Natural Competence Is Common among Clinical Isolates of Veillonella parvula and Is Useful for Genetic Manipulation of This Key Member of the Oral Microbiome.
Natural Competence Is Common among Clinical Isolates of Veillonella parvula and Is Useful for Genetic Manipulation of This Key Member of the Oral Microbiome.
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DOI：
10.3389/fcimb.2017.00139
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发表时间：
2017
影响因子：
5.7
通讯作者：
Merritt J
中科院分区：
文献类型：
--
作者：
Knapp S;Brodal C;Peterson J;Qi F;Kreth J;Merritt J
在人类口腔中发现的六种细孔菌是口腔菌群中最丰富的成员之一，发生在龈上和龈下牙菌斑以及口腔黏膜上。流行病学数据也表明，这些物种与最常见口腔疾病的发生有关。尽管它们无处不在，数量丰富，具有重要的生态意义，但令人惊讶的是，人们对细孔菌的生物学知之甚少，这主要是由于与它们的基因操作相关的困难。为了改进微细孔菌的遗传分析，我们从临床斑块样本中分离了一组微细孔菌，并使用新开发的转化方案筛选自然能力。许多小苗弧菌菌株表现出受培养基组成高度影响的自然能力。通过利用这种能力，我们能够利用克隆无关的等位基因交换诱变来确定与II型分泌系统（T2SS）同源的基因座内DNA摄取机制的可能来源。有趣的是，我们发现小叶弧菌的自然能力对不同来源的DNA表现出明显的偏好等级（质粒< PCR产物<基因组DNA），这与大多数自然能力物种不同。基因组与其他成员的比较表明，自然能力很可能广泛分布在这个群体中。据我们所知，本研究首次在整个细孔菌科中展示了自然能力和靶向等位基因交换诱变，为细孔菌遗传学研究提供了一种简单快速的方法。
The six Veillonella species found in the human oral cavity are among the most abundant members of the oral flora, occurring in both supra- and subgingival dental plaque as well as on the oral mucosa. Epidemiological data have also implicated these species in the development of the most common oral diseases. Despite their ubiquity, abundance, and ecological significance, surprisingly little is known about Veillonella biology, largely due to the difficulties associated with their genetic manipulation. In an effort to improve genetic analyses of Veillonella species, we isolated a collection of veillonellae from clinical plaque samples and screened for natural competence using a newly developed transformation protocol. Numerous strains of V. parvula were found to exhibit a natural competence ability that was highly influenced by growth medium composition. By exploiting this ability, we were able to utilize cloning-independent allelic exchange mutagenesis to identify the likely source of DNA uptake machinery within a locus homologous to type II secretion systems (T2SS). Interestingly, V. parvula natural competence was found to exhibit a clear hierarchy of preference for different sources of DNA (plasmid < PCR product < genomic DNA), which is unlike most naturally competent species. Genomic comparisons with other members of the Veillonellaceae family suggest that natural competence is likely to be widely distributed within this group. To the best of our knowledge, this study is the first demonstration of natural competence and targeted allelic exchange mutagenesis within the entire Veillonellaceae family and demonstrates a simple and rapid method to study Veillonella genetics.