自然青贮多花黑麦草优良乳酸菌的筛选及对多花黑麦草青贮品质的影响

为了筛选出适用于多花黑麦草青贮发酵的优良乳酸菌菌株，从自然青贮多花黑麦草中分离乳酸菌，筛选并鉴定生长快速且产酸效率高的菌株，分析其生理生化特征以及对多花黑麦草青贮品质的影响。利用传统的平板培养法，从自然青贮多花黑麦草中分离获得180株乳酸菌。其中，5株乳酸菌生长快速且产酸能力强，均能够在NaCl浓度为3.0%，温度为15~35 ℃，pH为3.5~7.0的MRS液体培养基中良好生长。经16S rRNA基因序列比对鉴定，菌株PR_LAB_9、PR_LAB_34、PR_LAB_67和PR_LAB_86为植物乳杆菌，PR_LAB_76为戊糖片球菌。对菌株进行生长曲线和产酸性能测定，该5株乳酸菌均能够在12 h内快速繁殖和产酸，菌株PR_LAB_76在24 h内生长速率和产酸效率均优于其他4株乳酸菌。进一步分析添加该5株乳酸菌对多花黑麦草青贮发酵品质、营养成分、微生物数量的影响，发现该5株乳酸菌均能提高多花黑麦草的青贮品质，包括显著降低青贮料pH和氨态氮含量，减少干物质损失，提高可溶性碳水化合物含量。其中，菌株PR_LAB_76处理组多花黑麦草青贮品质最优，干物质含量、粗蛋白含量、乳酸含量、乳酸菌数量显著高于其他4株乳酸菌处理组，且pH值、酵母菌数量显著低于其他4株乳酸菌处理组。综上，戊糖片球菌PR_LAB_76可作为多花黑麦草青贮的备选菌株。

Screening of superior lactic acid bacteria from natural Lolium multiflorum silage and their effects on silage quality

The aim of this study was to isolate superior lactic acid bacteria （LAB） strains suitable for Italian ryegrass silage fermentation. LAB were isolated from naturally fermenting Italian ryegrass and strains with rapid growth rate and high acid production efficiency were identified and selected. Subsequently the physiological and biochemical characteristics of these strains were analyzed and we then investigated the influence of these strains on the quality of Italian ryegrass silage. Using the traditional plate culture method， 180 strains of LAB were isolated from naturally fermenting Italian ryegrass silage in this study. Among them， five strains of LAB grew rapidly and had strong acid production ability. These five strains grew well in MRS liquid medium with a NaCl concentration of 3.0%， temperature of 15-35 ℃ and pH of 3.5-7.0. According to 16S rRNA gene sequence alignment and identification， four of the strains （designated PR_LAB_9， PR_LAB_34， PR_LAB_67 and PR_LAB_86） were Lactobacillus plantarum， and a fifth （PR_LAB_76） was Pediococcus pentosaceus. The growth curve and acid production ability of these five strains were determined. All these five LAB strains were able to reproduce and produce acid in 12 hours， and strain PR_LAB_76 had faster growth rate and better acid production efficiency in 24 h than the other four LAB strains. Fermentation quality， nutrient composition and microbial counts of Italian ryegrass ensiled without or with the five LAB strains were also investigated in the present study. All five LAB strains improved the quality of Italian ryegrass silage， including significant reduction in the pH， ammonia nitrogen content and dry matter loss， and increase in the water soluble carbohydrate concentration of ensiled forage. The dry matter content， crude protein content， lactic acid content and LAB counts in the group treated with PR_LAB_76 were significantly higher， and the pH value and yeast counts in the PR_LAB_76 treated silage were significantly lower than those of the four groups treated with other LAB strains. In conclusion， P. pentosaceus PR_LAB_76 was identified in this study as a superior inoculum for industry use in the production of Italian ryegrass silage.