Exploring natural antimicrobial properties: cell-free culture filtrates of Aspergillus niger and Rhizomucor pusillus
Year 2025,
Volume: 34 Issue: 2, 64 - 70
Derya Berikten
,
Sukran Gunaydın
Abstract
Fungi are rich sources of medicinal bioactive metabolites. The fungi used in this study were identified as Rhizomucor pusillus and Aspergillus niger through multilocus gene sequencing. HPLC was used to analyze the cell-free culture filtrate (CFCF) of the fungi. The most prevalent organic acid in A. niger was tartaric acid, whereas the most predominant acid in R. pusillus was succinic acid. Furthermore, chlorogenic acid was the most abundant phenolic compound in both CFCFs. The impacts of the CFCFs as antibacterial agents against Gram (+) bacteria Staphylococcus aureus, Enterococcus faecalis, methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-resistant coagulase-negative Staphylococcus sp. (MRCoNS) and Gram (-) bacteria Pseudomonas aeruginosa, Escherichia coli, a yeast Candida parapsilosis and a filamentous fungus A. niger were also investigated. CFCF of A. niger was found to exhibit significant antimicrobial activity by acting on a wider range of microorganisms than CFCF of R. pusillus. Especially, CFCFs of A. niger and R. pusillus demonstrated the highest levels of activity at a concentration of 0.125 mL/mL against MRSA and C. parapsilosis, respectively. This study offers the first comparison of organic acids and phenolics in R. pusillus and A. niger CFCFs, linking their distinct antimicrobial profiles to potential use in fungal-based antimicrobial formulations.
Ethical Statement
This study does not require any ethical clearance.
Thanks
We would like to thank Dr. Ahmet TÜLEK for HPLC analysis studies at Iğdır University.
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CLSI Guidelines. (2017). Reference Method for Broth Dilution Antifungal Susceptibility Testing of Yeasts. Clinical and Laboratory Standards Institute (CLSI) Document M27, 4th Edition.
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CLSI Guidelines. (2023). Performance Standards for Antimicrobial Susceptibility Testing. Clinical and Laboratory Standards Institute (CLSI) Document M100, 33rd Edition.
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Crozier, A., Jaganath, I. B., & Clifford, M. N. (2009). Dietary phenolics: chemistry, bioavailability and effects on health. Natural Product Reports, 26(8), 1001. https://doi.org/10.1039/b802662a
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Dibner, J. J., & Buttin, P. (2002). Use of Organic Acids as a Model to Study the Impact of Gut Microflora on Nutrition and Metabolism. Journal of Applied Poultry Research, 11(4), 453–463. https://doi.org/10.1093/japr/11.4.453
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Ferreira, F. V., & Musumeci, M. A. (2021). Trichoderma as biological control agent: scope and prospects to improve efficacy. World Journal of Microbiology and Biotechnology, 37(5), 90. https://doi.org/10.1007/s11274-021-03058-7
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Kenany, M. A., Youssef, M. M., & Gebreil, A. S. (2024). Antimicrobial activity of phytase extracted from a thermophilic fungus, Rhizomucor pusillus. Mansoura Journal of Chemistry, 66(1), 57–67. https://doi.org/10.21608/mjcc.2024.411481
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A comparison between the homocyclic aromatic metabolic pathways from plant-derived compounds by bacteria and fungi. Biotechnology Advances, 37(7), 107396. https://doi.org/10.1016/j.biotechadv.2019.05.002
