Mechanisms of Action of Luteolin Against Single- and Dual-Species of Escherichia coli and Enterobacter cloacae and Its Antibiofilm Activities

Abstract

Escherichia coli and Enterobacter cloacae are major foodborne pathogens and can form challenging single/mixed biofilms. A recent study demonstrated that luteolin (LUT) exhibits antibacterial activities against some pathogens; however, the mechanisms underlying the effects of LUT on planktonic and biofilm bacteria have never been fully elucidated. This study aimed to determine the antibacterial activity and its mechanism of action against E. coli and E. cloacae. Here, the antimicrobial mode of LUT was explored by evaluating alterations in both cell membrane integrity and cell morphology, and the antibiofilm activity of LUT was investigated using quantitative and qualitative assays. The results showed that minimal inhibitory concentration and minimum bactericidal concentration values of LUT against E. coli were 64 and 128 μg/mL and 128 and 256 μg/mL for E. cloacae mono- and dual-species, respectively. LUT impaired cell membrane integrity, as demonstrated by the remarkable increase in the number of membrane-damaged cells and definite variations in cell morphology. Moreover, LUT presented robust inhibitory effects on biofilm formation and the capacity to kill mono- and dual-species biofilm cells. Overall, these data show the potential benefit of using a natural antimicrobial and/or preservative in the food industry, LUT, to control mono- and mixed-species or biofilm-associated infections.

This is a preview of subscription content, access via your institution.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8

References

  1. 1.

    Alegbeleye, O. O., Singleton, I., & Sant'Ana, A. S. (2018). Sources and contamination routes of microbial pathogens to fresh produce during field cultivation: a review. Food Microbiology, 73, 177–208.

    Article  Google Scholar 

  2. 2.

    Kadariya, J., Smith, T. C., & Thapaliya, D. (2014). Staphylococcus aureus and staphylococcal food-borne disease: an ongoing challenge in public health. BioMed Research International, 2014(827965), 1–9.

    Article  Google Scholar 

  3. 3.

    Bennani, M., Badri, S., Baibai, T., Oubrim, N., Hassar, M., Cohen, N., & Amarouch, H. (2011). First detection of Shiga toxin-producing Escherichia coli in shellfish and coastal environments of Morocco. Applied Biochemistry and Biotechnology, 165(1), 290–299.

    CAS  Article  Google Scholar 

  4. 4.

    Peters, K. E., Chang, Y. C., Salem, A., Sultan, A., Doiphode, S., Ibrahim, E. E., & Mohammed, H. O. (2017). Risk of foodborne pathogens in various food products at retail in Qatar. Journal of Food Safety and Hygiene, 3, 27–33.

    Google Scholar 

  5. 5.

    Gao, X., Zhang, H., & Jiang, Q. (2019). Enterobacter cloacae associated with mass mortality in zoea of giant freshwater prawns Macrobrachium rosenbergii and control with specific chicken egg yolk immunoglobulins (IgY). Aquaculture, 501, 331–337.

    CAS  Article  Google Scholar 

  6. 6.

    Amrutha, B., Sundar, K., & Shetty, P. H. (2017). Study on E. coli and Salmonella biofilms from fresh fruits and vegetables. Journal of Food Science and Technology, 54, 1091–1097.

    CAS  Article  Google Scholar 

  7. 7.

    Zurob, E., Dennett, G., Gentil, D., Montero-Silva, F., Gerber, U., Naulin, P., Gomez, A., Fuentes, R., Lascano, S., Rodrigues da Cunha, T. H., et al. (2019). Inhibition of wild Enterobacter cloacae biofilm formation by nanostructured graphene- and hexagonal boron nitride-coated surfaces. Nanomaterials (Basel), 9, 49.

    Article  Google Scholar 

  8. 8.

    Yin, W., Wang, Y. T., Liu, L., & He, J. (2019). Biofilms: the microbial “protective clothing” in extreme environments. International Journal of Molecular Sciences, 3423, 14–20.

    Google Scholar 

  9. 9.

    Donlan, R. M. (2001). Biofilm formation: a clinically relevant microbiological process. Clinical Infectious Diseases, 33(8), 1387–1392.

