Strain Chromobacterium Violaceum Research Articles

Antimicrobial and Anti-quorum Sensing Activity of Geranium sp.

Aim: Antibiotic resistance, which has increased rapidly in recent years, is one of the leading public health threats. Alternative methods are being investigated for effective antibiotics in the fight against resistance. Geranium species are widely used for constipation, digestive disorders, and diabetes. It is also known that the plant is used in various public health applications. In line with this information, it was aimed to investigate the antimicrobial and anti-quorum sensing properties of the Geranium sp. Material and Methods: In this study it was aimed to prepare methanol, ethyl acetate, ethanol, and hexane extracts of Geranium sp. flower parts belonging to Rize province and investigate their antimicrobial activities by agar well diffusion method against various Gram-negative and Gram-positive bacteria and two fungal species. Anti-quorum sensing activity was determined using Chromobacterium violaceum strain. Results: Methanol extract of Geranium sp. showed antimicrobial activity against seven species whereas extraction with ethyl acetate showed antimicrobial activity against eight species. The ethanol extracted samples have the lowest antimicrobial activity so that only six species were affected. Additionally, methanol extract of Geranium sp. had violacein inhibition activity. Conclusions: As a result, it was determined that the plant, especially its methanol extract, has antimicrobial and anti-quorum sensing properties.

Comparative in vitro Study of Antimicrobial, Antibiofilm and Quorum Sensing Inhibitory Activities of Hypericum calycinum L. and Parietaria officinalis L. Extracts

Aim: This study aims to elucidate both the anti-virulence and antimicrobial effects of ethanol extracts from Hypericum calycinum L. and Parietaria officinalis L. Material and Methods: Antimicrobial activity was evaluated using the well diffusion method against five bacteria and two yeast isolates involved in human urinary tract infections (UTIs). The potential of the extracts to inhibit quorum sensing (QS), was assessed using the biosensor strain Chromobacterium violaceum ATCC 12472. Additionally, the antibiofilm activities were investigated using a microplate biofilm assay on Escherichia coli and Pseudomonas aeruginosa. Results: H. calycinum exhibited the highest inhibitory effect at a concentration of 100 mg/mL against Candida albicans with an inhibition zone of 24.5±0.71 mm, while P. officinalis showed its highest effect at the same concentration against E. coli with an inhibition zone of 15.5±0.71 mm. Overall, H. calycinum demonstrated stronger antimicrobial activity compared to P. officinalis. Both plant extracts inhibited QS at similar levels, with inhibition zones ranging between 10-12 mm. The antibiofilm effect varied depending on the bacterial species, but notably, P. officinalis extract exhibited over 80% antibiofilm efficacy against E. coli at all concentrations. Conclusion: This study demonstrates that H. calycinum L. and P. officinalis L. are potent antimicrobial agents against UTI pathogens. While their anti-QS efficacy is not exceptional, the significant inhibition of E. coli biofilm formation underscores their potential as formidable agents. Designed as a fundamental study, it highlights the promising antimicrobial properties of these plant extracts and marks the first investigation into their capabilities as QS and biofilm-preventive agents.

Discovery of β-nitrostyrene derivatives as potential quorum sensing inhibitors for biofilm inhibition and antivirulence factor therapeutics against Serratia marcescens.

