Antimicrobial Efficacy of Four Thieves Vinegar Against Pneumonia-Associated Respiratory Pathogens: A Sustainable and Edible Disinfectant Approach

Abstract

The aim of this study was to assess the antimicrobial efficacy of different Four Thieves Vinegar (FTV) variants, including annual, monthly and commercial FTV, against pneumonia-associated respiratory pathogens (PARPs) isolates. The well diffusion agar method was used to test the antimicrobial activity against 23 PARPs isolates such as Staphylococcus aureus (3 isolates), Acinetobacter baumannii (9 isolates), Klebsiella pneumoniae (6 isolates), and Pseudomonas aeruginosa (5 isolates). FTV was prepared by fermenting apple cider vinegar with selected several medicinal and aromatic herbs such as sage, rosemary, cinnamon, mint and lavender and others in a 3:2 ratio of apple cider vinegar to water under controlled conditions for monthly and annual incubation periods. Samples were tested in pure (100%) and 50% diluted forms. Among the variants tested, annual FTV showed the strongest inhibitory effect. The inhibition zones ranged from 19.52 mm to 16.70 mm for K. pneumoniae and 19.65 mm to 14.71 mm for A. baumannii. In contrast, monthly FTV and Apple-V showed moderate antimicrobial activity, while Commercial FTV showed the lowest efficacy, indicating that traditional fermentation enhances antimicrobial potency. The pure (100%) FTV samples generally showed larger inhibition zones than the 50% diluted samples, confirming the concentration-dependent efficacy of FTV. The antimicrobial effects varied with fermentation time and vinegar composition, with longer fermentation times correlating with stronger inhibitory activity. FTV showed strong antimicrobial potential against PARPs, making it a natural alternative to chemical disinfectants and highlighting the value of traditional methods. Further research should optimise formulations and assess stability against other hospital-acquired multidrug-resistant pathogens.

Keywords

• Antimicrobial activity
• four thieves vinegar
• disinfectant
• plant mixed vinegar
• pneumonia-associated respiratory pathogens

References

1. Abiola, R.R., Okoro, E.K., Sokunbi, O., 2022. Lactic acid bacteria and the food industry-A comprehensive review. International Journal of Health Sciences and Research, 12(5): 128-142.
2. Akerele, O., 1993. Nature’s medicinal bounty: don’t throw it away. World Health Forum, 14(4): 390-395.
3. Anlı, R.E., Çapar, E., 2024. Vinegar, olive olive and picklepickle. In: R.E. Anlı and P. Şanlıbaba (Eds.), Fermented Foods, 1. Edn, Nobel, Turkey, pp. 195-225.
4. Atanasov, A.G., Waltenberger, B., Pferschy-Wenzig, E.M., Linder, T., Wawrosch, C., Uhrin, P., Temml, V., Wang, L., Schwaiger, S., Heiss, E.H., Rollinger, J.M., 2015. Discovery and resupply of pharmacologically active plant-derived natural products: A review. Biotechnology Advances, 33(8): 1582-1614.
5. Bangar, S.P., Suri, S., Trif, M., Ozogul, F., 2022. Organic acids production from lactic acid bacteria: A preservation approach. Food Bioscience, 46: 101615.
6. Bhattacharya, D., Nanda, P.K., Pateiro, M., Lorenzo, J.M., Dhar, P., Das, A.K., 2022. Lactic acid bacteria and bacteriocins: Novel biotechnological approach for biopreservation of meat and meat products. Microorganisms, 10(10): 2058.
7. Boncan, D.A.T., Tsang, S.S., Li, C., Lee, I.H., Lam, H.M., Chan, T.F., Hui, J.H., 2020. Terpenes and terpenoids in plants: Interactions with environment and insects. International Journal of Molecular Sciences, 21(19): 73-82.
8. Buncic, S., Nychas, G.-J., Lee, M.R.F., Koutsoumanis, K., Hébraud, M., Desvaux, M., Chorianopoulos, N., Bolton, D., Blagojevic, B., Antic, D., 2014. Microbial pathogen control in the Beef Chain: Recent research advances. Meat Science, 97(3): 288-297.

