四川传统烟区烤烟叶面可培养细菌的遗传多样性分析

Genetic diversity of cultural phyllospheric bacteria in traditional tobacco cultivating areas of Sichuan province

摘要:

为认识烤烟(Flue-cured tobaccos)叶面可培养细菌的多样性，挖掘叶面可培养细菌资源.本研究以四川省各传统烟区旺长期烤烟叶片为材料，通过纯培养法及测定菌株的产吲哚乙酸(IAA)能力、溶磷溶钾特性、纤维素降解活性、拮抗病原菌等，并通过反转录重复因子扩增(BOXA1R-PCR)技术、16S rRNA基因测序和系统发育分析研究叶面可培养细菌的遗传多样性和功能特征.结果表明：分离得到的86株叶面细菌中有12株菌株(占总分离菌株的13.95%)具有产IAA的能力，4株产量较高(>57 μg/mL)；有9株(占总分离菌株的10.47%)表现溶磷活性，8株(占总分离菌株的9.30%)表现溶钾活性.基于BOXA1R-PCR图谱选取12株代表菌株进行16S rRNA基因序列测定，系统发育分析显示，86株菌株分别属于节杆菌属( Arthrobacter )、短小芽孢杆菌属( Lysinibacillus )、寡养单胞菌属( Stenotrophomonas )、无色杆菌属( Achromobacter )和假单胞菌属( Pseudomona )，其中以假单胞菌属( Pseudomonas )为优势菌属.表明四川传统烟区烤烟叶面存在着丰富的细菌资源.

Abstract:

To study the diversity of phyllospheric bacteria cultured in flue-cured tobacco leaves, and explore the resource of phyllospheric bacteria, this research collected flue-cured tobacco leaves in their fast growing stage from different traditional cultivation areas of Sichuan province, China. Phyllospheric bacteria strains were isolated by pure culture method, and their genetic diversity and functional characteristics were studied via determination of strains of indole 3-acetic acid (IAA) production ability, phosphorus and potassium solubilizing activities, cellulose degradation activity and antibacterial ability, as well as by transcription factor amplification technology (BOXA1R-PCR), 16S rRNA gene sequencing and phylogeny analysis. The results showed that among the 86 isolates, 13.95% (12/86) of the strains had the ability to produce IAA, 4of them had a higher yield (>57μg/mL); 10.47% (9/86) had phosphate solubilizing activities; 9.30% (8/86) showed solubilizing potassium activities. Based on the BOXA1R-PCR patterns, 12 representative strains were selected for 16S rRNA gene sequencing. Phylogenetic analysis based on the 16S rRNA gene sequencing showed that the isolates belonged to Arthrobacter, Lysinibacillus, Stenotrophomonas, Achromobacter, and Pseudomona, in which Pseudomona was the dominant genus. All results indicated that abundant culturable phyllospheric bacteria existed in flue-cured tobacco leaves in traditional tobacco areas in Sichuan province.