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Epiphytic yeasts, which colonize plant surfaces, may possess activity that can be harnessed to help plants defend themselves against various pathogens. Due to their unique characteristics, epiphytic yeasts belonging to the genus Pseudozyma hold great potential for use as biocontrol agents. We identified a unique, biologically active isolate of the epiphytic yeast Pseudozyma aphidis that is capable of inhibiting Botrytis cinerea via a dual mode of action, namely induced resistance and antibiosis. Here, we show that strain L12 of P. aphidis can reduce the severity of powdery mildew caused by Podosphaera xanthii on cucumber plants with an efficacy of 75%. Confocal and scanning electron microscopy analyses demonstrated P. aphidis proliferation on infected tissue and its production of long hyphae that parasitize the powdery mildew hyphae and spores as an ectoparasite. We also show that crude extract of P. aphidis metabolites can inhibit P. xanthii spore germination in planta. Our results suggest that in addition to its antibiosis as mode of action, P. aphidis may also act as an ectoparasite on P. xanthii. These results indicate that P. aphidis strain L12 has the potential to control powdery mildew.

Pseudozyma spp. are yeast-like fungi, classified in the Ustilaginales, which are mostly epiphytic or saprophytic and are not pathogenic to plants. Several Pseudozyma species have been reported to exhibit biological activity against powdery mildews. However, previous studies have reported that Pseudozyma aphidis, which can colonize plant surfaces, is not associated with the collapse of powdery mildew colonies. In this report, we describe a novel P. aphidis strain and study its interactions with its plant host and the plant pathogen Botrytis cinerea. This isolate was found to secrete extracellular metabolites that inhibit various fungal pathogens in vitro and significantly reduce B. cinerea infection in vivo. Moreover, P. aphidis sensitized Arabidopsis (Arabidopsis thaliana) plants' defense machinery via local and systemic induction of PATHOGENESIS-RELATED1 (PR1) and PLANT DEFENSIN1.2 (PDF1.2) expression. P. aphidis also reduced B. cinerea infection, locally and systemically, in Arabidopsis mutants impaired in jasmonic acid (JA) or salicylic acid (SA) signaling. Thus, in addition to direct inhibition, P. aphidis may inhibit B. cinerea infection via induced resistance in a manner independent of SA, JA, and Nonexpressor of PR1 (NPR1). P. aphidis primed the plant defense machinery and induced stronger activation of PDF1.2 after B. cinerea infection. Finally, P. aphidis fully or partially reconstituted PR1 and PDF1.2 expression in npr1-1 mutant and in plants with the SA hydroxylase NahG transgene, but not in a jasmonate resistant1-1 mutant, after B. cinerea infection, suggesting that P. aphidis can bypass the SA/NPR1, but not JA, pathway to activate PR genes. Thus, either partial gene activation is sufficient to induce resistance, or the resistance is not directed solely through PR1 and PDF1.2 but probably through other pathogen-resistance genes or pathways as well.

The ability of plant pathogens to rapidly develop resistance to commonly used pesticides challenges efforts to maximize crop production. Fungal biocontrol agents have become an important alternative to chemical fungicides as a result of environmental concerns regarding conventional pesticides, including resistance issues. The complex mode of action of biocontrol agents reduces the likelihood that pathogens will develop resistance to them. We recently isolated a unique, biologically active isolate of the epiphytic fungus Pseudozyma aphidis. We show that the extracellular metabolites secreted by our P. aphidis isolate can inhibit Xanthomonas campestris pv. vesicatoria, X. campestris pv. campestris, Pseudomonas syringae pv. tomato, Erwinia amylovora, Clavibacter michiganensis, and Agrobacterium tumefaciens in vitro. Moreover, application of Pseudozyma aphidis spores on tomato plants in the greenhouse significantly reduced (by 60%) the incidence of bacterial wilt and canker disease caused by C. michiganensis subsp. michiganensis on those plants as well as disease severity by 35%. Furthermore, infected plants treated with P. aphidis were 25% taller than control infected plants. We found that P. aphidis activates PR1a—and other pathogenesis-related genes in tomato plants—and can trigger an induced-resistance response against C. michiganensis that proceeds in a salicylic-acid-independent manner, as shown using NahG-transgenic tomato plants.

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