Synergistic action of synthetic peptides and amphotericin B causes disruption of the plasma membrane and cell wall in Candida albicans.

The objective of this work was to evaluate the combination of synthetic peptides based on the γ-core motif of defensin PvD1 with amphotericin B (AmB) at different concentrations against Candida albicans. We applied the checkerboard assay using different concentrations of the commercial drug AmB and the synthetic peptides γ31-45PvD1++ and γ33-41PvD1++ against C. albicans, aiming to find combinations with synergistic interactions. Between these two interactions involving γ31-45PvD1++ and AmB, an additive effect was observed. One such interaction occurred at concentrations of 0.009 µM of peptide γ31-45PvD1++ and 13.23 µM of AmB and another condition of 0.019 µM of peptide γ31-45PvD1++ and 6.61 µM of AmB. The other two concentrations of the interaction showed a synergistic effect in the combination of synthetic peptide γ31-45PvD1++ and AmB, where the concentrations were 1.40 µM peptide γ31-45PvD1++ and 0.004 µM AmB and 0.70 µM γ31-45PvD1++ peptide and 0.002 µM AmB. We proceeded with analysis of the mechanism of action involving synergistic effects. This examination unveiled a range of impactful outcomes, including the impairment of mitochondrial functionality, compromise of cell wall integrity, DNA degradation, and a consequential decline in cell viability. We also observed that both synergistic combinations were capable of causing damage to the plasma membrane and cell wall, causing leakage of intracellular components. This discovery demonstrates for the first time that the synergistic combinations found between the synthetic peptide γ31-45PvD1++ and AmB have an antifungal effect against C. albicans, acting on the integrity of the plasma membrane and cell wall.

Potent Anti-Candida Fraction Isolated from Capsicum chinense Fruits Contains an Antimicrobial Peptide That is Similar to Plant Defensin and is Able to Inhibit the Activity of Different α-Amylase Enzymes.

Antimicrobial peptides (AMPs) are molecules present in several life forms, possess broad-spectrum of inhibitory activity against pathogenic microorganisms, and are a promising alternative to combat the multidrug resistant pathogens. The aim of this work was to identify and characterize AMPs from Capsicum chinense fruits and to evaluate their inhibitory activities against yeasts of the genus Candida and α-amylases. Initially, after protein extraction from fruits, the extract was submitted to anion exchange chromatography resulting two fractions. Fraction D1 was further fractionated by molecular exclusion chromatography, and three fractions were obtained. These fractions showed low molecular mass peptides, and in fraction F3, only two protein bands of approximately 6.5 kDa were observed. Through mass spectrometry, we identified that the lowest molecular mass protein band of fraction F3 showed similarity with AMPs from plant defensin family. We named this peptide CcDef3 (Capsicum chinense defensin 3). The antifungal activity of these fractions was analyzed against yeasts of the genus Candida. At 200 µg/mL, fraction F1 inhibited the growth of C. tropicalis by 26%, fraction F2 inhibited 35% of the growth of C. buinensis, and fraction F3 inhibited all tested yeasts, exhibiting greater inhibition activity on the growth of the yeast C. albicans (86%) followed by C. buinensis (69%) and C. tropicalis (21%). Fractions F1 and F2 promoted membrane permeabilization of all tested yeasts and increased the endogenous induction of reactive oxygen species (ROS) in C. buinensis and C. tropicalis, respectively. We also observed that fraction F3 at a concentration of 50 µg/mL inhibited the α-amylase activities of Tenebrio molitor larvae by 96% and human salivary by 100%. Thus, our results show that fraction F3, which contains CcDef3, is a very promising protein fraction because it has antifungal potential and is able to inhibit the activity of different α-amylase enzymes.

Coffea canephora Peptides in Combinatorial Treatment with Fluconazole: Antimicrobial Activity against Phytopathogenic Fungus.

The objective of the present study was to evaluate the antimicrobial activity of the Cc-LTP2 and Cc-GRP peptides isolated from Coffea canephora seeds and their possible synergistic activity with the azole drug fluconazole and characterize their mechanisms of action on cells of pathogenic fungi. Cc-LTP2 and Cc-GRP alone or in combination with 20 µg/mL of fluconazole were evaluated for their antimicrobial activity on the fungus Fusarium solani, and the effects of these peptides on the permeability of membranes and the induction of oxidative stress were determined. Our results show that these peptides at a concentration of 400 µg/mL combined with 20 µg/mL of fluconazole were able to inhibit the growth of the tested fungi, promote changes in their growth pattern, permeabilize the membrane, and induce reactive oxygen species (ROS). Some of these results were also observed with the peptides alone or with fluconazole alone, suggesting that the peptides act synergistically, promoting the potentiation of antimicrobial action. In this study, it was shown that Cc-LTP2 and Cc-GRP in combination with fluconazole were able to inhibit the growth of the fungus F. solani, to promote permeabilization of its membrane, and to induce the production of ROS, suggesting a combinatorial activity between the peptides and fluconazole.