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Antimicrobial resistance (AMR) is an urgent worldwide health concern, requiring the exploration for novel antimicrobial interventions. A Gram-negative bacterium, Chromobacterium violaceum, synthesizes quorum-sensing-regulated violacein pigment, develops resilient biofilms, and is often used for the screening of anti-infective drugs. The aim of this work is to assess the antibacterial and antibiofilm properties of three polyphenols: 1,4-naphthoquinone, caffeic acid, and piperine. The determination of antibacterial activity was conducted by the agar overlay and broth microdilution techniques. Analysis of membrane rupture was conducted by crystal violet uptake and β-galactosidase assay. Inhibition of biofilm was evaluated using a 96-well microtiter plate assay. Biofilm structures were visualized using light, scanning electron microscopy (SEM), and confocal laser scanning electron microscopy (CLSM). Among the phytochemicals, 1,4-naphthoquinone exhibited the highest antibacterial action (25 mm zone of inhibition). The minimum inhibitory concentration of 1,4-naphthoquinone was determined to be 405 µM. Outer and inner membrane permeability was enhanced by 52.01% and 1.28 absorbance, respectively. Violacein production was reduced by 74.85%, and biofilm formation was suppressed by 63.25% at sub-MIC levels (202.5 µM). Microscopic analyses confirmed reduced adhesion on surfaces. Hemolytic activity of 1,4-naphthoquinone showed a concentration-dependent effect, with 32.16% haemolysis at 202.5 µM. Molecular docking revealed significant interactions of 1,4-naphthoquinone with DNA gyrase followed by CviR. These findings highlight 1,4-naphthoquinone's potent antibacterial efficacy against C. violaceum, proposing its use as a surface coating agent to prevent biofilm formation on medical devices, thereby offering a promising strategy to combat bacterial infections., Competing Interests: Declarations. Conflict of interest: The authors declare that they have no conflict of interest in the publication. Ethical approval: The research does not include experiments involving animals or human participants., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

A total of 52 Pasteurella multocida strains of capsular serogroups (A, B and D) were screened for anti-quorum sensing activity against Chromobacterium violaceum. Of which, 12 strains of serogroups A were found to possess anti-quorum sensing activity. Inhibition activity was highest for strain NIVEDIPm9 and lowest for strain NIVEDIPm30 based on zone of pigment inhibition. Further, cell free extract of NIVEDIPm9 strain showed highest anti-biofilm activity in reference E. coli strain and concentration dependent degradation activity of C6-AHL molecule. In whole genome sequence annotation of NIVEDIPm9 strain predicted the presence of four metallo-β-lactamases (MBL) fold metallo-hydrolase proteins. In docking studies, MBL1 and MBL3 proteins showed high binding affinity with autoinduce signalling molecules AHL compound of OH-C10, binding energy value were -6.3 and -6.2 kcal/mol. Interaction study of VAF and quorum sensing molecules showed that OmpA and HgbA proteins were stimulated by all the ten molecules (C4-AHLs, C6-AHLs, C10-AHLs, C14-AHLs, 3-oxo-C10-AHLs, 3OH-C10-HSL, C8-HSL, C10-HSL, C12-HSL, C14-HSL), while toxA gene was stimulated by OH-C10-AHL molecule, sodC gene was stimulated by none. In conclusion, we described the anti-quorum sensing activities of diverse P. multocida strains causing Pasteurellosis in livestock., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Sathish Bhadravati Shivachandra reports financial support was provided by ICAR-National Institute of Veterinary Epidemiology and Disease Informatics (NIVEDI). If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Objectives: Food products are often contaminated by pathogens and spoilage bacteria. Most of them can form biofilms, a community of cells embedded in protective extracellular matrix layers resistant to harsh conditions, including antibiotics. Therefore, alternative antibiofilm agents are required to overcome biofilm formation. This study aims to determine and quantify the antibiofilm activity of supernatants from plant-associated bacteria against biofilms of foodborne pathogen and food spoilage bacterium, namely Bacillus cereus and Bacillus subtilis., Results: Plant-associated bacteria (PAB) have shown promising antibiofilm activities against biofilm-forming pathogens in previous studies. Thirteen PAB isolated from Ternate, Indonesia were used in this study. Supernatants of PAB were subjected to antimicrobial activity and quorum quenching detection, both using the well diffusion method. Four supernatants inhibited the growth of B. subtilis, but none affected the growth of B. cereus. Eight supernatants were able to disrupt the quorum sensing system of an indicator bacterium, wild-type Chromobacterium violaceum. Biofilm inhibition and destruction were quantified using 96-well microplates. The highest biofilm inhibition and destruction activities of PAB supernatants against each of B. cereus and B. subtilis biofilms were > 76%, and were later confirmed by light microscope and scanning electron microscope. Brine shrimp lethality assay (BSLA) was conducted and revealed that the selected PAB supernatants were non-toxic. The 16S rRNA gene of PAB were sequenced and they showed similarities to Bacillus, Priestia, and Chryseobacterium. Compounds in the supernatants were determined by GC-MS which revealed contents of fatty acids, ethyl esters, and diketopiperazines. Therefore, PAB supernatants have potential as antibiofilm agents against biofilm formed by Bacillus cereus and Bacillus subtilis., Competing Interests: Declarations Ethics approval and consent to participate Not applicable. Consent for publication Not applicable. Competing interests The authors declare no competing interests., (© 2024. The Author(s).)

Objectives: This research aimed to identify and quantify the antibiofilm activity of bioactive compounds from bacteria isolated from rhizosphere and nodule butterfly pea (Clitoria ternatea), rhizosphere clove afo 3 (Syzygium aromaticum), nodule mimosa (Mimosa pudica L.), and soil from gold mining land which were recovered from Ternate, Tidore, Obi Island, and Marotai Island, Eastern part of Indonesia., Results: Eight supernatants from soil and plant-associated bacteria were found to have quorum quenching activity against Chromobacterium violaceum. All supernatants exhibited antibiofilm activity against biofilm formed by Aeromonas hydrophila and Vibrio harveyi. The supernatant of FT5 showed the highest activity in disrupting (66.59%) and inhibiting (85.63%) the biofilm of A. hydrophila. For V. harveyi, the supernatant of PTM3 showed the highest disruption activity (72.61%), whileRCA7 showed the highest inhibition activity(75.68%). The Gas Chromatography-Mass Spectrometry (GC-MS) identified fatty acids, ester, and diketopiperazine as the compounds related to the antibiofilm activity. Molecular identification revealed that the isolates belong to the genera Bacillus, Priestia, and Chryseobacterium., (© 2024. The Author(s).)

Quorum sensing (QS) is pivotal in coordinating virulence factors and biofilm formation in various pathogenic bacteria, making it a prime target for disrupting bacterial communication. Pseudomonas aeruginosa is a member of the "ESKAPE" group of bacterial pathogens known for their association with antimicrobial resistance and biofilm formation. The current antibiotic arsenal falls short of addressing biofilm-related infections effectively, highlighting the urgent need for novel therapeutic agents. In this study, we explored the anti-QS and anti-biofilm properties of theophylline against two significant pathogens, Chromobacterium violaceum and P. aeruginosa. The production of violacein, pyocyanin, rhamnolipid, and protease was carried out, along with the evaluation of biofilm formation through methods including crystal violet staining, triphenyl tetrazolium chloride assay, and fluorescence microscopy. Furthermore, computational analyses were conducted to predict the targets of theophylline in the QS pathways of P. aeruginosa and C. violaceum. Our study demonstrated that theophylline effectively inhibits QS activity and biofilm formation in C. violaceum and P. aeruginosa. In P. aeruginosa, theophylline inhibited the production of key virulence factors, including pyocyanin, rhamnolipid, protease, and biofilm formation. The computational analyses suggest that theophylline exhibits robust binding affinity to CviR in C. violaceum and RhlR in P. aeruginosa, key participants in the QS-mediated biofilm pathways. Furthermore, theophylline also displays promising interactions with LasR and QscR in P. aeruginosa. Our study highlights theophylline as a versatile anti-QS agent and offers a promising avenue for future research to develop novel therapeutic strategies against biofilm-associated infections., (© 2024. The Author(s), under exclusive licence to Springer Nature Switzerland AG.)

