SCREENING FOR ANTI-QUORUM SENSING AND ANTI-BIOFILM ACTIVITY IN Viscum album L. EXTRACTS AND ITS BIOCHEMICAL COMPOSITION

Fırsatçı patojenik bakteriler, virülans ifadelerini koordine etmek için Quorum Sensing (QS) sistemini kullanır. Dolayısıyla QS sistemi, bakteriyel enfeksiyonların kontrolü için yeni bir yaklaşım olarak tercih edilebilir. Bu çalışmanın amacı, 2 Gram-pozitif, 7 Gram-negatif patojenik bakteri üzerinde analiz edilen Viscum album L. bitkisinin gövde, yaprak ve meyve gibi bölümlerine ait etanol, kloroform ve diklorometan:metanol ekstraktlarının antimikrobiyal, anti-biyofilm ve anti-quorum sensing aktivitelerinin değerlendirilmesidir. Kullanılan bitki parçası ekstraktına ve test mikroorganizmasına bağlı olarak, 50-250mg/ml arasında değişen konsantrasyonlarda ekstraklar test edildi. 50mg/ml’lik konsantrasyonda antimikrobiyal aktivite görülmediği için anti-quorum sensing ve antibiyofilm çalışmalarında bu konsantrasyon kullanılmıştır. En iyi biyolojik aktivitenin görüldüğü çözücünün ise diklorometan:metanol olduğu saptandı. Ekstraktların anti-quorum sensing aktiviteleri, Chromobacterium violaceum CV026 biyosensör suşu ve sinyal molekülü N- (β-ketokaproil) -L-homoserin lakton (3-okso-C6-HSL)’nün bulunduğu besiyerinde agar difüzyon deneyi kullanılarak da tespit edilmiştir. Biyofilm, mikrotiter plaka testi ve kristal viyole kullanılarak ölçülmüştür. Yaprak ve gövde kısımlarının antimikrobiyal, anti-biyofilm ve anti-quorum sensing aktivitesi, meyve ekstraktına göre daha yüksek verimlilik göstermiştir. V. album özütlerinin, biyofilm ya da QS inhibisyonu yoluyla mikrobiyal enfeksiyonları tedavi etme potansiyeline sahip olduğu kanısına varılmıştır.

SCREENING FOR ANTI-QUORUM SENSING AND ANTI-BIOFILM ACTIVITY IN Viscum album L. EXTRACTS AND ITS BIOCHEMICAL COMPOSITION

Many opportunistic pathogenic bacteria use the Quorum Sensing (QS) system to coordinate their virulence expressions. Thus, QS can likely be a new approach to control bacterial infections. The aim of this study was to evaluate the antimicrobial, anti-biofilm, and anti-quorum sensing activities of ethanol, chloroform, and dichloromethane: methanol extracts of leaf, stem, and fruits of the European mistletoe Viscum album L. on 2 Gram-positive and 7 Gram-negative pathogenic bacteria. The extracts at concentrations ranging from 50 to 250mg/ml were tested depending on the extracts of the plant parts and the test bacteria. The extract with 50mg/ml concentration, in which no antimicrobial activity was observed, was used for anti-quorum sensing and antibiofilm studies. The dichloromethane: methanol extracts were found to show the highest biological activities. QS activities of the plant extracts were also determined using the recently established Chromobacterium violaceum CV026 reporter strain and the signaling molecule N-(ß-ketocaproyl)-L-homoserine lactone (3-oxo-C6-HSL) agar well diffusion assay. Biofilm was quantified using the microtiter plate test and the crystal violet assay. Anti-microbial, anti-biofilm, and anti-quorum sensing activity of leaf and stem extracts showed higher efficiency than fruit extracts. It was concluded that the extracts of V. album had the potential to treat microbial infections by biofilm inhibition or inhibition of QS.

___

  • 1. Adaramoye, O., Amanlou, M., Habibi-Rezaei, M., Pasalar, P. & Ali, M.-M. 2012. Methanolic extract of African mistletoe (Viscum album) improves carbohydrate metabolism and hyperlipidemia in streptozotocin-induced diabetic rats. Asian Pacific Journal of Tropical Medicine, 5(6): 427-433.
