Antibiotic resistance is spreading at a faster rate than new antibiotic agents applied for clinical remedies. It is an urgent need to discover potential compounds to combat multidrug-resistant (MDR) bacteria. Marine fungi offer a promising avenue for mining antibiotic-like molecules with chemical diversity. To discover structurally novel and antibiotic metabolites, we screened the in-house marine fungus genome library and found a fungus Stephanonectria keithii LZD-10-1 containing a non-ribosomal peptide synthetase (NRPS) cluster with 18 modules to synthesize a new subfamily of peptaibols with effective eradication against MDR pathogens. Targeting isolation of the cultured fungus afforded six new peptaibols, which exhibit the ability to kill MDR bacteria by targeting bacterial membrane phospholipids, especially phosphatidylglycerol (PG), leading to the dysfunction of bacterial membranes. Furthermore, their efficacies against methicillin-resistant Staphylococcus aureus (MRSA) in both Galleria mellonella and mouse wound infection models were observed. This study underscores the significance of employing genome-guided approaches to identify untapped marine fungi as potential sources for novel antibiotic candidates with unique scaffolds.

Dermatophytosis, an infectious disease caused by several fungi, can affect the hair, nails, and/or superficial layers of the skin and is of global significance. The most common dermatophytes in cats and dogs are Microsporum canis and Trichophyton mentagrophytes. Wood’s lamp examination, microscopic identification, and fungal culture are the conventional clinical diagnostic methods, while PCR (Polymerase Chain Reaction) and qPCR (Quantitative PCR) are playing an increasingly important role in the identification of dermatophytes. However, none of these methods could be applied to point-of-care testing (POCT). The recent development of the CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) based diagnostic platform promises a rapid, accurate, and portable diagnostic tool. In this paper, we present a Cas12a-fluorescence assay to detect and differentiate the main dermatophytes in clinical samples with high specificity and sensitivity. The Cas12a-based assay was performed with a combination of recombinase polymerase amplification (RPA). The results could be directly visualized by naked eyes under blue light, and all tested samples were consistent with fungal culture and sequencing results. Compared with traditional methods, the RPA-Cas12a-fluorescence assay requires less time (about 30 min) and less complicated equipment, and the visual changes can be clearly observed with naked eyes, which is suitable for on-site clinical diagnosis.

Background: Fungal infections pose an increasingly predominant threat to human and animal health. Modified compounds derived from chemo-diverse natural products offer enhanced therapeutic efficacies and promising approaches to combat life-threatening fungal pathogens. Methods: We performed biosynthetic gene clusters analysis of 2,4-diacetylchloroglucoside (DAPG) in 4292 shotgun metagenomes samples from the healthy and diseased skin. Then, we assessed the antifungal activity of DAPG and the derivative 2,4-diproylphloroglucinol (DPPG) against pathogenic fungi by minimum inhibitory concentrations. The inhibitory effects of DPPG were measured using hyphal growth assay and spore germination assay. Concurrently, the mechanism of DPPG on Aspergillus fumigatus was investigated in membrane permeability and fluidity. The therapeutic efficacy was evaluated in a Galleria mellonella infection model. Results: We observed a significantly higher abundance of bacteria harboring DAPG biosynthetic clusters on healthy skin compared to diseased skin. Further, we designed and synthesized a series of phloroglucinol derivatives based on DAPG and obtained an antifungal candidate DPPG. DPPG not only exhibited robust antifungal activity against Aspergillus spp. and Candida spp. but also impaired hyphal growth and spore germination of A. fumigatus in vitro. A mechanism study showed that DPPG reduced membrane fluidity and increased the leakage of cellular contents, resulting in membrane perturbation and fungal death. Lastly, the therapeutic efficacy of DPPG was confirmed in a G. mellonella infection model. Conclusions: Our study demonstrates that DPPG is a potent scaffold to combat invasive fungal infections.