The leaves of Piper betle L., known as betel leaf, have immense medicinal properties. It possesses potent antimicrobial efficacies and can be a valuable tool to combat drug-resistant microorganisms. Quorum sensing (QS) inhibition is one of the best strategies to combat drug resistance. The present study investigates the anti-quorum sensing and biofilm inhibitory potential of Piper betle L. leaf extract against two bacterial strains, Chromobacterium violaceum and Pseudomonas aeruginosa. The extract produced substantial QS-inhibition zones in a biosensor strain of C. violaceum (CV026), indicating interference with quorum-sensing signals. The Results demonstrated significant inhibition in biofilm formation and different QS-regulated virulence factors (violacein, exopolysaccharides, pyocyanin, pyoverdine, elastase) in both C. violaceum and P. aeruginosa at sub-MIC concentrations of the extract and tetracycline, an antibiotic with known anti-QS activity. The quantitative real-time PCR (qRT-PCR) revealed decreased gene expression in different QS-related genes in C. violaceum (cviI, cviR, and vioA) and P. aeruginosa (lasI, lasR, lasB, rhlI, rhlR, and rhlA) strains after treatment. Gas Chromatography-Mass Spectrometry (GC-MS) analysis identified the significant phytocompounds, mainly derivatives of chavicol and eugenol, in the extract. Of these compounds, chavicol acetate (affinity: -7.00 kcal/mol) and acetoxy chavicol acetate (affinity: -7.87 kcal/mol) showed the highest potential to bind with the CviR and LasR protein, respectively, as evident from the in-silico molecular docking experiment. The findings of this endeavour highlight the promising role of Piper betle L. as a source of natural compounds with anti-quorum sensing properties against pathogenic bacteria, opening avenues for developing novel therapeutic agents to combat bacterial infections., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. Bratati Sikdar reports financial support was provided by University Grants Commission. Sourav Mukherjee reports financial support was provided by University Grants Commission. Adarsha Raj reports financial support was provided by University Grants Commission. Debarati Banerjee reports financial support was provided by University Grants Commission. All the authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Inter-cellular signaling, referred to as quorum sensing (QS), regulates the production of virulence factors in numerous gram-negative bacteria, such as the human pathogens Pseudomonas aeruginosa and Chromobacterium violaceum. QS inhibition may provide an opportunity for the treatment of bacterial infections. This represents the initial study to examine the antibiofilm and antivirulence capabilities of rose absolute and its primary component, phenylethyl alcohol. QS inhibition was assessed by examining extracellular exopolysaccharide synthesis, biofilm development, and swarming motility in P. aeruginosa PAO1, along with violacein production in C. violaceum ATCC 12472. Molecular docking analysis was conducted to explore the mechanism by which PEA inhibits QS. Our results indicate that rose absolute and PEA caused decrease in EPS production (60.5-33.5%), swarming motility (94.7-64.5%), and biofilm formation (98.53-55.5%) in the human pathogen P. aeruginosa PAO1. Violacein production decreased by 98.1% and 62.5% with an absolute (0.5 v/v %) and PEA (2 mM). Moreover, the molecular docking analysis revealed a promising competitive interaction between PEA and AHLs. Consequently, this study offers valuable insights into the potential of rose absolute and PEA as inhibitors of QS in P. aeruginosa and C. violaceum., (© 2024. The Author(s).)

In last two decades, the world has seen an exponential increase in the antimicrobial resistance (AMR), making the issue a serious threat to human health. The mortality caused by AMR is one of the leading causes of human death worldwide. Till the end of the twentieth century, a tremendous success in the discovery of new antibiotics was seen, but in last two decades, there is negligible progress in this direction. The increase in AMR combined with slow progress of antibiotic drug discovery has created an urgent demand to search for newer methods of intervention to combat infectious diseases. One of such approach is to look for biofilm and quorum sensing (QS) inhibitors. Plants are excellent source of wide class compounds that can be harnessed to look for the compounds with such properties. This study proves a broad-spectrum biofilm and QS inhibitory potential of umbelliferone. More than 85% reduction in violacein production Chromobacterium violaceum 12472 was found. All tested virulent traits of Pseudomonas aeruginosa PAO1 and Serratia marcescens MTCC 97 were remarkably inhibited that ranged from 56.62% to 86.24%. Umbelliferone also successfully prevented the biofilm of test bacteria at least by 67.68%. Umbelliferone interacted at the active site of many proteins of QS circuit, which led to the mitigation of virulent traits. The stable nature of complexes of umbelliferone with proteins further strengthens in vitro results. After examining the toxicological profile and other drug-like properties, umbelliferone could be potentially developed as new drug to target the infections caused by Gram - ve bacteria.Communicated by Ramaswamy H. Sarma.

Natural products have a long history of success in treating bacterial infections, making them a promising source for novel antibacterial medications. Curcumin, an essential component of turmeric, has shown potential in treating bacterial infections and in this study, we covalently immobilized curcumin (Cur) onto chitosan (CS) using glutaraldehyde and tannic acid (TA), resulting in the fabrication of novel biocomposites with varying CS/Cur/TA ratios. Comprehensive characterization of these ternary biocomposites was conducted using FTIR, SEM, XPS, and XRD to assess their morphology, functional groups, and chemical structures. The inhibitory efficacy of these novel biocomposites (n = 4) against the growth and viability of Pseudomonas aeruginosa (ATCC27853) and Chromobacterium violaceum (ATCC12472) was evaluated and the most promising composite (C3) was investigated for its impact on quorum sensing (QS) and biofilm formation in these bacteria. Remarkably, this biocomposite significantly disrupted QS circuits and effectively curtailed biofilm formation in the tested pathogens without inducing appreciable toxicity. These findings underscore its potential for future in vivo studies, positioning it as a promising candidate for the development of biofilm disrupting antibacterial agents., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