Quorum sensing (QS) inhibition has emerged as a promising target for directed drug design, providing an appealing strategy for developing antimicrobials, particularly against infections caused by drug-resistant pathogens. In this study, we designed and synthesized a total of 33 β-nitrostyrene derivatives using 1-nitro-2-phenylethane (NPe) as the lead compound, to target the facultative anaerobic bacterial pathogen Serratia marcescens. The QS-inhibitory effects of these compounds were evaluated using S. marcescens NJ01 and the reporter strain Chromobacterium violaceum CV026. Among the 33 new β-nitrostyrene derivatives, (E)-1-methyl-4-(2-nitrovinyl)benzene (m-NPe, compound 28) was proven to be a potent inhibitor that reduced biofilm formation of S. marcescens NJ01 by 79%. Scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM) results revealed that treatment with m-NPe (50 μg/ml) not only enhanced the susceptibility of the formed biofilms but also disrupted the architecture of biofilms by 84%. m-NPe (50 μg/ml) decreased virulence factors in S. marcescens NJ01, reducing the activity of protease, prodigiosin, and extracellular polysaccharide (EPS) by 36%, 72%, and 52%, respectively. In S. marcescens 4547, the activities of hemolysin and EPS were reduced by 28% and 40%, respectively, outperforming the positive control, vanillic acid (VAN). The study also found that the expression levels of QS- and biofilm-related genes (flhD, fimA, fimC, sodB, bsmB, pigA, pigC, and shlA) were downregulated by 1.21- to 2.32-fold. Molecular dynamics analysis showed that m-NPe could bind stably to SmaR, RhlI, RhlR, LasR, and CviR proteins in a 0.1 M sodium chloride solution. Importantly, a microscale thermophoresis (MST) test revealed that SmaR could be a target protein for the screening of a quorum sensing inhibitor (QSI) against S. marcescens. Overall, this study highlights the efficacy of m-NPe in suppressing the virulence factors of S. marcescens, identifying it as a new potential QSI and antibiofilm agent capable of restoring or improving antimicrobial drug sensitivity.

Comparative Study on Quorum Modulatory Effect of Selected Medicinal Plants on Chromobacterium violaceum ATCC 12472 (MTCC 2656)

Among the main global health concern is the rampant rise in antibiotic resistant bacteria. One of the appealing and promising strategies to combat this menace is to target the adaptive mechanism called quorum sensing (QS) used by bacteria to survive. Exploratory research on anti-QS compounds derived from natural products has been a promising area. The present study investigated methanolic extracts from 26 plants to compare their anti-QS activity using the QS biosensor strain Chromobacterium violaceum American Type Culture Collection (ATCC 12472) (Microbial Type Culture Collection MTCC2656). QS-mediated violacein pigment inhibition was carried out using agar well diffusion method with concentrations ranging from 10 mg/ml to 100 mg/ml. Leaf extracts of Mangifera indica and Pimenta dioica and peel extract of Punica granatum were the only three plants found to exhibit violacein inhibitory potential till 10 mg/ml. The result of the minimum inhibitory concentration (MIC) showed 1.6 mg/ml for M. indica and P. dioica and 6.25 mg/ml for P. granatum. Further, violacein inhibitory properties of these extracts at and below MIC were evaluated by well diffusion assay (qualitative) and by flask incubation assay (quantitative). The zone of inhibition (well diffusion assay) was found to be 14.51 ± 0.63 mm to 10.37 ± 0.68mm for M. indica, 15.23 ± 0.57 mm to 9.62 ± 1.29 mm for P. granatum and 17.01 ± 0.1 mm to 13.14 ± 0.18 mm for P. dioica. The inhibitory effect of the plant extracts via quantitative assay on violacein ranged from 83-49%, 89-81%, and 89-49% for M. indica, P. granatum, and P. dioica respectively. Our findings suggested the potential of M. indica, P. granatum, and P. dioica methanolic extracts as a source of effective inhibitors of QS-mediated violacein production.

Pseudomonas aeruginosa is an Effective Indicator for Screening of Quorum Sensing Inhibition by Plant Extracts

