Vanilla modulates the activity of antibiotics and inhibits efflux pumps in drug-resistant Pseudomonas aeruginosa


Vanilla, a popular flavour extracted from Vanilla planifolia pods was tested for its antibiotic modulatory activity against extensively drug-resistant (XDR) Pseudomonas aeruginosa clinical isolates. Cured vanilla pod extract (VPE) was found non-bactericidal even at high doses (> 2000 µg/mL), however, it modulated the activity of several antibiotics at a sub-inhibitory concentration of 500 µg/mL. This modulation activity of VPE was observed for last line antibiotic options such as meropenem and tigecycline, as well as commonly used antibiotics like ciprofloxacin, levofloxacin and chloramphenicol. Further, it was observed that VPE inhibited the activity of efflux pumps in XDR P. aeruginosa clinical isolates. GC-MS spectral analysis revealed the dominance of vanillin, furfuran and some short-chain fatty acids in VPE. Therefore, further studies on the constituent ingredients in VPE are recommended to identify the active compounds and use them as antibiotic modulators and efflux pump inhibitors.

Graphical abstract

Vanilla extract inhibits dominant efflux pump in Pseudomonas aeruginosa and modulates the activity of last line and commonly used antibiotics

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Vanilla pod extract


Pseudomonas aeruginosa




Extensively drug-resistant


Ethidium bromide


Extended spectrum β-lactamases




Klebsiella pneumoniae Carbapenemases


Minimum inhibitory concentration


  1. Anacarso I, Quartieri A, De Leo R, Pulvirenti A (2018) Evaluation of the antimicrobial activity of a blend of monoglycerides against Escherichia coli and Enterococci with multiple drug resistance. Arch Microbiol 200:85–89.

    CAS  Article  PubMed  Google Scholar 

  2. Arya SS, Sharma MM, Das RK, Rookes J, Cahill D, Lenka SK (2019) Vanillin mediated green synthesis and application of gold nanoparticles for reversal of antimicrobial resistance in Pseudomonas aeruginosa clinical isolates. Heliyon 5:e02021.

    Article  PubMed  PubMed Central  Google Scholar 

  3. Betoni JEC, Mantovani RP, Barbosa LN, Di Stasi LC, Fernandes Junior A (2006) Synergism between plant extract and antimicrobial drugs used on Staphylococcus aureus diseases. Mem Inst Oswaldo Cruz 101:387–390.

    Article  PubMed  Google Scholar 

  4. Bezerra DP, Soares AKN, de Sousa DP (2016) Overview of the role of vanillin on redox status and cancer development. Oxid Med Cell Long 2016;2016:9734816.

  5. Bezerra CF, Camilo CJ, do Nascimento Silva MK, de Freitas TS, Ribeiro-Filho J, Coutinho HDM (2017) Vanillin selectively modulates the action of antibiotics against resistant bacteria. Microb Pathog 113:265–268.

    CAS  Article  PubMed  Google Scholar 

  6. Breidenstein EB, de la Fuente-Núñez C, Hancock RE (2011) Pseudomonas aeruginosa: all roads lead to resistance. Trends Microbiol 19:419–426.

    CAS  Article  PubMed  Google Scholar 

  7. Cabot G, Zamorano L, Moyà B, Juan C, Navas A, Blázquez J, Oliver A (2016) Evolution of Pseudomonas aeruginosa antimicrobial resistance and fitness under low and high mutation supply rates. Antimicrob Agents Chemother 02676 – 02615.

  8. CDC (2014) Antibiotic resistance threats in the United States, 2013. CDC, Atlanta. 2013

  9. Choi WH, Jiang M (2014) Evaluation of antibacterial activity of hexanedioic acid isolated from Hermetia illucens larvae. J Appl Biomed 1;12(3):179 – 89.

  10. Choo J, Rukayadi Y, Hwang JK (2006) Inhibition of bacterial quorum sensing by vanilla extract. Lett Appl Microbiol 42:637–641.

    CAS  Article  PubMed  Google Scholar 

  11. Christena LR, Subramaniam S, Vidhyalakshmi M, Mahadevan V, Sivasubramanian A, Nagarajan S (2015) Dual role of pinostrobin-a flavonoid nutraceutical as an efflux pump inhibitor and antibiofilm agent to mitigate food borne pathogens. RSC Adv 5:61881–61887.

    CAS  Article  Google Scholar 

  12. CLSI (2017) Performance Standards for Antimicrobial Susceptibility Testing; 27th ed. CLSI supplement M100. Clinical and Laboratory Standards Institute, Wayne

  13. Dreier J, Ruggerone P (2015) Interaction of antibacterial compounds with RND efflux pumps in Pseudomonas aeruginosa. Front Microbiol 6:660.