Pseudomonas aeruginosa causes life-threatening diseases and is resistant to almost all conventional antibiotics. The quorum sensing (QS) system of P. aeruginosa contributes to many pathogenic factors some of which are pigment production, motility, and biofilm. The disruption of quorum sensing system may be an impactful strategy to deal with infections. The present study investigates the anti-quorum sensing property of a bioactive molecule extracted from marine epibiotic bacteria present on the surface of seaweeds. Among all the isolates tested against monitor strain Chromobacterium violaceum (MTCC 2656), the one with the highest activity was identified as Bacillus zhangzhouensis SK4. The culture supernatant was extracted with chloroform which was then partially purified by TLC and column chromatography. The probable anti-QS compound was identified as 1,2-benzenedicarboxylic acid, bis (2-methylpropyl ester) by GC-MS and NMR analysis. The treatment of P. aeruginosa MCC 3457 with the lead compound resulted in the reduced production of pyocyanin, rhamnolipids, exopolysaccharide, biofilm, and motility. The observations of light and scanning electron microscopy also supported the biofilm inhibition. The lead compound showed synergism with the meropenem antibiotic and significantly reduced MIC. The molecular docking and pharmacokinetics study predicted 1, 2-benzenedicarboxylic acid, bis (2-methylpropyl ester), a phthalate derivative as a good drug candidate. The molecular dynamics study was also performed to check the stability of the lead compound and LasR complex. Further, lead compounds did not exhibit any cytotoxicity when tested on human embryonic kidney cells. As per our knowledge, this is the first report on the anti-QS activity of B. zhangzhouensis SK4, indicating that epibiotic bacteria can be a possible source of novel compounds to deal with the multidrug resistance phenomenon., Competing Interests: Declaration of competing interest There is no conflict of interest., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Pseudomonas aeruginosa is an opportunistic bacterium causing several health problems and having many virulence factors like biofilm formation on different surfaces. There is a significant need to develop new antimicrobials due to the spreading resistance to the commonly used antibiotics, partly attributed to biofilm formation. Consequently, this study aimed to investigate the anti-biofilm and anti-quorum sensing activities of Dioon spinulosum, Dyer Ex Eichler extract (DSE), against Pseudomonas aeruginosa clinical isolates. DSE exhibited a reduction in the biofilm formation by P. aeruginosa isolates both in vitro and in vivo rat models. It also resulted in a decrease in cell surface hydrophobicity and exopolysaccharide quantity of P. aeruginosa isolates. Both bright field and scanning electron microscopes provided evidence for the inhibiting ability of DSE on biofilm formation. Moreover, it reduced violacein production by Chromobacterium violaceum (ATCC 12,472). It decreased the relative expression of 4 quorum sensing genes (lasI, lasR, rhlI, rhlR) and the biofilm gene (ndvB) using qRT-PCR. Furthermore, DSE presented a cytotoxic activity with IC 50 of 4.36 ± 0.52 µg/ml against human skin fibroblast cell lines. For the first time, this study reports that DSE is a promising resource of anti-biofilm and anti-quorum sensing agents., (© 2022. The Author(s).)

With an upsurge in multidrug resistant bacteria backed by biofilm defence armours, there is a desperate need of new antibiotics with a non-traditional mechanism of action. Targeting bacteria by misguiding them or halting their communication is a new approach that could offer a new way to combat the multidrug resistance problem. Quorum sensing is considered to be the achilles heel of bacteria that has a lot to offer. Since, both quorum sensing and biofilm formation have been related to drug resistance and pathogenicity, in this study we synthesised new derivatives of citral with antiquorum sensing and biofilm disrupting properties. We previously reported antimicrobial and antiquorum sensing activity of citral and herein we report the synthesis and evaluation of citral and its derivatives (CD1-CD3) for antibacterial, antibiofilm and antiquorum sensing potential against Chromobacterium violaceum using standard methods. Preliminary results revealed that CD1 is the most active of all the derivatives. Qualitative and quantitative evaluation of antiquorum sensing activity at sub-inhibitory concentrations of these compounds also revealed high activity for CD1 followed by CD2, CD3 and citral. These compounds also inhibit biofilm formation at subinhibitory concentrations without causing any bacterial growth inhibition. These results were replicated by RT-qPCR with down regulation of the quorum sensing genes when C. violaceum was treated with these test compounds. Overall, the results are quite encouraging, revealing that biofilm and quorum sensing are interrelated processes and also indicating the potential of these derivatives to impede bacterial communication and biofilm formation.

The essential oil (EO) from the roots of Plectranthus barbatus Andr. (Syn. Coleus forskohlii Briq.) was evaluated for quorum sensing (QS) inhibitory activity. P. barbatus EO was screened for inhibition of QS regulated violacein production in Chromobacterium violaceum (ATCC 12472) wild-type strain. At inhibitory (6.25% v/v) and sub-inhibitory concentrations (3.125% v/v) of the EO, dose-dependent response in the inhibition of violacein production was observed in C. violaceum. Similarly, sub-MIC (6.25% v/v) of P. barbatus EO disrupted QS regulated biofilm formation by 27.87% and inhibited swarming and twitching motility in Pseudomonas aeruginosa PA01 implying its anti-infective and QS modulatory activity. Fluorescence microscopy studies confirmed the disruption of biofilm formation by EO in P. aeruginosa PAO1. Promising antibacterial activity was recorded at concentrations as low as 3.12% v/v for Listeria monocytogenes (ATCC 13932) and at 6.25% v/v for both Salmonella enterica subsp. enterica serovar Typhimurium (ATCC 25241) and Escherichia coli (ATCC 11775). Furthermore, significant dose-dependent inhibition was observed for biofilm formation and motility in all the tested pathogens in different treated concentrations. GC-MS analysis revealed α-pinene, endo-borneol, bornyl acetate, 1-Hexyl-2-Nitrocyclohexane as the major phytoconstituents. P. barbatus EO or its constituent compounds with QS modulatory, antimicrobial and biofilm inhibitory property could be potential new-age dietary source based intervention and preservation technologies.

Pseudomonas aeruginosa is one of the vulnerable opportunistic pathogens associated with nosocomial infections, cystic fibrosis, burn wounds and surgical site infections. Several studies have reported that quorum sensing (QS) systems are controlled the P. aeruginosa pathogenicity. Hence, the targeting of QS considered as an alternative approach to control P. aeruginosa infections. This study aimed to evaluate the anti-quorum sensing and antibiofilm inhibitory potential of Musa paradisiaca against Chromobacterium violaceum (ATCC 12472) and Pseudomonas aeruginosa. The methanol extract of M. paradisiacsa exhibits that better antibiofilm potential against P. aeruginosa. Then, the crude methanol extract was subjected to purify by column chromatography and collected the fractions. The mass-spectrometric analysis of a methanol extract of M. paradisiaca revealed that 1,8-cineole is the major compounds. 1, 8-cineole significantly inhibited the QS regulated violacein production in C. violaceum. Moreover, 1,8-cineole significantly inhibited the QS mediated virulence production and biofilm formation of P. aeruginosa without affecting their growth. The real-time PCR analysis showed the downregulation of autoinducer synthase and transcriptional regulator genes upon 1,8-cineole treatment. The findings of the present study strongly suggested that metabolite of M. paradisiaca impedes P. aeruginosa QS system and associated virulence productions.

Background: Indonesia is the third largest producer of fish and other aquaculture products in the world, making this industry a major contributor in the economy of Indonesia. However, this industry continually overcome challenges, one of them are bacterial outbreaks. In addition, the emergence of these bacterial outbreaks were worsen due to the biofilm produced by many significant pathogenic bacteria and the impact of increased antibiotic resistance. These issues have become a global concern, because antibiotics are currently one of the main treatments available to overcome this problems. Therefore, studies aimed at finding and characterizing bioactive compounds to combat these issues. In this study actinomycetes isolates were screened and characterized for their bioactive compounds produced which have inhibitory and destructive activity and also QS inhibitors against biofilm structure of aquatic pathogenic bacteria, such as Vibrio harveyi, A. hydrophila, and S. agalactiae., Result: Extracts (20 mg/mL) produced by sixteen Actinomycetes isolates showed anti-quorum sensing activity towards reporter stain Chromobacterium violaceum wild-type. Most of these extracts showed better inhibitory activity on all of the pathogenic bacteria biofilm structure tested than the destructive activity on the preformed of those biofilm structure. Subsequently, we also performed characterization of bioactive compound and found that in this study, polysaccharide is the most common antibiofilm agents, which were responsible to their antibiofilm activity. Finally, we found that the value of LC 50 of all extracts tested were more than 1 mg/mL, thereby all of extracts tested did not show cyto-toxic effect against Artemia salina., Conclusion: All of the extracts of Actinomycetes isolates showed promising inhibitory activity towards biofilm structure of pathogenic bacteria tested. So far, all of the extracts are potential to be QS inhibitors and antibiofilm agents of all pathogenic bacteria tested.