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Mäkelä, M. R., Marinović, M., Nousiainen, P., Liwanag, A. J. M., Benoit, I., Sipilä, J., Hatakka, A., de Vries, R. P., & Hildén, K. S. (2015). Aromatic Metabolism of Filamentous Fungi in Relation to the Presence of Aromatic Compounds in Plant Biomass (pp. 63–137). https://doi.org/10.1016/bs.aambs.2014.12.001
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Mokhtar, F. Y., Abo-El Nasr, A., Elaasser, M. M., & Elsaba, Y. M. (2022). Bioactive Secondary Metabolites from Aspergillus fumigatus ON428521 Isolated from Wadi El Rayan, El Fayum Governorate. Egyptian Journal of Botany, 0–0. https://doi.org/10.21608/ejbo.2022.152366.2058
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Nikaido, H., & Pagès, J.-M. (2012). Broad-specificity efflux pumps and their role in multidrug resistance of Gram-negative bacteria. FEMS Microbiology Reviews, 36(2), 340–363. https://doi.org/10.1111/j.1574-6976.2011.00290.x
-
Ren, M., Wang, X., Tian, C., Li, X., Zhang, B., Song, X., & Zhang, J. (2017). Characterization of Organic Acids and Phenolic Compounds of Cereal Vinegars and Fruit Vinegars in China. Journal of Food Processing and Preservation, 41(3), e12937. https://doi.org/10.1111/jfpp.12937
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Sabado, Jamille B., David Eden, Valentino, M. J. G. (2018). Mycochemicals and bioactivities of Aspergillus niger and Rhizomucor pusillus associated with vermicast. International Journal of Biosciences (IJB), 13(03), 306–313. https://doi.org/10.12692/ijb/13.3.306-313
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Samson, R. A., Visagie, C. M., Houbraken, J., Hong, S.-B., Hubka, V., Klaassen, C. H. W., Perrone, G., Seifert, K. A., Susca, A., Tanney, J. B., Varga, J., Kocsubé, S., Szigeti, G., Yaguchi, T., & Frisvad, J. C. (2014). Phylogeny, identification and nomenclature of the genus Aspergillus. Studies in Mycology, 78(1), 141–173. https://doi.org/10.1016/j.simyco.2014.07.004
-
Santra, H. K., Dutta, R., & Banerjee, D. (2024). Antifungal activity of bio-active cell-free culture extracts and volatile organic compounds (VOCs) synthesised by endophytic fungal isolates of Garden Nasturtium. Scientific Reports, 14(1), 11228. https://doi.org/10.1038/s41598-024-60948-0
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Scaife, K., Vo, T. D., Dommels, Y., Leune, E., Albermann, K., & Pařenicová, L. (2023). In silico and in vitro safety assessment of a fungal biomass from Rhizomucor pusillus for use as a novel food ingredient. Food and Chemical Toxicology, 179, 113972. https://doi.org/10.1016/j.fct.2023.113972
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-
Sharma, M., Chadha, B. S., Kaur, M., Ghatora, S. K., & Saini, H. S. (2008). Molecular characterization of multiple xylanase producing thermophilic/thermotolerant fungi isolated from composting materials. Letters in Applied Microbiology, 46(5), 526–535. https://doi.org/10.1111/j.1472-765X.2008.02357.x
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Subhash, S., Babu, P., Vijayakumar, A., Suresh, R. A., Madhavan, A., Nair, B. G., & Pal, S. (2022). Aspergillus niger Culture Filtrate (ACF) Mediated Biocontrol of Enteric Pathogens in Wastewater. Water, 14(1), 119. https://doi.org/10.3390/w14010119
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Tel-Çayan, G., Deveci, E., & Çayan, F. (2023). Study on Phenolic and Organic Acid Compositions and Antioxidant and Enzyme Inhibition Activities of Agaricomycetes Mushroom Species from Turkey. International Journal of Medicinal Mushrooms, 25(11), 11–25. https://doi.org/10.1615/IntJMedMushrooms.2023050127
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Year 2025,
