Inhibition of Pseudomonas aeruginosa Biofilm Formation and Quorum Sensing System by Extracts of Prunus avium Stalk

Öz

Recently, misuse or overuse of antibiotics has led to the antibiotic resistance problem, a global healthcare problem. Most virulence factors and biofilm formation in Pseudomonas aeruginosa are controlled by quorum sensing (QS). The inhibition of QS system by inhibitor molecules has been suggested as a novel alternative antivirulence approach in which no need to kill the bacteria. In the present study, QS and biofilm inhibitory potentials of the methanol and acetone extracts of Prunus avium stalk against P. aeruginosa were evaluated. The extracts were tested at the concentrations of 240, 120, and 60 μg/ml. lasB-gfp, rhlA-gfp, pqsA-gfp biosensor strains and P. aeruginosa PAO1 were used to monitor QS and biofilm inhibition, respectively. Fluorescence and absorbance measurements were performed on Cytation 3 multimode microplate reader. QS inhibition ratios for las, rhl, and pqs systems and biofilm inhibition ratios of the acetone extracts were recorded as 70.43%, 47.25%, 76.31%, and 47.76% (±6,60) and of the methanol extracts as 74.96%, 40.10%, 71.89%, and 38.54% (±3,56) at a certain concentration of 240 μg/ml, respectively. As a result, anti-QS and anti-biofilm properties of acetone extracts were better than that of methanol extracts. Further investigations are needed to discover inhibitor compounds of P. avium and also their effects on human cells and then these compounds may be used in new drug discoveries.

Anahtar Kelimeler

• Pseudomonas aeruginosa,quorum sensing,biofilm,quorum quenching,Prunus avium stalk,sweet cherry

Prunus avium Sapından Elde Edilen Özütlerle Pseudomonas aeruginosa’ya ait Quorum Sensing Sisteminin ve Biyofilm Oluşumunun İnhibisyonu

Öz

Öz Son zamanlarda, antibiyotiklerin yanlış veya aşırı kullanımı, küresel bir sağlık sorunu olan antibiyotik direnci sorununa yol açmıştır. Pseudomonas aeruginosa'da çoğu virülans faktörü ve biyofilm oluşumu, quorum sensing (QS) ile kontrol edilir. QS sisteminin inhibitör moleküller tarafından inhibisyonu, bakterileri öldürmeye gerek olmayan yeni bir alternatif antivirulens yaklaşımı olarak önerilmiştir. Bu çalışmada, Prunus avium sapından elde edilen metanol ve aseton özütlerinin P. aeruginosa'ya karşı QSI (QS inhibitörleri) ve anti-biyofilm potansiyellerini değerlendirdik. Ekstraktlar 240, 120 ve 60 μg/ml'lik konsantrasyonlarda test edilmiştir. QS ve biyofilm inhibisyonunu izlemek için lasB-gfp, rhlA-gfp, ve pqsA biyosensör suşları ve P. aeruginosa PAO1 kullanıldı. Floresans ve absorbans ölçümleri Cytation 3 çok modlu mikroplaka okuyucu üzerinde gerçekleştirildi. 240 μg/ml konsantrasyonunda aseton özütlerinin las, rhl ve pqs sistemleri üzerine QS ve biyofilm inhibisyon oranları sırasıyla % 70.43, % 47.25, % 76.31 ve % 47.76 (± 6,60) ve metanol özütlerinin sırasıyla % 74.96, % 40.10, % 71.89 ve % 38.54 (± 3,56) olarak kaydedilmiştir. Sonuç olarak, aseton özütlerinin anti-QS ve anti-biyofilm özellikleri metanol özütlerinden daha başarılı olmuştur. P. avium’un inhibitör bileşikleri ve bu bileşiklerin insan hücreleri üzerindeki etkilerini keşfetmek için daha fazla araştırmaya ihtiyaç vardır ve daha sonra bu bileşikler yeni ilaç keşiflerinde kullanılabilir.