Antimicrobial resistance among pathogenic bacteria has become a global threat to human health. Due to poor progress in development of new antimicrobial drugs, there is a need for the development of novel alternative strategies to combat the problem of multidrug resistance. Moreover, there is focus on ecofriendly approach for the synthesis nanoparticles having efficient medicinal properties including antivirulence properties to tackle the emergence of multi-drug resistance. Targeting quorum sensing controlled virulence factors and biofilms has come out to be a novel anti-infective drug target. The silver nanoparticles (Ag@CC-NPs) were synthesized from aqueous extract of Carum copticum and characterized using UV-vis absorption spectroscopy, fourier-transform infrared spectroscopy (FTIR), X-ray diffraction (XRD), transmission electron microscopy (TEM), and scanning electron microscopy (SEM). Ag@CC-NPs were checked for its ability to inhibit quorum sensing-mediated virulence factors and biofilms against three test pathogens at sub-MIC values. There was ~75% inhibition of violacein production by Ag@CC-NPs against C. violaceum. The P. aeruginosa virulence factors such as pyocyanin production, pyoverdin production, exoprotease activity, elastase activity, swimming motility and rhamnolipid production were inhibited by 76.9, 49.0, 71.1, 53.3, 89.5, and 60.0% at sub-MIC. Moreover, virulence factors of S. marcescens viz. prodigiosin production, exoprotease activity, and swarming motility was reduced by 78.4, 67.8, and 90.7%. Ag@CC-NPs also exhibited broad-spectrum antibiofilm activity with 77.6, 86.3, and 75.1% inhibition of biofilms of P. aeruginosa, S. marcescens, and C. violaceum respectively. The biofilm formation on glass coverslip was reduced remarkably as evident from SEM and CLSM analysis. The findings revealed the in vitro efficacy of Ag@CC-NPs against bacterial pathogens and can be exploited in the development of alternative therapeutic agent in management of bacterial infections for topical application, mainly wound infection, or coating of surfaces to prevent bacterial adherence on medical devices., Competing Interests: Declaration of competing interest The authors declare that there is no conflict of interest., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

The present study evaluated the efficacy of Y 2 O 3 :Tb (core) and Y 2 O 3 :Tb@SiO 2 nanospheres (core/shell NSs) against virulence functions regulated by quorum sensing (QS) and biofilm formation in pathogenic bacteria. Scanning electron microscope (SEM) images were used to study the size, shape, and morphology. The images clearly displayed spherical shaped, mono-dispersed particles with narrow size distribution and an average grain size of 110-130 nm. The chemical composition of the samples was determined by using energy dispersive X-ray (EDX) and X-ray photoelectron spectroscopy (XPS). We determined the impact of core and core/shell NSs on QS using sensor strains of Chromobacterium violaceum CVO26 and Pseudomonas aeruginosa PAO1 in a comparative study. Sub-MICs of core and core/shell NSs substantially suppressed QS-controlled violacein production in C. violaceum. Similar concentration-dependent effect of sub-MICs of synthesized core and core/shell NSs was observed in the QS-regulated virulence functions (elastase, total protease, pyocyanin production, swarming motility, and exopolysaccharide production) in PAO1. A concentration-dependent decrease (14-60%) was recorded in the biofilm forming capability of PAO1, upon treatment with core and core/shell NSs. Moreover, core/shell NSs were more effective in inhibiting biofilm at higher tested concentrations as compared to core-NSs. The synthesized NSs demonstrated significantly impaired attachment of cells to the microtiter plate indicating that NSs target biofilm inhibition at the attachment stage. Based on these results, we predict that core and core/shell NSs may be an alternative to combat the threat of drug-resistant pathogenic bacteria.

In this study, 10 different sulfide flavor compounds commonly used as food additives were screened for antiquorum-sensing activity. Among these, diallyl disulfide (DADS) and methyl 2-methyl-3-furyl disulfide (MMFDS) were found to exert the strongest inhibition against violacein production in Chromobacterium violaceum 026, the tested biosensor strain. DADS and MMFDS also inhibited the growth of Hafnia alvei H4, yielding MIC values of 48 and 41.6 mM, respectively. In addition, DADS and MMFDS also inhibited the ability of H. alvei H4 to produce acyl-homoserine lactone as demonstrated by the reduced level of C6-HSL in the supernatant of DADS-treated culture. At concentrations corresponding to 1/4 MIC, DADS, and MMFDS inhibited the swarming ability of H. alvei H4 by 73.50% and 76.43%, respectively, while having virtually no effect on cell growth. The same concentrations of DADS and MMFDS also completely inhibited the formation of biofilm. These antiquorum sensing effects of DADS and MMFDS involved changes in the expression of the quorum-sensing genes luxI and luxR. Quantitative RT-PCR analysis showed that the mRNA levels of both genes were significantly reduced by DADS and MMDFS at concentrations below their MICs. However, further test using a mutant strain of H. alvei lacking luxR (ΔluxR) revealed significant reduction in luxI mRNA level upon treatment of the strain with DADS or MMDFS, but no change in luxR mRNA level occurred when a luxI-lacking mutant (ΔluxI) was treated with these compounds. The result therefore suggested that the antiquorum-sensing effect of DADS and MMFDS against H. alvei H4 might operate mainly through the inhibition of luxI expression in the cells., Practical Application: The sulfide flavors compounds used in this paper are commonly used in food processing in China and are listed in the national standard of Chinese food additives GB2760-2014. The application of sulfide flavors in food processing can enhance aroma and prevent food spoilage., (© 2018 Institute of Food Technologists®.)

In most bacteria, a global level of regulation, termed quorum sensing (QS), exists involving intercellular communication via the production and response to cell density-dependent signal molecules. QS has been associated with a number of important features in bacteria including virulence regulation and biofilm formation. Consequently, there is considerable interest in understanding, detecting, and inhibiting QS. N-acylated homoserine lactones (AHLs) are used as extracellular QS signals by a variety of Gram-negative bacteria. Chromobacterium violaceum, commonly found in soil and water, produces the characteristic purple pigment violacein, regulated by AHL-mediated QS. Based on this readily observed pigmentation phenotype, C. violaceum strains can be used to detect various aspects of AHL-mediated QS activity. In another commonly used bioassay organism, Agrobacterium tumefaciens, QS can be detected by the use of a reporter gene such as lacZ. Here, we describe several commonly used approaches incorporating C. violaceum and A. tumefaciens that can be used to detect AHL and QS inhibitors. Due to the inherent low susceptibility of biofilm bacteria to antimicrobial agents, biofilm dispersion, whereby bacteria reenter the planktonic community, is another increasingly important area of research. At least one signal, distinct from traditional QS, has been identified and there are a variety of other environmental factors that also trigger dispersion. We describe a microtiter-based experimental strategy whereby potential biofilm dispersion compounds can be screened.

The present study explores the non-bactericidal anti-virulence efficacy of green synthesized silver nanoparticles (AgNPs) from Gelidiella acerosa against multi-drug resistant Vibrio spp. Spectral characterization of AgNPs was performed through UV-Visible, FT-IR, and energy-dispersive spectroscopic techniques followed by X-ray crystallography and zeta potential analysis. Further, the structural characterization was done by electron and atomic force microscopic techniques. AgNPs profoundly quelled the quorum sensing mediated violacein production in Chromobacterium violaceum and CV026. Characterized AgNPs at 100 μg mL -1 concentrations depicted a phenomenal anti-biofilm efficacy against Vibrio parahaemolyticus (71%) and Vibrio vulnificus (83%) biofilms, which was further confirmed through light, confocal, and scanning electron microscopic analyses. In vitro bioassays revealed the remarkable inhibitory values of AgNPs, by inhibiting the exopolysaccharide production, hydrophobicity, and motility. In vivo studies using Artemia franciscana larvae also confirmed the anti-infective proficiency, as the AgNPs effectively reduced the bacterial colonization and enhanced the survival rate of larvae up to 100% without any toxicity effect. Graphical abstract Rapid biosynthesized AgNPs from Gelidiella acerosa quench quorum sensing controlled virulence traits in vibrios.

