南海东海岛沉积物分离细菌的鉴定及其抑菌活性

Diversity and antimicrobial activities of cultivable bacteria isolated from Donghai island

摘要:

[背景] 海洋微生物在活性物质开发方面具有巨大的应用前景，而目前有关南海东海岛微生物的研究鲜少。[方法] 对从东海岛沉积物中分离纯化的海洋细菌，采用形态学观察、生理生化以及16S rRNA基因序列的系统发育分析方法进行鉴定；以大肠杆菌（Escherichia coli）、枯草芽孢杆菌（Bacillus subtilis）和金黄色葡萄球菌（Staphylococcus aureus）作为指示菌，测定其抑菌活性；对具有抑菌活性的菌株扩增聚酮合酶（Polyketide synthase I，PKSI）基因，并与已知的PKSI氨基酸序列比对；选择具有PKSI基因的代表菌株，检测菌株及其发酵抑菌物的稳定性。[结果] 分离纯化到25株海洋细菌，分属于不动杆菌属（Acinetobacter）、交替单胞菌属（Alteromonas）、芽孢杆菌属（Bacillus）、嗜冷杆菌属（Psychrobacter）、假交替单胞菌属（Pseudo-alteromonas）、海洋单胞菌属（Oceanimonas）、葡萄球菌属（Staphylococcus）、微球菌属（Micrococcus）和海杆菌属（Marinobacter）。12株菌株通过基因筛选检测到PKSI编码基因，其中6株菌株具有抑菌活性和PKSI编码基因，并分属于芽孢杆菌属和交替单胞菌属；PKSI氨基酸序列同源性分析推测菌株DHD-15和DHD-a可能产生新的I型聚酮合酶结构。菌株DHD-15和DHD-L生长温度范围为15-40℃，可耐受10% NaCl高盐以及pH 3和pH 11的酸碱条件，但不耐高温；菌株DHD-15产生的抑菌物质可耐受100℃和pH 11的高温碱性条件，在50℃、pH 9条件下制备和室温保藏条件下抑菌活性较高，其稳定性较好。[结论] 南海东海岛沉积物筛选的细菌种具有抑菌活性，具有产聚酮类活性物质的潜力。

Abstract:

[Background] Marine microorganisms have a great application potential in developing bioactive substances, but few researches reported so far on microorganisms in the east island of the south China sea. [Methods] Marine culturable bacteria isolated and purified from sediments from Donghai island were identified by morphological observation, physiological and biochemical analysis, and phylogenetic analysis of 16S rRNA gene sequence. Escherichia coli, Bacillus subtilis and Staphylococcus aureus were used as indicators to determine their antibacterial activities. Based on the study on bacteriostasis of sediment bacteria, Polyketide synthase I (PKSI) gene was screened for strains with bacteriostasis activity, and amino acid sequence alignment of PKSI positive bacteria was performed. PKSI positive representative strain was selected to test the stability of the strain and its fermentation bacteriostasis. [Results] In total 25 bacteria were isolated and purified, belonging to 9 genera: Acinetobacter, Alteromonas, Bacillus, Psychrobacter, Pseudoalteromonas, Oceanimonas, Staphylococcus, Micrococcus and Marinobacter. Through the screening of bacteriostatic activity and PKSI gene, 6 strains had bacteriostatic activity, and their bacteriostatic substances all contained ketone compounds. It was speculated that DHD-15 and DHD-a might contain novel structure of type I polyketide synthase. DHD-15 and DHD-L can grow normally between 15 and 40 ℃, and grow best at 1% salt concentration and pH between 4 and 6 in seawater. [Conclusion] There are relatively rich culture-able bacterial resources in the sediments of Donghai island, south China sea.