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Quorum sensing (QS) plays a crucial role in different stages of biofilm development, virulence production, and subsequently to the growth of bacteria in food environments. Biofilm mediated spoilage of food is one of the ongoing challenge faced by the food industry worldwide as it incurs substantial economic losses and leads to various health issues. In the present investigation, we studied the interference of quorum sensing, its regulated virulence functions, and biofilm in food-associated bacteria by colorant azorubine. In vitro bioassays demonstrated significant inhibition of QS and its coordinated virulence functions in Chromobacterium violaceum 12472 (violacein) and Pseudomonas aeruginosa PAO1 (elastase, protease, pyocyanin, and alginate). Further, the decrease in the production EPS (49–63%) and swarming motility (61–83%) of the pathogens was also recorded at sub-MICs. Azorubine demonstrated broad-spectrum biofilm inhibitory potency (50–65%) against Chromobacterium violaceum, Pseudomonas aeruginosa, E. coli O157:H7, Serratia marcescens, and Listeria monocytogenes. ROS generation due to the interaction between bacteria and azorubine could be responsible for the biofilm inhibitory action of the food colorant. Findings of the in vitro studies were well supported by molecular docking and simulation analysis of azorubine and QS virulence proteins. Azorubine showed strong binding to PqsA as compared to other virulent proteins (LasR, Vfr, and QscR). Thus, it is concluded that azorubine is a promising candidate to ensure food safety by curbing the menace of bacterial QS and biofilm-based spoilage of food and reduce economic losses. Keywords: Azorubine, Quorum sensing, Biofilm, Virulence, Food, Molecular docking

Novel, safe, and effective antilisterial agents are required in order to prevent Listeria monocytogenes infections and maintain food safety. This study synthesized silver nanoparticles (AgNPs) from the shoot extract of in vitro-grown Tamarix nilotica (TN) and characterized them using X-ray diffraction, Fourier transform infrared spectroscopy, UV-visible spectroscopy, dynamic light scattering, scanning electron microscopy (SEM), energy-dispersive X-ray spectroscopy (EDXS), and transmission electron microscopy (TEM). We also assessed the antilisterial potential of the synthesized TN-AgNPs by determining the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) against two strains of L. monocytogenes and L. innocua. TN-AgNPs (2×MICs) showed a significant decrease in growth in all Listeria test strains. Release of cellular content and cell morphology analysis of TN-AgNP-treated bacterial cells demonstrated the mechanism of bactericidal activity of AgNPs. In addition, TN-AgNPs induced a significant decrease in swimming motility (62–71%), biofilm formation (57–64%), and preformed biofilms (48–58%) in all Listeria test strains at sub-inhibitory concentrations. Microtitre plate assay results for biofilm inhibition were confirmed by SEM and CLSM visualization of TN-AgNP-treated and TN-AgNP-untreated Listeria test strains. TN-AgNPs also showed wound-healing activity in MCF-7 cells by inhibiting cell migration in a scratch plate assay. TN-AgNP-induced enhanced reactive oxygen species generation in treated cells could be a plausible reason for the biofilm inhibitory activity of AgNPs. TN-AgNPs having antilisterial, antibiofilm, and wound-healing properties can be effectively used to prevent L. monocytogenes infections in the food industry and healthcare.

Foodborne pathogens are one of the major cause of food-related diseases and food poisoning. Bacterial biofilms and quorum sensing (QS) mechanism of cell–cell communication have also been found to be associated with several outbreaks of foodborne diseases and are great threat to food safety. Therefore, In the present study, we investigated the activity of three tetrahedrally coordinated copper(I) complexes against quorum sensing and biofilms of foodborne bacteria. All the three complexes demonstrated similar antimicrobial properties against the selected pathogens. Concentration below the MIC i.e. at sub-MICs all the three complexes interfered significantly with the quorum sensing regulated functions in C. violaceum (violacein), P. aeruginosa (elastase, pyocyanin and alginate production) and S. marcescens (prodigiosin). The complexes demonstrated potent broad-spectrum biofilm inhibition in Pseudomonas aeruginosa, E. coli, Chromobacterium violaceum, Serratia marcescens, Klebsiella pneumoniae and Listeria monocytogenes. Biofilm inhibition was visualized using SEM and CLSM images. Action of the copper(I) complexes on two key QS regulated functions contributing to biofilm formation i.e. EPS production and swarming motility was also studied and statistically significant reduction was recorded. These results could form the basis for development of safe anti-QS and anti-biofilm agents that can be utilized in the food industry as well as healthcare sector to prevent food-associated diseases. Keywords: Copper compounds, Biofilm, Quorum sensing, Food safety, Pathogens

