Endophytic Bacterial Diversity in Grapevine (Vitis vinifera L.) Leaves Described by 16S rRNA Gene Sequence Analysis and Length Heterogeneity-PCR
Endophytic Bacterial Diversity in Grapevine (Vitis vinifera L.) Leaves Described by 16S rRNA Gene Sequence Analysis and Length Heterogeneity-PCR
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DOI：
10.1007/s12275-009-0082-1
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发表时间：
2009-08-01
影响因子：
3
通讯作者：
Bianco, Piero Attilio
中科院分区：
文献类型：
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作者：
Bulgari, Daniela;Casati, Paola;Bianco, Piero Attilio
通过培养和非培养方法分析了葡萄藤叶片组织内生细菌的多样性。为了直接从宏基因组中鉴定细菌内生菌，优化了细菌富集和DNA提取的方案。16 S rRNA基因文库的序列分析强调了五个不同的操作分类单位（OTU），显示最佳序列匹配的γ-变形菌，家庭肠杆菌科，泛菌属的优势。通过培养的细菌分离揭示了六个OTU的存在，显示出与放线菌属、短杆菌属以及与厚壁菌属芽孢杆菌属和肠球菌属的最佳序列匹配。使用来自单个细菌克隆的长度异质性-PCR（LH-PCR）电泳峰来建立代表与葡萄藤组织相关联地鉴定的细菌内生菌的数据库。对健康和植原体感染的葡萄藤植株的分析表明，LH-PCR可以作为一种有用的补充工具，用于检测细菌内生菌的多样性，特别是对大量样品的多样性调查。
Diversity of bacterial endophytes associated with grapevine leaf tissues was analyzed by cultivation and cultivation-independent methods. In order to identify bacterial endophytes directly from metagenome, a protocol for bacteria enrichment and DNA extraction was optimized. Sequence analysis of 16S rRNA gene libraries underscored five diverse Operational Taxonomic Units (OTUs), showing best sequence matches with gamma-Proteobacteria, family Enterobacteriaceae, with a dominance of the genus Pantoea. Bacteria isolation through cultivation revealed the presence of six OTUs, showing best sequence matches with Actinobacteria, genus Curtobacterium, and with Firmicutes genera Bacillus and Enterococcus. Length Heterogeneity-PCR (LH-PCR) electrophoretic peaks from single bacterial clones were used to setup a database representing the bacterial endophytes identified in association with grapevine tissues. Analysis of healthy and phytoplasma-infected grapevine plants showed that LH-PCR could be a useful complementary tool for examining the diversity of bacterial endophytes especially for diversity survey on a large number of samples.