Penicillium expansum, a common spoilage organism in postharvest fruits, can cause fruit decay and deterioration and endanger human health. It is of great significance to investigate the antimicrobial mechanism of the antimicrobial peptide from Paenibacillus ehimensis on P. expansum spores. The antimicrobial activity of the antimicrobial peptide against P. expansum spores was determined by using the two-fold dilution method as well as measuring the time-killing curve. Scanning electron microscopy (SEM) and transmission electron microscopy (TEM) were used to evaluate the effect of the antimicrobial peptide on the ultrastructure of P. expansum spores. The effects of the antimicrobial peptide on the cell membrane and reactive oxygen species (ROS) accumulation of P. expansum were analyzed by fluorescence probes. The results showed that the minimum inhibitory concentration (MIC) of the antimicrobial peptide against P. expansum spores was 3.5 AU/mL. The spore germination rate was significantly decreased by 28.30%, 84.57% and 100% by the antimicrobial peptide at concentrations of 0.5 MIC, 1 MIC and 2 MIC compared with the blank control (P < 0.05). After treatment with the antimicrobial peptide, the spores appeared seriously sunken, the intracellular contents were leaked out, and the morphology and structure were changed. The antimicrobial peptide damaged the cell wall of P. expansum, resulting in the leakage of alkaline phosphatase. The antimicrobial peptide depolarized the cell membrane potential in a dose-dependent manner, and increased the cell membrane permeability, leading to K⁺ leakage. The fluidity of the cell membrane was increased, which in turn resulted in a significant decrease in DPH fluorescence intensity (P < 0.05). The integrity of the cell membrane was damaged by the antimicrobial peptide, so the fluorescence intensity of SYTOX-Green and the contamination rate of PI were increased. Moreover, the antimicrobial peptide at 1 MIC and 2 MIC increased the fluorescence intensity of DCFH-DA significantly (P < 0.05) and resulted in ROS accumulation, which affected the physiology and metabolism of P. expansum spores. This study indicated that the target sites of the antimicrobial peptide against P. expansum spores were mainly the cell membrane and ROS metabolism.

This study investigated the inhibitory effect of antimicrobial peptide from Paenibacillus ehimensis on the biofilms of Candida albicans by the microdilution and time-kill curve assays. Microscopic observation was conducted to assess the impact of the peptide on the formation of C. albicans germ tube and hyphae. Additionally, the 2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide (XTT) assay was employed to investigate the influence on the formation of C. albicans biofilms, pre-formed biofilms, and the clearance rate of mature biofilms. Fluorescent probes were utilized to observe the alteration in the structure of biofilms and the status of intracellular fungal entities. The number of viable cells in biofilms was measured by the spread plate method, and real-time polymerase chain reaction (PCR) was employed to determine the expression levels of genes associated with biofilm formation. The results showed that the minimum inhibitory concentration of the antimicrobial peptide against C. albicans was 8.28 AU/mL. This peptide reduced the rate of germ tube formation in C. albicans, and prevented the formation of hyphae, making it exist in the form of yeasts. Furthermore, the antimicrobial peptide influenced both the formation and eradication of biofilms and disrupted the structural integrity of biofilms after a short duration, leading to damage and even death of C. albicans, and reducing cell counts in biofilms. The antimicrobial peptide reduced the expression levels of multiple genes related to biofilm formation (such as ALS1, ALS3, HWP1, EFG1, ECE1 and UME6), therefore inhibiting biofilm formation. The above findings demonstrated that antimicrobial peptide effectively inhibited the biofilm formation, mature biofilm clearance and the expression of biofilm-forming genes in C. albicans. This study lays a theoretical foundation for the development of novel antimicrobial agents against C. albicans and provides a basis for the prevention and control of foodborne pathogen contamination to ensure food quality and safety.