  • 2. Adonizio, A.L., Downum, K., Bennett, B.C. & Mathee, K. 2006. Anti-quorum sensing activity of medicinal plants in southern Florida. Journal of Ethnopharmacology, 105(3): 427-435.
  • 3. Amabeoku, G.J., Leng, M.J. & Syce, J.A. 1998. Antimicrobial and anticonvulsant activities of Viscum capense. Journal of Ethnopharmacology, 61(3): 237-241.
  • 4. Antunes, L.C.M., Ferreira, R.B., Buckner, M.M. & Finlay, B.B. 2010. Quorum sensing in bacterial virulence. Microbiology, 156(8): 2271-2282.
  • 5. Arda, N., Onay, E., Koz, O. & Kirmizigul, S. 2003. Monosaccharides and polyols from mistletoes (Viscum album L) growing on two different host species. Biologia-Bratislava, 58(6): 1037-1042.
  • 6. Bazargani, M.M. & Rohloff, J. 2016. Antibiofilm activity of essential oils and plant extracts against Staphylococcus aureus and Escherichia coli biofilms. Food Control, 61: 156-164.
  • 7. Bezek, K., Kurinčič, M., Knauder, E., Klančnik, A., Raspor, P., Bucar, F. & Smole Možina, S. 2016. Attenuation of Adhesion, Biofilm Formation and Quorum Sensing of Campylobacter jejuni by Euodia ruticarpa. Phytotherapy research, 30(9): 1527-1532.
  • 8. Brackman, G. & Coenye, T. 2015. Quorum sensing inhibitors as anti-biofilm agents. Current Pharmaceutical Design, 21(1): 5-11.
  • 9. Bussing, A. 2003. Mistletoe: The Genus Viscum. Harwood Academic Publishers, Amsterdam.
  • 10. Büssing, A. & Schietzel, M. 1999. Apoptosis-inducing properties of Viscum album L. extracts from different host trees, correlate with their content of toxic mistletoe lectins. Anticancer research, 19(1A): 23-28.
  • 11. Chandrashekhara, Niranjan Raj, S., Manjunath, G., Deepak, S. & Shekar Shetty, H. 2010. Seed treatment with aqueous extract of Viscum album induces resistance to pearl millet downy mildew pathogen. Journal of Plant Interactions, 5(4): 283-291.
  • 12. Chung, P.Y. & Toh, Y.S. 2014. Anti-biofilm agents: recent breakthrough against multi-drug resistant Staphylococcus aureus. Pathogens and disease, 70(3): 231-239.
  • 13. Davis, P.H. 1965. Flora of Turkey, Vol 1-10. Edinburgh University Press, Edinburgh. 14. de Araujo, A.R., Quelemes, P.V., Perfeito, M.L.G., de Lima, L.I., Sá, M.C., Nunes, P.H.M., Joanitti, G.A., Eaton, P., Soares, M.J.d.S. & de Souza de Almeida Leite, J.R. 2015. Antibacterial, antibiofilm and cytotoxic activities of Terminalia fagifolia Mart. extract and fractions. Annals of Clinical Microbiology and Antimicrobials, 14(1): 25.
  • 15. Deliorman, D., Çalış, I. & Ergun, F. 2001. A new acyclic monoterpene glucoside from Viscum album ssp. album. Fitoterapia, 72(2): 101-105.
  • 16. Dulger, B. & Gonuz, A. 2004. Antimicrobial activity of some Turkish medicinal plants. Pakistan Journal of Biological Sciences, 7(9): 1559-1562.
  • 17. Erdonmez, D., Kenar, N. & Turkmen, K.E. 2016. Evaluation of anti-quorum sensing activity of Elaeagnus angustifolia L. extracts from Central Anatolia. Journal of Biotechnology, 231: 34.
  • 18. Ergun, F. & Deliorman, D. 1995. Viscum album L. (Ökse otu) Bitkisinin Kimyasal Bileşimi. Journal of Faculty of Pharmacy of Ankara University, 24(2): 21-33.
  • 19. Eroğlu, M. & Usta, M. 1993. Viscum album L.'un Sarıçam Artımına Odunun Kimyasal ve Morfolojik Özelliklerine Etkisinin Araştırılması. II Ulusal Orman Ürünleri Endüstrisi Kongresi, 1(1): 116-122.
