Fig 1.
Graphical abstract of anti-microbial activity of ethyl pyruvate.
Fingerprints of non-disinfected hands on blood agar containing ethyl pyruvate (middle) ethyl lactate (right) and medium (left). The microbes displayed at the circumferences of the agar plates depict a selection of microbes investigated in the present study. Plates were incubated at 37°C for 48 h.
Fig 2.
Effect of ethyl pyruvate on growth of planktonic pathobionts.
(a) Representative diagram of MIC determination for E. coli, Lactobacillus spp. and Candida spp. in 96-well plate. The final EP concentration ranging from 0.39 to 200 mM; growth control (GC), and sterility control (SC). White spots in the wells depict the growth of tested isolates. (b) Selective anti-microbial activity of EP against pathobionts, e.g. Candida spp. (n = 108), Escherichia coli (n = 8), and Lactobacillus spp. (n = 31) (***P< 0.0001).
Table 1.
Distribution of identified vaginal isolates and their MIC, MBC and MFC values.
Fig 3.
Inhibition by ethyl pyruvate of the growth of a wide panel of fungal and bacterial pathobionts.
(a) Cultures of Trichophyton mentagrophytes (left, white) and Trichophyton tonsurans I (right, yellow) were treated with different concentration of EP (0, 1 and 5 mM) for 24 hours. Cultures of Aspergillus fumigatus (b) and Candida albicans (c) were treated by EP at 0, 1, 5, and 10 concentrations (mM). (d) Minimum (Min) and maximum (Max) inhibitory concentration of EP for different bacteria, fungi and moulds (S1 Fig).
Table 2.
Effect of ethyl pyruvate on dermatophytes.
Fig 4.
Ethyl pyruvate inhibits formation and dispersion of Candida albicans biofilms.
Biofilms were grown in a 96-well tissue culture plates in the presence of ethyl pyruvate (EP) (a), ethyl lactate (EL) (b) and Amphotericin B (AmpB) (c). The corresponding MIC-values were obtained from planktonic cell studies. Fluorescence microscopic analysis revealed the effect of EP on Candida albicans biofilms formation (d-g). Non-treated biofilms showing dense yeast cells (single arrow) and hyphae (triple arrow) and the extracellular matrix (double arrow) casing the yeast cells (d). Treatment of cells by 10, 25, and 50 mM EP is shown in e, f and g, respectively. Calcofluor staining was used to generate the images at x10 magnification using fluorescence microscopy. Dispersion of matured Candida albicans biofilm by EP (h), EL (i) and AmpB (j). An inverted light microscopic analysis of non-treated pre-formed Candida albicans biofilms shows filamentous biofilms containing both hyphae and yeast cells (k). Destructed biofilm by EP (4xMIC) displays no cells and hyphae (l). Results are mean ± standard error of the mean of three independent experiments. *p<0.05; **p<0.01, ***p<0.001.
Fig 5.
Destruction by ethyl pyruvate of biofilms prepared under dynamic fluid condition.
(a) Colony forming units derivable from biofilms before and after treatment with ethyl pyruvate (EP) in silicone tubes. (b) Treatment of matured biofilms in silicone tubes by 50 mM EP and 50 mM ethyl lactate (EL) as well as Penicillin/Streptomycin (0.28/0.17 mM) as a control. Destruction of biofilm in the presence of antibacterial compounds was evaluated at different time intervals. Scanning electron microscopy clearly shows well-structured matured biofilm, containing microorganisms encased in extracellular matrix (c, d). EP treatment (50mM) displays few cells, cell debris and dissolution of extracellular matrix (e, f). Representative profile of MALDI-TOF mass spectrum of the identified Sphingomonas paucimobilis from the silicone tube biofilm (g).
Fig 6.
Growth inhibition of fluconazole-resistant and susceptible Candida albicans by ethyl pyruvate.
The growth of both resistant and non-resistant Candida isolates in the presence of glucose was inhibited by EP (a, b), whereas when gluconeogenesis was initiated with 3% glycerol/ethanol (GE) medium (c, d).