Inhibition of agronomically relevant fungal phytopathogens by tyrocidines, cyclic antimicrobial peptides isolated from Bacillus aneurinolyticus

Abstract

The tyrocidines, a complex of analogous cyclic decapeptides produced by Bacillus aneurinolyticus, exhibited noteworthy activity against a range of phytopathogenic fungi, including Fusarium verticillioides, Fusarium solani and Botrytis cinerea. The activity of the tyrocidine peptide complex (Trc mixture) and purified tyrocidines exhibited minimum inhibition concentrations below 13 mg ml "1 (~10 mM) and was significantly more potent than that of the commercial imidazole fungicide, bifonazole. Although the tyrocidines' activity was negatively influenced by the presence of Ca 2+, it remained unaffected by the presence of Mg 2+ , Na + and K + . Microscopic analysis revealed significant impact on the morphology of F. solani and Bot. cinerea including retarded germination and hyperbranching of hyphae. Studies with membrane-impermeable dyes, SYTOX green and propidium iodide suggested that the main mode of action of tyrocidines involves the disruption of fungal membrane integrity. Because of the tyrocidines' broad spectrum and potent antifungal activity, possible multiple targets reducing the risk of overt resistance and general salt tolerance, they are promising candidates that warrant further investigation as bio-fungicides.

Antifungal membranolytic activity of the tyrocidines against filamentous plant fungi

“…There is a possibility that the antifungal action of the tyrocidines may not only be the result of membranolytic activity, particularly as fungal spores treated with tyrocidines tended to form non-viable hyperbranched structures, which eludes to an alternative target or mode of action [5]. The major tyrocidines have good activity against the planktonic form of pathogenic yeast C. albicans, as well as activity to prevent its biofilms [5]. This study also indicated that membrane leakage is part of the tyrocidine mode of anti-candida action.…”

“…Lytic activity has been observed for the tyrocidines against Gram-positive bacteria [3], but not against P. falciparum and it was proposed that the tyrocidines interfere with parasite development [6]. A natural extract from tyrothricin, the tyrocidine mixture (Trc mixture) containing the six major tyrocidines, has significant antifungal activity against a broad array of filamentous fungi, many of which are plant pathogens, but also a number of human pathogens [5]. Fluorescent microscopy of fungal hyphae treated with the tyrocidines alluded to the likelihood that antifungal activity leads to membrane leakage [5].…”

“…Fluorescent microscopy of fungal hyphae treated with the tyrocidines alluded to the likelihood that antifungal activity leads to membrane leakage [5]. There is a possibility that the antifungal action of the tyrocidines may not only be the result of membranolytic activity, particularly as fungal spores treated with tyrocidines tended to form non-viable hyperbranched structures, which eludes to an alternative target or mode of action [5]. The major tyrocidines have good activity against the planktonic form of pathogenic yeast C. albicans, as well as activity to prevent its biofilms [5].…”

“…The tyrocidines and analogues exhibit a broad spectrum of low micromolar activity against Gram-positive bacteria such as the food pathogen Listeria monocytogenes [3,4], filamentous fungi such as Aspergillus fumigates [5] as well as nanomolar activity against the malaria parasite Plasmodium falciparum [6]. The tyrocidines were also found to be active against the pathogenic yeast Candida albicans [7,8], with tyrocidine A (TrcA) being synergistic with the membrane active antifungal drug, amphotericin B, in the eradication of mature C. albicans biofilms [8].…”

“…A natural extract from tyrothricin, the tyrocidine mixture (Trc mixture) containing the six major tyrocidines, has significant antifungal activity against a broad array of filamentous fungi, many of which are plant pathogens, but also a number of human pathogens [5]. Fluorescent microscopy of fungal hyphae treated with the tyrocidines alluded to the likelihood that antifungal activity leads to membrane leakage [5]. There is a possibility that the antifungal action of the tyrocidines may not only be the result of membranolytic activity, particularly as fungal spores treated with tyrocidines tended to form non-viable hyperbranched structures, which eludes to an alternative target or mode of action [5].…”