Low concentrations of local honey modulate ETA expression, and quorum sensing related virulence in drug-resistant Pseudomonas aeruginosa recovered from infected burn wounds

Document Type : Original Article


Department of Biology, College of Science, Salahaddin University-Erbil, Erbil, Iraq


Objective(s): Honey’s ability to kill microorganisms and even eradication of chronic infections with drug-resistant pathogens has been documented by numerous studies. The present study is focused on the action of honey in its sub-inhibitory levels to impact on the pathogens coordinated behaviors rather than killing them.
Materials and Methods: The impact of local honey on the quorum sensing related virulence of multidrug resistant Pseudomonas aeruginosa burn isolates was investigated by detection its effect on the virulence, biofilm formation and expression of quorum sensing related and exotoxin A genes.
Results: Experiments to characterise and quantify the impact of honey on the P. aeruginosa quorum sensing networks showed that the expression of exotoxin A ( ETA), las and rhl glucons reduced by low concentrations of honey including the associated virulence factors.
Conclusion: Our results indicated that honey fights infections either by its bactericidal components which vigorously kill cells or by weakening bacterial coordination and virulence through interruption of quorum sensing system.


Main Subjects

1. Geddes A. Infection in the twenty-first century: predictions and postulates. J Antimicrob Chemoth  2000;46: 873–877.
2. Bjarnsholt T, Jensen PO, Rasmussen TB, Christophersen L, Calum H, Hentzer M, et al. Garlic blocks quorum sensing and promotes rapid clearing of pulmonary Pseudomonas aeruginosa infections. Microbiology. 2005;151:3873-3880.
3. Rasmussen TB, Bjarnsholt T, Skindersoe ME, Hentzer M, Kristoffersen P, Kote M, et al. Screening for quorum-sensing inhibitors (QSI) by use of a novel genetic system, the QSI selector. J Bacteriol. 2005;187:1799-1814.
4. Rasmussen TB, Givskov M. Quorum sensing inhibitors: a bargain of effects. Microbiology. 2006;152:895-904.
5. Rasko DA, Sperandio V. Anti-virulence strategies to combat bacteria-mediated disease. Nat Rev Drug Discov. 2010;9:117-128.
6. Bouyahya A, Dakka N, Et-Touys A, Abrini J, Bakri Y. Medicinal plant products targeting quorum sensing for combating bacterial infections. Asian Pac J Trop Med. 2017;10:729-743.
7. Hong KW, Koh CL, Sam CK, Yin WF, Chan KG. Quorum quenching revisited--from signal decays to signalling confusion. Sensors (Basel). 2012;12:4661-4696.
8. Fu B, Wu Q, Dang M, Bai D, Guo Q, Shen L, et al. Inhibition of Pseudomonas aeruginosa Biofilm Formation by Traditional Chinese Medicinal Herb Herba patriniae. BioMed Res Int. 2017;2017:1-10.
9. Smith RS, Iglewski BH. P. aeruginosa quorum-sensing systems and virulence. Curr Opin Microbiol. 2003;6(1):56-60.
10. Antunes LC, Ferreira RB, Buckner MM, Finlay BB. Quorum sensing in bacterial virulence. Microbiology. 2010;156:2271-2282.
11. Jadaun V, Prateeksha P, Singh BR, Paliya BS, Upreti DK, Rao CV, et al. Honey enhances the anti-quorum sensing activity and anti-biofilm potential of curcumin. RSC Advances. 2015;5:71060-71070.
12. Maddocks SE, Jenkin RE. Honey: a sweet solution to the growing problem of antimicrobial resistance? Future Microbiol. 2013;8:1419-1429.
13. Tille PM. Bailey & Scott’s Diagnostic Microbiology. 13 ed. China: Elsevier/Mosby; 2014.
14. Roberts AE, Maddocks SE, Cooper RA. Manuka honey is bactericidal against Pseudomonas aeruginosa and results in differential expression of oprF and algD. Microbiology. 2012;158:3005-3013.
