蓝狐源约氏乳杆菌菌株ZJBF004分离鉴定与体外益生潜能的评价

Isolation and Identification of Lactobacillus johnsonii Strain ZJBF004 from Alopex lagopus and Its Probiotic Potential Evaluation in Vitro

为获得犬科动物源乳酸菌并对其益生潜能进行评价,本研究通过测定生长曲线、产酸能力、对低pH和高胆盐环境的耐受性、在体外模拟胃肠道溶液中的存活能力、抑菌能力、抗生素抗性、对蓝狐小肠上皮原代细胞的黏附能力及抗氧化性评价了菌株ZJBF004的益生特性。结果表明:利用固体乳酸细菌培养基从蓝狐的肠道内容物中分离到1株优势乳酸菌,将其命名为菌株ZJBF004,经菌落菌体形态、生理生化特性和16S rRNA基因序列分析鉴定其为约氏乳杆菌(Lactobacillus johnsonii)。菌株ZJBF004产酸能力较强且在37℃培养时能迅速进入对数生长期;在pH=5的培养基中耐受6 h时仍能保持较好的生长情况,在胆盐浓度为3 g/L的条件下至少可耐受6 h,且存活率高于70%,在人工模拟胃液和肠液中耐受3 h时存活率高于90%;对大肠杆菌、沙门氏菌、金黄色葡萄球菌具有良好的抑菌效果;对蓝狐小肠上皮原代细胞的黏附能力为(18.67±3.12)个/细胞;1,1-二苯基-2-三硝基苯肼(DPPH)自由基清除率为28.50%,表明菌株ZJBF004具有一定的抗氧化活性。由此可见,新分离的蓝狐源约氏乳杆菌菌株ZJBF004具有优良的体外益生性能,可作为潜在犬科动物用益生菌进一步研究。

In order to obtain a canine source of lactic acid bacteria and evaluate its probiotic potential,in this study,the probiotic characteristics of strain ZJBF004 were evaluated by measuring its growth curve,acid production capacity,tolerance to low pH and high concentration of bile salt,viability in vitro simulated gastrointestinal solution,antibacterial capacity,antibiotic resistance,adhesion to primary Alopex lagopus intestinal epithelial cells and antioxidant activity.The results showed that a dominant lactic acid bacteria was isolated from Alopex lagopus’s intestinal tract by solid Lactobacillus culture medium and it was named strain ZJBF004.The strain ZJBF004 was identified as Lactobacillus johnsonii by the analysis of colony morphology,physiological and biochemical characteristics and 16S rRNA sequence analysis.The strain ZJBF004 had strong acid-producing ability and was able to enter logarithmic growth phase rapidly in 37℃;the strain ZJBF004 could maintain good growth in the medium containing with pH 5.0,70%bacteria were survived in tolerant to 3 g/L bile salt in 6 hours,and 90%bacteria were survived in simulated gastric and intestinal juices in 6 hours;the antimicrobial effect of strain ZJBF004 on pathogens which Escherichia coli,Salmonella and Staphylococcus aureus was good.Moreover,the adhesion ability of strain ZJBF004 to Alopex lagopus intestinal primary epithelial cells was(18.67±3.12)bacterias/cell,and the 1,1-diphenyl-2-trinitrophenyl hydrazine(DPPH)radical scavenge rate was 28.50%,which showed the strain ZJBF004 had a certain antioxidant activity.In conclusion,the newly isolated Lactobacillus johnsonii strain ZJBF004 has good beneficial effects in vitro and can be further studied as candidate probiotics for feeding canines.