Quorum Sensing and Biofilms

Quorum Sensing and Biofilms

  • Mesrop Ayrapetyan added an answer:
    Are any quorum sensing inhibitors available for medicinal use in Europe?
    I am involved in an ongoing study looking at biofilm infection in patients. To my knowledge QSI are not yet used but may be advantageous in the fight against chronic bacterial infection. Any advice in this area gratefully received.
    Mesrop Ayrapetyan · University of North Carolina at Chapel Hill

    I don't have any direct insight into what is available in Europe (or anywhere for that matter), as far as QSI's go. But I can say that although this is a well-conceived idea since QS induces many virulence genes in a wide range of pathogens, there maybe potential shortcomings to this approach. One such idea is that the use of QS inhibitors may reduce the growth rate of bacteria during infection. This is a problem because reducing the growth rate will reduce the efficacy of many bactericidal antibiotics. It is almost like using a bacteriostatic and bactericidal antibiotic in combination, which is not an effective strategy. This is especially relevant for chronic and recurrent infections. In infections such as TB or recurrent urinary tract infections or respiratory infections in patients with cystic fibrosis, the bacteria will reduce in number during treatment, but some cells will enter a slow or non-growing "persistent" or "viable but nonculturable" phenotype. It may be feasible to reverse such dormant states in combination with antibiotic therapy to eradicate the cells. Of course, there is ethical concern with this approach (i.e. the reactivation of a dormant pathogen within a host has the potential to harm a patient). In any case, these are simply hypothetical situations I am proposing. But it's definitely an interesting topic and one that deserves more attention. 

  • Ines el ghali added an answer:
    Can Quorum sensing autoinducers such as AHLs and AI-2 destroyed or affected by high temperature?

    Quorum sensing autoinducers

    Ines el ghali · National Institute of Livestock and Grassland Science

    Thank you Dear Christian Utpatel : practically saved my life :)

  • Alireza Japoni-Nejad added an answer:
    Are efflux pumps of bacteria the means by which all secretions from the bacterium take place?
    The first response of bacteria to a noxious agent such as an antibiotic is the induced over expression of efflux pumps. Once the threat of the noxious agent passes, the level of efflux pump expression returns to its normal baseline.
    Material secreted by bacteria such as those involved in quorum sensing (QS) and biofilm take place via the efflux pump system of the bacterium (Varga ZG, Armada A, Cerca P, Amaral L, Mior Ahmad Subki MA, Savka MA, Szegedi E, Kawase M, Motohashi N, Molnár J. Inhibition of quorum sensing and efflux pump system by trifluoromethyl ketone proton pump inhibitors. In Vivo. 2012). Because diffusion of other materials does not take place readily across the plasma membrane or outer membrane of bacteria, the means by which these materials (toxins, inducers, inhibitors, etc) are secreted may represent targets that may be exploited for the therapy of infections whose success lies in the materials secreted.
    I would like for the members of RG to present their views on this subject for the purpose of expanding the possibility of new targets to be exploited, especially for the therapy of Gram-negative infections.
    Alireza Japoni-Nejad · Pasteur Institute of Iran (IPI)

    Dear Dr Leonard Amaral 

    ATP binding cassette (ABC pumps) transporters is Type one secretion systems (T1SS) . ABC are heterotrimeric complexes consisting of an IM ABC exporter, a membrane fusion protein (MFP) and a pore-forming, outermembrane protein (OMP). T1SS allow the secretion of a wide range of substrates (proteinaceous and nonproteinaceous) from the cytoplasm to the extracellular space in a single step, without a stable periplasmic intermediate.

  • Fatma Allam added an answer:
    What indicator organisms can I use for simple detection of Quorum Sensing Inhibitors?

    I want to screen for QSI from environmental bacterial isolates and plant fragments. I am currently working with Chromobacterium ATCC 12472 by inoculating it on soft agar overlayed on test organisms or plant fragments.

    Right now I feel the need to add more indicator organisms to further confirm QSI activity. 

    Can anyone suggest or recommend other indicator bacterial strains? And can I ask where or how to obtain them?

    Fatma Allam · Alexandria University

    Dr. Ankita Pagedar,

    Could you please add the publications of your answer.

    Thanks

  • Putri Sari added an answer:
    Does anyone have Chromobacterium violaceum CV026 and VIR07?
    I need these cultures for my research.
    Putri Sari · Bogor Agricultural University

    Dear Mrs. Hafizah Chenia

    Would you mind to send me your reprint, i need CV026 too.

    my email address : putriekasari@apps.ipb.ac.id 

    thank you so much for your kind help

    SIncerely

    Putri Eka Sari-Indonsia

  • Hafizah Chenia asked a question:
    Does anyone have access to the Vibrio harveyi Autoinducer 2 biosensor set..BB120, BB152 etc?