Emergence of antibiotic resistance among pathogenic bacteria encourages us to search for new molecules as an alternative treatment. The aim of this study was to evaluate the antiquorum sensing (anti-QS) and antibiofilm potential of Salvadora persica L. methanolic extracts to prevent the infections due to Staphylococcus as an alternate to antibiotics. The methanolic extracts of S. persica L. fruit, leaves and stems was assessed for their activity in inhibiting QS-depedent phenomenon such as violacein pigment production in Chromobacterium violaceum, swarming motility of Pseudomonas aeruginosa PAO1 and biofilm formation in oral Staphylococcus strains on polymethylmetacrylate (PMMA). Methanolic fruit extract of S. persica L. showed a high degree of anti-biofilm formation on PMMA and on violacein inhibition with a percentage of reduction equal to 90% when MIC value (20 mg/ml) was used. 100 μg/ml of S. persica L. leaves exhibited inhibition in swarming motility of PAO1 at 29.17%. Because the methanolic extracts of S. persica L. demonstrated anti-QS and antibiofilm activity at very low concentrations, it could be further exploited for novel molecules to treat oral Staphylococcus infections., (Published by Elsevier Ltd.)

Multidrug-resistance bacteria commonly use cell-to-cell communication that leads to biofilm formation as one of the mechanisms for developing resistance. Quorum sensing inhibition (QSI) is an effective approach for the prevention of biofilm formation. A Gram-negative bacterium, Delftia tsuruhatensis SJ01, was isolated from the rhizosphere of a species of sedge ( Cyperus laevigatus ) grown along the coastal-saline area. The isolate SJ01 culture and bacterial crude extract showed QSI activity in the biosensor plate containing the reference strain Chromobacterium violaceum CV026. A decrease in the violacein production of approximately 98% was detected with the reference strain C. violaceum CV026. The bacterial extract (strain SJ01) exhibited anti-quorum sensing activity and inhibited the biofilm formation of clinical isolates wild-type Pseudomonas aeruginosa PAO1 and P. aeruginosa PAH. A non-toxic effect of the bacterial extract (SJ01) was detected on the cell growth of the reference strains as P. aeruginosa viable cells were present within the biofilm. It is hypothesized that the extract (SJ01) may change the topography of the biofilm and thus prevent bacterial adherence on the biofilm surface. The extract also inhibits the motility, virulence factors (pyocyanin and rhamnolipid) and activity (elastase and protease) in P. aeruginosa treated with SJ01 extract. The potential active compound present was identified as 1,2-benzenedicarboxylic acid, diisooctyl ester. Microarray and transcript expression analysis unveiled differential expression of quorum sensing regulatory genes. The key regulatory genes, LasI, LasR, RhlI , and RhlR were down-regulated in the P. aeruginosa analyzed by quantitative RT-PCR. A hypothetical model was generated of the transcriptional regulatory mechanism inferred in P. aeruginosa for quorum sensing, which will provide useful insight to develop preventive strategies against the biofilm formation. The potential active compound identified, 1,2-benzenedicarboxylic acid, diisooctyl ester, has the potential to be used as an anti-pathogenic drug for the treatment of biofilm-forming pathogenic bacteria. For that, a detailed study is needed to investigate the possible applications.

The present study investigates the role of quorum sensing (QS) molecules expressed by C. sakazakii in biofilm formation and extracellular polysaccharide expression. The QS signaling was detected using Chromobacterium violaceum 026 and Agrobacterium tumefaciens NTL4(pZLR4) based bioassay. Long chain N-acyl-homoserine lactones (AHLs) with C6- C18 chain length were identified using High Performance Liquid Chromatography and Liquid Chromatography-High Resolution Mass Spectrometry. A higher Specific Biofilm Formation (SBF) index (p < 0.05) with the presence of genes associated with cellulose biosynthesis (bcsA, bcsC and bcsG) was observed in the strains. AHLs and their mechanisms can serve as novel targets for developing technologies to eradicate and prevent biofilm formation by C. sakazakii.

Quorum sensing (QS) is a cell density dependent expression of species in bacteria mediated by compounds called autoinducers (AI). Several processes responsible for successful establishment of bacterial infection are mediated by QS. Inhibition of QS is therefore being considered as a new target for antimicrobial chemotherapy. Flavonoid compounds are strong antioxidant and antimicrobial agents but their applications are limited due to their poor dissolution and bioavailability. Our objective was to investigate the effect of kaempferol loaded chitosan nanoparticles on modulating QS mediated by AI in model bioassay test systems. For this purpose, kaempferol loaded nanoparticles were synthesized and characterized in terms of hydrodynamic diameter, hydrogen bonding, amorphous transformation and antioxidant activity. QS inhibition in time dependent manner of nanoparticles was measured in violacein pigment producing using the biosensor strain Chromobacterium violaceum CV026 mediated by AI known as acylated homoserine lactone (AHL). Our results indicated that the average kaempferol loaded chitosan/TPP nanoparticle size and zeta potential were 192.27±13.6nm and +35mV, respectively. The loading and encapsulation efficiency of kaempferol into chitosan/TPP nanoparticles presented higher values between 78 and 93%. Kaempferol loaded chitosan/TPP nanoparticle during the 30 storage days significantly inhibited the production of violacein pigment in Chromobacterium violaceum CV026. The observation that kaempferol encapsulated chitosan nanoparticles can inhibit QS related processes opens up an exciting new strategy for antimicrobial chemotherapy as stable QS-based anti-biofilm agents., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Many bacteria use small diffusible signaling molecules to communicate each other termed as quorum sensing (QS). Most Gram-negative bacteria use acyl homoserine lactone (AHL) as QS signal molecules. Using these signaling molecules, bacteria are able to express specific genes in response to population density. This work aimed to detect the production of QS signal molecules and biofilm formation in Ralstonia solanacearum isolated from various diseased tomato plants with symptoms of bacterial wilt. A total of 30 R. solanacearum strains were investigated for the production of QS signal molecules using Chromobacterium violaceum CV026 and Agrobacterium tumefaciens NT1 (pZLR4) biosensor systems. All 30 bacterial isolates from various bacterial wilt-affected tomato plants produced AHL molecules that induced the biosensor. The microtiter plate assay demonstrated that of the 30 bacterial isolates, 60 % formed biofilm, among which four isolates exhibited a higher degree of biofilm formation. The biofilm-inducing factor was purified from these four culture supernatants. The structure of the responsible molecule was solved using nuclear magnetic resonance and mass spectroscopy and was determined to be 2-hydroxy-4-((methylamino)(phenyl)methyl) cyclopentanone (HMCP), which was confirmed by chemical synthesis and NMR. The Confocal laser scanning microscopic analysis showed well-developed biofilm architecture of bacteria when treated with HMCP. The knowledge we obtained from this study will be useful for further researcher on the role of HMCP molecule in biofilm formation.

Pomegranate rind has been found to inhibit numerous pathogens, mostly attributed to its tannin fraction. The present study was conducted to investigate the quorum sensing (QS) inhibition effect of tannin-rich fraction from pomegranate rind (TFPR) by using an indicator strain Chromobacterium violaceum. Meanwhile, its effect on biofilm formation and motility of Escherichia coli was evaluated. It was shown that TFPR inhibited QS-regulated violacein pigment production. Biofilm formation and motility of E. coli were also hindered by TFPR. Transcriptional analysis further showed that TFPR repressed expressions of curli genes (csgB and csgD) and various motility genes (fimA, fimH, flhD, motB, qseB, and qseC). Our findings indicated that TFPR has potential application for controlling E. coli contaminations or biofilms in the food industry.

