation were enhanced by quorum sensing signal molecules AHLs. In conclusion, SdiA plays a crucial role in Escherichia coli biofilm formation by regulating the expression of non-coding RNA CsrB. Our study provides new insights into SdiA-non-coding RNA regulatory network involved in Escherichia coli biofilm formation.

Various Biomimetics, Including Peptides as Antifungals.

Biomimetics, which are similar to natural compounds that play an important role in the metabolism, manifestation of functional activity and reproduction of various fungi, have a pronounced attraction in the current search for new effective antifungals. Actual trends in the development of this area of research indicate that unnatural amino acids can be used as such biomimetics, including those containing halogen atoms; compounds similar to nitrogenous bases embedded in the nucleic acids synthesized by fungi; peptides imitating fungal analogs; molecules similar to natural substrates of numerous fungal enzymes and quorum-sensing signaling molecules of fungi and yeast, etc. Most parts of this review are devoted to the analysis of semi-synthetic and synthetic antifungal peptides and their targets of action. This review is aimed at combining and systematizing the current scientific information accumulating in this area of research, developing various antifungals with an assessment of the effectiveness of the created biomimetics and the possibility of combining them with other antimicrobial substances to reduce cell resistance and improve antifungal effects.

Antibiotic adjuvant activities of quorum sensing signal molecules DSF and BDSF against mature biofilms of Staphylococci

Among promising antibiofilm compounds, quorum-sensing (QS) molecules that regulate biological processes such as biofilm formation and intra- or interspecies communication appear to be good candidates. The invitro antibiotic-adjuvant effects of QS molecules diffusible signal factor (DSF) and B. cenocepacia producing-DSF (BDSF) were investigated against mature Staphylococcal biofilms. Broth microdilution methods were used for the determinations of MIC, MBC, MBIC, and MBEC, and bactericidal activities were determined by TKC method. The lowest MICs were obtained with ciprofloxacin and gentamicin, and MBECs with ciprofloxacin. DSF and BDSF at 0.5 µM decreased the MICs as 2–8, and 2–32 fold, respectively. In TKC studies, –cidal activities were achieved by BDSF + gentamycin, or ciprofloxacin, and DSF + daptomycin, vancomycin, meropenem or gentamycin combinations. Synergistic effects were generally obtained with BDSF + gentamicin combinations, followed by DSF + daptomycin against most S. aureus; while BDSF + gentamicin or ciprofloxacin, and DSF + vancomycin or meropenem were synergist against some S. epidermidis biofilms. Also, the antagonist effects were observed with BDSF + meropenem or ciprofloxacin against each MSSE and MSSA. It is estimated that these QS molecules, although it was strain dependent, generally enhanced the antibiotic activity, and would be a new and effective treatment strategy for biofilm control, either alone or as an antibiotic adjuvant.

Whole-Cell Biosensors for Qualitative and Quantitative Analysis of Quorum Sensing Signal Molecules and the Investigation of Quorum Quenching Agents.

In Pseudomonas aeruginosa relevant features including virulence and biofilm formation are controlled by quorum sensing (QS), a cell density-dependent intercellular communication system based on the production and response to signal molecules. P. aeruginosa has evolved chemically distinct compounds employed as QS signal molecules (QSSMs) that can be detected and quantified through rapid, sensitive, and low-cost methods based on whole-cell biosensors. Here, we present a series of protocols based on whole-cell biosensors for qualitative and quantitative analysis of QSSMs produced by P. aeruginosa. These protocols can be used to investigate the impact of environmental conditions, genetic modifications, or quorum quenching agents on the production of QSSMs in P. aeruginosa.

Exogenous N-hexanoyl-l-homoserine lactone mitigates acid rain stress effects through modulation of structural and functional changes in Triticum aestivum leaf

Acid rain (AR) is a global environmental problem that causes a number of morphological, physiological and molecular changes in plants. N-acyl homoserine lactones, as bacterial quorum sensing signal molecules, mediate a number of physiological processes in plants. In this study, we have demonstrated for the first time the stress-protective role of AHL foliar treatment under conditions of abiotic stress - acid rain. The effects of simulated acid rain (SAR), treatment of plants with N-hexanoyl-l-homoserine lactone (C6-HSL) solution, and a combination of these two factors on the microstructure of the leaf surface, photosynthetic pigments, secondary metabolites, and lipoxygenase activity in winter wheat plants were analyzed. It was established that on the fourth day following treatment of 14-day-old plants with C6-HSL, the thickness of the outer cell wall of the leaf epidermis, together with the cuticle layer, increased by 15 %. SAR treatment resulted in a significant thinning of the cell wall (33 %), while pretreatment with C6-HSL followed by SAR reduced cell wall thickness by only 9 %. Under SAR, the cuticular wax layer was destroyed and there were unequal wax lamellae on the epidermal surface, while in C6-HSL treated plants only partial cracking of the cuticular wax layer, slight destruction of the wax lamellae, and formation of wax crusts took place. C6-HSL induced a significant increase in total chlorophyll content under non-SAR conditions, and significantly mitigated the decrease of total chlorophyll content caused by SAR. Lipoxygenase activity, the content of total phenols, flavonoids, and proline increased after treatment of plants with C6-HSL solution alone and in combination with SAR. This is the first report on the possibility of using C6-HSL as an ecological biostimulator under conditions of AR.