宜春富硒温泉中产蛋白酶嗜热菌株的筛选与鉴定

Isolation and identification of producing-protease thermophiles from hot springs se-enriched in Yichun

目的:从宜春富硒温泉古井水中筛选分离产蛋白酶嗜热菌,并进行初步的分类鉴定。方法:通过脱脂牛奶平板,55℃高温培养的方法筛选产蛋白酶嗜热菌。通过形态观察、生理生化鉴定和生长特性的研究,以及菌株16S r RNA基因序列的分析,确定其种属和系统进化关系。结果:从水样中分离筛选到23株产蛋白酶嗜热菌,生长特性实验表明,这23株菌株生长p H范围为p H5.0~9.0,生长的温度范围25~65℃,除N27外,其余菌株最适生长温度为45~55℃。通过生理生化和16S r RNA基因序列分析,结果表明:这些菌株分别属于厌氧芽孢杆菌属(Anoxybacillus)、硫胺素芽孢杆菌属(Aneurinibacillus)、芽孢杆菌属(Bacillus)和短芽孢杆菌属(Brevibacillus)4个属。其中地衣芽孢杆菌(Bacillus licheniformis)2株,波斯坦芽孢杆菌为(Brevibacillus borstelensis)4株。7株厌氧芽孢杆菌属(Anoxybacillus)菌株中,N9、N13、N18、N19、N28初步鉴定为Anoxybacillus rupiensis,而N2、N12初步鉴定为Anoxybacillus sp,剩余的10株(除N8外)均嗜热气硫胺素芽孢杆菌(Aneurinibacillus thermoaerophilus),占全部分离菌株总数的39.1%,为宜春富硒温泉水样中产蛋白酶嗜热菌的优势菌群。结论:获得4个属产蛋白酶嗜热菌株,丰富了产蛋白酶菌株的资源。

Objective： Screening and isolating producing-protease thermophiles from hot springs se-enriched in Yichun and taken the species classification and identification. Methods： By Skim milk plates and 55 ℃,the producing-protease thermophiles were isolated and taken the morphological observation,physiological and biochemical characterististics,growth characteristics,and the sequence analysis based on 16 Sr RNA gene,to identify their species and phylogenetic relationships.Results： 23 strains of producing-protease thermophiles were isolated from the sample.The growth characteristics experiment of23 strains of producing-protease thermophiles show that the p H range for growth was 5.0～9.0,the temperature range for growth was 25～65 ℃,optimum at 45～55 ℃（ except N27）.The physiological and biochemical and 16 S r RNA gene sequence analysis showed that these strains belong to Anoxybacillus,Aneurinibacillus,Bacillus and Brevibacillus 4 genera. Among them,2 strains Bacillus licheniformis,4 strains Brevibacillus,borstelensis）.in the 7 strains Anoxybacillus,N9、N13、N18、N19、N28 were identified as Anoxybacillus rupiensis,and N2,N12 as Anoxybacillus sp,The remaining 10 strains（ except N8） were Aneurinibacillus thermoaerophilus,accounting for 39.1% of the total number of all isolated strains. It could be regarded as an dominant group of producing-protease thermophiles in the hot spring. Conclusion： Four genera of producing-protease thermophiles were obtained,which enriched the diversity of producing-protease thermophiles.