There is grave necessity to counter the menace of drug-resistant biofilms of pathogens using nanomaterials. Moreover, we need to produce nanoparticles (NPs) using inexpensive clean biological approaches that demonstrate broad-spectrum inhibition of microbial biofilms and cytotoxicity against HepG2 cell lines. In the current research work, titanium dioxide (TiO2) NPs were fabricated through an environmentally friendly green process using the root extract of Withania somnifera as the stabilizing and reducing agent to examine its antibiofilm and anticancer potential. Further, X-ray diffraction (XRD), Fourier transform infrared (FTIR), scanning electron microscopy (SEM), transmission electron micrograph (TEM), energy-dispersive X-ray spectroscopy (EDS), dynamic light scattering (DLS), thermogravimetric analysis (TGA), and Brunauer-Emmett-Teller (BET) techniques were used for determining the crystallinity, functional groups involved, shape, size, thermal behavior, surface area, and porosity measurement, respectively, of the synthesized TiO2 NPs. Antimicrobial potential of the TiO2 NPs was determined by evaluating the minimum inhibitory concentration (MIC) against Escherichia coli, Pseudomonas aeruginosa, methicillin-resistant Staphylococcus aureus, Listeria monocytogenes, Serratia marcescens, and Candida albicans. Furthermore, at levels below the MIC (0.5 × MIC), TiO2 NPs demonstrated significant inhibition of biofilm formation (43–71%) and mature biofilms (24–64%) in all test pathogens. Cell death due to enhanced reactive oxygen species (ROS) production could be responsible for the impaired biofilm production in TiO2 NP–treated pathogens. The synthesized NPs induced considerable reduction in the viability of HepG2 in vitro and could prove effective in controlling liver cancer. In summary, the green synthesized TiO2 NPs demonstrate multifarious biological properties and could be used as an anti-infective agent to treat biofilm-based infections and cancer.

Probiotics are live microorganisms or bacterial cultures that can have beneficial effects for the host when ingested. Probiotics must survive stressful conditions of the gastrointestinal tract by tolerating acid, bile, and gastric enzymes and must adhere to intestinal epithelial cells to colonize the gut. Moreover, probiotics should have antimicrobial effects against pathogenic microorganisms and desirable antibiotic susceptibility patterns. Many gastrointestinal diseases, such as diarrhea, irritable bowel syndrome, and chronic inflammatory bowel disease are caused by intestinal microflora imbalance, which is an important factor in bacterial translocation and infection. The current treatment of intestinal microbiota imbalance is using antibiotics; however, misuse or overuse of antibiotics contributes to resistance, which is one of the major public health problems worldwide. Another concern is the decreasing efficacy of antibiotics in treating human and animal infections because of the biofilm formation of pathogenic bacteria. Bacterial cells in biofilms are highly protected, less subjected to mutation, represent low metabolic activity, and become resistant to antibiotics. Therefore, the probiotic strains having both antimicrobial and antibiofilm properties may be expected to be therapeutically more effective. The antimicrobial probiotics having biofilm-dispersive properties can yield better clinical benefits as a therapeutic agent. Keeping in focus the promise shown by probiotics, current review is aimed to highlight the antibiofilm potential of probiotic microorganisms against drug-resistant pathogens.