    My panel from ATCC died due to biofreezer malfunction. I want to test for AI-2 production.

  • Trinish Sarkar added an answer:
    Which specific nutrient stress induces stationary phase in an E.coli population growing in Luria-Bertani medium?
    Among several stress conditions, low levels of carbon, nitrogen or phosphorus, as well as amino acid starvation, trigger Rpos cascade,in turn contributing to stationary phase induction.
    Trinish Sarkar · Indian Institute of Science Education and Research Kolkata
    Thank you Surendra for recommending the paper.It was very helpful.
  • Atena Eslami added an answer:
    Can you recommend some simple software for working with phylogenetic trees?
    I am just starting with phylogenetic trees. Does anyone know a free and not too complicate online platform for creating such trees? I have got environmental samples from biofilms.
    Suggestions? Any help would be priceless.
    Atena Eslami · Lorestan University
    Hi there are many ways to have a nice phylogenetic trees such as NTYSIS, Genealex, ClustalW, MEGA4 and 5, BEAST softwares.
  • Daniel Deng added an answer:
    How do you distinguish/separate dormant bacteria from active bacteria?
    In bacteria biofilm, a lot of bacteria become metabolically dormant while some are still actively growing. How do you separate them?
    Daniel Deng · University of Southern Mississippi
    Dr. Flemming, I would like to have a look at those papers. Please send me PDFs to danielttkl@gmail.com. Thank you very much.
  • Ganga Viswanath added an answer:
    How can we determine AHLs from Biofilm in Microtitre plates?
    I want to grow pesudomonas aeruginosa PA01 biofilm in microtitre plate, can I measure AHLs or AI-2 in microtitre plate?
    Ganga Viswanath · M S Swamininathan Research Foundation
    you cannot measure it directly on microtitre plate. In the plate you can just observe whether AHL is positive or negative. here are some papers which might be helpful for you.
    Extraction of violacein from Chromobacterium violaceum provides
    a new quantitative bioassay for N-acyl homoserine lactone
    autoinducers
    Isolation and characterization of acyl homoserine
    lactone–producing bacteria during an urban
    river biofilm formation
    http://jb.asm.org/content/187/10/3477
  • Biofilms in water systems - reasons and solutions?
    What are the major reasons for using biofilms in water systems and what are possible solutions?
    Mohammad Mahfuz Ali Khan Shawan · International Centre for Diarrhoeal Disease Research, Bangladesh
    Hi, Najmul,

    Micro-organisms thrive in the presence of water and nutrients. The movement of water creates an unstable environment for survival. Consequently, micro-organisms have evolved in a manner that allows them to adhere to a substrate with great tenacity. When they find a suitable surface they attach themselves and then initiate multiplication. Growth can be rapid, with new cells attaching to each other as well as extending laterally across the surface.

    There are a variety of nutrient sources within water systems. These include scale, sediment, corrosion products and other trapped organic and inorganic matter. The movement of the water ensures that colonies receive a constant supply of nutrients. When this is combined with specific temperature conditions, which are often found inside heating systems such as calorifiers and heat exchangers, the perfect conditions exist for rapid microbial growth. The population will predominantly consist of aerobic (oxygen seeking) and anaerobic (living without oxygen) bacteria.

    Using Chlorine Dioxide as a part of a water hygiene management programme will eliminate biofilm and maintain a safe, efficient and cost effective water system.

    Have a nice day.........
  • Joseph Molnar added an answer:
    If bacteria use 'Quorum Sensing' to engage in intra-and-inter-species communication, how can we leverage on this phenomenon for antibiotics?
    Different chemicals are produced by bacteria for inter and intra species communication, how do we use these chemicals to our advantage in developing antibiotics?
    Joseph Molnar · University of Szeged
    The QS can be a potential target of inhibitor phytochemicals or synthetic compounds. QS inhibitors are able to modify the effect of chemotherapy through blocking acyl-homoserin lactone synthase, other mediators, or second messenger e.g. cyclic-di-GMP resulting in changes in active efflux or diffusion, changes in cell to cell signal transport, secretion of biofilm, swarming, efflux pump systems, PMF, cell adhesion, cohesion etc.
  • Mohamed ahmed Sebak added an answer:
    Do you know any protocols to detect metabolic activity in bacterial biofilm?
    I grow my bacteria in a define culture medium (C-limited) in 96-well plates. I tried TTC assay, but it doesn't seem to work well. I have to add extra glucose to intensify the reaction and the red precipitate is hard to dissolve for measurement.
    Mohamed ahmed Sebak · Beni Suef University
    I think XTT may be useful

