Research Article
BibTex RIS Cite

Precise Demonstration of Salt-Stress Induced Antifungal Activity of Origanum onites Essential Oil and Taraxacum officinale Extract Against Drug Resistant Isolates of Candida albicans and Aspergillus fumigatus Using Micro-Colony Method

Year 2025, Volume: 46 Issue: 2, 212 - 224, 30.06.2025

Esra Seyran

https://doi.org/10.17776/csj.1522316

Abstract

The rise of drug-resistant fungal pathogens has intensified the need for novel antifungal agents. Plants are a significant source, although effective concentrations in extracts are often low. Accurate in vitro assays are essential for validating these compounds. This study uses the micro-colony method, measuring hyphal growth and cell diameter under a microscope with digital imaging, to assess antifungal activity quickly and precisely. We evaluated Origanum onites essential oil and Taraxacum officinale methanol extract against drug-resistant Candida albicans and Aspergillus fumigatus strains. Yeast cell pigmentation was also assessed using image processing tools. To enhance compound penetration, mono and divalent salts (100mM KCl, NaCl, CaCl2) were added to the media. In salt-free media, Origanum onites essential oil inhibited Candida albicans (MIC: 0.3 μl/ml; MFC: 0.03 μl/ml) and Aspergillus fumigatus (MIC: 0.15 μl/ml; MFC: 0.03 μl/ml), while Taraxacum officinale extract was ineffective. Salt stress increased Origanum onites activity against Aspergillus fumigatus (MIC: 0.075 μl/ml) but had minimal impact on Candida albicans. Salt stress enabled Taraxacum officinale extract to inhibit Candida albicans (EC50: 12.71 μg/ml) and reduced its pigmentation dose-dependently without affecting toxicity against Aspergillus fumigatus. These results demonstrate that the micro-colony assay effectively evaluates plant-derived antifungal compounds and detects subtle dose-response variations in pathogenic fungi

Keywords

Antifungal drug resistance Micro-colony method Plant metabolites Salt stress

Ethical Statement

There are no conflicts of interest in this work.

Supporting Institution

Sivas Cumhuriyet Üniversitesi

Thanks

The author thanks to Assoc. Prof. Dr. Serap Çetinkaya her support and critical review of this study.