    Article  PubMed  PubMed Central  Google Scholar 

  14. Dunne WM, Hardin DJ (2005) Use of several inducer and substrate antibiotic combinations in a disk approximation assay format to screen for AmpC induction in patient isolates of Pseudomonas aeruginosa, Enterobacter spp., Citrobacter spp., and Serratia spp. J Clin Microbiol 43:5945–5949.

  15. Dzotam JK, Kuete V (2017) Antibacterial and antibiotic-modifying activity of methanol extracts from six Cameroonian food plants against multidrug-resistant enteric bacteria. BioMed Res Int 2017;2017:1583510.

  16. Fankam AG, Kuiate J-R, Kuete V (2017) Antibacterial and antibiotic resistance modulatory activities of leaves and bark extracts of Recinodindron heudelotii (Euphorbiaceae) against multidrug-resistant Gram-negative bacteria. BMC Complem Altern Med 17:168.

    CAS  Article  Google Scholar 

  17. Fitzgerald DJ, Stratford M, Gasson MJ, Ueckert J, Bos A, Narbad A (2004) Mode of antimicrobial action of vanillin against Escherichia coli, Lactobacillus plantarum and Listeria innocua. J Appl Microbiol 97(1):104–113.

    CAS  Article  PubMed  Google Scholar 

  18. Gallage NJ, Møller BL (2018) Vanilla: The most popular flavour. In: Biotechnology of Natural Products. Springer, Berlin, pp 3–24.

  19. Garvey MI, Rahman MM, Gibbons S, Piddock LJ (2011) Medicinal plant extracts with efflux inhibitory activity against Gram-negative bacteria. Int J Antimicrob Agents 37:145–151.

    CAS  Article  PubMed  Google Scholar 

  20. Ginovyan M, Trchounian A (2019) Novel approach to combat antibiotic resistance: evaluation of some Armenian herb crude extracts for their antibiotic modulatory and antiviral properties. J Appl Microbiol 127(2):472–480.

    CAS  Article  PubMed  Google Scholar 

  21. Giske C, Martinez-Martinez L, Cantón R, Stefani S (2018) EUCAST guidelines for detection of resistance mechanisms and specific resistances of clinical and/or epidemiological importance, pp 4–12. Accessed 15 Sept 2019

  22. González-Bello C (2017) Antibiotic adjuvants - a strategy to unlock bacterial resistance to antibiotics. Bioorg Med Chem Lett 27:4221–4228.

    CAS  Article  PubMed  Google Scholar 

  23. Grassme H et al (2003) Host defense against Pseudomonas aeruginosa requires ceramide-rich membrane rafts. Nat Med 9:322.

    CAS  Article  PubMed  Google Scholar 

  24. Jadhav D, Rekha B, Gogate PR, Rathod VK (2009) Extraction of vanillin from vanilla pods: A comparison study of conventional soxhlet and ultrasound assisted extraction. J Food Eng 93:421–426.

    Article  Google Scholar 

  25. Jantrapanukorn B, Pongpraritt S, Powthong P, Pheungphu T (2017) The study of antibacterial activity in enteric pathogens of roselle (Hibiscus sabdariffa Linn.) by Broth micro-dilution method. J Appl Pharm Sci 7(5):119–122.

    CAS  Article  Google Scholar 

  26. Jones CA, Davis JS, Looke D (2017) Death from an untreatable infection may signal the start of the post-antibiotic era. Med J Aust 206:292e293.

    Article  Google Scholar 

  27. Kim T, Silva J, Jung Y (2009) Antibacterial activity of fresh and processed red muscadine juice and the role of their polar compounds on Escherichia coli O157: H7. J Appl Microbiol 107:533–539.

    CAS  Article  PubMed  Google Scholar 

  28. Kovac J, Gavaric N, Bucar F, Mozina SS (2014) Antimicrobial and resistance modulatory activity of Alpinia katsumadai seed phenolic extract, essential oil and post-distillation extract. Food Technol Biotechnol 52:248.

  29. Lee K, Lim Y, Yong D, Yum J, Chong Y (2003) Evaluation of the Hodge test and the imipenem-EDTA double-disk synergy test for differentiating metallo-β-lactamase-producing isolates of Pseudomonas spp. and Acinetobacter spp. J Clin Microbiol 41:4623–4629.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  30. Levy SB (2002) The antibiotic paradox: how the misuse of antibiotics destroys their curative power, 2nd edn. Int Microbiol 5:155–156.