Quorum sensing has been shown to play a crucial role in Pseudomonas aeruginosa pathogenesis where it activates expression of myriad genes that regulate the production of important virulence factors such as biofilm formation. Antagonism of quorum sensing is an excellent target for antimicrobial therapy and represents a novel approach to combat drug resistance. In this study, Chromobacterium violaceum biosensor strain was employed as a fast, sensitive, reliable, and easy to use tool for rapid screening of soil samples for Quorum Sensing Inhibitors (QSI) and the optimal conditions for maximal QSI production were scrutinized. Screening of 127 soil isolates showed that 43 isolates were able to breakdown the HHL signal. Out of the 43 isolates, 38 isolates were able to inhibit the violet color of the biosensor and to form easily detectable zones of color inhibition around their growth. A confirmatory bioassay was carried out after concentrating the putative positive cell-free lysates. Three different isolates that belonged to Bacillus cereus group were shown to have QSI activities and their QSI activities were optimized by changing their culture conditions. Further experiments revealed that the cell-free lysates of these isolates were able to inhibit biofilm formation by P. aeruginosa clinical isolates., (© 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Chromobacterium violaceum is an opportunistic pathogen that causes infections that are difficult to treat. The goal of this research was to evaluate the effect of selected tannins (tannic acid (TA) and gallic acid (GA)) on bacterial growth, motility, antibiotic (carbenicillin, tetracycline) susceptibility, and biofilm formation. Both tannins, particularly TA, impaired bacterial growth levels and swimming motilities at sub-minimum inhibitory concentrations (sub-MICs). In combination with tannins, antibiotics showed increased MICs, suggesting that tannins interfered with antibacterial activity. Sub-MICs of tetracycline or TA alone enhanced biofilm formation of C. violaceum; however, in combination, these compounds inhibited biofilm formation. In contrast, carbenicillin at sub-MICs was effective in inhibiting C. violaceum biofilm formation; however, in combination with lower concentrations of TA or GA, biofilms were enhanced. These results provide insights into the effects of tannins on C. violaceum growth and their varying interaction with antibiotics used to target C. violaceum infections.

Bacterial urinary tract infections (UTIs) are the most common nosocomial infections, accounting for about 40 % of all hospital-acquired infections. The bacterial spectrum of nosocomial UTIs is broad and the treatment of UTIs is becoming difficult owing to the emergence of drug resistance. Therefore, it is reasonable to investigate novel and alternative therapeutic strategies to treat UTIs. Since UTIs are caused by uropathogens with quorum sensing (QS)-dependent biofilm forming abilities, interruption of QS systems may be a novel approach to combat drug resistance. In the present study, a methanol extract (and hexane extract derived from it) of the medicinal plant Hyptis suaveolens (L.) were shown to have anti-QS activity against the biosensor strain Chromobacterium violaceum (ATCC 12472). Furthermore, the hexane extract of H. suaveolens (HEHS) inhibited biofilm formation by uropathogens such as Escherichia coli, Proteus vulgaris, Proteus mirabilis, Klebsiella pneumoniae and Serratia marcescens. HEHS promotes the loosening of biofilm architecture and strongly inhibits in vitro biofilm formation by uropathogens, which was more apparent from microscopic images. In addition to this, HEHS reduces the production of QS-dependent virulence factors like protease and hemolysin, along with motility. The partial purification and GC-MS analysis of the active fraction revealed the presence of several therapeutically important compounds which may synergistically act on the uropathogens and possibly reduce the QS-dependent phenotypes. These findings suggest HEHS as potential phytotherapeutic agent which can be employed to formulate protective strategies against biofilm linked infections caused by uropathogens.

The aim of this study was to evaluate the anti-biofilm and quorum sensing inhibitory (QSI) potential of tender coconut water (TCW) against Chromobacterium violaceum and Pseudomonas aeruginosa. TCW significantly inhibited the QS regulated violacein, virulence factors and biofilm production without affecting their growth. qRT-PCR analysis revealed the down-regulation of autoinducer synthase, transcriptional regulator and virulence genes. Mass-spectrometric analysis of a petroleum ether extract of the TCW hydrolyte revealed that 2-furaldehyde diethyl acetal (2FDA) and palmitic acid (PA) are the major compounds. In vitro bioassays confirmed the ability of 2FDA to inhibit the biofilm formation and virulence factors. In addition, the combination of PA with 2FDA resulted in potent inhibition of biofilm formation and virulence factors. The results obtained strongly suggest that TCW can be exploited as a base for designing a novel antipathogenic drug formulation to treat biofilm mediated infections caused by P. aeruginosa.

Growth of members of most of the studied genera of gram-positive (Dietzia, Kocuria, and Rhodo- coccus) and gram-negative bacteria (Pseudomonas and Chromobacterium) in biofilms exhibited higher resistance to an translation inhibitor, azithromycin compared to the growth of planktonic cultures of the same strains. Low concentrations of azithromycin were found to stimulate biofilm formation by the studied saprotrophic strains. The rate of synthesis of the polysaccharide matrix component exceeded the rate of cell growth, indicating implementation of the biofilm phenotype under these conditions. It was found that an alkylhydroxybenzene (AHB) compound 4-hexylresorcinol was capable of almost uniform suppression of growth of both planktonic cultures and biofilms of the saprotrophic strains under study. In some cases, combined action ofazithromycin and AHB resulted in an additive inhibitory effect and prevented the stimulation of biofilm growth by subinhibitory azithromycin concentrations. Thus, AHB may be considered a promising antibiofilm agent.

Chromobacterium violaceum abounds in soil and water ecosystems in tropical and subtropical regions and occasionally causes severe and often fatal human and animal infections. The quorum sensing (QS) system and biofilm formation are essential for C. violaceum's adaptability and pathogenicity, however, their interrelation is still unknown. C. violaceum's cell and biofilm morphology were examined by atomic force microscopy (AFM) in comparison with growth rates, QS-dependent violacein biosynthesis and biofilm biomass quantification. To evaluate QS regulation of these processes, the wild-type strain C. violaceum ATCC 31532 and its mini-Tn5 mutant C. violaceum NCTC 13274, cultivated with and without the QS autoinducer N-hexanoyl-L-homoserine lactone (C6-HSL), were used. We report for the first time the unusual morphological differentiation of C. violaceum cells, associated with biofilm development and directed by the QS autoinducer. AFM revealed numerous invaginations of the external cytoplasmic membrane of wild-type cells, which were repressed in the mutant strain and restored by exogenous C6-HSL. With increasing bacterial growth, polymer matrix extrusions formed in place of invaginations, whereas mutant cells were covered with a diffusely distributed extracellular substance. Thus, quorum sensing in C. violaceum involves a morphological differentiation that organises biofilm formation and leads to a highly differentiated matrix structure.

The effects of four alkylhydroxybenzene (AHB) homologs of different hydrocarbon chain lengths on synthesis of the pigment violacein induced by C6-homoserine lactone (HSL) and biofilm formation by Chromobacterium violaceum NCTC 13274 and on Escherichia coli pAL103 bioluminescence in the presence of C6-oxo-HSL were studied. It was shown that the inhibitory effect of alkylhydroxybenzenes on the growth of C. violaceum increased in the C5-AHB --> C12-AHB series in the absence of this activity in C1-AHB. Sub-inhibitory AHB concentrations reduced violacein production and suppressed biofilm formation. These effects were presented as individual and group regression dependencies between the analyzed parameters. It was shown using the bioluminescent model that the regulatory effects of AHBs are not associated with their direct competition with HSL and that they develop as a result of changes in the sensitivity of bacterial cells to the respective quorum sensing inducer.

