A Comprehensive in vitro and in silico Assessment of Eugenol Glycoconjugates against Azole and Amphotericin B Resistant Rhizopus delemar var arrhizus

Background: Rhizopus delemar var arrhizus is a major cause of mucormycosis, a severe infectious disease with high morbidity and mortality. Treatment is challenging due to rising antifungal resistance. Glycosylation is a crucial technique for enhancing the properties of phenolic compounds like eugenol. The present study tries to examine the antifungal efficacy of eugenol glycoconjugates against azole and amphotericin B-resistant R. delemar. Methods and Results: Out of 50 soil samples, 12 Mucor isolates were isolated with 7 identified as R. delemar via 18S ITS sequencing. Antifungal susceptibility testing (AST) revealed that all R. delemar isolates were resistant to amphotericin B (MIC >1 μg/mL). Most isolates also showed resistance to posaconazole (MIC >1 μg/mL) and itraconazole (MIC >2 μg/mL). AST of eugenol glycoconjugate (coded 6g) showed efficacy against resistant R. delemar isolates, with MIC values ranging from 6.25μg/mL to 25μg/mL. Flow cytometry confirmed its fungicidal activity, correlating with MIC data. Compound 6g significantly reduced conidial germination within 24h and exhibited no cytotoxicity on A549 lung cancer cells. In-silico analysis revealed a negative binding affinity of compound 6g for the spore coat protein CotH3, which could a potential antifungal target. Conclusions: Compound 6g could be an potential antifungal molecule against resistant R. delemar isolates, which requires further studies.