Volume: 34 Issue: 2, 64 - 70
Derya Berikten
,
Sukran Gunaydın
References
-
Agrawal, K., & Verma, P. (2021). Fungal metabolites: A recent trend and its potential biotechnological applications. In New and Future Developments in Microbial Biotechnology and Bioengineering (pp. 1–14). Elsevier. https://doi.org/10.1016/B978-0-12-821005-5.00001-6
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Aydi Ben Abdallah, R., Hassine, M., Jabnoun-Khiareddine, H., & Daami-Remadi, M. (2023). Exploration of non-phytopathogenic Aspergillus spp. isolates recovered from soil and compost as potential source of bioactive metabolites for potato Fusarium dry rot control. Brazilian Journal of Microbiology, 54(2), 1103–1113. https://doi.org/10.1007/s42770-023-00925-3
-
Baz, A. M., Elwy, E., Ahmed, W. A., & El-Sayed, H. (2024). Metabolic profiling, antimicrobial, anticancer, and in vitro and in silico immunomodulatory investigation of Aspergillus niger OR730979 isolated from the Western Desert, Egypt. International Microbiology, 27(6), 1677–1691. https://doi.org/10.1007/s10123-024-00503-z
-
Benoit, I., Navarro, D., Marnet, N., Rakotomanomana, N., Lesage-Meessen, L., Sigoillot, J.-C., Asther, M., & Asther, M. (2006). Feruloyl esterases as a tool for the release of phenolic compounds from agro-industrial by-products. Carbohydrate Research, 341(11), 1820–1827. https://doi.org/10.1016/j.carres.2006.04.020
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CLSI Guidelines. (2017). Reference Method for Broth Dilution Antifungal Susceptibility Testing of Yeasts. Clinical and Laboratory Standards Institute (CLSI) Document M27, 4th Edition.
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CLSI Guidelines. (2023). Performance Standards for Antimicrobial Susceptibility Testing. Clinical and Laboratory Standards Institute (CLSI) Document M100, 33rd Edition.
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Crozier, A., Jaganath, I. B., & Clifford, M. N. (2009). Dietary phenolics: chemistry, bioavailability and effects on health. Natural Product Reports, 26(8), 1001. https://doi.org/10.1039/b802662a
-
Dibner, J. J., & Buttin, P. (2002). Use of Organic Acids as a Model to Study the Impact of Gut Microflora on Nutrition and Metabolism. Journal of Applied Poultry Research, 11(4), 453–463. https://doi.org/10.1093/japr/11.4.453
-
El-hawary, S. S., Moawad, A. S., Bahr, H. S., Abdelmohsen, U. R., & Mohammed, R. (2020). Natural product diversity from the endophytic fungi of the genus Aspergillus. RSC Advances, 10(37), 22058–22079. https://doi.org/10.1039/D0RA04290K
-
Ferreira, F. V., & Musumeci, M. A. (2021). Trichoderma as biological control agent: scope and prospects to improve efficacy. World Journal of Microbiology and Biotechnology, 37(5), 90. https://doi.org/10.1007/s11274-021-03058-7
-
Gaurav, A., Bakht, P., Saini, M., Pandey, S., & Pathania, R. (2023). Role of bacterial efflux pumps in antibiotic resistance, virulence, and strategies to discover novel efflux pump inhibitors. Microbiology, 169(5). https://doi.org/10.1099/mic.0.001333
-
Gill, H., Sykes, E. M. E., Kumar, A., & Sorensen, J. L. (2023). Isolation of Bioactive Metabolites from Soil Derived Fungus-Aspergillus fumigatus. Microorganisms, 11(3), 590. https://doi.org/10.3390/microorganisms11030590
-
Gómez-García, M., Sol, C., de Nova, P. J. G., Puyalto, M., Mesas, L., Puente, H., Mencía-Ares, Ó., Miranda, R., Argüello, H., Rubio, P., & Carvajal, A. (2019). Antimicrobial activity of a selection of organic acids, their salts and essential oils against swine enteropathogenic bacteria. Porcine Health Management, 5(1), 32. https://doi.org/10.1186/s40813-019-0139-4