The formation of biofilm by bacteria confers resistance to biocides and presents problems in medical and veterinary clinical settings. Here we report the effect of carvacrol, one of the major antimicrobial components of oregano oil, on the formation of biofilms and its activity on existing biofilms. Assays were carried out in polystyrene microplates to observe (a) the effect of 0-0.8 mM carvacrol on the formation of biofilms by selected bacterial pathogens over 24 h and (b) the effect of 0-8 mM carvacrol on the stability of pre-formed biofilms. Carvacrol was able to inhibit the formation of biofilms of Chromobacterium violaceum ATCC 12472, Salmonella enterica subsp. Typhimurium DT104, and Staphylococcus aureus 0074, while it showed no effect on formation of Pseudomonas aeruginosa (field isolate) biofilms. This inhibitory effect of carvacrol was observed at sub-lethal concentrations (<0.5 mM) where no effect was seen on total bacterial numbers, indicating that carvacrol's bactericidal effect was not causing the observed inhibition of biofilm formation. In contrast, carvacrol had (up to 8 mM) very little or no activity against existing biofilms of the bacteria described, showing that formation of the biofilm also confers protection against this compound. Since quorum sensing is an essential part of biofilm formation, the effect of carvacrol on quorum sensing of C. violaceum was also studied. Sub-MIC concentrations of carvacrol reduced expression of cviI (a gene coding for the N-acyl-L-homoserine lactone synthase), production of violacein (pigmentation) and chitinase activity (both regulated by quorum sensing) at concentrations coinciding with carvacrol's inhibiting effect on biofilm formation. These results indicate that carvacrol's activity in inhibition of biofilm formation may be related to the disruption of quorum sensing.

Bacterial biofilms are responsible for many persistent infections by many clinically relevant pathogens such as Staphylococcus aureus and Pseudomonas aeruginosa. Biofilms are much more resistant to conventional antibiotics than their planktonic counterparts. Quorum sensing, an intercellular communication system, controls pathogenesis and biofilm formation in most bacterial species. Quorum sensing provides an important pharmacological target since its inhibition does not provide a selective pressure for resistance. In this study, we investigated the quorum sensing and biofilm inhibitory activities of 126 plant extracts from 71 species collected from neotropical rainforests in Costa Rica. Quorum sensing and biofilm interference were assessed using a modified disc diffusion bioassay with Chromobacterium violaceum ATCC 12,472 and a spectrophotometric bioassay with Pseudomonas aeruginosa PA14, respectively. Species with significant anti-quorum sensing and/or anti-biofilm activities belonged to the Meliaceae, Melastomataceae, Lepidobotryaceae, Sapindaceae, and Simaroubaceae families. IC50 values ranged from 45 to 266 µg/mL. Extracts of these active species could lead to future development of botanical treatments for biofilm-associated infections., (Georg Thieme Verlag KG Stuttgart · New York.)

Aims: The aim of this study was to investigate the effect of clary sage, juniper, lemon and marjoram essential oils (EOs) and their major components on the formation of bacterial and yeast biofilms and on the inhibition of AHL-mediated quorum sensing (QS)., Methods and Results: Biofilm formation was measured by crystal violet and resazurin staining, and QS inhibition was detected by paper disc diffusion assay. Marjoram EO inhibited Bacillus cereus, Pichia anomala, Pseudomonas putida and mixed-culture biofilm formation of Ps. putida and Escherichia coli and showed the best QS inhibitor effect on Chromobacterium violaceum. For B. cereus, all components showed better antibiofilm capacity than the parent EOs. Lemon EO inhibited E. coli and mixed-culture biofilms, and cinnamon was effective against the mixed forms. Scanning electron microscopy showed the loss of three-dimensional structures of biofilms., Conclusions: The EOs and components used seem to be good candidates for prevention of biofilm formation and inhibition of the AHL-mediated QS mechanism., Significance and Impact of the Study: Biofilm formation on foods and food industrial equipment is a serious problem causing food spoilage and emergence of foodborne diseases. This article highlights the importance of studying EOs as potential disinfectants and food preservatives., (© 2013 The Society for Applied Microbiology.)

Inhibition of quorum sensing (QS)-regulated virulence factors including biofilm is a recognized anti-pathogenic drug target. The search for safe and effective anti-QS agents is expected to be useful to combat diseases caused by multidrug-resistant bacteria. In this study, effect of a commonly used antibiotic, doxycycline on QS was evaluated using sensor strains of Chromobacterium violaceum (ATCC 12472 and CVO26) and Pseudomonas aeruginosa PAO1. Sub-MICs of doxycycline reduced QS-controlled violacein production in C. violaceum to a significant degree (70 %) and showed a significant reduction of LasB elastase (67.2 %), pyocyanin (69.1 %), chitinase (69.8 %) and protease (65 %) production and swarming motility (74 %) in P. aeruginosa PAO1 over untreated controls. Similar results were also recorded against a clinical strain of P. aeruginosa (PAF-79). Interestingly, doxycycline at respective sub-MICs (4 and 32 μg ml(-1)) significantly reduced the biofilm-forming capability and exopolysaccharide production in both the strains of P. aeruginosa (PAO1 and PAF-79) over untreated controls. The results of this study highlight the multiple actions of doxycycline against QS-linked traits/virulence factors and its potential to attenuate virulence of P. aeruginosa.

Increasing evidence has shown that antibiotics function as intermicrobial signaling molecules instead of killing weapons. However, mechanisms and key factors that are involved in such functions remain poorly understood. Earlier findings have associated antibiotic signaling with quorum sensing (QS); however, results varied among experiments, antibiotics, and bacterial strains. In this study, we found that antibiotics at subinhibitory concentrations improved the violacein-producing ability of Chromobacterium violaceum ATCC 12472. Quantitative real-time polymerase chain reaction of QS-associated gene transcripts and bioassay of violacein production in a QS mutant strain demonstrated that antibiotics enhanced the production of N-acyl-L-homoserine lactones (AHLs; QS signaling molecules) and increased AHL-inducing QS-mediated virulence, including chitinase production and biofilm formation. Moreover, a positive flagellar activity and an increased bacterial clustering ability were found, which are related to the antibiotic-induced biofilm formation. Our findings suggested that antibiotic-mediated interspecific signaling also occurs in C. violaceum, thereby expanding the knowledge and language of cell-to-cell communication., (© 2013 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.)

Dendrophthoe falcata is a hemiparasitic plant commonly used for ailments such as ulcers, asthma, impotence, paralysis, skin diseases, menstrual troubles, pulmonary tuberculosis, and wounds. In this context, the validations of the traditional claim that the leaf extract of D. falcata possesses antibiofilm and anti-quorum sensing activity against different bacterial pathogens were assessed. The bacterial biofilms were quantified by crystal violet staining. Among the 17 bacterial pathogens screened, the methanolic fraction of the leaf extract clearly demonstrated antibiofilm activity for Proteus mirabilis, Vibrio vulnificus, Aeromonas hydrophila, Shigella sonnei, Chromobacterium violaceum ATCC 12472, Vibrio parahaemolyticus, Vibrio harveyi, Vibrio alginolyticus, Vibrio cholerae, and Proteus vulgaris. At biofilm inhibitory concentrations, biofilm formation was reduced by up to 70-90 %. Furthermore, the potential quorum-sensing activity of the leaf extract was tested by agar well diffusion using Chromobacterium violaceum (ATCC 12472 & CV O26) reporter strains. The inhibition of violacein production may be due to direct or indirect interference on QS by active constituents or the interactive effect of different phytocompounds present in the extracts. This is the first report on antibiofilm and QS activity of D. falcata leaf extracts, signifying the scope for development of complementary medicine for biofilm-associated infections., (Georg Thieme Verlag KG Stuttgart · New York.)