    Try it

    Best wishes
  • Mesrop Ayrapetyan added an answer:
    How do bacteria regulate the production of quorum sensing molecules?
    Quorum sensing (QS) in bacteria regulates their gene expression in a population-density dependent manner, which controls population behaviors. Once the population reach a high density and so do the QS molecules, the expression of many genes in bacteria are triggered. Do bacteria still keep on producing QS signals once their population is high? How do they regulate QS production?
    Mesrop Ayrapetyan · University of North Carolina at Chapel Hill
    Several types of quorum sensing have been discovered to date. For example: there are species specific autoinducers (QS molecules) known as acyl-homoserine lactones which have been discovered in many bacteria. There is also interspecies quorum sensing through the autoinducer-2 molecule which has been found in several species as well. There are also other quorum sensing molecules that are peptides.

    In Vibrios for example, the quorum sensing master regulator (LuxR) which is activated by the presence of a high concentration of AI-2 molecules (QS molecules), regulates many genes (activates or represses). Interestingly, LuxR also increases transcription of the gene that codes for the enzyme that catalyzes the formation of AI-2. This may imply that at high cell density when there are more and more QS molecules around, even more is being produced.

    To get a more detailed answer about specific quorum sensing systems I would look at some of the literature. Here's a good paper to give you some idea

    http://everettsd.schoolwires.net/cms/lib07/WA01920133/Centricity/Domain/1010/Quorum%20Sensing%20-%20Bassler%20Article.pdf

    I hope this helps.

    -Mez
  • Richard H Bentham added an answer:
    Is bacterial colony grown on agar plate a biofilm?
    I know it is more or less a philosophical question, however quite intriquing. Bacterial cells on agar plate adhere to surface and majority of cells is covered by slime layers. Thus it matches the biofilm definition, however I haven't heard anyone who would define "agar" colonies such way.
    Richard H Bentham · Flinders University
    Hi All,

    I think a simple answer to this question is that some organisms grown in monoculture on an agar plate are incapable of forming biofilms on other nutrient depleted surfaces. If they can't grow as biofilm on a surface other than nutrient agar then I suppose that they are not truly biofilm forming organisms. Alternatively there are many biofilm organisms that will not grow on agar surfaces. I think we should understand that agar is , for the most part, a very artificial medium.

    Organisms like Pseuodmonads will grow happily on agar and are well known biofilm pioneers, others are secondary colonisers (for instance my interest Legionellae) or of course the predators (virus, protozoa and other eukarya). I agree with Andrew Spiers that there are a lot of terms floating around (pun intended!) but I think they tend to refer to physiological adaptation ecological niches. To that extent quorum sensing in response to environment may be a major determinant of biofilm / floc / slime/ granule etc formation within any given ecosystem.