Food handlers are important component in assessment and maintenance of food quality as they are carriers of food pathogens causing spoilage. Food spoilage is attributed to quorum sensing (QS) controlled development of biofilms. Therefore, there is an urgent need to develop novel QS and biofilm inhibitors to prevent spoilage of food products. In the present study, toxin producing biofilm forming methicillin-resistant Staphylococcus aureus (MRSA) were isolated from food handlers. Further, eugenol was screened for its QS and anti-biofilm properties. Analysis of nasal and hand swabs revealed the presence of seven toxigenic and biofilm forming MRSA strains. Eugenol demonstrated significant anti-QS activity in CVO26 and also reduced the QS-regulated production of elastase, protease, chitinase, pyocyanin and exopolysaccharide (EPS) in PAO1 considerably. Eugenol demonstrated 17%–86%, 24%–69%, 30%–91%, 9%–94% and 4%–89% reduction in biofilm biomass of S. aureus ATCC 25923 and MRSA strains FSA3, FSA11, FSA13 and FSA32, respectively. Sub-inhibitory concentrations of eugenol also decreased the metabolic activity in biofilm cells. Molecular docking analysis showed high binding affinity of eugenol that represents its biofilm inhibitory activity. This is the first report on the carriage of toxigenic drug-resistant biofilm forming S. aureus by food handlers and inhibition of their biofilms in the Kingdom of Saudi Arabia. The findings give a clear insight into the food safety hazards associated with the carriage of S. aureus and present eugenol as a broad-spectrum anti-QS and anti-biofilm agent.

Excessive and unabated use of antibiotics has led to the emergence of multi-drug resistant (MDR) bacteria. The ineffectiveness of current antibiotic therapy and the slow development of new drugs with novel modes of action have made the task of combating MDR infections even more difficult. The problem of multi-drug resistance among pathogens has prompted the scientific community to look for alternative strategies. One such approach, termed antipathogenic/antivirulence therapy, is considered to be a viable alternative. This strategy is focused on rendering the pathogen ineffective by inhibiting its virulence traits rather than killing it. Since antipathogenic/antivirulence compounds target bacterial virulence, the likelihood of developing resistance is also reduced considerably. The areas of major interest in the antivirulence approach include the inhibition of quorum sensing and biofilm formation. The pioneering discovery of halogenated furanones as quorum sensing inhibitors (QSIs) has prompted the scientific community to search for novel QSIs of both natural and synthetic origin. However, QSIs like furanones are not recommended for human use due to issues related to their toxicity and stability. Recently, researchers across the globe have turned their attention toward nanomaterials as potential anti-infective drugs, targeting QS and biofilm formation. Although there are numerous studies regarding the antibacterial effect of nanoparticles, current reports on the antiquorum sensing and biofilm inhibition properties are still scarce. Therefore, in this article we have made an honest attempt to summarize the reports on the anti-QS properties of nanoparticles and their future as novel anti-infective drugs.

Quorum sensing (QS) in bacteria is a density dependent communication system that regulates the expression of genes, including production of virulence factors in many pathogens. The emergence of antibiotic resistance among pathogenic bacteria represents a major threat in both hospitals as well as environmental settings. Interference of quorum sensing (QS)-regulated virulence factors and biofilms is a recognized anti-pathogenic therapy. Safe, stable and effective anti-QS agents are needed to combat diseases caused by multidrug-resistant bacteria. The present study was performed to assess the inhibitory effect of third generation antibiotic ceftazidime against Gram-negative bacterial pathogens. Sub-MICs of ceftazidime demonstrated dose dependent inhibition of QS regulated virulence traits and biofilm formation in various strains of Chromobacterium violaceum (CV12472 and CVO26), Pseudomonas aeruginosa (PAO1 and PAF79) and Aeromonas hydrophila (WAF38). β-galactosidase assay revealed ceftazidime inhibited the las and pqs QS systems in P. aeruginosa. Alongside, in vivo studies demonstrated enhanced survival of Caenorhabditis elegans after the treatment with the drug. Molecular docking analysis showed the high binding affinity of ceftazidime which represents its QS inhibitory activity. By highlighting the broad spectrum anti-quorum sensing and biofilm inhibiting activities against 3 different bacterial pathogens, ceftazidime seems a more potent candidate in counteracting the infections caused by drug resistant bacteria.

