贵州兴义喀斯特洞穴土可培养细菌多样性及其产蛋白酶、淀粉酶活性筛选

Studies on diversity of culturable bacteria in karst cave soil of Xingyi, Guizhou and their protease and amylase activities

摘要:

【目的】进一步了解贵州喀斯特洞穴土可培养细菌的物种多样性组成及其产蛋白酶、淀粉酶生物活性能力。【方法】选取11 种分离培养基,利用稀释直接涂布平板法对贵州黔西南兴义市喀斯特地区白碗窑镇魔家大溶洞洞内土壤进行可培养细菌分离;利用两种鉴定培养基对相关细菌进行生物活性判定。【结果】根据16S rRNA 基因序列的系统进化分析,将分离得到的217 株细菌分别归类到24 个属的63 个不同种类,其中红球菌属(Rhodococcus)和链霉菌属(Streptomyces)为该洞内土壤可培养细菌的优势菌群,分别占24.42%和21.66%。大多数菌株与已知典型菌株的16S rRNA 基因序列相似性为97.90%-99.99%,其中至少有4 株菌株(D3T01、D911、D961 和D502)为潜在的新分类单元。对217 株细菌进行蛋白酶和淀粉酶活性筛选,其中具有蛋白酶或淀粉酶活性的99 株,占分离菌株的45.62%,分别属于18 个属的38 个不同种;同时具有蛋白酶和淀粉酶活性的36 株,占具有酶活性菌株的36.36%,占分离菌株的16.59%。【结论】贵州兴义喀斯特洞穴土中存在丰富多样的细菌类群,且蕴藏着一定数量的潜在新物种资源;此外功能酶菌株在喀斯特洞穴土壤中大量存在,为工业应用奠定了资源基础,极具进一步发掘和研究的价值。

Abstract:

[Objective] The main focus for the current study is to investigate the diversity of culturable bacteria in soils of karst caves located in Xingyi county of Guizhou province and to analyze their potential for production of protease and amylase. [Methods] Isolation of the culturable bacteria was done by spread plate technique using 11 synthetic media. Phylogenetic relationship of the isolates based on their 16S rRNA gene sequences was analyzed. The isolates were then screened for production of protease and amylase using plate method. [Results] Based on the 16S rRNA gene sequences, a total of 217 isolates affiliated to 63 species of 24 genera were determined, of which the genera Rhodococcus and Streptomyces were the dominant bacteria (comprising 24.42% and 21.66% respectively). Majority of these strains had 16S rRNA gene sequence similarities ranging between 97.90% and 99.99% with related type strains, except the strains D3T01, D911, D961 and D502 which are potential new taxa. Of these culturable bacteria isolated, 99 strains belonging to 38 species of 18 genera were positive for protease and/or amylase activities. Among these, 36 strains had both amylase and protease activities, thus constituting about 36.36% of the bioactive strains and 16.59% of the total isolates. [Conclusion] The results indicated that soil of karst caves in southwest of Guizhou are promising sources for isolation of many bioactive strains, with potential for further exploration and research.