    hope this helps,

    Cheers,

    Richard
  • Marwan M. A. Mohammed added an answer:
    Biofilm Exopolysaccharide determination by fluorescence dye
    I am doing biofilm EPS determination using WGA and ConA alexa 488 fluorescence dye but even in positive control, I do not get any fluorescence under spectrofluorometer. I am using the concentration of dye as 10 micolite/mL as recommended by manufacturer of Invitrogen and paper.
    Does anybody know why fluorescence is not visible?
    Marwan M. A. Mohammed · University of Bergen
    I stained the proteins in the biofilm matrix using the SYPRO® Ruby Biofilm Matrix Stain, but I didn't try staining of Exopolysaccharide.
  • Giuseppe Gallo added an answer:
    How can I prepare cracking buffer for bacterial cell sonication for protein extraction?
    I am going to prepare cracking buffer pH 7.5, can I replace Dithiotheritol (DTT) with SDS for availability?
    Giuseppe Gallo · Università degli studi di Palermo
    DTT and SDS are used for different purposes in protein buffers so you could not replace them reciprocally. Standard methods to obtain native proteins imply the utilization of buffers at pH 7 or 7.5 essentially containing Tris-HCl and salt (like NaCl). If you are interested in obtaining a particular protein and have problem in getting it in your lysate you can try to change pH or salt concentration to improve solubility in the buffer. In any case, I would like to suggest you to add protease inhibitors (some people use DTT or other reductants at very very low concentrations to prevent protease activity in native lysis buffer but it would be best to work at low temperature and use other inhibitors like PMSF).
    All the best,
    GG
  • Ramona Khanum added an answer:
    What is the pathway for the formation of bio film by Staphylococcus aureus? What are the proteins and genes involved?
    I would like to do docking to see antimicrobial substance inhibitory activity on the proteins/molecules involved in the pathway for the production of Staphylococcus aureus bio film.
    Ramona Khanum · International Medical University (IMU)
    Hi Yan Yan, I have worked on S. epidermidis, hence I have more refs on their mechanism of biofilm formation (if you want to have read at them, do inform me) but I have a ref on S. aureus & their biofilm formation - hope it helps...
  • Gunjankumar Mehta added an answer:
    Can anyone provide me with A. tumefaciens KYC6 & C. violaceum ATCC 12475 or C. violaceum ATCC 31532 for my doctoral research?
    I am using Chromobacterium and Agrobacterium bioreporter systems for QS AHL detection bioassay. Bioassay with both of these reporter systems requires positive controls for justification of results and to establish truthfulness of my reporter strain. If anyone can help me, I would be really obliged. The support would be acknowledge at all appropriate places and events.
    Gunjankumar Mehta · Virani Science College
    Thank you Viraj, will surely do that.
  • Srinath Naganathan added an answer:
    What is the mechanism of Quorum sensing in skin disease related microbes like Candida?
    This field is a developing ground where the potential elements responsible for the activation and metabolism of compound there by causing a protective layer making it antibiotic resistant. We are here trying to understand the pathways involved in quorum sensing mechanism present in Candida, so that most of the problems related to skin can be tackled.
    Srinath Naganathan · Kongunadu Arts and Science College
    Thank you Daniel for your kind suggestion, I will try that but can you please send the details of principal supervisor or Scientist who is working in this QS aspect?
  • Vihang Ghalsasi added an answer:
    Does anyone know how to detach bacteria from cultured biofilm, such as colony biofilm, or biofilm attached on glass/plastic slides?
    I want to count bacteria from biofilm with DNA probes and study bacterial interactions in biofilm.
    Vihang Ghalsasi · Universität Heidelberg
    I am working with E. coli biofilms. I grow them in plastic petri plates for 24 or 48 h. Remove the planktonic cells, wash the plate gently and simply scrape off the attached cells with a spreader or cell scrapper by putting some buffer or medium on the plate. It works well if you want to harvest the cells for counting, fluorescence measurement or for any reason where they need to be viable. Sonication is likely to kill the cells. As correctly pointed out by Davide, it is important to characterize the setup to ensure that the cells are alive. I would suggest to use sonication bath. If you are growing biofilms in microtiter plates, you can wash the wells gently, put some buffer or medium, seal the plate thoroughly by parafilm and put it in the sonication bath. Pulses of 30 to 60 s will detach the cells from the surface. Viability can be checked by plating. As far as my experience says, the cells are mostly viable.
  • Pramal Biswa added an answer:
    Has anyone performed analysis for N-acylhomoserine lactones (AHLs) using TLC and LC-MS?
    I am working on bacterial quorum sensing and biofilm. I tried TLC for AHLs assay using Agrobacterium bioreporter, but it didn't work, although I got positive results with T-streak method. Does anybody know how to detect AHLs, by TLC and LC-MS?
    Pramal Biswa · Indian Institute of Technology Madras
    its all a matter of trail and error ......i guess sara. I use initital 0.1 OD and grow it till 48hours in xgal supplemented media at 25oC and then observe

    One more point incubate your culture at temperatures below 30 to get better results

    Feel free to ask for more doubts.will try helping
  • Alexandre Fugère added an answer:
    What is the best way to conclude that a microorganism is able to form biofilm?
    Any protocol, suggestion, or idea is welcome.
    Alexandre Fugère · Laboratoire M2
    Staining biofilm grown statically with Crystal Violet works very well. All of my biofilm and biofilm stimulation assays were done this way. Great way to easily quantify different levels of biofilm formation capacity by measuring resulting OD (560 nm). Especially if you're aiming at comparing different strains with each other.
  • Rakesh Yashroy added an answer:
    Can quorum sensing signals in gram-negative microbes be sent via pilus-like nanotubes & outer membrane vesicles as in EM pictures in this paper here?
    Microbe-microbe, microbe-host/target cell signalling may not just be confined to free signal molecules flow with dilution on the way.
    Rakesh Yashroy · Indian Veterinary Research Institute
    Thanks Mehvish for the reference.
  • Siddiqi MF asked a question:
    AHL and Autoinder-2
    How can we measure concentration of AHLs and Autoinducer-2 in biofilm?
  • Siddiqi MF asked a question:
    How to Quantify (concentration) of AHLs and AI-2 in the Biofilm?
    Any Spectrophotometric method ?
  • Siddiqi MF asked a question:
    How to quantify Acylhomoserine lactones in biofilm? Any Spectrophotometric method ?
    Quantification of AHLs and AI-2 in biofilm?

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