  • 20. Ertürk, Ö., Kati, H., Yayli, N. & Demirbağ, Z. 2004. Antimicrobial activity of Viscum album L. subsp. abietis (Wiesb). Turkish Journal of Biology, 27(4): 255-258.
  • 21. Ghuman, S., Ncube, B., Finnie, J.F., McGaw, L.J., Coopoosamy, R.M. & Van Staden, J. 2016. Antimicrobial Activity, Phenolic Content, and Cytotoxicity of Medicinal Plant Extracts Used for Treating Dermatological Diseases and Wound Healing in KwaZulu-Natal, South Africa. Frontiers in Pharmacology, 7, 320.
  • 22. Gray, A. & Flatt, P. 1999. Insulin-secreting activity of the traditional antidiabetic plant Viscum album (mistletoe). Journal of Endocrinology, 160(3): 409-414.
  • 23. Güner, A. 2012. Türkiye Bitkileri Listesi (Damarlı Bitkiler). Nezahat Gökyiğit Botanik Bahçesi ve Flora Araştırmaları Derneği Yayını, İstanbul.
  • 24. Habeck, M. 2003. Mistletoe compound enters clinical trials. Drug discovery today, 8(2): 52-53.
  • 25. Hoffman, L.R., D'Argenio, D.A., MacCoss, M.J., Zhang, Z., Jones, R.A. & Miller, S.I. 2005. Aminoglycoside antibiotics induce bacterial biofilm formation. Nature, 436(7054): 1171-1175.
  • 26. Hussain, M.A., Khan, M.Q., Hussain, N. & Habib, T. 2011. Antibacterial and antifungal potential of leaves and twigs of Viscum album L. Journal of Medicinal Plants Research, 5(23): 5545-5549.
  • 27. Karagöz, A., Önay, E., Arda, N. & Kuru, A. 2003. Antiviral potency of mistletoe (Viscum album ssp. album) extracts against human parainfluenza virus type 2 in Vero cells. Phytotherapy Research, 17(5): 560-562.
  • 28. Kaya, M., Sargin, I. & Erdonmez, D. 2016. Microbial biofilm activity and physicochemical characterization of biodegradable and edible cups obtained from abdominal exoskeleton of an insect. Innovative Food Science & Emerging Technologies, 36: 68-74.
  • 29. Khan, M.S.A., Zahin, M., Hasan, S., Husain, F. M., Ahmad, I. 2009. Inhibition of quorum sensing regulated bacterial functions by plant essential oils with special reference to clove oil. Letters in applied microbiology, 49(3): 354-360.
  • 30. Kotan, R., Okutucu, A., Ala Görmez, A., Karagoz, K., Dadasoglu, F., Karaman, I., Hasanekoglu, I. & Kordali, Ş. 2013. Parasitic bacteria and fungi on common mistletoe (Viscum album L.) and their potential application in biocontrol. Journal of Phytopathology, 161(3): 165-171.
  • 31. Martinelli, L., Rosa, J.M., Ferreira, C.D.S.B., Nascimento, G.M.D.L., Freitas, M.S., Pizato, L.C., Santos, W.D.O., Pires, R.F., Okura, M.H. & Malpass, G.R.P. 2017. Antimicrobial activity and chemical constituents of essential oils and oleoresins extracted from eight pepper species. Ciência Rural, 47(5): e20160899.
  • 32. McClean, K.H., Winson, M.K., Fish, L., Taylor, A., Chhabra, S.R., Camara, M., Daykin, M., Lamb, J.H., Swift, S., Bycroft, B.W. and Stewart, G.S., 1997. Quorum sensing and Chromobacterium violaceum: exploitation of violacein production and inhibition for the detection of N-acylhomoserine lactones. Microbiology, 143(12), 3703-3711.
  • 33. Musthafa, K., Ravi A., Annapoorani, A., Packiavathy, I.V. & Pandian, S. 2010. Evaluation of anti-quorum-sensing activity of edible plants and fruits through inhibition of the N-acyl-homoserine lactone system in Chromobacterium violaceum and Pseudomonas aeruginosa. Chemotherapy, 56(4): 333-339.