    CAS  Article  Google Scholar 

  10. 10.

    Wu, H., Moser, C., Wang, H. Z., Hoiby, N., & Song, Z. J. (2015). Strategies for combating bacterial biofilm infections. International Journal of Oral Science, 7(1), 1–7.

    Article  Google Scholar 

  11. 11.

    Hoiby, N., Bjarnsholt, T., Givskov, M., Molin, S., & Ciofu, O. (2010). Antibiotic resistance of bacterial biofilms. International Journal of Antimicrobial Agents, 35(4), 322–332.

    Article  Google Scholar 

  12. 12.

    Römling, U., & Balsalobre, C. (2012). Biofilm infections, their resilience to therapy and innovative treatment strategies. Journal of Internal Medicine, 272(6), 541–561.

    Article  Google Scholar 

  13. 13.

    Ren, D., Madsen, J. S., de la Cruz-Perera, C. I., Bergmark, L., Sorensen, S. J., & Burmolle, M. (2014). High-throughput screening of multispecies biofilm formation and quantitative PCR-based assessment of individual species proportions, useful for exploring interspecific bacterial interactions. Microbial Ecology, 68(1), 146–154.

    CAS  Article  Google Scholar 

  14. 14.

    Schwering, M., Song, J., Louie, M., Turner, R. J., & Ceri, H. (2013). Multi-species biofilms defined from drinking water microorganisms provide increased protection against chlorine disinfection. Biofouling., 29(8), 917–928.

    CAS  Article  Google Scholar 

  15. 15.

    Mahato, N., Sharma, K., Koteswararao, R., Sinha, M., Baral, E., & Cho, M. H. (2019). Citrus essential oils: extraction, authentication and application in food preservation. Critical Reviews in Food Science and Nutrition, 59(4), 611–625.

    CAS  Article  Google Scholar 

  16. 16.

    Tuorkey, M. J. (2016). Molecular targets of luteolin in cancer. European Journal of Cancer Prevention, 25(1), 65–76.

    CAS  Article  Google Scholar 

  17. 17.

    Luo, Y., Chen, S., & Zhou, J. (2019). Luteolin cocrystals: characterization, evaluation of solubility, oral bioavailability and theoretical calculation. Journal of Drug Delivery Science and Technology, 50, 248–254.

    CAS  Article  Google Scholar 

  18. 18.

    Zhang, Q., Yang, J., & Wang, J. (2016). Modulatory effect of luteolin on redox homeostasis and inflammatory cytokines in a mouse model of liver cancer. Oncology Letters, 12(6), 4767–4772.

    CAS  Article  Google Scholar 

  19. 19.

    Skroza, D., Simat, V., Smole Mozina, S., Katalinic, V., Boban, N., & Generalic, M. I. (2019). Interactions of resveratrol with other phenolics and activity against food-borne pathogens. Food Science & Nutrition, 7(7), 2312–2318.

    CAS  Article  Google Scholar 

  20. 20.

    Straza, T. R. A., Cottrell, M. T., & Ducklow, H. W. (2009). Geographic and phylogenetic variation in bacterial biovolume as revealed by protein and nucleic acid staining. Applied and Environmental Microbiology, 75(12), 4028–4034.

    CAS  Article  Google Scholar 

  21. 21.

    Jegal, U., Lee, J. H., Lee, J., Jeong, H., Kim, M. J., & Kim, K. H. (2019). Ultrasound-assisted gatifloxacin delivery in mouse cornea, in vivo. Scientific Reports, 9(1), 15532.

    Article  Google Scholar 

  22. 22.

    Salgado, H. R., & Lopes, C. C. (2006). Determination of gatifloxacin in bulk and tablet preparations by high-performance liquid chromatography. Journal of AOAC International, 89(3), 642–645.

    CAS  PubMed  Google Scholar 

  23. 23.

    Sanhueza, L., Melo, R., & Montero, R. (2017). Synergistic interactions between phenolic compounds identified in grape pomace extract with antibiotics of different classes against Staphylococcus aureus and Escherichia coli. PLoS One, 12(2), e0172273.

