Access to Anti-Biofilm Compounds from Endolichenic Fungi Using a Bioguided Networking Screening

Abstract

Endolichenic microorganisms represent a new source of bioactive natural compounds. Lichens, resulting from a symbiotic association between algae or cyanobacteria and fungi, constitute an original ecological niche for these microorganisms. Endolichenic fungi inhabiting inside the lichen thallus have been isolated and characterized. By cultivation on three different culture media, endolichenic fungi gave rise to a wide diversity of bioactive metabolites. A total of 38 extracts were screened for their anti-maturation effect on Candida albicans biofilms. The 10 most active ones, inducing at least 50% inhibition, were tested against 24 h preformed biofilms of C. albicans, using a reference strain and clinical isolates. The global molecular network was associated to bioactivity data in order to identify and priorize active natural product families. The MS-targeted isolation led to the identification of new oxygenated fatty acid in Preussia persica endowed with an interesting anti-biofilm activity against C. albicans yeasts.

Keywords: Candida biofilms; endolichenic fungi; fatty acids; metabolomic.

Figure 1

Total quantity of crude fungal extracts obtained from the three culture media.

Figure 2

Principal component analysis score plots for the 38 endolichenic fungal extracts investigated based on HPLC-MS/MS data figuring the 2 first principal components for the totality of the extracts (A) or after removal of signals from Coniochaeta and Biscognauxia extracts (B). Colors were attributed to the different fungal genus investigated in the present study (Anthostomella (red), Biscogniauxia (green), Coniochaeta (blue), Fusarium (cyan), Peziza (pink), Plectania (yellow), Preussia (purple)). Active extracts (*) inducing more than 50% adherence inhibition at 50, 100 or 200 µg/mL in 48 h are highlighted. They correspond to AP_SAB (Anthostomella pinea), AP_PDA (A. pinea), AP_MEA (A. pinea), CL_SAB (C. lignicola), CL_PDA (C. lignicola), CL_MEA (C. lignicola), PP_1_PDA (P. persica), PP_1_SAB (P. persica), PV_MEA (P. varia), PSP_MEA (Plectania sp.), CD_MEA, (Coniochaeta decumbens), CH_PDA (Coniochaeta hoffmannii), CH_MEA (C. hoffmannii), CH_SAB (C. hoffmanii), PP_2_MEA (Preussia persica), PP_2_PDA (P. persica), PP_2_SAB (P. persica), FA_PDA (Fusarium avenaceum), BM_1_PDA (Biscogniauxia mediterranea), BM_1_SAB (Biscogniauxia mediterranea).

Figure 3

Feature-based molecular network corresponding to Coniochaeta extracts. Nodes are filled with pie charts representing the mean peak areas for each feature in the 3 defined groups of activity (anti-maturation activity at 100 µg/mL in 48 h): low (red) for 0–40%, medium (orange) for 40–60% and high (green) for 60–100%. Node sizes are related to FiBiCo scores indicated in grey, highlighting compounds mostly linked to the activity observed. Nodes with corresponding hits in the GNPS MS/MS spectra library (with cosine > 0.65) are surrounded by a red circle, nodes which correspond to compounds isolated in the present study by a blue circle.

Figure 4

Feature-based molecular network corresponding to Biscogniauxia extracts. Nodes are filled with pie charts representing the mean peak areas for each feature in the 3 defined groups of activity (anti-maturation activity at 200 µg/mL in 48 h): low (red) for 0–30%, medium (orange) for 30–50% and high (green) for 50–100%. Node sizes are related to FiBiCo scores indicated in grey, highlighting compounds mostly linked to the activity observed.

Figure 5

Feature-based molecular network corresponding to Preussia extracts. Nodes are filled with pie charts representing the mean peak areas for each feature in the 3 defined groups of activity (anti-maturation activity at 50 µg/mL in 48 h): low (red) for 0–40%, medium (orange) for 40–50% and high (green) for 50–100%. Node sizes are related to FiBiCo scores indicated in grey, highlighting compounds mostly linked to the activity observed.

Figure 6

Feature-based molecular network corresponding to Anthostomella extracts. Nodes are filled with pie charts representing the mean peak areas for each feature in the different extracts investigated: MEA (light yellow), PDA (light green) and SAB (dark green).

Figure 7

Chemical structures of compounds identified as actives by the FiBiCo script. Emodin was biosynthezised by Coniochaeta species and the fatty acid was isolated from Preussia persica extracts.