Background: Emergence of antibiotic resistance among bacterial pathogens often leads to the failure of existing antibiotics to treat bacterial infections; thus, there is a need to seek alternative treatment measures. The aim of this study was to evaluate the anti-quorum sensing (anti-QS) and antibiofilm potential of Capparis spinosa to prevent the onset of bacterial infections as an alternate to antibiotics., Methods: The methanolic extract of the dried fruits of C. spinosa was assessed for its activity in inhibiting QS-depedent phenomenon such as violacein pigment production in Chromobacterium violaceum, biosurfactant production in Pseudomonas aeruginosa PAO1, swimming and swarming motility, exopolysaccharide production (EPS) and biofilm formation in Escherichia coli, Proteus mirabilis, Serratia marcescens and PAO1., Results: Extract of C. spinosa showed a higher degree of anti-QS activity in a dose dependent manner without affecting the bacterial growth. At 2 mg/mL, this extract significantly (p ≤0.005) inhibited the biofilm formation to 79, 75, 73, 70% and EPS production to 58, 46, 66 and 67% in S. marcescens, PAO1, E. coli and P. mirabilis, respectively. It also exhibited inhibition in swimming and swarming motility of bacterial pathogens. The non-enzymatic nature of the anti-QS compound in C. spinosa was confirmed by proteinase K and heat treatment., Conclusions: Because the methanolic extract of C. spinosa demonstrated anti-QS and antibiofilm activity at 0.5-2 mg/mL, it could be further exploited for novel molecules to treat the emerging infections of antibiotic resistant bacterial pathogens., (Copyright © 2011 IMSS. Published by Elsevier Inc. All rights reserved.)

Quorum-sensing bacteria communicate via small molecules called autoinducers to coordinate collective behaviors. Because quorum sensing controls virulence factor expression in many clinically relevant pathogens, membrane-permeable quorum sensing antagonists that prevent population-wide expression of virulence genes offer a potential route to novel antibacterial therapeutics. Here, we report a strategy for inhibiting quorum-sensing receptors of the widespread LuxR family. Structure-function studies with natural and synthetic ligands demonstrate that the dimeric LuxR-type transcription factor CviR from Chromobacterium violaceum is potently antagonized by molecules that bind in place of the native acylated homoserine lactone autoinducer, provided that they stabilize a closed conformation. In such conformations, each of the two DNA-binding domains interacts with the ligand-binding domain of the opposing monomer. Consequently, the DNA-binding helices are held apart by ∼60 Å, twice the ∼30 Å separation required for operator binding. This approach may represent a general strategy for the inhibition of multidomain proteins., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Aim of the Study: The study aimed to test the activity of Terminalia catappa L. against bacterial quorum sensing (QS) in order to provide a potential scientific basis for the traditional use of leaf extracts of this plant as an antiseptic., Materials and Methods: The anti-QS activity of the methanolic leaf extract of Terminalia catappa was detected through the inhibition of the QS-controlled violacein pigment production in Chromobacterium violaceum. Fractions resulting from size-exclusion chromatography were assayed. The most active fraction was characterized through qualitative phytochemical detection methods. The effect of this fraction on known QS-controlled phenotypes in test strains was assessed., Results: The fraction with the highest activity (labeled as TCF12) was characterized to be tannin-rich. It specifically inhibited QS-controlled violacein production in Chromobacterium violaceum with 50% reduction achieved at 62.5 μg mL(-1) without significantly affecting growth up to about 962 μg mL(-1). The assessment of its effects on LasA activity of Pseudomonas aeruginosa ATCC 10145 found that the production of this virulence determinant is reduced in a concentration dependent manner with about 50% reduction at 62.5 μg mL(-1). Furthermore, it was found that TCF12 was able to inhibit the maturation of biofilms of Pseudomonas aeruginosa, a phenotype that has also been known to be QS-regulated., Conclusion: Therefore, tannin-rich components of Terminalia catappa leaves are able to inhibit certain phenotypic expression of QS in the test strains used., (Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.)

Conventional disinfection and sterilization methods are often ineffective with biofilms, which are ubiquitous, hard-to-destroy microbial communities embedded in a matrix mostly composed of exopolysaccharides. The use of gas-discharge plasmas represents an alternative method, since plasmas contain a mixture of charged particles, chemically reactive species and UV radiation, whose decontamination potential for free-living, planktonic micro-organisms is well established. In this study, biofilms were produced using Chromobacterium violaceum, a Gram-negative bacterium present in soil and water and used in this study as a model organism. Biofilms were subjected to an atmospheric pressure plasma jet for different exposure times. Our results show that 99.6 % of culturable cells are inactivated after a 5 min treatment. The survivor curve shows double-slope kinetics with a rapid initial decline in c.f.u. ml(-1) followed by a much slower decline with D values that are longer than those for the inactivation of planktonic organisms, suggesting a more complex inactivation mechanism for biofilms. DNA and ATP determinations together with atomic force microscopy and fluorescence microscopy show that non-culturable cells are still alive after short plasma exposure times. These results indicate the potential of plasma for biofilm inactivation and suggest that cells go through a sequential set of physiological and morphological changes before inactivation.

The marine bacterium Pseudoalteromonas tunicata produces an antibacterial and autolytic protein, AlpP, which causes death of a subpopulation of cells during biofilm formation and mediates differentiation, dispersal, and phenotypic variation among dispersal cells. The AlpP homologue (LodA) in the marine bacterium Marinomonas mediterranea was recently identified as a lysine oxidase which mediates cell death through the production of hydrogen peroxide. Here we show that AlpP in P. tunicata also acts as a lysine oxidase and that the hydrogen peroxide generated is responsible for cell death within microcolonies during biofilm development in both M. mediterranea and P. tunicata. LodA-mediated biofilm cell death is shown to be linked to the generation of phenotypic variation in growth and biofilm formation among M. mediterranea biofilm dispersal cells. Moreover, AlpP homologues also occur in several other gram-negative bacteria from diverse environments. Our results show that subpopulations of cells in microcolonies also die during biofilm formation in two of these organisms, Chromobacterium violaceum and Caulobacter crescentus. In all organisms, hydrogen peroxide was implicated in biofilm cell death, because it could be detected at the same time as the killing occurred, and the addition of catalase significantly reduced biofilm killing. In C. violaceum the AlpP-homologue was clearly linked to biofilm cell death events since an isogenic mutant (CVMUR1) does not undergo biofilm cell death. We propose that biofilm killing through hydrogen peroxide can be linked to AlpP homologue activity and plays an important role in dispersal and colonization across a range of gram-negative bacteria.

We studied the effect of the quorum-sensing (QS) blockers 5-hydroxy-3[(1R)-1-hydroxypropyl]-4-methylfuran-2(5H)-one (FUR1), (5R)-3,4-dihydroxy-5-[(1S)-1,2-dihydroxyethyl]furan-2(5H)-one (FUR2) and triclosan (TRI) on the formation of bacterial biofilms, and the effect of these biofilms on the larval attachment of the polychaete Hydroides elegans and the bryozoan Bugula neritina. 14-day-old subtidal biofilms were harvested from artificial substrata and were allowed to develop in the laboratory with and without QS blockers. QS blockers inhibited the production of violacein by the QS reporter strain Chromobacterium violaceum CV026 and did not affect the metabolic activity of bacteria in multispecies biofilms. At a concentration of 10(-3) M all three tested compounds inhibited the establishment of microbial communities, but at one of 10(-4) M only FUR2 inhibited establishment. The tested QS blockers caused changes in bacterial density and bacterial community structure, as revealed by terminal restriction fragment length polymorphism and FISH. The groups most affected by QS blockers were Alphaproteobacteria, Gammaproteobacteria and the Cytophagales. Larvae of H. elegans and B. neritina avoided settling on biofilms that had developed in the presence of QS blockers. Our results suggest that QS blockers directly control the formation of multi-species biofilms, and indirectly - by means of biofilm properties - affect larval attachment on these modified biofilms.