15. Issac ASV, Palani A, Ramaswamy BR, Shunmugiah KP, Arumugam VR. Antiquorum sensing and antibiofilm potential of Capparis spinosa. Arch Med Res. 2011;42:658-668.
16. Chu W, Zhou S, Jiang Y, Zhu W, Zhuang X, Fu J. Effect of traditional Chinese herbal medicine with antiquorum sensing activity on Pseudomonas aeruginosa. Evid Based Complement Alternat Med. 2013;2013:1-7.
17. Kessler E, Safrin M, Olson JC, Ohman DE. Secreted LasA of Pseudomonas aeruginosa Is a Staphylolytic Protease. J Biol Chem. 1993; 268: 7503-7508.
18. Li H, Li X, Wang Z, Fu Y, Ai Q, Dong Y, et al. Autoinducer-2 regulates Pseudomonas aeruginosa PAO1 biofilm formation and virulence production in a dose-dependent manner. BMC Microbiol. 2015;15:192.
19. Deziel E, Comeau Y, Villemur R. Initiation of biofilm formation by Pseudomonas aeruginosa 57RP correlates with emergence of hyperpiliated and highly adherent phenotypic variants deficient in swimming, swarming, and twitching motilities. J Bacteriol. 2001;183:1195-1204.
20. Limban C, Luminita Marutescu, Chifiriuc MC. Synthesis, Spectroscopic properties and antipathogenic activity of new thiourea derivatives. Molecules 2011;16: 7593-7607.
21. Al-Shabib NA, Husain FM, Ahmed F, Khan RA, Ahmad I, Alsharaeh E, et al. Biogenic synthesis of Zinc oxide nanostructures from Nigella sativa seed: Prospective role as food packaging material inhibiting broad-spectrum quorum sensing and biofilm. Sci Rep. 2016;6:36761.
22. Schaber JA, Carty NL, McDonald NA, Graham ED, Cheluvappa R, Griswold JA, et al. Analysis of quorum sensing-deficient clinical isolates of Pseudomonas aeruginosa. J Med Microbiol. 2004;53:841-853.
23. Cotar A, DINU S, Chifiriuc M-C, Banu O, Iordache C, Larion C, et al. Screening of molecular markers of quorum sensing in Pseudomonas aeruginosa strains isolated from clinical infections. Roum Biotech Lett. 2008;13(3):3765-3771.
24. Qin X, Emerson J, Stapp J, Stapp L, Abe P, Burns JL. Use of real-time PCR with multiple targets to identify Pseudomonas aeruginosa and other nonfermenting Gram-negative bacilli from patients with cystic fibrosis. J Clin Microbiol. 2003;41:4312-4317.
25. Song KP, Chan TK, Ji ZL, Wong SW. Rapid identification of Pseudomonas aeruginosa from ocular isolates by PCR using exotoxin A-specific primers. Mol Cell Probes. 2000;14:199-204.
26. Deziel E, pine FoL, Sylvain Milot, Jianxin He, Michael N. Mindrinos, Ronald G. Tompkins, et al. Analysis of Pseudomonas aeruginosa 4-hydroxy-2alkylquinolines (HAQs) reveals a role for 4-hydroxy-2heptylquinoline in cell-to-cell communication Proc Natl Acad Sci USA. 2004;101:1339-1344.
27. Hazan R, He J, Xiao G, Dekimpe V, Apidianakis Y, Lesic B, et al. Homeostatic interplay between bacterial cell-cell signaling and iron in virulence. PLoS Pathog. 2010;6:1-14.
28. Defoirdt T, Brackman G, Coenye T. Quorum sensing inhibitors: how strong is the evidence? Trends Microbiol. 2013;21:619-624.
29. Wang M, Schaefer AL, Dandekar AA, Greenberg EP. Quorum sensing and policing of Pseudomonas aeruginosa social cheaters. Proceedings of the National Academy of Sciences of the United States of America. 2015;112(7):2187-2191.
30. Yang Q, Scheie AA, Benneche T, Defoirdt T. Specific quorum sensing-disrupting activity (A QSI) of thiophenones and their therapeutic potential. Sci Rep. 2015;5:18033.
31. Prateeksha, Singh BR, Shoeb M, Sharma S, Naqvi AH, Gupta VK, et al. Scaffold of selenium nanovectors and honey phytochemicals for inhibition of pseudomonas aeruginosa quorum sensing and biofilm formation. Front Cell Infect Microbiol. 2017;7:93.