In the present study, a facile environmentally friendly approach was described to prepare monodisperse iron oxide (Fe3O4) nanoparticles (IONPs) by low temperature solution route. The synthesized nanoparticles were characterized using x-ray diffraction spectroscopy (XRD), Raman spectroscopy, field emission scanning electron microscopy (FESEM) measurements, Fourier-Transform Infrared Spectroscopy (FTIR), and Thermogravimetric analysis (TGA) analyses. XRD patterns revealed high crystalline quality of the nanoparticles. SEM micrographs showed the monodispersed IONPs with size ranging from 6 to 9 nm. Synthesized nanoparticles demonstrated MICs of 32, 64, and 128 μg/ml against Gram negative bacteria i.e., Serratia marcescens, Escherichia coli, and Pseudomonas aeruginosa, respectively, and 32 μg/ml against Gram positive bacteria Listeria monocytogenes. IOPNs at its respective sub-MICs demonstrated significant reduction of alginate and exopolysaccharide production and subsequently demonstrated broad-spectrum inhibition of biofilm ranging from 16 to 88% in the test bacteria. Biofilm reduction was also examined using SEM and Confocal Laser Scanning Microscopy (CLSM). Interaction of IONPs with bacterial cells generated ROS contributing to reduced biofilm formation. The present study for the first time report that these IONPs were effective in obliterating pre-formed biofilms. Thus, it is envisaged that these nanoparticles with broad-spectrum biofilm inhibitory property could be exploited in the food industry as well as in medical settings to curtail biofilm based infections and losses.

The emergence of multi-drug resistance in pathogenic bacteria in clinical settings as well as food-borne infections has become a serious health concern. The problem of drug resistance necessitates the need for alternative novel therapeutic strategies to combat this menace. One such approach is targeting the quorum-sensing (QS) controlled virulence and biofilm formation. In this study, we first screened different fractions of Psoralea corylifolia (seed) for their anti-QS property in the Chromobacterium violaceum 12472 strain. The methanol fraction was found to be the most active fraction and was selected for further bioassays. At sub-inhibitory concentrations, the P. corylifolia methanol fraction (PCMF) reduced QS-regulated virulence functions in C. violaceum CVO26 (violacein); Pseudomonas aeruginosa (elastase, protease, pyocyanin, chitinase, exopolysaccharides (EPS), and swarming motility), A. hydrophila (protease, EPS), and Serratia marcescens (prodigiosin). Biofilm formation in all the test pathogens was reduced significantly (p ≤ 0.005) in a concentration-dependent manner. The β-galactosidase assay showed that the PCMF at 1,000 μg/ml downregulated las-controlled transcription in PAO1. In vivo studies with C. elegans demonstrated increased survival of the nematodes after treatment with the PCMF. Bakuchiol, a phytoconstituent of the extract, demonstrated significant inhibition of QS-regulated violacein production in C. violaceum and impaired biofilm formation in the test pathogens. The molecular docking results suggested that bakuchiol efficiently binds to the active pockets of LasR and RhlR, and the complexes were stabilized by several hydrophobic interactions. Additionally, the molecular dynamics simulation of LasR, LasR–bakuchiol, RhlR, and RhlR–bakuchiol complexes for 50 ns revealed that the binding of bakuchiol to LasR and RhlR was fairly stable. The study highlights the anti-infective potential of the PCMF and bakuchiol instead of bactericidal or bacteriostatic action, as the extract targets QS-controlled virulence and the biofilm.

Monophasic tin dioxide nanoflowers (TONFs) assembled by rod-like nanostructures were prepared by coprecipitation method using tin chloride and ammonia precipitators, as the starting materials, without using any surfactants or templates. The structural, compositional, optical, and morphological properties of TONFs were investigated by XRD, FT-IR, UV-vis, SEM-EDX, and TEM techniques. Synthesized TONFs demonstrated inhibition of quorum sensing- (QS-) regulated virulence in pathogens, viz., Chromobacterium violaceum, Pseudomonas aeruginosa, and Serratia marcescens. Significant reduction in biofilm formation in all test pathogens was also observed which was further validated by CLSM images illustrating disturbed biofilm architecture. Vital functions like EPS, swarming motility, and cell surface hydrophobicity that contribute to successful biofilm formation were reduced after addition of sub-MICs of TONFs. Significant eradication of preformed biofilms was recorded upon addition of subinhibitory concentrations of TONFs in all test pathogens. The study highlights the broad-spectrum QS and biofilm inhibition by TONFs that can be exploited in future to protect food from contamination and combatting persistent drug-resistant infections.

