Hydrophobins are required for conidial hydrophobicity and plant root colonization in the fungal biocontrol agent Clonostachys rosea.

Mukesh K Dubey,Dan Funck Jensen,Magnus Karlsson

doi:10.1186/1471-2180-14-18

Mukesh K Dubey, Dan Funck Jensen + Show 1 more

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https://doi.org/10.1186/1471-2180-14-18

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Abstract

BackgroundFilamentous fungi produce small cysteine rich surface active amphiphilic hydrophobins on the outer surface of cell walls that mediate interactions between the fungus and the environment. The role of hydrophobins in surface hydrophobicity, sporulation, fruit body formation, recognition and adhesion to host surface and virulence have been reported. The aim of the present study was to characterize the biological function of hydrophobins in the fungal biocontrol agent Clonostachys rosea in order to understand their potential roles in biocontrol mechanisms.ResultsBased on the presence of hydrophobin domains, cysteine spacing patterns and hydropathy plots, we identified three class II hydrophobin genes in C. rosea. Gene expression analysis showed basal expression of Hyd1, Hyd2 and Hyd3 in all conditions tested with the exception of induced Hyd1 expression in conidiating mycelium. Interestingly, up-regulation of Hyd1, Hyd2 and Hyd3 was found during C. rosea self interaction compared to interactions with the fungal plant pathogens Botrytis cinerea or Fusarium graminearum in dual culture assays. Phenotypic analysis of C. rosea deletion and complementation strains showed that Hyd1 and Hyd3 are jointly required for conidial hydrophobicity, although no difference in mycelia hydrophobicity was found between wild type (WT) and mutant strains. Interestingly, mutant strains showed increased growth rates, conidiation and enhanced tolerances of conidia to abiotic stresses. Antagonism tests using in vitro dual culture and detached leaf assays showed that the mutant strains were more aggressive towards B. cinerea, F. graminearum or Rhizoctonia solani, and that aggression was partly related to earlier conidial germination and enhanced tolerance of mutant strains to secreted fungal metabolites. Furthermore, in vitro Arabidopsis thaliana root colonization assays revealed reduced root colonization ability of the ΔHyd3 strain, but not for the ΔHyd1 strain. Furthermore, enhanced root colonization ability for the ΔHyd1ΔHyd3 strain was found in comparison to WT.ConclusionsThese results show a role for hydrophobins in conidial hydrophobicity, control of conidial germination under stress conditions, and in root colonization in C. rosea. However, functional studies of Hyd2 remains to be performed in order to fully assess the role of hydrophobins in C. rosea.

Highlights

Filamentous fungi produce small cysteine rich surface active amphiphilic hydrophobins on the outer surface of cell walls that mediate interactions between the fungus and the environment
The current study aims to understand the biological function of hydrophobins in C. rosea with emphasis on its role in fungal growth and development, antagonism, and interactions with plants
Identification and phylogenetic analysis of C. rosea hydrophobins Blast searches against a C. rosea strain IK726 draft genome database using a total of 35 class I, class Ia and class II hydrophobin amino acid sequences from Trichoderma spp. [29], identified three genes with an E-value ≤ 1 × 10-5

Summary

Introduction

Filamentous fungi produce small cysteine rich surface active amphiphilic hydrophobins on the outer surface of cell walls that mediate interactions between the fungus and the environment. Hydrophobins act as natural surfactants and reduce the surface tension of the medium, and perform a variety of biological functions in the life cycle of filamentous fungi These include formation of a protective layer surrounding the hyphae and sexual structures, development of aerial hyphae, sporulation and spore dispersal, and fruit body formation [1,2,3]. In the entomopathogenic fungus Beauveria bassiana, deletion of hydrophobin genes results in decreased spore hydrophobicity and adhesion, loss of water-mediated dispersal, and lowered virulence to insects [10]. In another entomopathogenic fungus, Metarhizium brunneum, characterization of hydrophobins showed their role in conidiation, hydrophobicity, pigmentation and virulence [11]. Similar results are reported for Botrytis cinerea where deletion mutants of hydrophobin genes does not display any phenotypic differences compared to the wild type (WT) strain [13]

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