The antimicrobial peptide BCE3 isolated from fermented Penaeus vannamei: inhibition mechanism on Bacillus cereus and application in rice

Abstract:

[Objective] To study the inhibitory effect and mechanism of a novel antimicrobial peptide (BCE3) isolated from Penaeus vannamei processing waste fermented with Bacillus subtilis against Bacillus cereus. [Methods] The small peptide sequences in the fermentation broth were identified by ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS), and the potential antimicrobial peptides were screened by bioinformatics. The minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC), and time-kill curve of BCE3 against B. cereus were determined by the microdilution method and plate colony counting method. Then, the alkaline phosphatase release assay, propidium iodide (PI) staining, nucleic acid and protein leakage assays, and flow cytometry were employed to examine the effects of BCE3 on the cell wall and cell membrane of B. cereus. The effect of BCE3 on bacterial DNA was explored by the gel retardation assay, fluorescence spectroscopy, and molecular docking. Finally, the antimicrobial effects of BCE3 in rice were evaluated by the colony counting method. [Results] The potential antimicrobial peptide BCE3 screened out showed the MIC of 62.5 μg/mL and MBC of 125.0 μg/mL against B. cereus. The time-kill curve revealed that BCE3 reduced the bacterial count by 86.0% within 3 h (62.5 μg/mL), outperforming nisin. BCE3 caused damage to the bacterial cell wall and membrane, leading to the leakage of cell contents. Moreover, it can bind with DNA to kill the bacteria. In addition, BCE3 (125.0 μg/mL) exerted a significant inhibitory effect on the growth of B. cereus in rice. [Conclusion] BCE3 inhibits B. cereus by altering the permeability of the cell membrane and binding to DNA, thus leading to bacterial death. These findings provide a theoretical basis for application of BCE3 in the control of B. cereus.

Key words:Penaeus vannamei processing waste;antimicrobial peptide BCE3;Bacillus cereus;antimicrobial mechanism