9. De Roos, J., De Vuyst, L., 2018. Acetic acid bacteria in fermented foods and beverages. Current Opinion in Biotechnology, 49: 115-119.
10. Dilimen, E., Ceyhan, T., Heperkan, Z.D., 2021. Determination of some chemical and microbiological properties of kiwi vinegar produced under different conditions. International Journal of Food Engineering Research, 7(1): 17-31.
11. Duarte, M.T., de Fátima Carrijo, K., 2014. Quantificação do teor de nitrito de sódio residual em linguiças cozidas tipo calabresa comercializadas no sul do estado do Rio De Janeiro, Brasil. Enciclopédia Biosfera, 10: 1606-1615. (In Portuguese).
12. Erhonyota, C., Edo, G.I., Onoharigho, F.O., 2023. Comparison of poison plate and agar well diffusion method determining the antifungal activity of protein fractions. Acta Ecologica Sinica, 43(4): 684-689.
13. Flores, M., Toldrá, F., 2021. Chemistry, safety, and regulatory considerations in the use of nitrite and nitrate from natural origin in meat products-invited review. Meat Science, 171: 108272.
14. Garcia, S., 2020. Pandemics and traditional plant-based remedies. A historical-botanical review in the era of COVID-19. Frontiers in Plant Science, 11: 571042.
15. Giudici, P., Lemmetti, F., Mazza, S., 2015. Sensorial properties and evaluation of balsamic vinegars. In: P. Giudici, F. Lemmetti and S. Mazza (Eds.), Balsamic Vinegars: Tradition, Technology, Trade, Springer Cham, New York, pp. 143-162.
16. Karthikeyan, G., Swamy, M.K., Viknesh, M.R., Shurya, R., Sudhakar, N., 2020. Bioactive Phytocompounds to fight against antimicrobial resistance. In: M.K. Swamy (Ed.), Plant-Derived Bioactives: Production, Properties and Therapeutic Applications, Springer, Singapore, pp. 335-381.
17. Kaveh, S., Mahoonak, A.S., Ghorbani, M., Jafari, S.M., 2022. Fenugreek Sseed (Trigonella foenum graecum) protein hydrolysate loaded in nanosized liposomes: Characteristic, storage stability, controlled release and retention of antioxidant activity. Industrial Crops and Products, 182: 114908.
18. Kelley, G., 2020. Doctor Beaky, the four thieves, and de fabulis pestis. Contemporary Legend Series 3, 10: 48-72. Kopaczyk, J.M., Warguła, J., Jelonek, T., 2020. The variability of terpenes in conifers under developmental and environmental stimuli. Environmental and Experimental Botany, 180: 104197.
19. Krapac, M., Major, N., Plavša, T., Jeromel, A., Tomaz, I., Poljuha, D., 2024. Enrichment of white wine vinegar with aromatic plants: The impact on aromatic, polyphenolic, and sensory profiles. Applied Sciences, 14(16): 6909.
20. Laukkanen-Ninios, R., Fredriksson-Ahomaa, M., Korkeala, H., 2014. Enteropathogenic Yersinia in the pork production chain: Cchallenges for control. Comprehensive Reviews in Food Science and Food Safety, 13(6): 1165-1191.
21. Ledermann-Dehnhardt, W., 2021. El abate Molina, la viruela…y también Darwin. Revista Chilena de Infectología, 38(2): 254-259. (In Spanish).
22. Li, S.Y., Chen, C., Zhang, H.Q., Guo, H.Y., Wang, H., Wang, L., Zhang, X., Hua, S.N., Yu, J., Xiao, P.G., Li, R.S., 2005. Identification of natural compounds with antiviral activities against SARS-associated coronavirus. Antiviral Research, 67(1): 18-23.
23. Mgomi, F.C., Yang, Y.R., Cheng, G., Yang, Z.Q., 2023. Lactic acid bacteria biofilms and their antimicrobial potential against pathogenic microorganisms. Biofilm, 5(1): 100118.
24. Mitjà, O., Corbacho-Monné, M., Ubals, M., Tebé, C., Peñafiel, J., Tobias, A., Ballana, E., Alemany, A., Riera-Martí, N., Pérez, C.A., Suñer, C., Laporte, P., Admella, P., Mitjà, J., Clua, M., Bertran, L., Sarquella, M., Gavilán, S., Ara, J., Argimon, J.M., Casabona, J., Cuatrecasas, G., Cañadas, P., Elizalde-Torrent, A., Fabregat, R., Farré, M., Forcada, A., Flores-Mateo, G., Muntada, E., Nadal, N., Narejos, S., Nieto, A., Prat, N., Puig, J., Quiñones, C., Reyes-Ureña, J., Ramírez-Viaplana, F., Ruiz, L., Riveira-Muñoz, E., Sierra, A., Velasco, C., Vivanco-Hidalgo, R.M., Sentís, A., G-Beiras, C., Clotet, B., Vall-Mayans, M., 2021. Hydroxychloroquine for early treatment of adults with mild coronavirus disease 2019: A randomized, controlled trial. Clinical Infectious Diseases, 73(11): 4073-4081.
25. Ninkuu, V., Zhang, L., Yan, J., Fu, Z., Yang, T., Zeng, H., 2021. Biochemistry of terpenes and recent advances in plant protection. International Journal of Molecular Sciences, 22(11): 5710.
26. Radi, M., Shadikhah, S., Sayadi, M., Kaveh, S., Amiri, S., Bagheri, F., 2023. Effect of Thymus vulgaris essential oil-loaded nanostructured lipid carriers in alginate-based edible coating on the postharvest quality of tangerine fruit. Food Bioprocess Technology, 16(1): 185-198.
27. Sakhare, P.Z., Sachindra, N.M., Yashoda, K.P., Narasimha Rao, D.N., 1999. Efficacy of intermittent decontamination treatments during processing in reducing the microbial load on the broiler chicken carcass. Food Control, 10(3): 189-194.
28. Sanwal, N., Gupta, A., Bareen, M.A., Sharma, N., Sahu, J.K., 2023. Kombucha fermentation: Recent trends in process dynamics, functional bioactivities, toxicity management, and potential applications. Food Chemistry Advances, 3: 100421.
29. Shelton, T.V., 2019. Nature’s own remedies: Chinese Medicine in progressive era America. Pacific Historical Review, 88(3): 378-409.
30. Solieri, L., Giudici, P., 2009. Vinegars of the world. In: L. Solieri and P. Giudici (Eds.), Vinegars Vinegars of the World, Springer Milano, Milan, pp. 1-16.
31. Van de Vel, E., Sampers, I., Raes, K., 2019. A review on influencing factors on the minimum inhibitory concentration of essential oils. Critical Reviews in Food Science and Nutrition, 59(3): 357-378.
32. Xu, H., Hong, J.H., Kim, D., Jin, Y.H., Pawluk, A.M., Mah, J.H., 2022. Evaluation of bioactive compounds and antioxidative activity of fermented green tea produced via one- and two-step fermentation. Antioxidants, 11(8): 1425.
33. Zapaśnik, A., Sokołowska, B., Bryła, M., 2022. Role of lactic acid bacteria in food preservation and safety. Foods, 11(9): 1283.