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Analysis of the effect of Bacillus velezensis culture filtrate on the growth and proteome of Cladobotryum mycophilum

ABSTRACT: Cladobotryum mycophilum, the causative agent of cobweb disease on Agaricus bisporus results in significant crop losses for mushroom growers worldwide. Cobweb disease is treated through strict hygiene control methods and the application of chemical fungicides but an increase in fungicide resistant Cladobotryum strains has resulted in a need to develop alternative biocontrol treatment methods. The aim of the work presented here was to evaluate the response of C. mycophilum to a Bacillus velezensis isolate to assess its potential as a novel biocontrol agent. Exposure of 48 hr C. mycophilum cultures to 25% v/v 96h B. velezensis culture filtrate resulted in a 57% reduction in biomass (P < 0.0002), a disruption in hyphal structure and morphology, and the appearance of aurofusarin in culture medium. Proteomic analysis of B. velezensis culture filtrate revealed the presence of peptidase 8 (subtilisin), peptide deformylase and probable cytosol aminopeptidase which are known to induce cell disruption. Characterisation of the proteomic response of C. mycophilum following exposure to B. velezensis culture filtrate revealed an increase in the abundance of a variety of proteins associated with stress response (ISWI chromatin-remodelling complex ATPase ISW2 (+24 fold), carboxypeptidase Y precursor (+3 fold) and calmodulin (+2 fold). There was also a decrease in the abundance of proteins associated with transcription (40S ribosomal protein S30 (-26 fold), 40S ribosomal protein S21 (-3 fold) and carbohydrate metabolism, (L-xylulose reductase (-10 fold). The results presented here indicate that B. velezensis culture filtrate is capable of inhibiting the growth of C. mycophilum and inducing a stress response, thus indicating its potential to control this important pathogen of mushrooms.

INSTRUMENT(S): Q Exactive HF

ORGANISM(S): Trichoderma Sp. 1 Yw-2011

SUBMITTER: Joy Clarke

LAB HEAD: Kevin Kavanagh

PROVIDER: PXD026467 | Pride | 2021-10-12

REPOSITORIES: Pride

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Project description:Aspergillus fumigatus is an opportunistic fungal pathogen of the lungs resulting in both chronic and acute infection. The fungus interacts with numerous bacteria that compose the microflora of the lung including Pseudomonas aeruginosa and Klebsiella pneumoniae, both common isolates from Cystic fibrosis sputum. It is well documented that bacterial and fungal interactions both physically and through production of secreted products which can alter the growth and virulence of other species. The interaction with P. aeruginosa has been characterised previously indicating decreased growth and increased gliotoxin production in the presence of cells and the opposite when only exposed to culture filtrate. In our analysis exposure of cell free culture filtrate inhibited fungal growth and increased gliotoxin production. This was supported with proteomic analysis indicating reduction in 1,3-beta-glucanosyltransferase (-3.97 fold), methyl sterol monooxygenase erg25B (-2.9 fold) and Calcium/calmodulin dependent protein kinase (-4.2 fold) involved in fungal development. Coupled with increased expression of Glutathione S-transferase gliG (6.17 fold), non-ribosomal peptide synthase gliP (3.67 fold), O-methyltransferase gliM (3.5 fold), Gamma-glutamyl acyltransferase gliK (2.89 fold) and thioredoxin reductase gliT (2.33 fold) involved in Gliotoxin production indicate a possible shift towards secondary metabolism. Qualitative proteomic analysis of the bacterial supernatant also elucidates possible mechanisms for these alterations including physical interaction by Outer membrane protein A, High-affinity zinc uptake system protein ZnuA and Iron uptake system component EfeO reducing essential nutrient availability, enzymatic activity including Periplasmic serine endoprotease DegP-like and redox active proteins including Alkyl hydroperoxide reductase C. These proteomic alterations to the fungus could have implications for the severity of virulence in the context of coinfection and may impact disease progression.

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Analysis of the effect of Bacillus velezensis culture filtrate on the growth and proteome of Cladobotryum mycophilum

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