References

  • [1] Warrell D.A., Cox T.M., Weatherall D., Benz Jr E.J., Firth J.D, Oxford textbook of Medicine, Oxford university press., (2003).
  • [2] Chandrasekar P.H., Manavathu E.K., Antifungal resistance: aspergillus. In: Antimicrobial Drug Resistance: Clinical and Epidemiological Aspects, Totowa, NJ: Humana Press., (2009) 953–65.
  • [3] Akins R.A., Sobel J.D., Antifungal targets, mechanisms of action, and resistance in Candida albicans, In: Antimicrobial Drug Resistance: Mechanisms of Drug Resistance., (2009) 347–407.
  • [4] Vicente M.F., Basilio A., Cabello A., Peláez F., Microbial natural products as a source of antifungals, Clin. Microbiol. Infect., 9(1) (2003)15–32.
  • [5] Evans W.C., Trease and Evans Pharmacognosy, 15th ed. W.B. Saunders., (2002) 585.
  • [6] European Pharmacopoeia Commission, European Directorate for the Quality of Medicines & Healthcare, European pharmacopoeia, Council of Europe., 1 (2010).
  • [7] Ahmad A., Khan A., Akhtar F., Yousuf S., Xess I., Khan L.A., Fungicidal activity of thymol and carvacrol by disrupting ergosterol biosynthesis and membrane integrity against Candida, Eur. J. Clin. Microbiol. Infect. Dis ., 30 (2011) 41–50.
  • [8] Kim J.H., Campbell B.C., Mahoney N., Chan K.L., May G.S., Targeting antioxidative signal transduction and stress response system: control of pathogenic Aspergillus with phenolics that inhibit mitochondrial function, J. Appl. Microbiol., 101 (2006) 181–9.
  • [9] Koroishi A.M., Foss S.R., Cortez D.A., Ueda-Nakamura T., Nakamura C.V., Dias Filho B.P., In vitro antifungal activity of extracts and neolignans from Piper regnellii against dermatophytes, J. Ethnopharm., 117 (2008) 270–7.
  • [10] Seyran M., Development of rapid in vitro assays and current status of fungicide sensitivity in the pecan scab pathogen Fusicladium effusum [Doctoral dissertation], University of Georgia., (2008).
  • [11] Qui J., Detection and mechanism of resistance to sterol demethylation inhibiting fungicides in Cercospora arachidicola [MSc Thesis], University of Georgia., Department of Plant Pathology., (2010).
  • [12] Seyran M., Brenneman T.B., Stevenson K.L., In vitro toxicity of alternative oxidase inhibitors salicylhydroxamic acid and propyl gallate on Fusicladium effusum, J. Pest Sci., 83 (2010) 421–7.
  • [13] Hosono K., Effect of nystatin on the release of glycerol from salt-stressed cells of the salt-tolerant yeast Zygosaccharomyces rouxii, Arch. Microbiol., 173 (2000) 284–7.
  • [14] Farwick M., Siewe R.M., Krämer R., Glycine betaine uptake after hyperosmotic shift in Corynebacterium glutamicum, J. Bacteriol., 177 (1995) 4690–5.
  • [15] Kulikova N.A., Perminova I.V., Badun G.A., Chernysheva M.G., Koroleva O.V., Tsvetkova E.A., Estimation of uptake of humic substances from different sources by Escherichia coli cells under optimum and salt stress conditions by use of tritium-labeled humic materials, Appl. Environ. Microbiol., 76 (2010) 6223–30.
  • [16] Woo J.H., Kamei Y., Antifungal mechanism of an anti-Pythium protein (SAP) from the marine bacterium Streptomyces sp. strain AP77 is specific for Pythium porphyrae, a causative agent of red rot disease in Porphyra spp, Appl. Microbiol. Biotech., 62 (2003) 407–13.
  • [17] Zore G.B., Thakre A.D., Rathod V., Karuppayil S.M., Evaluation of anti‐Candida potential of geranium oil constituents against clinical isolates of Candida albicans differentially sensitive to fluconazole: inhibition of growth, dimorphism and sensitization, Mycoses., 54 (2011) 99-109.
  • [18] Katiyar S.K., Gordon V.R., McLaughlin G.L., Edlind T.D., Antiprotozoal activities of benzimidazoles and correlations with beta-tubulin sequence, Antimicrob. Agents Chemother., 38 (1994) 2086–90.
  • [19] Maresova L., Muend S., Zhang Y.Q., Sychrova H., Rao R., Membrane hyperpolarization drives cation influx and fungicidal activity of amiodarone, J. Biol. Chem., 284 (2009) 2795–802.
  • [20] Hammer Ø., Harper D.A., Past: paleontological statistics software package for education and data analysis, Palaeontol. Electron., 4 (2001)1.
  • [21] Jacobson E.S., Hove E., Emery H.S., Antioxidant function of melanin in black fungi, Infect. Immun., 63 (1995) 4944–5.
  • [22] Ben Arfa A., Combes S., Preziosi‐Belloy L., Gontard N., Chalier P., Antimicrobial activity of carvacrol related to its chemical structure, Lett. Appl. Microbiol., 43 (2006) 149–54.
  • [23] Rao A., Zhang Y., Muend S., Rao R., Mechanism of antifungal activity of terpenoid phenols resembles calcium stress and inhibition of the TOR pathway, Antimicrob. Agents Chemother., 54 (2010) 5062–9.
  • [24] Bakkali F., Averbeck S., Averbeck D., Idaomar M., Biological effects of essential oils–a review, Food Chem. Toxicol., 46 (2008) 446–75.
  • [25] Seyran M., Brenneman T.B., Stevenson K.L., A rapid method to monitor fungicide sensitivity in the pecan scab pathogen, Fusicladium effusum, Crop Prot., 29 (2010)1257–63.
  • [26] National Committee for Clinical Laboratory Standards (NCCLS), Performance standards for antimicrobial susceptibility testing: eight international supplement M100-S14, Wayne, PA: NCCLS., (2004).
  • [27] Hosono K., Effect of nystatin on the release of glycerol from salt-stressed cells of the salt-tolerant yeast Zygosaccharomyces rouxii, Arch. Microbiol. 173 (2000) 284–7.

