獭兔肠黏膜乳杆菌的分离鉴定及其体外益生特性研究

Identification and in vitro probiotic potential of Lactobacillus strains isolated from small intestinal mucosa of Rex rabbit

【目的】分离筛选适于在獭兔肠道中存活并定殖的高黏附性乳酸菌株。【方法】通过乳酸菌选择性培养基从5和10日龄哺乳獭兔肠黏膜中分离具有抑制大肠杆菌活性的菌株,经16SrRNA基因序列分析进行鉴定,并检测其耐人工胃液、耐胆盐及黏附特性、表面疏水性等益生特性。【结果】分离筛选出3株具有较强抑制大肠杆菌活性的菌株,分别鉴定为短乳杆菌(L1)、植物乳杆菌(L2)、干酪乳杆菌(L3),其抑菌活性大小为L3>L2>L1。3株菌株均能耐受pH=4.0的人工胃液;在pH=3.0人工胃液中,L2和L3的活菌含量未出现明显变化(P>0.05),而L1的活菌含量显著下降(P<0.05);3株菌株在pH=2.0和pH=1.8人工胃液中的活菌含量均显著降低(P<0.01)。L1和L2均能耐受2.0g/L胆盐;虽然L1和L2在3.0g/L胆盐中的活菌量出现极显著(P<0.01)和显著(P<0.05)下降,但二者仍保留较高活菌含量;胆盐对L3的存活有明显抑制作用,随着胆盐含量的升高,L3的活菌含量极显著降低(P<0.01)。L2和L3对黏蛋白的黏附率无显著差异(P>0.05),均显著高于L1(P<0.05)。L3的表面疏水性最高,L2次之,二者之间无显著差异(P>0.05);L1的表面疏水性低于L2(P=0.092),显著低于L3(P<0.05)。【结论】分离所获得的3株乳杆菌均具备益生菌特性,其中以植物乳杆菌L2最佳。

[Objective] This study aimed at isolating and screening Lactobacillus strains with high adhesion rate from intestinal tracts of Rex rabbit, [Method] Lactobacillus strains that could inhibit Esche- richla coli were isolated and screened from small intestinal mueosa of 5 and 10 day-old Rex rabbits using Lactobacillus selective medium. They were identified by 16S rRNA sequence analysis, and the probiotic properties including tolerances to artificial gastric juice and bile salts, adhesion rates to small intestinal mu- cin,and hydrophobicity were evaluated, fResultsl Three Lactobacillus strains that could inhibit E. coli K88 were isolated and screened, and were identified as Lactobacillus brevis （L1） ,Lactobacillus plantarum （L2） and Lactobacillus casei （L3）. Their antimicrobial properties were in the order of L3〉L2〉L1. The three strains showed great resistance to simulated gastric juice with pH of 4.0. In the simulated gastric juice with pH of 3.0,the viable counts of L2 and L3 appeared insignificant changes （P〈0. 05）, while that of L1 dropped significantly （P〈0.05）. The viable counts of all strains decreased extremely significantly （P〈0.01） when pH decreased to 2.0 and 1.8. L1 and L2 showed great resistance to 2.0 g/L bile salt. Although the viable counts of L1 and L2 showed extremely significant （P〈0.01） and significant （P〈0.05） reduction in 3.0 g/L bile salt,both of them remained high viable bacterium counts. Bile salts showed inhibitory effect against L2. The viable counts of L3 reduced extremely significantly （P〈0.01） with the increase of bile salts content. There were no significant differences between adhesion rates of L2 and L3 （P〉0.05）, and adhesion rates of L2 and L3 were significantly higher than that of L1 （P〈0.05）. L3 showed the high-est hydrophobicity, followed by L2 without significant difference （P〉0.05）. L1 had significantly lower hydrophobicity than the other two strains （P〈0.05）. [Conclusion] All the three obtained strains had pro-biotic properties and L2 was the best.