  • 34. Nawrot, R., Barylski, J., Nowicki, G., Broniarczyk, J., Buchwald, W. & Goździcka-Józefiak, A. 2014. Plant antimicrobial peptides. Folia microbiologica, 59(3): 181-196.
  • 35. Nazaruk, J. & Orlikowski, P. 2016. Phytochemical profile and therapeutic potential of Viscum album L. Natural Product Research, 30(4): 373-385.
  • 36. O'Toole, G.A. 2011. Microtiter dish biofilm formation assay. Journal of Visualized Experiments, (47): e2437-e2437.
  • 37. Ogunmefun, O., Fasola, T., Saba, A. & Oridupa, O. 2013. The ethnobotanical, phytochemical and mineral analyses of Phragmanthera incana (Klotzsch), a species of mistletoe growing on three plant hosts in South-Western Nigeria. International journal of biomedical science, 9(1): 33.
  • 38. Oliveira, B.D.Á., Rodrigues, A.C., Cardoso, B.M.I., Ramos, A.L.C.C., Bertoldi, M.C., Taylor, J.G., da Cunha, L.R. & Pinto, U.M. 2016. Antioxidant, antimicrobial and anti-quorum sensing activities of Rubus rosaefolius phenolic extract. Industrial Crops and Products, 84: 59-66.
  • 39. Rumbaugh, K.P., Diggle, S.P., Watters, C.M., Ross-Gillespie, A., Griffin, A.S. & West, S.A. 2009. Quorum sensing and the social evolution of bacterial virulence. Current Biology, 19(4): 341-345.
  • 40. Rutherford, S.T. & Bassler, B.L. 2012. Bacterial quorum sensing: its role in virulence and possibilities for its control. Cold Spring Harbor perspectives in medicine, 2(11): a012427.
  • 41. Sadananda, T., Govindappa, M. & Ramachandra, Y. 2014. In vitro antioxidant activity of lectin from different endophytic fungi of Viscum album L. British Journal of Pharmaceutical Research, 4(5): 626.
  • 42. Sánchez, E., Rivas Morales, C., Castillo, S., Leos-Rivas, C., García-Becerra, L. & Ortiz Martínez, D.M. 2016. Antibacterial and Antibiofilm Activity of Methanolic Plant Extracts against Nosocomial Microorganisms. Evidence-Based Complementary and Alternative Medicine, 2016: 1572697.
  • 43. Sandasi, M., Leonard, C.M. & Viljoen, A.M. 2008. The effect of five common essential oil components on Listeria monocytogenes biofilms. Food Control, 19(11): 1070-1075.
  • 44. Sengul, M., Yildiz, H., Gungor, N., Cetin, B., Eser, Z. & Ercisli, S. 2009. Total phenolic content, antioxidant and antimicrobial activities of some medicinal plants. Pakistan Journal of Pharmaceutical Sciences, 22(1): 102-106.
  • 45. Stepanović, S., Vuković, D., Dakić, I., Savić, B. & Švabić-Vlahović, M. 2000. A modified microtiter-plate test for quantification of staphylococcal biofilm formation. Journal of microbiological methods, 40(2): 175-179.
  • 46. Stoss, M. & Gorter, R.W. 1998. No evidence of IFN-γ increase in the serum of HIV-positive and healthy subjects after subcutaneous injection of a non-fermented Viscum album L. extract. Natural immunity, 16(4): 157-164.
  • 47. Turker, A.U., Yıldırım, A.B. & Karakas, F.P. 2012. Antitumor and antibacterial activities of Viscum album L. grown on different host trees. Spatula DD, 2: 229-236.
  • 48. Tusenius, K., Spoek, J. & Kramers, C. 2001. Iscador Qu for chronic hepatitis C: an exploratory study. Complementary therapies in medicine, 9(1): 12-16.
  • 49. Vuong, C., Kocianova, S., Voyich, J.M., Yao, Y., Fischer, E.R., DeLeo, F.R. & Otto, M. 2004. A crucial role for exopolysaccharide modification in bacterial biofilm formation, immune evasion, and virulence. Journal of Biological Chemistry, 279(52): 54881-54886.
  • 50. Waters, C.M. & Bassler, B.L. 2005. Quorum sensing: cell-to-cell communication in bacteria. Annual review of cell and developmental biology, 21: 319-346.