High-throughput fluorescence screening assay for the identification and comparison of antimicrobial peptides’ activity on various yeast species

Variations in yeast plasma‐membrane lipid composition affect killing activity of three families of insect antifungal peptides

“…b). In contrast to our previous studies (Kodedová & Sychrová, , ), where we used the C. parapsilosis CBS 604 strain, which, like the C. glabrata DSY565 strain, was markedly less susceptible to the fungicidal effect of peptides than other Candida species, we found the C. parapsilosis CP69 strain to be considerably more sensitive to the action of the peptides (Fig. a).…”

“…The rate of uptake of the fluorescence probe diS‐C 3 (3) in control cells of different species (Figs a and ) was primarily caused by their different membrane potentials, permeability of cell surface structures for the probe and presence of MDR pumps that export the probe from the cells. The diS‐C 3 (3) fluorescence method is very good for studying the interaction of antifungal peptides with the yeast plasma membrane because their action is accompanied by a loss of plasma membrane potential, documented by dramatic changes in the staining level (position of the emission maximum λ max and intensity I max ) of yeast cells with this potentiometric fluorescence probe (Kodedová & Sychrová, , ). Mild damage to the plasma membrane is exhibited as a slow and gradual increase in λ max (a relative hyperpolarisation of the plasma membrane) in the diS‐C 3 (3) assay, and the cells are able to recover from this nonlethal damage.…”

“…Mild damage to the plasma membrane is exhibited as a slow and gradual increase in λ max (a relative hyperpolarisation of the plasma membrane) in the diS‐C 3 (3) assay, and the cells are able to recover from this nonlethal damage. On the other hand, a rapid shift of the position of the emission maximum λ max to a level near 580 nm is typical for permeabilised and dead cells (Kodedová & Sychrová, ). All representative peptides of the three structural families effectively permeabilised the cells of six pathogenic Candida species, as well as S. cerevisiae , but their killing activity was dependent of concentration and exposure time.…”

“…). The observed generally lower levels of diS‐C 3 (3) staining of the DSY565 strain than those of the cdr1 Δ mutant are a consequence of fluorescence‐probe efflux via Cdr1 (Kodedová & Sychrová, ). Nevertheless, the peptides 0.7 μM HAL‐I and 1 μM HYL‐23 caused an identical increase in the diS‐C 3 (3) staining of DSY565 and DSY1041 strains (highlighted by colour arrows in Fig.…”

Characterization, Preparation, and Purification of Marine Bioactive Peptides

“…The agar diffusion technique is usually used for determining the minimum inhibitory concentration in solid media [81]. Furthermore, there are some other assays to evaluate the antimicrobial activity like the disc diffusion assay [82], broth dilution [83], high throughput fluorescence screening assay [84], and so forth. The growing problem of resistance to conventional antibiotics and the need for new antibiotics has stimulated interest in the development of antimicrobial peptides as human therapeutics [72].…”