Quorum sensing is a well-known mechanism used by many bacteria to control important virulence factors through the production and subsequent response to N-acylhomoserine lactones (AHLs). Quorum sensing inhibition (QSI), targeting AHL-dependent signalling, has been reported as a strategy for the control of bacterial pathogenicity. Thus, this study aimed at investigating the capability of Pseudomonas aeruginosa as an indicator to rapidly screen for potential QSI caused by plant extracts based on pigment inhibition. Four ethanolic plant extracts including: dried flower buds of clove (Syzygium aromaticum L), bark of cinnamon (Cinnamomum cassia L), leaves of moringa (Moringa oleifera Lam), and Al-Gutub (Tribulus terrestris L) dried fruits were investigated for their QSI ability in comparison with furanone (the known QS inhibitor). Agar well diffusion assay was used, in which the indicator isolate, P. aeruginosa, was inoculated onto Pseudomonas special agar medium containing 8 mm wells to load different concentrations of the plant extracts along with furanone or phosphate buffered saline (negative control). The results showed that furanone was successful in inhibiting the pyocyanin pigment of P. aeruginosa without interfering with the bacterial growth, this verified the ability of this bacterium as a potent QS indicator organism. Importantly, clove extract at different concentrations caused complete inhibition of the pyocyanin pigment with little impact on the bacterial growth. On the use of cinnamon, only the high concentrations showed clear dye inhibition of the indicator organism. Moringa also caused certain degree of dye inhibition but only at high concentrations. Conversely, Al-Gutub neither affected on the pyocyanin production nor on the bacterial viability. In conclusion, P. aeruginosa isolate, pyocyanin producer, was successful in the screening of QS inhibitors. This simple study recommends the use of such isolate without relying on the import of certain strains of Chromobacterium violaceum, Pseudomonas aureofaciens, or Agrobacterium tumefaciens‎‎‎‎‎.

Roles of transcriptional factor PsrA in the regulation of quorum sensing in Pseudomonas aeruginosa PAO1.

The transcription factor PsrA regulates fatty acid metabolism, the type III secretion system, and quinolone signaling quorum sensing system in Pseudomonas aeruginosa. To explore additional roles of PsrA in P. aeruginosa, this study engineered a P. aeruginosa PAO1 strain to carry a recombinant plasmid with the psrA gene (pMMBpsrA) and examined the impact of elevated psrA expression to the bacterium. Transcriptomic analysis revealed that PsrA significantly downregulated genes encoding the master quorum-sensing regulators, RhlR and LasR, and influenced many quorum-sensing-associated genes. The role of PsrA in quorum sensing was further corroborated by testing autoinducer synthesis in PAO1 [pMMBpsrA] using two reporter bacteria strains Chromobacterium violaceum CV026 and Escherichia coli [pSB1075], which respond to short- and long-chain acyl homoserine lactones, respectively. Phenotypic comparisons of isogenic ΔpsrA, ΔlasR, and ΔpsrAΔlasR mutants revealed that the reduced elastase, caseinase, and swarming activity in PAO1 [pMMBpsrA] were likely mediated through LasR. Additionally, electrophoretic mobility shift assays demonstrated that recombinant PsrA could bind to the lasR promoter at a 5'-AAACGTTTGCTT-3' sequence, which displays moderate similarity to the previously reported consensus PsrA binding motif. Furthermore, the PsrA effector molecule oleic acid inhibited PsrA binding to the lasR promoter and restored several quorum sensing-related phenotypes to wild-type levels. These findings suggest that PsrA regulates certain quorum-sensing phenotypes by negatively regulating lasR expression, with oleic acid acting as a crucial signaling molecule.

Ocimum sanctum as a Source of Quorum Sensing Inhibitors to Combat Antibiotic Resistance of Human and Aquaculture Pathogens.

Biofilms play a decisive role in the infectious process and the development of antibiotic resistance. The establishment of bacterial biofilms is regulated by a signal-mediated cell-cell communication process called "quorum sensing" (QS). The identification of quorum sensing inhibitors (QSI) to mitigate the QS process may facilitate the development of novel treatment strategies for biofilm-based infections. In this study, the traditional medicinal plant Ocimum sanctum was screened for QS inhibitory potential. Sub-MICs of the extract significantly affected the secretion of EPS in Gram-negative human pathogens such as Escherichia coli, Pseudomonas aeruginosa PAO1, Proteus mirabilis, and Serratia marcescens, as well as aquaculture pathogens Vibrio harveyi, V. parahaemolyticus, and V. vulnificus, which render the bacteria more sensitive, leading to a loss of bacterial biomass from the substratum. The observed inhibitory activity of the O. sanctum extract might be attributed to the presence of eugenol, as evidenced through ultraviolet (UV)-visible, gas chromatography-mass spectroscopy (GC-MS), Fourier transformer infrared (FTIR) spectroscopy analyses, and computational studies. Additionally, the QSI potential of eugenol was corroborated through in vitro studies using the marker strain Chromobacterium violaceum.

