Competition assays and physiological experiments of soil and phyllosphere yeasts identify Candida subhashii as a novel antagonist of filamentous fungi

Abstract

BackgroundWhile recent advances in next generation sequencing technologies have enabled researchers to readily identify countless microbial species in soil, rhizosphere, and phyllosphere microbiomes, the biological functions of the majority of these species are unknown. Functional studies are therefore urgently needed in order to characterize the plethora of microorganisms that are being identified and to point out species that may be used for biotechnology or plant protection. Here, we used a dual culture assay and growth analyses to characterise yeasts (40 different isolates) and their antagonistic effect on 16 filamentous fungi; comprising plant pathogens, antagonists, and saprophytes.ResultsOverall, this competition screen of 640 pairwise combinations revealed a broad range of outcomes, ranging from small stimulatory effects of some yeasts up to a growth inhibition of more than 80% by individual species. On average, yeasts isolated from soil suppressed filamentous fungi more strongly than phyllosphere yeasts and the antagonistic activity was a species-/isolate-specific property and not dependent on the filamentous fungus a yeast was interacting with. The isolates with the strongest antagonistic activity were Metschnikowia pulcherrima, Hanseniaspora sp., Cyberlindnera sargentensis, Aureobasidium pullulans, Candida subhashii, and Pichia kluyveri. Among these, the soil yeasts (C. sargentensis, A. pullulans, C. subhashii) assimilated and/or oxidized more di-, tri- and tetrasaccharides and organic acids than yeasts from the phyllosphere. Only the two yeasts C. subhashii and M. pulcherrima were able to grow with N-acetyl-glucosamine as carbon source.ConclusionsThe competition assays and physiological experiments described here identified known antagonists that have been implicated in the biological control of plant pathogenic fungi in the past, but also little characterised species such as C. subhashii. Overall, soil yeasts were more antagonistic and metabolically versatile than yeasts from the phyllosphere. Noteworthy was the strong antagonistic activity of the soil yeast C. subhashii, which had so far only been described from a clinical sample and not been studied with respect to biocontrol. Based on binary competition assays and growth analyses (e.g., on different carbon sources, growth in root exudates), C. subhashii was identified as a competitive and antagonistic soil yeast with potential as a novel biocontrol agent against plant pathogenic fungi.Electronic supplementary materialThe online version of this article (doi:10.1186/s12866-016-0908-z) contains supplementary material, which is available to authorized users.

Characterization of antagonistic yeasts for biocontrol applications on apples or in soil by quantitative analyses of synthetic yeast communities

“…Candida subhashii also inhibited the growth of T. basicola more strongly on agar plates containing NAG than on media supplemented with glucose or maltose. These findings may indicate, as suggested previously (Hilber‐Bodmer et al, ), that C. subhashii is adapted to grow on insect cuticle and/or fungal cell walls, of either dead or living organisms. Based on these findings, including NAG or chitin in a C. subhashii formulation may stimulate the antagonism and thus the biocontrol activity of this yeast.…”

“…Six yeast isolates from the apple phyllosphere or soil ( A. pullulans , C. sargentensis , C. subhashii , Hanseniaspora sp., M. pulcherrima and P. kluyveri ), identified as overall strong antagonists of a broad range of different filamentous fungi (Hilber‐Bodmer et al, ), were used in the present study.…”

“…One example of at least two experiments that resulted in similar outcomes is shown. (b) Binary competition assays of the six yeasts against the soilborne plant pathogen Thielaviopsis basicola were carried out on YNB medium with glucose, maltose, or NAG as carbon sources and otherwise performed as previously described (Hilber‐Bodmer et al, ). The mean relative growth (and standard error) of T. basicola in the presence of the six yeasts and on the control plate is shown (four replicates were used) [Colour figure can be viewed at wileyonlinelibrary.com]…”

“…The six yeasts included in the synthetic community studied here were isolated from soil ( A. pullulans , C. subhashii , C. sargentensis ) or the apple phyllosphere ( Hanseniaspora sp., M. pulcherrima , P. kluyveri ; Hilber‐Bodmer et al, ). Despite its origin, the isolate NBB 7.2.1 of A. pullulans was a strong competitor on apple.…”

“…(APC 12.1), Cyberlindnera sargentensis (SHA 17.2), Aureobasidium pullulans (NBB 7.2.1), Candida subhashii (FGA 2.2) and Pichia kluyveri (APC 11.10B). These six isolates were previously characterized and identified as strong antagonists of a range of filamentous fungi (Hilber‐Bodmer, Schmid, Ahrens, & Freimoser, ). The soilborne fungal pathogen Thielaviopsis basicola (DCM 1603a; SH200142.07) was isolated from an infected carrot in Switzerland.…”