Biofilms are complex aggregation of cells that are embedded in EPS matrix. These microcolonies are highly resistant to drugs and are associated with various diseases. Biofilms have greatly affected the food safety by causing severe losses due to food contamination and spoilage. Therefore, novel antibiofilm agents are needed. This study investigates the antibiofilm and protein binding activity of zinc nanoparticles (ZnNPs) synthesized from leaf extract ofOchradenus baccatus. Standard physical techniques, including UV-visible spectroscopy Fourier transform infrared spectroscopy and X-ray diffraction and transmission electron microscopy, were used to characterize the synthesized OB-ZnNPs. Synthesized OB-ZnNPs demonstrated significant biofilm inhibition in human and food-borne pathogens (Chromobacterium violaceum,Escherichia coli,P. aeruginosa,Klebsiella pneumoniae,Serratia marcescens, andListeria monocytogenes) at subinhibitory concentrations. OB-ZnNPs significantly reduced the virulence factors like violacein, prodigiosin, and alginate and impaired swarming migration and EPS production. OB-ZnNPs demonstrated efficient binding with HSA protein and no change in their structure or stability was observed. In addition,in vivotoxicity evaluation confirmed that OB-ZnNPs possessed no serious toxic effect even at higher doses. Moreover, they were found to have excellent antioxidant properties that can be employed in the fields of food safety and medicine. Hence, it is envisaged that the OB-ZnNPs can be used as potential nanomaterials to combat drug resistant bacterial infections and prevent contamination/spoilage of food.

The resistance and pathogenesis of bacteria could be related to their ability to sense and respond to population density, termed quorum sensing (QS). Inhibition of the QS system is considered as a novel strategy for the development of antipathogenic agents, especially for combating drug-resistant bacterial infections. In the present study, the anti-QS activity of Onion peel ethylacetate fraction (ONE) was tested against Chromobacterium violaceum CV12472 and Pseudomonas aeruginosa PAO1. ONE inhibit the QS-mediated virulence factors production such as violacein in C. violaceum and elastase, pyocyanin in P. aeruginosa. Further, the treatment with sub-MICs of ONE significantly inhibited the QS-mediated biofilm formation, EPS (Extracellular polymeric substances) production and swarming motility. Further, quercetin 4′-O-β-D glucopyranoside (QGP) was isolated from ONE and its anti-QS potential was confirmed after observing significant inhibition of QS-controlled virulence factors such as violacein, elastase, pyocyanin and biofilm formation in test pathogens. Molecular docking analysis predicted that QGP should be able to bind at the active sites of Vfr and LasR, and if so blocks the entry of active sites in Vfr and LasR.

Bacterial spoilage of food products is regulated by density dependent communication system called quorum sensing (QS). QS control biofilm formation in numerous food pathogens and Biofilms formed on food surfaces act as carriers of bacterial contamination leading to spoilage of food and health hazards. Agents inhibiting or interfering with bacterial QS and biofilm are gaining importance as a novel class of next-generation food preservatives/packaging material. In the present study, Zinc nanostructures were synthesised using Nigella sativa seed extract (NS-ZnNPs). Synthesized nanostructures were characterized hexagonal wurtzite structure of size ~24 nm by UV-visible, XRD, FTIR and TEM. NS-ZnNPs demonstrated broad-spectrum QS inhibition in C. violaceum and P. aeruginosa biosensor strains. Synthesized nanostructures inhibited QS regulated functions of C. violaceum CVO26 (violacein) and elastase, protease, pyocyanin and alginate production in PAO1 significantly. NS-ZnNPs at sub-inhibitory concentrations inhibited the biofilm formation of four-food pathogens viz. C. violaceum 12472, PAO1, L. monocytogenes, E. coli. Moreover, NS-ZnNPs was found effective in inhibiting pre-formed mature biofilms of the four pathogens. Therefore, the broad-spectrum inhibition of QS and biofilm by biogenic Zinc oxide nanoparticles and it is envisaged that these nontoxic bioactive nanostructures can be used as food packaging material and/or as food preservative.