Effect of a phthalate derivative purified from Bacillus zhangzhouensis SK4 on quorum sensing regulated virulence factors of Pseudomonas aeruginosa

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.

A novel sequential pretreatment coupled with statistically optimized bioleaching for highly effective biorecovery of platinum group metals from spent catalyst waste

The biorecovery of platinum group metals (PGM) from spent catalyst waste is gaining widespread interest as an alternative to conventional recovery processes. A challenging aspect of the biorecovery of PGM from spent automotive catalyst (SAC) is the low biomobilization of PGM due to the complex nature of SAC. This work discusses statistical optimization of the bioleaching parameters using a central composite design for the biorecovery of PGM from SAC. A novel approach is presented where PGM from sequentially pretreated SAC is optimally recovered using heat oxidation, sonication, and reduction prior to bioleaching. An engineered strain of Chromobacterium violaceum was able to leach Pt (76 %), Pd (81 %), and Rh (100 %) from SAC at a pulp density of 0.5 % w/v following three days of spent medium leaching. Under the same conditions, pretreatment of SAC using formic before leaching enhanced the leaching efficiency to Pt (91 %), Pd (95 %), and Rh (100 %) following two days of spent medium leaching. Overall, this study demonstrates the effectiveness of sequential pretreatment before bioleaching and statistical optimization of bioleaching for enhanced biorecovery of PGM.

EVALUATION OF ANTI-QUORUM SENSING ACTIVITY OF-HEXADECANOIC ACID PRODUCED BY PSEUDOMONAS STUTZERI SJ4 – A MARINE EPIBIOTIC BACTERIUM

Quorum sensing (QS) mechanism is cell communication that plays vital role in the development of infection by many pathogenic microorganisms. It controls multiple virulence factors such as pigmentation, biofilm formation, swarming motility, resistance towards antibiotics, extracellular polysaccharide production (EPS) and expression of several collective traits. The disruption of quorum sensing mechanism can be a solution to the emerging problem of multi-drug resistance among pathogenic bacteria. The effector molecule for Quorum sensing inhibition may be enzymatic or non- enzymatic in nature termed as Quorum Quenching (QQ) or Quorum sensing inhibitory (QSIs)/anti –QS compound, respectively. We used marine epibiotic bacteria as a source to obtained novel bacterial strain as QSI producer. One of the potent isolate, SJ4 was identified as Pseudomonas stutzeri SJ4, it is a short rod, gram negative bacterium. The ethyl acetate extract from P. stutzeri SJ4, showed highest QSI activity against monitor strain Chromobacterium violaceum (MTCC 5526). The extracted compound was tested against P. aeruginosa PAO1 at minimum inhibitiory concentration (MIC) and sub-MIC to study the effect on virulence factors. The significant inhibition of pycocyanin pigment, EPS production, rhamnolipid production and reduced swimming and swarming motility was observed. In addition, biofilm formation was notably inhibited which was confirmed by staining and spectometric method. Based on this observation, QS interption by extract which contain QSI remarkebly reduced the virulence of pathogen hence, can be use as therapeutic agents. The Thin Layer Chomatography (TLC) and Gas chrmoatography-Mass Spectrometry (GC-MS) identified major compound as n-Hexadecanoic acid. However, further research is required on purification of compound and its potential applications for the treatement of infections.

Comparison of the Efficiency of Selected Disinfectants against Planktonic and Biofilm Populations of Escherichia coli and Staphylococcus aureus.