32. Camplin AL, Maddocks SE. Manuka honey treatment of biofilms of Pseudomonas aeruginosa results in the emergence of isolates with increased honey resistance. Ann Clin Microbiol Antimicrob. 2014;13:19.
33. Eteraf-Oskouei T, Najafi M. Traditional and modern uses of natural honey in human diseases: A Review. Iran J Basic Med Sci. 2013;16:731-742.
34. Cooper RA, Halas E, Molan PC. The efficacy of honey in inhibiting strains of Pseudomonas aeruginosa from infected burns. J Burn Care Rehabil. 2002;23:366-370.
35. Lee JH, Park JH, Kim JA, Neupane GP, Cho MH, Lee CS, et al. Low concentrations of honey reduce biofilm formation, quorum sensing, and virulence in Escherichia coli O157:H7. Biofouling. 2011;27:1095-1104.
36. Wang R, Starkey M, Hazan R, Rahme LG. Honey’s ability to counter bacterial infections arises from both bactericidal compounds and QS inhibition. Front Microbiol. 2012;3:144.
37. Hentzer M, Hong Wu, Jens Bo Andersen, Kathrin Riedel, Thomas B.Rasmussen, Niels Bagge, et al. Attenuation of Pseudomonas aeruginosa virulence by quorum sensing inhibitors. EMBO J. 2003;22:3803-3815.
38. Kharazmi A. Interactions of Pseudomonas aeruginosa proteases with the cells of the immune system. Antibiot Chemother 1989;42:42-49.
39. Fothergill JL, Panagea S, Hart CA, Walshaw MJ, Pitt TL, Winstanley C. Widespread pyocyanin over-production among isolates of a cystic fibrosis epidemic strain. BMC Microbiol. 2007;7:1-10.
40. Pratt LA, Kolter R. Genetic analysis of Escherichia coli biofilm formation: roles of flagella, motility, chemotaxis and type I pili. Mol Microbiol. 1998;30:285-293.
41. Hoiby N, Bjarnsholt T, Givskov M, Molin S, Ciofu O. Antibiotic resistance of bacterial biofilms. Int J Antimicrob Agents. 2010;35:322-332.
42. Potera C. Forging a Link Between Biofilms and Disease. Science 1999;283:1837–1839.
43. Costerton JW, Philip S. Stewart, Greenberg EP. Bacterial biofilms: A common cause of persistent infections. Science     1999;284:1318-1322.
44. Lee K, Yoon SS. Pseudomonas aeruginosa Biofilm, a programmed bacterial life for fitness. J Microbiol Biotechnol. 2017;27:1053-1064.
45. Caiazza NC, Merritt JH, Brothers KM, O’Toole GA. Inverse regulation of biofilm formation and swarming motility by Pseudomonas aeruginosa PA14. J Bacteriol. 2007;189:3603-3612.
46. Patriquin GM, Banin E, Gilmour C, Tuchman R, Greenberg EP, Poole K. Influence of quorum sensing and iron on twitching motility and biofilm formation in Pseudomonas aeruginosa. J Bacteriol. 2008;190:662-671.
47. Taormina PJ, Brendan A. N., Beuchat LR. Inhibitory activity of honey against foodborne pathogens as influenced by the presence of hydrogen peroxide and level of antioxidant power. International Journal of Food Microbiology 2001;69  217–225.
48. Kwakman PH, te Velde AA, de Boer L, Speijer D, Vandenbroucke-Grauls CM, Zaat SA. How honey kills bacteria. FASEB J. 2010;24:2576-2582.
49. Balasubramanian D, Schneper L, Kumari H, Mathee K. A dynamic and intricate regulatory network determines Pseudomonas aeruginosa virulence. Nucleic Acids Res. 2013;41:1-20.
50. Liu PV. Extracellular Toxins of Pseudomonas aeruginosa. J Infect Dis 1974;130:S94-S99.
51. Bjorn MJ, Michael L. Vasil, Jerald C. Sadoff, IglewskiI BH. Incidence of exotoxin production by Pseudomonas species. Infect Immun. 1977;16: 362-366.
52. Hessler JL, Kreitman RJ. An early step in Pseudomonas exotoxin action is removal of the terminal lysine residue, which allows binding to the KDEL receptor. Biochemistry 1997;36:14577-14582.