  31. Linares JF, López JA, Camafeita E, Albar JP, Rojo F, Martínez JL (2005) Overexpression of the multidrug efflux pumps MexCD-OprJ and MexEF-OprN is associated with a reduction of type III secretion in Pseudomonas aeruginosa. J Bacteriol 187:1384–1391.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  32. Lu WJ, Lin HJ, Hsu PH, Lai M, Chiu JY, Lin HT (2019) Brown and red seaweeds serve as potential efflux pump inhibitors for drug-resistant Escherichia coli. Evid Based Complement Alternat Med 2019:1836982.

    Article  PubMed  PubMed Central  Google Scholar 

  33. Magiorakos AP, Srinivasan A, Carey RB, Carmeli Y, Falagas ME, Giske CG, Harbarth S, Hindler JF, Kahlmeter G, Olsson-Liljequist B, Paterson DL (2012) Multidrug-resistant, extensively drug-resistant and pandrug-resistant bacteria: an international expert proposal for interim standard definitions for acquired resistance. Clin Microbiol Infect 18(3):268–281.

    CAS  Article  PubMed  Google Scholar 

  34. Mahamoud A, Chevalier J, Alibert-Franco S, Kern WV, Pagès J-M (2007) Antibiotic efflux pumps in Gram-negative bacteria: the inhibitor response strategy. J Antimicrob Chemother 59:1223–1229.

    CAS  Article  PubMed  Google Scholar 

  35. Malchione MD, Torres LM, Hartley DM, Koch M, Goodman JL (2019) Carbapenem and colistin resistance in Enterobacteriaceae in Southeast Asia: review and mapping of emerging and overlapping challenges. Int J Antimicrob Agents 54(4):381–399.

    CAS  Article  PubMed  Google Scholar 

  36. Martins M, McCusker MP, Viveiros M, Couto I, Fanning S, Pagès J-M, Amaral L (2013) A simple method for assessment of MDR bacteria for over-expressed efflux pumps. Open Microbiol J 7:72.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  37. Masuda N, Sakagawa E, Ohya S, Gotoh N, Tsujimoto H, Nishino T (2000) Substrate specificities of MexAB-OprM, MexCD-OprJ, and MexXY-oprM efflux pumps in Pseudomonas aeruginosa. Antimicrob Agents Chemother 44:3322–3327.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  38. Miladi H, Zmantar T, Chaabouni Y, Fedhila K, Bakhrouf A, Mahdouani K, Chaieb K (2016) Antibacterial and efflux pump inhibitors of thymol and carvacrol against food-borne pathogens. Microb Pathog 99:95–100.

    CAS  Article  PubMed  Google Scholar 

  39. Mittal A, Kumar N, Chauhan NS (2019) Curcumin Encapsulated PEGylated Nanoliposomes: A Potential Anti-Infective Therapeutic Agent. Indian J Microbiol 59:336–343.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  40. Moon K, Cha J (2020) Enhancement of antioxidant and antibacterial activities of Salvia miltiorrhiza roots fermented with Aspergillus oryzae. Foods 9(1):34.

    CAS  Article  PubMed Central  Google Scholar 

  41. Ngongang FC, Fankam AG, Mbaveng AT, Wamba BE, Nayim P, Beng VP, Kuete V (2020) Methanol Extracts from Manilkara zapota with Moderate Antibacterial Activity Displayed Strong Antibiotic-Modulating Effects against Multidrug-Resistant Phenotypes. Pharmacol 3(1):37.

    Article  Google Scholar 

  42. Olivares Pacheco J, Alvarez-Ortega C, Rico MA, Martínez JL (2017) Metabolic compensation of fitness costs is a general outcome for antibiotic-resistant Pseudomonas aeruginosa mutants overexpressing efflux pumps. MBio 8:e00500–e00517.

    Article  PubMed  PubMed Central  Google Scholar 

  43. Piddock LJ (2006) Clinically relevant chromosomally encoded multidrug resistance efflux pumps in bacteria. Clin Microbiol Rev 19:382–402.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  44. Pinheiro RE, Chaves TP, Melo ES, Ali S, Ali SW, Umer M, Gama GS, Lira DN, Souza JS, Soares MJ, Santos AS (2020) Modulatory-antibiotic activity of the essential oil from Eucalyptus citriodora against MDR bacterial strains. Cell Mol Biol (Noisy-le-Grand. France) 66(4):60–64

    Google Scholar 

  45. Poole K (2007) Efflux pumps as antimicrobial resistance mechanisms. Ann Med 39:162–176.

    CAS  Article  PubMed  Google Scholar 

  46. Pournaras S, Poulou A, Tsakris A (2010) Inhibitor-based methods for the detection of KPC carbapenemase-producing Enterobacteriaceae in clinical practice by using boronic acid compounds. J Antimicrob Chemother 65:1319–1321.