The aim of this study is to compare the efficacy of selected food disinfectants on planktonic populations of Staphylococcus aureus and Escherichia coli and on the same microorganisms (MOs) incorporated in a biofilm. Two disinfectants were used for treatment: peracetic acid-based disinfectant (P) and benzalkonium chloride-based disinfectant (D). Testing of their efficacy on the selected MO populations was performed using a quantitative suspension test. The standard colony counting procedure was used to determine their efficacy on bacterial suspensions in tryptone soy agar (TSA). The germicidal effect (GE) of the disinfectants was determined based on the decimal reduction ratio. For both MOs, 100% GE was achieved at the lowest concentration (0.1%) and after the shortest exposure time (5 min). Biofilm production was confirmed with a crystal violet test on microtitre plates. Both E. coli and S. aureus showed strong biofilm production at 25 °C with E. coli showing significantly higher adherence capacity. Both disinfectants show a significantly weaker GE on 48 h biofilms compared to the GE observed after application of the same concentrations on planktonic cells of the same MOs. Complete destruction of the viable cells of the biofilms was observed after 5 min of exposure to the highest concentration tested (2%) for both disinfectants and MOs tested. The anti-quorum sensing activity (anti-QS) of disinfectants P and D was determined via a qualitative disc diffusion method applied to the biosensor bacterial strain Chromobacterium violaceum CV026. The results obtained indicate that the disinfectants studied have no anti-QS effect. The inhibition zones around the disc therefore only represent their antimicrobial effect.

Probiotic potential and anti-quorum sensing activity of Enterococcus faecalis and Lactobacillus kunkeei isolates from Apis mellifera

One health is an approach that gains importance in the protection of public health and advocates that the environment-animal-human health is linked. Microbial resistance occurring in any of these three factors circulates directly or indirectly between species and affects health. In this study, antimicrobial, anti-quorum sensing activity and potential probiotic properties of Lactobacillus acidophilus and Enterococcus faecalis 1 and 2 isolates from Apis mellifera gut were investigated. Results showed that isolates were found to be resistant to gentamicin, streptomycin, erythromycin, and clindamycin tested. It was determined that isolates were gamma haemolytic and resistant to acid and enzymatic environment conditions. Enterococcus faecalis isolates had antimicrobial activity on Escherichia coli ATCC 25922, Enterococcus faecalis ATCC 29212 and Pseudomonas aeruginosa ATCC 27853 and anti-quorum sensing activity of the culture supernatants of all isolates was detected by a violacein pigment inhibition of Chromobacterium violaceum strain.

Cloning and characterization of Aiia, an acylhomoserine lactonase from Bacillus cereus RC1 to control soft rot causing pathogen Lelliottia amnigena RCE.

Bacterial pathogenesis-associated characteristics such as biofilm formation, synthesis of hydrolyzing enzymes, and toxins are regulated by Acyl Homoserine Lactones (AHLs), small peptides and diffusing signal factors (DSF). Lelliottia amnigena is gram negative bacteria and its pathogenicity is regulated by the luxR andluxIclass of quorum sensing. The signaling molecules and their concentrations are essential for the virulence of the pathogenic bacterium. To suppresses the pathogenicity; the concentration of signalling molecules must be controlled or degraded. The lactonase have the ability to hydrolyze lactonesof different chain length. The present study deals with a newer approach to control the pathogenesis of Lelliottia amnigena through isolation and characterization of Aiia lactonase from Bacillus cereus RC1. Aiia lactonase specific primers were used to amplify the gene, and the sequence thus obtained was submitted to the Genbank database under accession # OK643884.1. The gene was cloned in pBE-S shuttle vector and transformed in the recombinant host. The expressed and purified protein had a molecular weight of 28.00 KDa and exhibited its optimum activity at 37℃ by inhibiting the violacein pigment of the monitor strain Chromobacterium violaceum MTCC 2656. The proteinaceous nature of the purified molecule was confirmed by incubating it in the presence of proteinase K for 1h. The activity of the pathogenesis-related protein, polygalacturonase was drastically reduced in the presence of the purified Aiia protein. The purified protein also showed a zone of inhibition when plated together with Lelliottia amnigena RCE (MZ712952.1). Searches of the Conserved Domain Database suggested that this protein belonged to the Metallo-beta-lactamase superfamily and is closely related to Aiia from B. thuringiensis serovar kurstaki. Modeling of the protein structure was done using I-TASSER; a C-score of 0.55 suggested that the model was of good quality. To be used commercially, this recombinant protein needs to be purified at an industrial scale; it can then be used to repress the growth of soft rot causing bacteria in horticultural crops during their storage period.