-
Halkai, K. R. (2017). Biosynthesis, Characterization and Antibacterial Efficacy of Silver Nanoparticles Derived from Endophytic Fungi against P. gingivalis. JOURNAL OF CLINICAL AND DIAGNOSTIC RESEARCH. https://doi.org/10.7860/JCDR/2017/29434.10681
-
Hameed, A., Hussain, S. A., Yang, J., Ijaz, M. U., Liu, Q., Suleria, H. A. R., & Song, Y. (2017). Antioxidants Potential of the Filamentous Fungi (Mucor circinelloides). Nutrients, 9(10), 1101. https://doi.org/10.3390/nu9101101
-
Hashem, A. H., Attia, M. S., Kandil, E. K., Fawzi, M. M., Abdelrahman, A. S., Khader, M. S., Khodaira, M. A., Emam, A. E., Goma, M. A., & Abdelaziz, A. M. (2023). Bioactive compounds and biomedical applications of endophytic fungi: a recent review. Microbial Cell Factories, 22(1), 107. https://doi.org/10.1186/s12934-023-02118-x
-
Kenany, M. A., Youssef, M. M., & Gebreil, A. S. (2024). Antimicrobial activity of phytase extracted from a thermophilic fungus, Rhizomucor pusillus. Mansoura Journal of Chemistry, 66(1), 57–67. https://doi.org/10.21608/mjcc.2024.411481
-
Kumar, N., & Goel, N. (2019). Phenolic acids: Natural versatile molecules with promising therapeutic applications. Biotechnology Reports, 24, e00370. https://doi.org/10.1016/j.btre.2019.e00370
-
Lesage-Meessen, L., Stentelaire, C., Lomascolo, A., Couteau, D., Asther, M., Moukha, S., Record, E., Sigoillot, J.-C., & Asther, M. (1999). Fungal transformation of ferulic acid from sugar beet pulp to natural vanillin. Journal of the Science of Food and Agriculture, 79(3), 487–490. https://doi.org/10.1002/(SICI)1097-0010(19990301)79:3<487::AID-JSFA273>3.0.CO;2-8
-
Lou, Z., Wang, H., Zhu, S., Ma, C., & Wang, Z. (2011). Antibacterial Activity and Mechanism of Action of Chlorogenic Acid. Journal of Food Science, 76(6). https://doi.org/10.1111/j.1750-3841.2011.02213.x
Lubbers, R. J. M., Dilokpimol, A., Visser, J., Mäkelä, M. R., Hildén, K. S., & de Vries, R. P. (2019).
-
A comparison between the homocyclic aromatic metabolic pathways from plant-derived compounds by bacteria and fungi. Biotechnology Advances, 37(7), 107396. https://doi.org/10.1016/j.biotechadv.2019.05.002
-
Mäkelä, M. R., Marinović, M., Nousiainen, P., Liwanag, A. J. M., Benoit, I., Sipilä, J., Hatakka, A., de Vries, R. P., & Hildén, K. S. (2015). Aromatic Metabolism of Filamentous Fungi in Relation to the Presence of Aromatic Compounds in Plant Biomass (pp. 63–137). https://doi.org/10.1016/bs.aambs.2014.12.001
-
Mokhtar, F. Y., Abo-El Nasr, A., Elaasser, M. M., & Elsaba, Y. M. (2022). Bioactive Secondary Metabolites from Aspergillus fumigatus ON428521 Isolated from Wadi El Rayan, El Fayum Governorate. Egyptian Journal of Botany, 0–0. https://doi.org/10.21608/ejbo.2022.152366.2058
-
Nikaido, H., & Pagès, J.-M. (2012). Broad-specificity efflux pumps and their role in multidrug resistance of Gram-negative bacteria. FEMS Microbiology Reviews, 36(2), 340–363. https://doi.org/10.1111/j.1574-6976.2011.00290.x
-
Ren, M., Wang, X., Tian, C., Li, X., Zhang, B., Song, X., & Zhang, J. (2017). Characterization of Organic Acids and Phenolic Compounds of Cereal Vinegars and Fruit Vinegars in China. Journal of Food Processing and Preservation, 41(3), e12937. https://doi.org/10.1111/jfpp.12937
-
Sabado, Jamille B., David Eden, Valentino, M. J. G. (2018). Mycochemicals and bioactivities of Aspergillus niger and Rhizomucor pusillus associated with vermicast. International Journal of Biosciences (IJB), 13(03), 306–313. https://doi.org/10.12692/ijb/13.3.306-313
-
Samson, R. A., Houbraken, J., Thrane, U., Frisvad, J. C., & Andersen, B. (2010). Food and indoor fungi. CBS-KNAW Fungal Biodiversity Centre.
