一株海藻多糖降解菌的分离鉴定及产酶条件优化

Isolation identification and enzyme-producing conditions analysis of a seaweed polysaccharides-degrading bacteria

从大型褐藻藻体分离获得一株具有高效降解琼胶和褐藻胶能力的革兰氏阴性菌菌株ST-6。16S rDNA序列分析结果表明,该菌株与海绵假单胞菌的相似度达到99%,NJ法构建系统进化树也与海绵假单胞菌归为一类,鉴定为海绵假单胞菌Pseudomonas pachastrellae ST-6。在2216E培养基、30℃培养条件下,海绵假单胞菌ST-6的生长曲线表明,接种10~48 h为菌株的指数生长期,48~72 h为生长稳定期。产酶结果表明,菌株ST-6在指数生长期时菌液的胞外琼胶酶相对酶活力较高,在接种48 h时,菌液的琼胶酶相对酶活力最高为249.15 U/m L。此外,发现菌株ST-6的琼胶酶和褐藻胶酶的分泌类型分别为非诱导型和诱导型。采用单因素分析法对其生长和产酶条件进行分析,结果表明,海绵假单胞菌ST-6最适生长条件为:温度25~35℃,33值5~9;最适产琼胶酶条件为:温度30℃,33值为7,琼胶浓度0.3%。最适产褐藻胶酶条件为:温度35℃,33值为9。在温度30℃、339和褐藻酸钠浓度0.15%的培养条件下,海绵假单胞菌ST-6获得最高胞外褐藻胶酶相对酶活力为135.54 U/m L。

In this study,we isolated a marine agar and alginate-degrading Gram-negative bacterial strain ST-6 from brown seaweed.The results of our 16 S rDNA sequence analysis indicate that the similarity was 99% between the strain ST-6 and the strains of Pseudomonas pachastrellae.We classified these strains as a class in phylogenetic trees using the NJ method and identified the strain as P.pachastrellae ST-6.After culturing in a 2216 E medium at 30℃,we simultaneously analyzed the biomass accumulation and enzyme production of P.pachastrellae ST-6.The growth curve results indicate the exponential and stationary growth phases to be 10-48 h and 48-72 h after inoculation,respectively.The enzyme production results indicate that the enzyme activity was higher in the exponential growth phase and that the highest extracelluar agarase relative activity was 249.15 U/mL 48 h after inoculation.In addition,the secretion types of agarase and alginate lyase of P.pachastrellae ST-6 were non-inducible and inducible,respectively.The optimum growth conditions were a culture temperature of 25-35 ℃ and initial pH values of 5 and 9,respectively.The optimum agarase-producing conditions were a culture temperature of 30℃,an initial pH value of7,and an initial agar concentration of 0.30%.The optimum alginate lyase-producing conditions were a culture temperature of 35℃,an initial pH value of 9,and an alginate concentration of 0.30%.We obtained a maximum relative extracelluar alginate lyase activity of 135.54 U/mL in culture conditions with a temperature of 30℃,a pH value of 7,and an alginate concentration of 0.30%.