Ketoprofen, piroxicam and indomethacin-suppressed quorum sensing and virulence factors in Acinetobacter baumannii.

Quorum sensing (QS) inhibition is a promising strategy to suppress bacterial virulence and control infection caused by Gram-negative and Gram-positive bacteria. This study explores the QS inhibiting activity of the non-steroidal anti-inflammatory drugs (NSAIDs) in Acinetobacter baumannii. Ketoprofen, piroxicam and indomethacin revealed QS inhibition via elimination of violacein production of the reporter strain Chromobacterium violaceum ATCC 12472 without affecting bacterial growth. The minimal inhibitory concentration (MIC) of ketoprofen, piroxicam and indomethacin was determined against A. baumannii strains ATCC 17978, ATCC 19606, A1, A11 and A27 by the microbroth dilution method. The MICs of ketoprofen against tested isolates were 0.7-6.25 mg ml-1 , piroxicam MICs were 1.25-2.5mg ml-1 , and indomethacin MICs were 3.12-12.5mg ml-1 . Those compounds significantly inhibited QS-associated virulence factors such as biofilm formation, and surface motility, as well as, significantly increased bacterial tolerance to oxidative stress without affecting bacterial growth. On the molecular level, the three compounds significantly inhibited the transcription of QS regulatory genes abaI/abaR and biofilm-regulated genes cusD and pgaB. Molecular docking analysis revealed the potent binding affinity of the three compounds with AbaI via hydrogen and/or hydrophobic bonds. These results indicate that NSAIDs, ketoprofen, piroxicam and indomethacin, could be potential inhibitors of the QS and could suppress the QS-related virulence factors of A. baumannii. Ketoprofen, piroxicam and indomethacin could provide promising implications and strategies for combating the virulence and pathogenesis of A. baumannii.

Synthesis and characterization of bismuth(III) complex with an EDTA-based phenylene ligand and its potential as anti-virulence agent

A new acyclic bismuth complex (Bi-edtabz) was synthesized from a mixture of solutions of the ligand (EDTA-based phenylene) and bismuth under acidic conditions. Its anti-virulence properties were evaluated against Escherichia coli O157: H7, Listeria monocytogenes, Pseudomonas aeruginosa, Salmonella enterica sub. enterica serovar Typhimurium and Staphylococcus aureus. The bismuth complex was characterized by NMR, UV-Vis, FTIR, ESI/MS and TG. Furthermore, Bi-edtabz complex at 0.25–1 mM presented better antibiofilm properties against E. coli O157: H7 and S. aureus with values of biomass reduction of 30.1–57.1% and 37.8–55.5%, respectively, compared with the ligand edtabz. While biofilm formation of L. monocytogenes, P. aeruginosa and Salmonella Typhimurium was most impaired by edtabz (biomass reduction of 66.1–100%, 66.4–88.0% and 50.9–67.1%), respectively. Additionally, Bi-edtabz inhibited the swimming motility of E. coli O157: H7 (12.5%) and colony spread of S. aureus (47.2%) at 1 mM and inhibited violacein production, a quorum-sensing related pigment of the biosensor strain Chromobacterium violaceum. Hence, edtabz and the Bi-edtabz complex can be used as novel anti-virulence agents against pathogenic bacteria.

Synthesis and Evaluation of Quorum Quenching activity of Ag, TiO2 and ZnO metal nanoparticles against wild type and mutant strains of Chromobacterium violaceum