Year 2025, Volume: 46 Issue: 2, 212 - 224, 30.06.2025

Esra Seyran

https://doi.org/10.17776/csj.1522316

Abstract

References

  • [1] Warrell D.A., Cox T.M., Weatherall D., Benz Jr E.J., Firth J.D, Oxford textbook of Medicine, Oxford university press., (2003).
  • [2] Chandrasekar P.H., Manavathu E.K., Antifungal resistance: aspergillus. In: Antimicrobial Drug Resistance: Clinical and Epidemiological Aspects, Totowa, NJ: Humana Press., (2009) 953–65.
  • [3] Akins R.A., Sobel J.D., Antifungal targets, mechanisms of action, and resistance in Candida albicans, In: Antimicrobial Drug Resistance: Mechanisms of Drug Resistance., (2009) 347–407.
  • [4] Vicente M.F., Basilio A., Cabello A., Peláez F., Microbial natural products as a source of antifungals, Clin. Microbiol. Infect., 9(1) (2003)15–32.
  • [5] Evans W.C., Trease and Evans Pharmacognosy, 15th ed. W.B. Saunders., (2002) 585.
  • [6] European Pharmacopoeia Commission, European Directorate for the Quality of Medicines & Healthcare, European pharmacopoeia, Council of Europe., 1 (2010).
  • [7] Ahmad A., Khan A., Akhtar F., Yousuf S., Xess I., Khan L.A., Fungicidal activity of thymol and carvacrol by disrupting ergosterol biosynthesis and membrane integrity against Candida, Eur. J. Clin. Microbiol. Infect. Dis ., 30 (2011) 41–50.
  • [8] Kim J.H., Campbell B.C., Mahoney N., Chan K.L., May G.S., Targeting antioxidative signal transduction and stress response system: control of pathogenic Aspergillus with phenolics that inhibit mitochondrial function, J. Appl. Microbiol., 101 (2006) 181–9.
  • [9] Koroishi A.M., Foss S.R., Cortez D.A., Ueda-Nakamura T., Nakamura C.V., Dias Filho B.P., In vitro antifungal activity of extracts and neolignans from Piper regnellii against dermatophytes, J. Ethnopharm., 117 (2008) 270–7.
  • [10] Seyran M., Development of rapid in vitro assays and current status of fungicide sensitivity in the pecan scab pathogen Fusicladium effusum [Doctoral dissertation], University of Georgia., (2008).
  • [11] Qui J., Detection and mechanism of resistance to sterol demethylation inhibiting fungicides in Cercospora arachidicola [MSc Thesis], University of Georgia., Department of Plant Pathology., (2010).
  • [12] Seyran M., Brenneman T.B., Stevenson K.L., In vitro toxicity of alternative oxidase inhibitors salicylhydroxamic acid and propyl gallate on Fusicladium effusum, J. Pest Sci., 83 (2010) 421–7.
  • [13] Hosono K., Effect of nystatin on the release of glycerol from salt-stressed cells of the salt-tolerant yeast Zygosaccharomyces rouxii, Arch. Microbiol., 173 (2000) 284–7.
  • [14] Farwick M., Siewe R.M., Krämer R., Glycine betaine uptake after hyperosmotic shift in Corynebacterium glutamicum, J. Bacteriol., 177 (1995) 4690–5.
  • [15] Kulikova N.A., Perminova I.V., Badun G.A., Chernysheva M.G., Koroleva O.V., Tsvetkova E.A., Estimation of uptake of humic substances from different sources by Escherichia coli cells under optimum and salt stress conditions by use of tritium-labeled humic materials, Appl. Environ. Microbiol., 76 (2010) 6223–30.