    Article  Google Scholar 

  24. 24.

    Basile, A., Sorbo, S., Giordano, S., Ricciardi, L., Ferrara, S., Montesano, D., Castaldo Cobianchi, R., Vuotto, M. L., & Ferrara, L. (2000). Antibacterial and allelopathic activity of extract from Castanea sativa leaves. Fitoterapia, 71, S110–S116.

    CAS  Article  Google Scholar 

  25. 25.

    Stiefel, P., Schmidt-Emrich, S., & Maniura-Weber, K. (2015). Critical aspects of using bacterial cell viability assays with the fluorophores SYTO9 and propidium iodide. BMC Microbiology, 15(1), 36.

    Article  Google Scholar 

  26. 26.

    Lee, K. A., Moon, S. H., & Kim, K. T. (2010). Antimicrobial effects of various flavonoids on Escherichia coli O157:H7 cell growth and lipopolysaccharide production. Food Science and Biotechnology, 19(1), 257–261.

    CAS  Article  Google Scholar 

  27. 27.

    Khan, I., Bahuguna, A., Kumar, P., Bajpai, V. K., & Kang, S. C. (2017). Antimicrobial potential of carvacrol against uropathogenic Escherichia coli via membrane disruption, depolarization, and reactive oxygen species generation. Frontiers in Microbiology, 8, 2421.

    Article  Google Scholar 

  28. 28.

    He, M., Wu, T., & Pan, S. (2014). Antimicrobial mechanism of flavonoids against Escherichia coli ATCC 25922 by model membrane study. Applied Surface Science, 305, 515–521.

    CAS  Article  Google Scholar 

  29. 29.

    Stratakos, A. C., Linton, M., Ward, P., Campbell, M., Kelly, C., Pinkerton, L., Stef, L., Pet, I., Stef, D., Iancu, T., Theodoridou, K., Gundogdu, O., & Corcionivoschi, N. (2019). The antimicrobial effect of a commercial mixture of natural antimicrobials against Escherichia coli O157:H7. Foodborne Pathogens and Disease, 16(2), 119–129.

    CAS  Article  Google Scholar 

  30. 30.

    Liu, N. T., Nou, X., Lefcourt, A. M., Shelton, D. R., & Lo, Y. M. (2014). Dual-species biofilm formation by Escherichia coli O157:H7 and environmental bacteria isolated from fresh-cut processing facilities. International Journal of Food Microbiology, 171, 15–20.

    Article  Google Scholar 

  31. 31.

    Olszewska, M. A., Nynca, A., Bialobrzewski, I., Kocot, A. M., & Laguna, J. (2019). Assessment of the bacterial viability of chlorine- and quaternary ammonium compounds-treated Lactobacillus cells via a multi-method approach. Journal of Applied Microbiology, 126(4), 1070–1080.

    CAS  Article  Google Scholar 

  32. 32.

    Qian, W., Zhang, J., Wang, W., Wang, T., Liu, M., Yang, M., Sun, Z., Li, X., & Li, Y. (2020). Antimicrobial and antibiofilm activities of paeoniflorin against carbapenem-resistant Klebsiella pneumoniae. Journal of Applied Microbiology, 128(2), 401–413.

    CAS  Article  Google Scholar 

Download references

Funding

This work was supported by the Science and Technology Program of China Selenium Industry Research Institute (2018FXZX03-15) and the Xi’an Weiyang District Science and Technology Project [201926].

Author information

Affiliations

Authors

Corresponding author

Correspondence to Weidong Qian.

Ethics declarations

Conflict of Interest

The authors declare that they have no conflict of interest.

Additional information

Publisher’s Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Qian, W., Fu, Y., Liu, M. et al. Mechanisms of Action of Luteolin Against Single- and Dual-Species of Escherichia coli and Enterobacter cloacae and Its Antibiofilm Activities. Appl Biochem Biotechnol 193, 1397–1414 (2021). https://doi.org/10.1007/s12010-020-03330-w

Download citation

Keywords

  • Luteolin
  • Escherichia coli
  • Enterobacter cloacae
  • Cell membrane damage
  • Antibiofilm formation