    CAS  Article  PubMed  Google Scholar 

  47. Rao M, Padyana S, Dipin K, Kumar S, Nayak B, Varela MF (2018) Antimicrobial compounds of plant origin as efflux pump inhibitors: new avenues for controlling multidrug resistant pathogens. J Antimicrob Agents 4:1–6.

    Article  Google Scholar 

  48. Seasotiya L, Dalal S (2014) Screening of Indian medicinal plants as efflux pump inhibitors of fluoroquinolones. J Pharmacogn Phytochem 3:235–241

    Google Scholar 

  49. Selvi AT, Brindha V, Vedaraman N, Kanagaraj J, Sundar VJ, Khambhaty Y, Saravanan P (2020) Eco-friendly curing of hides/skins using phyto based Citrus limon leaves paste. J Clean Prod :247:119117.

    CAS  Article  Google Scholar 

  50. Seukep AJ, Kuete V, Nahar L, Sarker SD, Guo M (2020) Plant-derived secondary metabolites as the main source of efflux pump inhibitors and methods for identification. J Pharm Anal 7:277–290.

    Article  Google Scholar 

  51. Shyamala B, Naidu MM, Sulochanamma G, Srinivas P (2007) Studies on the antioxidant activities of natural vanilla extract and its constituent compounds through in vitro models. J Agric Food Chem 55:7738–7743.

    CAS  Article  PubMed  Google Scholar 

  52. Simoes M, Bennett RN, Rosa EA (2009) Understanding antimicrobial activities of phytochemicals against multidrug resistant bacteria and biofilms. Nat Prod Rep 26:746–757.

    CAS  Article  PubMed  Google Scholar 

  53. Siriyong T, Srimanote P, Chusri S, Yingyongnarongkul B-e, Suaisom C, Tipmanee V, Voravuthikunchai SP (2017) Conessine as a novel inhibitor of multidrug efflux pump systems in Pseudomonas aeruginosa. BMC Complem Altern Med 17:405.

    CAS  Article  Google Scholar 

  54. Smith DJ et al (2016) Pseudomonas aeruginosa antibiotic resistance in A ustralian cystic fibrosis centres. Respirol 21:329–337.

    Article  Google Scholar 

  55. Torres CA, Nuñez M, Isla MI, Castro M, González AM, Zampini IC (2017) Antibacterial synergism of extracts from climbers belonging to Bignoniaceae family and commercial antibiotics against multi-resistant bacteria. J Her Med 8:24–30.

    Article  Google Scholar 

  56. Webber M, Piddock L (2003) The importance of efflux pumps in bacterial antibiotic resistance. J Antimicrob Chemother 51:9–11.

    CAS  Article  PubMed  Google Scholar 

  57. WHO (2017) WHO publishes list of bacteria for which new antibiotics are urgently needed. WHO: Geneva. Accessed 12 Aug 2019

  58. Yonezawa H et al (2012) Destructive effects of butyrate on the cell envelope of Helicobacter pylori. J Med Microbiol 61:582–589.

    CAS  Article  PubMed  Google Scholar 

  59. Zhang S, Mueller C (2012) Comparative analysis of volatiles in traditionally cured bourbon and ugandan Vanilla bean (Vanilla planifolia) extracts. J Agric Food Chem 60 (42):10433–10444.

  60. Zhang H, Zhu F, Fu Q, Zhang X, Zhu X (2019) Mechanical properties of renewable plasticizer based on ricinoleic acid for PVC. Polym Test 76:199–206.

    CAS  Article  Google Scholar 

  61. Zmantar T et al (2016) Use of juglone as antibacterial and potential efflux pump inhibitors in Staphylococcus aureus isolated from the oral cavity. Microb Pathog 101:44–49.

    CAS  Article  PubMed  Google Scholar 

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The authors extend sincere thanks to Dr.Renu Bharadwaj, Head of Department, Microbiology, B. J. Govt. Medical College, Pune – 411001, India and Golwilkar Metropolis Health Services India, Pvt. Ltd., Pune – 411004, India for providing clinical isolates. Sagar Arya is the recipient of a Deakin University (Australia) post-graduate scholarship.

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Correspondence to Sangram K. Lenka.

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Arya, S.S., Sharma, M.M., Rookes, J.E. et al. Vanilla modulates the activity of antibiotics and inhibits efflux pumps in drug-resistant Pseudomonas aeruginosa. Biologia 76, 781–791 (2021).

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  • Vanilla pod extract
  • Extensively drug-resistant
  • Pseudomonas aeruginosa
  • Antibiotic modulatory activity
  • GC-MS
  • Efflux pumps