Anahtar Kelimeler

• Pseudomonas aeruginosa,quorum sensing,biofilm,quorum quenching,Prunus avium stalk,sweet cherry

Kaynakça

1. [1] Peterson, J.W. (1996). Bacterial pathogenesis. In Medical microbiology 4th edition, S. Baron (ed.), University of Texas Medical Branch, Galveston.
2. [2] Rasko, D.A. and Sperandio, V. (2010). Anti-virulence strategies to combat bacteria-mediated disease. Nat. Rev. Drug Discov., 9(2): 117-128.
3. [3] Dickey, S.W., Cheung, G.Y. and Otto, M. (2017). Different drugs for bad bugs: Antivirulence strategies in the age of antibiotic resistance. Nat. Rev. Drug Discov., 16: 457-471.
4. [4]World Health Organization (2017). Prioritization Pathogens Infographic. http://www.who.int/medicines/prioritization-pathogens-infographic.pdf?ua=1.
5. [5] Driscoll, J.A., Brody, S.L. and Kollef, M.H. (2007). The epidemiology, pathogenesis and treatment of Pseudomonas aeruginosa infections. Drugs, 67: 351-368.
6. [6] Schwarz, C., Bouchara, J.P., Buzina, W., Chrenkova, V., Dmeńska, H., de la Pedrosa, E.G.G., Canton, R., Fiscarelli, E., Le Govic, Y., Kondori, N., Matos, T., Romanowska, E., Ziesing, S. and Sedlacek, L. (2018). Organization of patient management and fungal epidemiology in cystic fibrosis. Mycopathologia, 183: 7-19.
7. [7] Singh, P.K., Schaefer, A.L., Parsek, M.R., Moninger, T.O., Welsh, M.J. and Greenberg, E.P. (2000). Quorum-sensing signals indicate that cystic fibrosis lungs are infected with bacterial biofilms. Nature, 407(6805): 762-764.
8. [8] Smith, R.S. and Iglewski, B.H. (2003). Pseudomonas aeruginosa quorum sensing as a potential antimicrobial target. J. Clin. Invest., 112(10): 1460-1465.

