Waterfield Nicholas R, Sanchez-Contreras Maria, Eleftherianos Ioannis, Dowling Andrea, Yang Guowei, Wilkinson Paul, Parkhill Julian, Thomson Nicholas, Reynolds Stuart E, Bode Helge B, Dorus Steven, Ffrench-Constant Richard H
Department of Biology and Biochemistry, University of Bath, Bath BA2 7AY, United Kingdom.
Proc Natl Acad Sci U S A. 2008 Oct 14;105(41):15967-72. doi: 10.1073/pnas.0711114105. Epub 2008 Oct 6.
Current sequence databases now contain numerous whole genome sequences of pathogenic bacteria. However, many of the predicted genes lack any functional annotation. We describe an assumption-free approach, Rapid Virulence Annotation (RVA), for the high-throughput parallel screening of genomic libraries against four different taxa: insects, nematodes, amoeba, and mammalian macrophages. These hosts represent different aspects of both the vertebrate and invertebrate immune system. Here, we apply RVA to the emerging human pathogen Photorhabdus asymbiotica using "gain of toxicity" assays of recombinant Escherichia coli clones. We describe a wealth of potential virulence loci and attribute biological function to several putative genomic islands, which may then be further characterized using conventional molecular techniques. The application of RVA to other pathogen genomes promises to ascribe biological function to otherwise uncharacterized virulence genes.
当前的序列数据库现已包含大量病原菌的全基因组序列。然而，许多预测基因缺乏任何功能注释。我们描述了一种无需假设的方法——快速毒力注释（RVA），用于针对四种不同分类群（昆虫、线虫、变形虫和哺乳动物巨噬细胞）对基因组文库进行高通量平行筛选。这些宿主代表了脊椎动物和无脊椎动物免疫系统的不同方面。在这里，我们使用重组大肠杆菌克隆的“毒性增强”试验，将RVA应用于新出现的人类病原体共生光杆菌。我们描述了大量潜在的毒力位点，并将生物学功能归因于几个假定的基因组岛，然后可以使用传统分子技术对其进行进一步表征。将RVA应用于其他病原体基因组有望为原本未表征的毒力基因赋予生物学功能。