  • [16] Woo J.H., Kamei Y., Antifungal mechanism of an anti-Pythium protein (SAP) from the marine bacterium Streptomyces sp. strain AP77 is specific for Pythium porphyrae, a causative agent of red rot disease in Porphyra spp, Appl. Microbiol. Biotech., 62 (2003) 407–13.
  • [17] Zore G.B., Thakre A.D., Rathod V., Karuppayil S.M., Evaluation of anti‐Candida potential of geranium oil constituents against clinical isolates of Candida albicans differentially sensitive to fluconazole: inhibition of growth, dimorphism and sensitization, Mycoses., 54 (2011) 99-109.
  • [18] Katiyar S.K., Gordon V.R., McLaughlin G.L., Edlind T.D., Antiprotozoal activities of benzimidazoles and correlations with beta-tubulin sequence, Antimicrob. Agents Chemother., 38 (1994) 2086–90.
  • [19] Maresova L., Muend S., Zhang Y.Q., Sychrova H., Rao R., Membrane hyperpolarization drives cation influx and fungicidal activity of amiodarone, J. Biol. Chem., 284 (2009) 2795–802.
  • [20] Hammer Ø., Harper D.A., Past: paleontological statistics software package for education and data analysis, Palaeontol. Electron., 4 (2001)1.
  • [21] Jacobson E.S., Hove E., Emery H.S., Antioxidant function of melanin in black fungi, Infect. Immun., 63 (1995) 4944–5.
  • [22] Ben Arfa A., Combes S., Preziosi‐Belloy L., Gontard N., Chalier P., Antimicrobial activity of carvacrol related to its chemical structure, Lett. Appl. Microbiol., 43 (2006) 149–54.
  • [23] Rao A., Zhang Y., Muend S., Rao R., Mechanism of antifungal activity of terpenoid phenols resembles calcium stress and inhibition of the TOR pathway, Antimicrob. Agents Chemother., 54 (2010) 5062–9.
  • [24] Bakkali F., Averbeck S., Averbeck D., Idaomar M., Biological effects of essential oils–a review, Food Chem. Toxicol., 46 (2008) 446–75.
  • [25] Seyran M., Brenneman T.B., Stevenson K.L., A rapid method to monitor fungicide sensitivity in the pecan scab pathogen, Fusicladium effusum, Crop Prot., 29 (2010)1257–63.
  • [26] National Committee for Clinical Laboratory Standards (NCCLS), Performance standards for antimicrobial susceptibility testing: eight international supplement M100-S14, Wayne, PA: NCCLS., (2004).
  • [27] Hosono K., Effect of nystatin on the release of glycerol from salt-stressed cells of the salt-tolerant yeast Zygosaccharomyces rouxii, Arch. Microbiol. 173 (2000) 284–7.
There are 27 citations in total.

Details

Primary Language English
Subjects Plant Biochemistry, Mycology
Journal Section Natural Sciences
Authors

Esra Seyran 0000-0002-0384-4300

Publication Date June 30, 2025
Submission Date July 25, 2024
Acceptance Date April 2, 2025
Published in Issue Year 2025Volume: 46 Issue: 2

Cite

APA Seyran, E. (2025). Precise Demonstration of Salt-Stress Induced Antifungal Activity of Origanum onites Essential Oil and Taraxacum officinale Extract Against Drug Resistant Isolates of Candida albicans and Aspergillus fumigatus Using Micro-Colony Method. Cumhuriyet Science Journal, 46(2), 212-224. https://doi.org/10.17776/csj.1522316