Anticandidal activity of ethanolic root extract of Juglans regia (L.): Effect on growth, cell morphology, and key virulence factors

Summary The emergence of drug-resistant strains has encouraged several studies on natural products with antifungal activity and low toxicity. In this study, the antifungal effect of methanolic root extract of Juglans regia (JRE) was investigated against 9 strains of Candida (one reference and 8 clinical strains) through MIC 90 and spot assays. To gain insight into the mechanism of antifungal action, we carried out confocal scanning laser microscopy (CLSM), transmission electron microscopy (TEM), and then examined the effect of JRE on hydrolytic enzyme secretion. Additionally, JRE was subjected to various phytochemical tests, chemically characterized by gas chromatography-mass spectrometry analysis (GC-MS) and its toxicity was tested against H9c2 rat cardiac myoblasts. The phytochemical tests showed the presence of phenols, alkaloids, steroids, saponins, and tannins in JRE. In the GC-MS analysis, a total of 40 compounds were identified. JRE was found to be effective in liquid media with MICs ranging from 300 to 700 μg/mL. Spot assay results revealed that Candida cells show increased sensitivity to JRE. CSLM experiments showed that cells exposed to JRE (MIC) exhibited cell membrane disruption. TEM micrograph of treated cells showed extensive breakage in the cell wall and cell membrane. Average inhibition of proteinase and phospholipase secretion (of five C. albicans strains) at MIC/2 values of JRE was 45.17%, and 34.29%, respectively. Cellular toxicity of JRE against H9c2 rat cardiac myoblasts was less than 10% at the highest MIC value. These findings encourage further development of JRE.