Fungi treated with small chemicals exhibit increased antimicrobial activity against facultative bacterial and yeast pathogens

Abstract

For decades, fungi have been the main source for the discovery of novel antimicrobial drugs. Recent sequencing efforts revealed a still high number of so far unknown "cryptic" secondary metabolites. The production of these metabolites is presumably epigenetically silenced under standard laboratory conditions. In this study, we investigated the effect of six small mass chemicals, of which some are known to act as epigenetic modulators, on the production of antimicrobial compounds in 54 spore forming fungi. The antimicrobial effect of fungal samples was tested against clinically facultative pathogens and multiresistant clinical isolates. In total, 30 samples of treated fungi belonging to six different genera reduced significantly growth of different test organisms compared to the untreated fungal sample (growth log reduction 0.3-4.3). For instance, the pellet of Penicillium restrictum grown in the presence of butyrate revealed significant higher antimicrobial activity against Staphylococcus (S.) aureus and multiresistant S. aureus strains and displayed no cytotoxicity against human cells, thus making it an ideal candidate for antimicrobial compound discovery. Our study shows that every presumable fungus, even well described fungi, has the potential to produce novel antimicrobial compounds and that our approach is capable of rapidly filling the pipeline for yet undiscovered antimicrobial substances.

Figure 1

Antimicrobial effects of the AZA treated (+AZA) and untreated (−AZA) supernatant of A. clavatus (L19) on the growth of S. aureus ((a): OD₆₀₀, (b): CFU/mL). Antimicrobial effects of the NO treated (+NO) and untreated (−NO) supernatant of H. koningii (NG_14 BRE-1P1) on the growth of P. aeruginosa ((c): OD₆₀₀, (d): CFU/mL). Control comprises S.aureus or P. aeruginosa grown without fungal sample. Data is presented as mean values ± standard deviations of three biological replicates performed in triplicate. ∗indicates significant difference (P < 0.05).

Figure 2

Antimicrobial activity of the pellet of P. restrictum (PRF-18) grown in the absence (−butyrate) and presence (+butyrate) of butyrate against S. aureus ((a): OD₆₀₀, (b): CFU/mL) and the MRSA strain B337919 ((c): OD₆₀₀). Control comprises S. aureus or MRSA grown without fungal samples. Data is represented as mean values ± standard deviations of three biological replicates performed in triplicate. ∗indicates significant difference (P < 0.05).

Figure 3

Antimicrobial activity against C. albicans ((a): OD₆₀₀, (b): CFU/mL) and E. dermatitidis ((c): OD₆₀₀) in the extracted pellet of P. crustosum (D_D27) grown with (+TSA) and without TSA (−TSA). Controls comprise C. albicans and E. dermatitidis grown without fungal sample. Data comprises mean values ± standard deviations of three biological replicates performed in triplicate. ∗indicates significant difference (P < 0.05).

Figure 4

Effect of the GlcNAc treated (+GlcNAc) and untreated (−GlcNAc) supernatant of P. spinulosum (Li0102XIX) on the growth of S. aureus measured by OD₆₀₀ (a) and CFU/mL (b). Control comprises S. aureus grown without fungal sample. Data are represented as mean values ± standard deviations of three biological replicates performed in triplicate. ∗indicates significant differences (P < 0.05).

Figure 5

Cytotoxic effects of the supernatant of A. clavatus (L19), the supernatant of H. koningii (NG_14 BRE-1P1), the pellet of P. restrictum (PRF-18), and the extracted pellet of P. crustosum (D_D27) incubated with (treated) and without (untreated) the respective SC using human intestinal epithelial Caco2 (a) and hepatocytic HepG2 (b) cells. Control comprises Moser medium. Values, given as % of dead cells, represent mean values ± standard deviations of three biological replicates performed in triplicate.