-
Samson, R. A., Seifert, K. A., Kuijpers, A. F., Houbraken, J. A. M. P., & Frisvad, J. C. (2004). Phylogenetic analysis of Penicillium subgenus Penicillium using partial β-tubulin sequences. Studies in Mycology, 49(1), 175–200.
-
Samson, R. A., Visagie, C. M., Houbraken, J., Hong, S.-B., Hubka, V., Klaassen, C. H. W., Perrone, G., Seifert, K. A., Susca, A., Tanney, J. B., Varga, J., Kocsubé, S., Szigeti, G., Yaguchi, T., & Frisvad, J. C. (2014). Phylogeny, identification and nomenclature of the genus Aspergillus. Studies in Mycology, 78(1), 141–173. https://doi.org/10.1016/j.simyco.2014.07.004
-
Santra, H. K., Dutta, R., & Banerjee, D. (2024). Antifungal activity of bio-active cell-free culture extracts and volatile organic compounds (VOCs) synthesised by endophytic fungal isolates of Garden Nasturtium. Scientific Reports, 14(1), 11228. https://doi.org/10.1038/s41598-024-60948-0
-
Scaife, K., Vo, T. D., Dommels, Y., Leune, E., Albermann, K., & Pařenicová, L. (2023). In silico and in vitro safety assessment of a fungal biomass from Rhizomucor pusillus for use as a novel food ingredient. Food and Chemical Toxicology, 179, 113972. https://doi.org/10.1016/j.fct.2023.113972
-
Serra, R., Cabañes, F. J., Perrone, G., Castellá, G., Venâncio, A., Mulè, G., & Kozakiewicz, Z. (2006). Aspergillus ibericus : a new species of section Nigri isolated from grapes. Mycologia, 98(2), 295–306. https://doi.org/10.1080/15572536.2006.11832702
-
Sharma, M., Chadha, B. S., Kaur, M., Ghatora, S. K., & Saini, H. S. (2008). Molecular characterization of multiple xylanase producing thermophilic/thermotolerant fungi isolated from composting materials. Letters in Applied Microbiology, 46(5), 526–535. https://doi.org/10.1111/j.1472-765X.2008.02357.x
-
Subhash, S., Babu, P., Vijayakumar, A., Suresh, R. A., Madhavan, A., Nair, B. G., & Pal, S. (2022). Aspergillus niger Culture Filtrate (ACF) Mediated Biocontrol of Enteric Pathogens in Wastewater. Water, 14(1), 119. https://doi.org/10.3390/w14010119
-
Sułkowska-Ziaja, K., Trepa, M., Olechowska-Jarząb, A., Nowak, P., Ziaja, M., Kała, K., & Muszyńska, B. (2023). Natural Compounds of Fungal Origin with Antimicrobial Activity—Potential Cosmetics Applications. Pharmaceuticals, 16(9), 1200. https://doi.org/10.3390/ph16091200
-
Tel-Çayan, G., Deveci, E., & Çayan, F. (2023). Study on Phenolic and Organic Acid Compositions and Antioxidant and Enzyme Inhibition Activities of Agaricomycetes Mushroom Species from Turkey. International Journal of Medicinal Mushrooms, 25(11), 11–25. https://doi.org/10.1615/IntJMedMushrooms.2023050127
-
Tumilaar, S. G., Hardianto, A., Dohi, H., & Kurnia, D. (2024). A Comprehensive Review of Free Radicals, Oxidative Stress, and Antioxidants: Overview, Clinical Applications, Global Perspectives, Future Directions, and Mechanisms of Antioxidant Activity of Flavonoid Compounds. Journal of Chemistry, 2024, 1–21. https://doi.org/10.1155/2024/5594386
-
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