Toxoflavin Produced by Burkholderia gladioli from Lycoris aurea Is a New Broad-Spectrum Fungicide

Abstract

Fungal infections not only cause extensive agricultural damage but also result in serious diseases in the immunodeficient populations of human beings. Moreover, the increasing emergence of drug resistance has led to a decrease in the efficacy of current antifungals. Thus, screening of new antifungal agents is imperative in the fight against antifungal drug resistance. In this study, we show that an endophytic bacterium, Burkholderia gladioli HDXY-02, isolated from the medicinal plant Lycoris aurea, showed broad-spectrum antifungal activity against plant and human fungal pathogens. An antifungal ability assay indicated that the bioactive component was produced from strain HDXY-02 having an extracellular secreted component with a molecular weight lower than 1,000 Da. In addition, we found that this new antifungal could be produced effectively by liquid fermentation of HDXY-02. Furthermore, the purified component contributing to the antifungal activity was identified to be toxoflavin, a yellow compound possessing a pyrimido[5,4-e][1,2,4]triazine ring. In vitro bioactivity studies demonstrated that purified toxoflavin from B. gladioli HDXY-02 cultures had a significant antifungal activity against the human fungal pathogen Aspergillus fumigatus, resulting in abolished germination of conidia. More importantly, the growth inhibition by toxoflavin was observed in both wild-type and drug-resistant mutants (cyp51A and non-cyp51A) of A. fumigatus Finally, an optimized protocol for the large-scale production of toxoflavin (1,533 mg/liter) has been developed. Taken together, our findings provide a promising biosynthetic resource for producing a new antifungal reagent, toxoflavin, from isolates of the endophytic bacterium B. gladioliIMPORTANCE Human fungal infections are a growing problem associated with increased morbidity and mortality. Moreover, a growing number of antifungal-resistant fungal isolates have been reported over the past decade. Thus, the need for novel antifungal agents is imperative. In this study, we show that an endophytic bacterium, Burkholderia gladioli, isolated from the medicinal plant Lycoris aurea, is able to abundantly secrete a compound, toxoflavin, which has a strong fungicidal activity not only against plant fungal pathogens but also against human fungal pathogens Aspergillus fumigatus and Candida albicans, Cryptococcus neoformans, and the model filamentous fungus Aspergillus nidulans More importantly, toxoflavin also displays an efficacious inhibitory effect against azole antifungal-resistant mutants of A. fumigatus Consequently, our findings provide a promising approach to abundantly produce toxoflavin, which has novel broad-spectrum antifungal activity, especially against those currently problematic drug-resistant isolates.

Keywords: Aspergillus; Burkholderia gladioli; antifungal; endophytic bacterium; fungal pathogens; toxoflavin.