Diosgenin producing Bacillus sp. strain IRMC27M2 as a genome-mined weapon against multidrug-resistant Candidozyma (Candida) auris.

Abstract

Candidozyma auris (Candida auris) is an emerging multidrug-resistant (MDR) fungal pathogen prioritised by the World Health Organisation that poses a significant global health threat due to high mortality. Discovering novel antifungal drugs is crucial for effective treatment. This study identifies and describes a native bacterial isolate, Bacillus sp. strain IRMC27M2, with anti-C. auris activity. An integrated approach was used, including 16S rRNA gene sequencing to identify the bacterial isolate, followed by whole-genome sequencing, antifungal analysis, cytotoxicity testing, gas chromatography-mass spectrometry (GC-MS) analysis and comparative genomics. The IRMC27M2 genome was sequenced using nanopore long-read sequencing and the resulting genome (3,87,328 bp) is phylogenetically related to Bacillus amyloliquefaciens and Bacillus velezensis. Biosynthesis-related gene clusters (BGCs) were identified in the IRMC27M2 genome. Media optimisation protocols (FRC6 and REM3) were performed to enhance secondary metabolite production and the resulting ethyl acetate fractions were analysed by UV spectrophotometry and GC-MS. Antifungal analysis with metabolites from Bacillus sp. strain IRMC27M2 significantly reduced cell size and induced crushed phenotypes in C. albicans and C. auris. Collapse of cell membranes and lysis of cells were observed. Whole-genome sequencing revealed 10 BGCs potentially involved in antifungal compound biosynthesis. The metabolites produced using FRC6 and REM3 protocols showed no cytotoxic effects. GC-MS analysis of the ethyl acetate fraction revealed a range of metabolites, with diosgenin being the most abundant. Manual and reverse verification confirmed the presence of genes linked to the methylerythritol phosphate biosynthesis pathway and confirmed the capability of IRMC27M2 for diosgenin production. In conclusion, the findings highlight the significant potential of Bacillus sp. strain IRMC27M2 as a biofactory for the production of diosgenin. This signifies promising future research for developing treatments against multidrug-resistant C. albicans and C. auris which are prioritised by the WHO. Further research is necessary to confirm if Bacillus sp. strain IRMC27M2 represents a novel subspecies.