9. [9] Rasmussen, T.B. and Givskov, M. (2006). Quorum-sensing inhibitors as anti-pathogenic drugs. Int. J. Med. Microbiol., 296(2-3): 149-161.
10. [10] Rutherford, S.T. and Bassler, B.L. (2012). Bacterial quorum sensing: its role in virulence and possibilities for its control. Cold Spring Harb. Perspect. Med., 2(11):a012427.
11. [11] Khan F., Javaid, A., Kim, Y.M., (2019). Functional diversity of quorum sensing receptors in pathogenic bacteria: Interspecies, intraspecies and interkingdom level. Curr. Drug Targets, 20(6): 655-667.
12. [12] Lee, J. and Zhang, L. (2015). The hierarchy quorum sensing network in Pseudomonas aeruginosa. Protein & Cell, 6(1): 26-41.
13. [13] Moradali, M.F., Ghods, S. and Rehm, B.H. (2017). Pseudomonas aeruginosa lifestyle: a paradigm for adaptation, survival, and persistence. Front. Cell Infect. Microbiol., 7:39.
14. [14] Olson, M.E., Ceri, H., Morck, D.W., Buret, A.G. and Read, R.R. (2002). Biofilm bacteria: formation and comparative susceptibility to antibiotics. Can. J. Vet. Res., 66(2): 86.
15. [15] Tang, K. and Zhang, X.H. (2014). Quorum quenching agents: resources for antivirulence therapy. Mar. Drugs, 12: 3245-3282.
16. [16] Kalia, V.C. (2013). Quorum sensing inhibitors: an overview. Biotechnol. Adv., 31(2):224-245.
17. [17] Fowler, M.W. (2006). Plants, medicines and man. J. Sci. Food Agric., 86 (12): 1797-1804.
18. [18] Koh, C.L., Sam, C.K., Yin, W.F., Tan, L., Krishnan. T., Chong. Y. and Chan, K.G. (2013). Plant-derived natural products as sources of anti-quorum sensing compounds. Sensors, 13(5): 6217-6228.
19. [19] Al-Moghrabi, R.S., Abdel-Gaber, A.M. and Rahal, H.T. (2018). A comparative study on the inhibitive effect of Crataegus oxyacantha and Prunus avium plant leaf extracts on the corrosion of mild steel in hydrochloric acid solution. Int. J. Ind. Chem., 9(3):255-263.
20. [20] Ademović, Z., Hodžić, S., Zahirović, Z.H., Husejnagić, D., Džananović, J., Šarić-Kundalić, B. and Suljagić, J. (2017). Phenolic compounds, antioxidant and antimicrobial properties of the wild cherry (Prunus avium L.) stem. Acta Period. Technol., 48: 1-13.
21. [21] Kim, D.O., Heo, H.J., Kim, Y.J., Yang, H.S. and Lee, C.Y. (2005). Sweet and sour cherry phenolics and their protective effects on neuronal cells. J. Agric. Food Chem., 53(26): 9921-9927.
22. [22] McCune, L.M., Kubota. C., Stendell-Hollis, N.R. and Thomson, C.A. (2010). Cherries and health: a review. Crit. Rev. Food Sci. Nutr., 51: 1-12.
23. [23] Ahn, S.M., Ryu, H.Y., Kang, D.K., Jung, I.C. and Sohn, H.Y. (2009). Antimicrobial and antioxidant activity of the fruit of Prunus avium L. Microbiology and Biotechnology Letters, 37(4): 371-376.
24. [24] Ördögh, L. (2010). Antioxidant and antimicrobial activities of fruit juices and pomace extracts against acne-inducing bacteria. Acta Biol. Szeged., 54(1): 45-49.
25. [25] Hussain, M.A. (2011). Isolation and identification of an anthocyanin compound from cherry fruit (Prunus avium L.) and study of its antibacterial activity. Tikrit Journal of Pure Science, 16(2): 26-30.
26. [26] Rovčanin, B.R., Ćebović, T., Stešević, D., Kekić, D. and Ristić, M. (2015). Antibacterial effect of Herniaria hirsuta, Prunus avium, Rubia tinctorum and Sempervivum tectorum plant extracts on multiple antibiotic resistant Escherichia coli. Bioscience Journal, 31(6): 1852-1861.
27. [27] Gonçalves, B., Landbo, A.K., Let, M., Silva, A.P., Rosa, E. and Meyer, A.S. (2004). Storage affects the phenolic profiles and antioxidant activities of cherries (Prunus avium L) on human low‐density lipoproteins. J. Sci. Food Agr., 84(9): 1013-1020.
28. [28] Serrano, M., Guillén, F., Martínez-Romero, D., Castillo, S. and Valero, D. (2005). Chemical constituents and antioxidant activity of sweet cherry at different ripening stages. J. Agric. Food Chem., 53(7): 2741-2745.
29. [29] Zhang, Y., Neogi, T., Chen, C., Chaisson, C., Hunter, D.J. and Choi, H.K. (2012). Cherry consumption and decreased risk of recurrent gout attacks. Arthritis & Rheumatism, 64(12): 4004-4011.
