Microbial pathogenesis · 2025
Designing of eucalyptol nanoemulsion drug delivery system loaded with amphotericin B for Candida albicans biofilm control: in vitro and in vivo assessment using the Galleria mellonella model.
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Keymaram Mahyar, Lotfali Ensieh, Mousazadeh Marziyeh, Nikkhah Maryam, Raiesi Omid, Yadegari Mohammad Hossein
Аннотация
OBJECTIVE: (s): Candida species can form biofilm in various clinical settings, making them challenging to treat due to their resistance to antifungal drugs. The present study aimed to evaluate the effects of eucalyptol/amphotericin B nanoemulsion (NEA) on Candida albicans (C. albicans) biofilm.
MATERIALS AND METHODS: In this study, NEA was synthesized and its capacity to combat biofilms generated by four clinical isolates and a reference strain of C. albicans was investigated according to CLSI M27-A3 guidelines. The NEA was prepared using eucalyptol oil, Tween 80, and ethanol, and characterized by measuring particle size, Z-potential, and encapsulation efficiency (EE %). The cytotoxicity study was performed on myeloblast KG1 cells and human RBCs. The gene assessment and molecular docking were conducted by Real-Time PCR and AutoDock Vina software, respectively. The in vivo stage was conducted on Galleria mellonella (G. mellonella) larvae, and ultimately, histopathological analysis was executed using H and E Staining.
RESULTS: Based on the results obtained, the NEA exhibited EE of 80 % for Amphotericin B (AmB), displaying an average size of 39 nm and zeta potential of -2 mV. Minimum inhibitory concentration (MIC) and minimum biofilm inhibitory concentration (MBIC) revealed more efficiency to inhibit biofilms (MIC Geomean = 4.25 μg/mL, MBIC Geomean = 19.58 μg/mL) compared to AmB (MIC Geomean = 17.05 μg/mL, MBIC Geomean = 59.37 μg/mL). While ALS1 and HWP1 gene expression remained unchanged, molecular docking suggested the binding of eucalyptol to ALS1 and HWP1 proteins. The results of FE-SEM demonstrated the biofilm destruction through yeast bursting and cytoplasm leakage. The NEA, in comparison to AmB, decreased melanization in tissue sections and also enhanced the survival of G. melonella larvae.
CONCLUSION: The NEA exhibited superior biocompatibility toward the KG1 myeloblasts cell line and RBCs cells additionally enhances the survival rate of G. mellonella larvae compared to AmB. Overall, NEA proved an efficient nano-formulated product against C. albicans biofilms.
MATERIALS AND METHODS: In this study, NEA was synthesized and its capacity to combat biofilms generated by four clinical isolates and a reference strain of C. albicans was investigated according to CLSI M27-A3 guidelines. The NEA was prepared using eucalyptol oil, Tween 80, and ethanol, and characterized by measuring particle size, Z-potential, and encapsulation efficiency (EE %). The cytotoxicity study was performed on myeloblast KG1 cells and human RBCs. The gene assessment and molecular docking were conducted by Real-Time PCR and AutoDock Vina software, respectively. The in vivo stage was conducted on Galleria mellonella (G. mellonella) larvae, and ultimately, histopathological analysis was executed using H and E Staining.
RESULTS: Based on the results obtained, the NEA exhibited EE of 80 % for Amphotericin B (AmB), displaying an average size of 39 nm and zeta potential of -2 mV. Minimum inhibitory concentration (MIC) and minimum biofilm inhibitory concentration (MBIC) revealed more efficiency to inhibit biofilms (MIC Geomean = 4.25 μg/mL, MBIC Geomean = 19.58 μg/mL) compared to AmB (MIC Geomean = 17.05 μg/mL, MBIC Geomean = 59.37 μg/mL). While ALS1 and HWP1 gene expression remained unchanged, molecular docking suggested the binding of eucalyptol to ALS1 and HWP1 proteins. The results of FE-SEM demonstrated the biofilm destruction through yeast bursting and cytoplasm leakage. The NEA, in comparison to AmB, decreased melanization in tissue sections and also enhanced the survival of G. melonella larvae.
CONCLUSION: The NEA exhibited superior biocompatibility toward the KG1 myeloblasts cell line and RBCs cells additionally enhances the survival rate of G. mellonella larvae compared to AmB. Overall, NEA proved an efficient nano-formulated product against C. albicans biofilms.