Made available in DSpace on 2015-10-22T06:29:19Z (GMT). No. of bitstreams: 0 Previous issue date: 2015-05-13. Added 1 bitstream(s) on 2015-10-22T09:46:03Z : No. of bitstreams: 1 WOS000355832500001.pdf: 8351305 bytes, checksum: e895a7b4d27ef3b9a80dbdda4bc6d728 (MD5) Deanship of Scientific Research at KSU Bacterial quorum sensing (QS) is a density dependent communication system that regulates the expression of certain genes including production of virulence factors in many pathogens. Bioactive plant extract/compounds inhibiting QS regulated gene expression may be a potential candidate as antipathogenic drug. In this study anti-QS activity of peppermint (Menthe piperita) oil was first tested using the Chromobacterium violaceum CVO26 biosensor. Further, the findings of the present investigation revealed that peppermint oil (PMO) at sub-Minimum Inhibitory Concentrations (sub-MICs) strongly interfered with acyl homoserine lactone (AHL) regulated virulence factors and biofilm formation in Pseudomonas aeruginosa and Aeromonas hydrophila. The result of molecular docking analysis attributed the QS inhibitory activity exhibited by PMO to menthol. Assessment of ability of menthol to interfere with QS systems of various Gram-negative pathogens comprising diverse AHL molecules revealed that it reduced the AHL dependent production of violacein, virulence factors, and biofilm formation indicating broad-spectrum anti-QS activity. Using two Escherichia colt biosensors, MG4/pKDT17 and pEAL08-2, we also confirmed that menthol inhibited both the las and pqs QS systems. Further, findings of the in vivo studies with menthol on nematode model Caenorhabditis elegans showed significantly enhanced survival of the nematode. Our data identified menthol as a novel broad spectrum QS inhibitor. Department of Agricultural Microbiology, Aligarh Muslim University, Aligarh, India Department of Food Science and Nutrition, College of Food and Agricultural Sciences, King Saud University, Riyadh, Saudi Arabia Department of Zoology, Aligarh Muslim University, Aligarh, India Department of Biochemistry, Protein Research Chair, College of Science, King Saud University, Riyadh, Saudi Arabia School of Pharmaceutical Sciences, Sao Paulo State University, Araraquara, Brazil Deanship of Scientific Research at KSU: RGP-215

Trigonella foenum-graecumL. (Fenugreek) is an important plant of the Leguminosae family known to have medicinal properties. However, fraction based antiquorum sensing and antibiofilm activities have not been reported from this plant. In the present studyT. foenum-graecumseed extract was sequentially fractionated and sub-MICs were tested for above activities. The methanol fraction of the extract demonstrated significant inhibition of AHL regulated virulence factors: protease, LasB elastase, pyocyanin production, chitinase, EPS, and swarming motility inPseudomonas aeruginosaPAO1 and PAF79. Further, QS dependent virulence factor in the aquatic pathogenAeromonas hydrophilaWAF38 was also reduced. Application ofT. foenum-graecumseed extract to PAO1, PAF79, and WAF38 decreased the biofilm forming abilities of the pathogens by significant levels. The extract also exhibited reduced AHL levels and subsequent downregulation oflasBgene.In vivostudy showed an enhanced survival of PAO1-preinfectedC. elegansafter treatment with extract at 1 mg/mL. Further, the major compound detected by GC-MS, caffeine, reduced the production of QS regulated virulence factors and biofilm at 200 µg/mL concentration indicating its role in the activity of the methanol extract. The results of the present study reveal the potential anti-QS and antibiofilm property ofT. foenum-graceumextract and caffeine.