The conventional strategies to combat pathogenic microbes seem to be inefficient to curb the menace of pathogens. The bacterial biofilm is a product of the quorum sensing (QS) process mediated by acylhomoserine lactones (AHL) in many pathogens. The QS inhibitors or quorum quenching (QQ) molecules that disrupt bacterial communication offer an alternative promising strategy to lower antibiotic resistance in pathogens. The nanoparticles can exhibit QQ potential that interrupts cell-to-cell communication. In the present study, laboratory synthesized metal-based nanoparticles such as silver (Ag), titanium oxide(TiO2) and zinc oxide (ZnO) were characterized using powder X-Ray diffractometer, UV-Visible spectroscopy, EDS, Cyclic voltammetry and TEM imaging technique. Ag and TiO2 nanoparticles were synthesized by the green synthesis method in the form of quantum dots, which was evidenced from TEM images on a less than 7nm scale. Silver quantum dots with size variation from 2.4nm to 4.2nm and TiO2 with size ranging from 4.9nm to 8.5nm were detected. The synthesized nanoparticles were elucidated for their potential as QQ by employing both wild and mutant strains of Chromobacterium violaceum. Nanoparticles affected violacein pigment production in both the strains of C. violaceum. Lower concentrations of NPs did not show any considerable effect on the AHLdependent Quorum sensing system, however, at moderate to higher concentrations, metal NPs exhibit Quorum Q activity. Higher concentrations of AgNPs and ZnONPs (i.e. 250 ppm and above) show cell inhibitory activity. In the percent pigment(violacein) inhibition assay, maximum pigment inhibition was detected at a concentration range of 100-125ppm in both the strains of C. violaceum without inhibiting cell growth. In the case of TiO2NPs, maximum pigment inhibition was detected at 500ppm without affecting cell growth in both the strains of C. violaceum. The percent pigment inhibition observed in all nanoparticles was nearly similar in both strains of C. violaceum indicating that the mechanism of action of pigment inhibition is due to interference at the reception of the AHL signaling molecule.

Aromatherapeutic essential oils and their pharmaceutical combinations: Tools for inhibition of quorum sensing activity and biofilm formation of human pathogens

Background and Aims: Aromatherapy, as one of the complementary therapies, uses essential oils as the main therapeutic agents to treat several diseases. In the present study, it was aimed to investigate inhibition of quorum sensing (QS) and biofilm formation of aromatherapeutic essential oils (AEOs) and their pharmaceutical combinations (PC-I and PC-II). Methods: The anti-QS potential of AEOs were determined using the biosensor strains Chromobacterium violaceum ATCC 12472 and Pseudomonas aeruginosa PAO1. Anti-QS activity was detected by agar-well diffusion and violacein pigment inhibition assays. Blocking of PAO1 swim and swarm motilities and biofilm formation was also performed. Results: Most of the AEOs demonstrated highly active (>95%) violacein pigment inhibition. Additionally, they inhibited swarming (40.34%-72.80%) and swimming (20.06%-50.08%) motilities of PAO1. Moreover, the majority of AEOs also decreased the biofilm formation, particularly on P. aeruginosa and S. aureus. Conclusion: Consequently, aromatherapeutic formulations might be a complementary or prophylactic cure for infectious disease by their anti-QS and antibiofilm activities rather than just antimicrobial effects.

Inhibition of Quorum Sensing Regulated Virulence Factors and Biofilm Formation by Eucalyptus globulus against Multidrug-Resistant Pseudomonas aeruginosa.

ObjectivesThe quorum-sensing–inhibitory and anti-biofilm activities of the methanol extract of E. globulus leaves were determined against clinically isolated multidrug-resistant Pseudomonas aeruginosa.MethodsThe preliminary anti-quorum–sensing (AQS) activity of eucalyptus was investigated against a biosensor strain Chromobacterium violaceum ATCC 12472 (CV12472) by using the agar well diffusion method. The effect of sub-minimum inhibitory concentrations (sub-MICs) of the methanol extract of eucalyptus on different quorum-sensing–regulated virulence factors, such as swarming motility, pyocyanin pigment, exopolysaccharide (EPS), and biofilm formation, against clinical isolates (CIs 2, 3, and 4) and reference PA01 of Pseudomonas aeruginosa were determined using the swarm diameter (mm)-measurement method, chloroform extraction method, phenol (5%)-sulphuric acid (concentrated) method, and the microtiter plate assay respectively, and the inhibition (%) in formation were calculated.ResultsThe preliminary AQS activity (violacein pigment inhibition) of eucalyptus was confirmed against Chromobacterium violaceum ATCC 12472 (CV12472). The eucalyptus extract also showed concentration-dependent inhibition (%) of swarming motility, pyocyanin pigment, EPS, and biofilm formation in different CIs and PA01 of P. aeruginosa.ConclusionOur results revealed the effectiveness of the E. globulus extract for the regulation of quorum-sensing–dependent virulence factors and biofilm formation at a reduced dose (sub-MICs) and suggest that E. globulus may be a therapeutic agent for curing and controlling bacterial infection and thereby reducing the possibility of resistance development in pathogenic strains.

