关键词: 大菱鲆肠道, 广谱拮抗性乳酸菌, 筛选与鉴定, 细菌素

Abstract: Strain LP1-4, with strong antagonistic activity against both Gram-negative bacteria and Gram-positive bacteria, was screened out of lactic acid bacteria (LAB) isolated from the turbot intestine using the Oxford cup method. LP1-4 was identified as Lactobacillus plantarum according to physiological and biochemical characteristics and 16S rRNA sequence analysis. The antimicrobial substances in the cell-free supernatant (CFS) of LP1-4 were heat stable and effective in the pH range of 2.5–5.0, but were sensitive to protease. These substances were identified as bacteriocins. The antibacterial activity of LP1-4 reached the peak after 24 h of culture, and the diameter of inhibition zone of the ethyl acetate extract was 25.42 mm. A bacteriocin with molecular mass less than 1.0 ku was purified from the extract by ultrafiltration using Labscale TFF system and Sephadex G-25 column chromatography. The retention time of the target peak in high performance liquid chromatography (HPLC) was 5.0 min. The molecular mass of the bacteriocin was estimated to be 0.854 ku by matrix-assisted laser desorption ionisation/time-of-flight/time-of-flight mass spectrometry (MALDI-TOF/TOF MS). The amino acid sequence of this bacteriocin was Glu-Asn-Lys-Pro-Ala-Ala-Pro-Lys. In conclusion, LP1-4 has potential applications in controlling spoilage and pathogenic bacteria in aquatic products as a biological preservative.

Key words: turbot intestine, broad-spectrum antagonistic lactic acid bacteria, screening and identification, bacteriocin