Division of Soil and Water Management, Katholieke Universiteit Leuven, Leuven, Belgium.
FEMS Microbiol Ecol. 2011 Apr;76(1):14-25. doi: 10.1111/j.1574-6941.2010.01028.x. Epub 2011 Jan 12.
Real-time PCR and PCR-denaturing gradient gel electrophoresis (DGGE) approaches that specifically target the Variovorax 16S rRNA gene were developed to estimate the number and diversity of Variovorax in environmental ecosystems. PCR primers suitable for both methods were selected as such that the enclosed sequence showed maximum polymorphism. PCR specificity was maximized by combining PCR with a targeted endonuclease treatment of template DNA to eliminate 16S rRNA genes of the closely related Acidovorax. DGGE allowed the grouping of PCR amplicons according to the phylogenetic grouping within the genus Variovorax. The toolbox was used to assess the Variovorax community dynamics in agricultural soil microcosms (SMs) exposed to the phenylurea herbicide linuron. Exposure to linuron resulted in an increased abundance within the Variovorax community of a subgroup previously linked to linuron degradation through cultivation-dependent isolation. SMs that were treated only once with linuron reverted to the initial community composition 70 days after linuron exposure. In contrast, SMs irrigated with linuron on a long-term base showed a significant increase in Variovorax number after 70 days. Our data support the hypothesis that the genus Variovorax is involved in linuron degradation in linuron-treated agricultural soils.
开发了针对 Vairovrrax 16S rRNA 基因的实时 PCR 和 PCR-变性梯度凝胶电泳（DGGE）方法，以估计环境生态系统中 Vairovrrax 的数量和多样性。选择了适合这两种方法的 PCR 引物，因为封闭序列显示出最大的多态性。通过将 PCR 与靶向内切酶处理模板 DNA 相结合，可以最大限度地提高 PCR 的特异性，从而消除与 Acidovorax 密切相关的 16S rRNA 基因。DGGE 允许根据 Variovorax 属内的系统发育分组对 PCR 扩增子进行分组。该工具包用于评估暴露于苯脲除草剂利谷隆的农业土壤微宇宙（SM）中的 Variovorax 群落动态。暴露于利谷隆会导致与利谷隆降解有关的亚群在 Variovorax 群落中的丰度增加，这是通过培养依赖性分离得到的。仅用利谷隆处理一次的 SM 在利谷隆暴露 70 天后恢复到初始群落组成。相比之下，用利谷隆长期灌溉的 SM 在 70 天后 Variovorax 的数量显著增加。我们的数据支持这样一种假设，即 Variovorax 属参与了利谷隆处理的农业土壤中利谷隆的降解。