30. [30] Hentzer, M., Riedel, K., Rasmussen, T.B., Heydorn, A., Andersen, J.B., Parsek, M. and Kjelleberg, S. (2002). Inhibition of quorum sensing in Pseudomonas aeruginosa biofilm bacteria by a halogenated furanone compound. Microbiology, 148(1): 87-102.
31. [31] Yang, L., Barken, K.B., Skindersoe, M.E., Christensen, A.B., Givskov, M. and Tolker-Nielsen, T. (2007). Effects of iron on DNA release and biofilm development by Pseudomonas aeruginosa. Microbiology, 153(5): 1318-1328.
32. [32] Yang, L., Rybtke, M.T., Jakobsen, T.H., Hentzer, M., Bjarnsholt, T., Givskov, M. and Tolker-Nielsen T. (2009). Computer-aided identification of recognized drugs as Pseudomonas aeruginosa quorum-sensing inhibitors. Antimicrob. Agents Chemother., 53(6): 2432-2443.
33. [33] Bjarnsholt, T., Van Gennip, M., Jakobsen, T.H., Christensen, L.D., Jensen, P.Ø. and Givskov, M. (2005). Garlic blocks quorum sensing and promotes rapid clearing of pulmonary Pseudomonas aeruginosa infections. Microbiology, 151(12): 3873-3880.
34. [34] Tateda, K., Comte, R., Pechere, J.C., Köhler, T., Yamaguchi, K. and Van Delden C. (2001). Azithromycin inhibits quorum sensing in Pseudomonas aeruginosa. Antimicrob. Agents Chemother., 45(6): 1930-1933.
35. [35] Di Cagno, R., Surico, R.F., Minervini, G., Rizzello, C.G., Lovino, R., Servili, M, Taticchi, A., Urbani, S. and Gobbett, M. (2011). Exploitation of sweet cherry (Prunus avium L.) puree added of stem infusion through fermentation by selected autochthonous lactic acid bacteria. Food Microbiology, 28: 900–909.
36. [36] Bursal, E., Köksal, E., Gülçin, İ., Bilsel, G. and Gören, A.C. (2013). Antioxidant activity and polyphenol content of cherry stem (Cerasus avium L.) determined by LC–MS/MS. Food Res. Int., 51(1): 66-74.
37. [37] Basanta, M.F., Plá, M.F.E., Raffo, M.D., Stortz, C.A. and Rojas, A.M. (2014). Cherry fibers isolated from harvest residues as valuable dietary fiber and functional food ingredients. J. Food Eng., 126: 149-155.
38. [38] Aires, A., Dias, C., Carvalho, R. and Saavedra, M.J. (2017). Analysis of glycosylated flavonoids extracted from sweet-cherry stems, as antibacterial agents against pathogenic Escherichia coli isolates. Acta Biochim. Pol., 64(2): 265-271.
39. [39] Oyetayo, A.M. and Bada, S.O. (2017). Phytochemical screening and antibacterial activity of Prunus avium extracts against selected human pathogens. J. Altern. Complement Med., 1-8.
40. [40] Duarte, A.P. and Silva, B.M. (2014). Nutritional and phytochemical potential of Prunus avium L. In Natural Products: Research Reviews, V.K. Gupta (ed.), Daya Publishing House, India, 185-202.
41. [41] Petković, B., Matoš, S., Gorgi, N. and Kukrić, Z. (2014). Analysis of antioxidant activity of different species of wild cherry (Prunus avium L.). Glo. Adv. Res. J. Agric. Sci ., 3(5): 128-135.
42. [42] Bastos, C., Barros, L., Dueñas, M., Calhelha, R.C., Queiroz, M.J.R., Santos-Buelga, C. and Ferreira, I.C. (2015). Chemical characterisation and bioactive properties of Prunus avium L.: The widely studied fruits and the unexplored stems. Food Chem., 173: 1045-1053.
43. [43] Ravichandiran, V., Shanmugam, K. and Princy Solomon A. (2013). Screening of SdiA inhibitors from Melia dubia seeds extracts towards the hold back of uropathogenic E. coli quorum sensing-regulated factors. Medicinal Chemistry, 9(6): 819-827.
44. [44] Abraham, S.V.P.I, Palani, A., Ramaswamy, B.R., Shunmugiah, K.P. and Arumugam, V.R. (2011). Antiquorum sensing and antibiofilm potential of Capparis spinosa. Arch. Med. Res., 42(8): 658-668.
45. [45] Sandasi, M., Leonard, C.M. and Viljoen, A.M. (2010). The in vitro antibiofilm activity of selected culinary herbs and medicinal plants against Listeria monocytogenes. Lett. Appl. Microbiol., 50(1): 30-35.
46. [46] da Silva Trentin, D., Giordani, R.B., Zimmer, K.R., da Silva, A.G., da Silva, M.V., dos Santos Correia, M.T, Baumvol I.J.R. and Macedo A.J. (2011). Potential of medicinal plants from the Brazilian semi-arid region (Caatinga) against Staphylococcus epidermidis planktonic and biofilm lifestyles. J. Ethnopharmacol., 137(1): 327-335.