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Antofine, a phenanthroindolizidine alkaloid, is a bioactive natural product isolated from milkweeds that exhibits numerous biological activities, including anticancer, antimicrobial, antiviral, and anti-inflammatory properties. However, the direct targets and mode of action of antofine have not been determined. In this report, we show that antofine displays antifungal properties against the phytopathogen Fusarium graminearum, the cause of Fusarium head blight disease (FHB). FHB does devastating damage to agriculture, causing billions of dollars in economic losses annually. We therefore sought to understand the mode of action of antofine in F. graminearum using insights from yeast chemical genomic screens. We used haploinsufficiency profiling (HIP) to identify putative targets of antofine in yeast and identified three candidate targets, two of which had homologs in F. graminearum. The Fusarium homologues of two targets, glutamate dehydrogenase (FgGDH) and resistance to rapamycin deletion 2 (FgRRD2), can bind antofine. Of the two genes, only the Fgrrd2 knockout displayed a loss of virulence in wheat, indicating that RRD2 is an antivirulence target of antofine in F. graminearum. Mechanistically, we demonstrate that antofine disrupts the interaction between FgRRD2 and FgTap42, which is part of the Tap42-phosphatase complex in the target of rapamycin (TOR) signaling pathway, a central regulator of cell growth in eukaryotes and a pathway of extensive study for controlling numerous pathologies. IMPORTANCE Fusarium head blight caused by the fungal pathogen Fusarium graminearum is a devastating disease of cereal crops worldwide, with limited effective chemical treatments available. Here we show that the natural alkaloid compound antofine can inhibit fusarium head blight in wheat. Using yeast genomic screening, we identified the TOR pathway component RRD2 as a target of antofine that is also required for F. graminearum pathogenicity.

Bioassay-guided fractionation of the crude extract (80% EtOH) of the leaves of Cestrum schlechtendahlii, a plant used by Q'eqchi' Maya healers for treatment of athlete's foot, resulted in the isolation and identification of two spirostanol saponins (1 and 2). Structure elucidation by MS, 1D-NMR, and 2D-NMR spectroscopic methods identified them to be the known saponin (25R)-1β,2α-dihydroxy-5α-spirostan-3-β-yl-O-α-L-rhamnopyranosyl-(1 → 2)-β-D-galactopyranoside (1) and new saponin (25R)-1β,2α-dihydroxy-5α-spirostan-3-β-yl-O-β-D-galactopyranoside (2). While 2 showed little or no antifungal activity at the highest concentration tested, 1 inhibited growth of Saccharomyces cerevisiae (minimum inhibitory concentration (MIC) of 15-25 μM), Candida albicans, Cryptococcus neoformans, and Fusarium graminearum (MIC of 132-198 μM).

The Marcgraviaceae is a neotropical plant family of lianas and shrubs distributed throughout Central and South America, mainly in evergreen and semi-evergreen tropical forests. In this study, extracts of 12 Marcgraviaceae species were assessed for inhibition of bacterial quorum sensing (QS) in Chromobacterium violaceum, bacterial biofilm formation in Pseudomonas aeruginosa PA14, and fungal growth in Saccharomyces cerevisiae. Six species showed promising inhibitory activities in at least one of the three bioassays (Marcgravia nervosa Triana & Planch., Marcgravia polyantha Delp., Marcgravia schippii Standl., Marcgraviastrum subsessile (Benth.) Bedell, Schwartzia brasiliensis (Choisy) Bedell ex Gir-Cañas, and Schwartzia costaricensis (Gilg.) Bedell). Analyses of the crude extract of the leaves of Marcgravia nervosa using HPLC–APCI–MS showed the presence of five pentacyclic triterpenes: ursolic acid (2), betulinic acid (3), α–amyrin (4), β–amyrin (5), lupeol (6). Bioassay-guided fractionation of this plant resulted in the isolation and identification of 2-methoxy-1,4-naphthoquinone (1) as the active principle with a minimum inhibitory concentration (MIC) of 85–100 μmol·L−1 against Saccharomyces cerevisiae BY4741 (haploid) and BY4743 (diploid).