Characterization of mechanisms underlying degradation of sclerotia of Sclerotinia sclerotiorum by Aspergillus aculeatus Asp-4 using a combined qRT-PCR and proteomic approach

Xiaojia Hu,Changbing Yu,Xing Liao,Lu Qin,Lei Hu,Lihua Xie,Yangmin Gong,Yinshui Li,Jude E Maul,Dilip K Lakshman,Xiangsheng Liao,Daniel P Roberts

doi:10.1186/s12864-017-4016-8

Abstract

BackgroundThe biological control agent Aspergillus aculeatus Asp-4 colonizes and degrades sclerotia of Sclerotinia sclerotiorum resulting in reduced germination and disease caused by this important plant pathogen. Molecular mechanisms of mycoparasites underlying colonization, degradation, and reduction of germination of sclerotia of this and other important plant pathogens remain poorly understood.ResultsAn RNA-Seq screen of Asp-4 growing on autoclaved, ground sclerotia of S. sclerotiorum for 48 h identified 997 up-regulated and 777 down-regulated genes relative to this mycoparasite growing on potato dextrose agar (PDA) for 48 h. qRT-PCR time course experiments characterized expression dynamics of select genes encoding enzymes functioning in degradation of sclerotial components and management of environmental conditions, including environmental stress. This analysis suggested co-temporal up-regulation of genes functioning in these two processes. Proteomic analysis of Asp-4 growing on this sclerotial material for 48 h identified 26 up-regulated and 6 down-regulated proteins relative to the PDA control. Certain proteins with increased abundance had putative functions in degradation of polymeric components of sclerotia and the mitigation of environmental stress.ConclusionsOur results suggest co-temporal up-regulation of genes involved in degradation of sclerotial compounds and mitigation of environmental stress. This study furthers the analysis of mycoparasitism of sclerotial pathogens by providing the basis for molecular characterization of a previously uncharacterized mycoparasite-sclerotial interaction.

Highlights

The biological control agent Aspergillus aculeatus Asp-4 colonizes and degrades sclerotia of Sclerotinia sclerotiorum resulting in reduced germination and disease caused by this important plant pathogen
Unless indicated otherwise isolate Asp-4 was cultured in potato dextrose broth (PDB) or potato dextrose agar (PDA) plus 50 μg/ mL hygromycin as this isolate was naturally resistant to that level of hygromycin
RNA-seq transcriptome sequencing, sequence assembly, and functional annotation of Aspergillus aculeatus Asp-4 Transcriptomes of isolate Asp-4 grown on ground sclerotia of S. sclerotiorum in 0.9% NaCl for 48 h at 28 °C and on the PDA control were determined from one experiment

Summary

Introduction

The biological control agent Aspergillus aculeatus Asp-4 colonizes and degrades sclerotia of Sclerotinia sclerotiorum resulting in reduced germination and disease caused by this important plant pathogen. The use of mycoparasites as biological control agents is an alternative strategy for control of S. sclerotiorum [3, 11, 12] These mycoparasites are applied to the field and expected to colonize and kill or weaken sclerotia resulting in the reduction of initial inoculum. Sclerotia, such as those produced by S. sclerotiorum, are hyphal aggregates that consist of a pigmented rind, a thin-walled cortex, and a large central medulla [13,14,15]. In addition to the daunting physical barrier due to the complexity of these interwoven polymers, mycoparasites are likely to encounter potentially stressful conditions as S. sclerotiorum produces toxic phenolic compounds and fatty acids [18], reactive oxygen species [19], organic acids such as oxalic acid, and various enzymes capable of producing toxic products [20]

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