Quorum quenching attenuation of rhizobacteria Chromobacterium violaceum virulence by Desmodium gangeticum root extract

Quorum sensing signal molecules used amongst bacteria for communicating cell density can be interfered with through quenching of the signal molecule, a process known as quorum quenching. Several bacteria depend upon a mechanism generally called as quorum sensing to identify their population density and initiate its colonization. The goal of this research was to test and optimize an anti-quorum sensing chemical derived from Desmodium gangeticum root extract against the biosensor strain Chromobacterium violaceum, a rhizosphere bacterium with an opportunistic pathogenic ability. Desmodium gangeticum root extract contained a chemical known as 1,6-octadien, 3,5-dimethyl-, cis, which had a promising bacterial silencing action. The studies suggested that the extract have a downregulating effect upon bacterial virulence. Response surface modelling (RSM) and the Box-Behnken technique were used to investigate the optimization of dependent variables such as bacterial density, rate of administration, and extract concentration. ANOVA and Cook's distance table were used to verify the mathematical model that resulted. The chemical exhibited anti-quorum sensing activity that is dose dependent, peaking at 300 μg/ml extract three times per day. The findings imply that the extract has no bactericidal effect but does affect the quorum sensing system.

EVALUATION OF SOME TRADITIONAL THERAPEUTIC PROPERTIES OF USNEA LONGISSIMA (ASCOMYCOTA, LICHENIZED FUNGI): ANTIMICROBIAL, ANTIQUORUM AND ANTIOXIDANT

Usnea longissima, fruticose lichen growing in the temperate region, is well known traditional medicinal herb and a reservoir of some unique secondary metabolites. Traditionally it has been commonly used as an antimicrobial agent for the treatment of ailments like tuberculosis, ulcers and skin diseases, etc. In our study, we evaluate antimicrobial, antiquorum sensing (QS) and antioxidant activity of Usnea longissima secondary metabolites extracted in acetone, methanol, and 70% hydroalcoholic solvents. Antimicrobial activity of extracts was evaluated against fungus (Aspergillus niger, Candida albicans) and six bacterial (Agrobacterium tumefaciens, Escherichia coli, Klebsiella pneumoniae, Pseudomonas aerogenosa, Staphylococcus aureus, Streptococcus mutans) strains. Whereas anti-QS activity was tested against biosensor strain Chromobacterium violaceum (CV) MCC 2290 and antibiofilm activity was checked in two gram-negative bacteria (P. aeruginosa and E. coli), two gram-positive bacteria (Bacillus cereus and Staphylococcus aureus) and fungus (Candida albicans). Results of experimental data indicated that methanolic extract exhibited maximum activity against A. tumefaciens (24±0.2 mm) and C. albicans (30±0.3 mm) with the minimum inhibitory concentration of 300µg/mL and 200µg/mL respectively. Interestingly, we also observed that all extracts effectively inhibited quorum sensing in C. violaceum and biofilm formation in Bacillus cereus, Escherichia coli, Pseudomonas aerogenosa, Staphylococcus aureus, at the concentration (100-300 µg/mL), for Candida albicans (50-150 µg/mL). Usnea longissima extract showed strong reducing power and hydroxyl radical activity and low DPPH scavenging activity. The present study suggested that U. longissima possesses potential antimicrobial and antiquorum sensing activity. These biological properties could be attributed to its significant antioxidant potential which validates its traditional use as an antimicrobial herbal source.