The broad-range detection and identification of bacterial DNA from clinical specimens are a foundational approach in the practice of molecular microbiology. However, there are circumstances under which conventional testing may yield false-negative or otherwise uninterpretable results, including the presence of multiple bacterial templates or degraded nucleic acids. Here, we describe an alternative, next-generation sequencing approach for the broad range detection of bacterial DNA using broad-range 16S rRNA gene hybrid capture (“16S Capture”). The method is able to deconvolute multiple bacterial species present in a specimen, is compatible with highly fragmented templates, and can be readily implemented when the overwhelming majority of nucleic acids in a specimen derive from the human host. We find that this approach is sensitive to detecting as few as 17 Staphylococcus aureus genomes from a background of 100 ng of human DNA, providing 19- to 189-fold greater sensitivity for identifying bacterial sequences than standard shotgun metagenomic sequencing, and is able to successfully recover organisms from across the eubacterial tree of life. Application of 16S Capture to a proof-of-principle case series demonstrated its ability to identify bacterial species that were consistent with histological evidence of infection, even when diagnosis could not be established using conventional broad range bacterial detection assays. 16S Capture provides a novel means for the efficient and sensitive detection of bacteria embedded in human tissues and for specimens containing highly fragmented template DNA.

中文翻译：

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通过广泛的16S rRNA基因富集来敏感鉴定临床标本中的细菌DNA

临床样本中细菌DNA的广泛检测和鉴定是分子微生物学实践的基础方法。但是，在某些情况下，常规测试可能会产生假阴性或其他无法解释的结果，包括存在多个细菌模板或降解的核酸。在这里，我们描述了一种使用下一代16S rRNA基因杂合捕获（“ 16S Capture”）进行细菌DNA的广泛检测的下一代测序方法。该方法能够使样品中存在的多种细菌种类解卷积，与高度碎片化的模板兼容，并且当样品中绝大多数核酸来源于人宿主时都可以轻松实施。来自100 ng人类DNA的金黄色葡萄球菌基因组，与标准散弹枪宏基因组测序相比，鉴定细菌序列的灵敏度提高了19到189倍，并且能够成功地从整个真细菌树中回收生物。16S Capture在原理证明案例系列中的应用证明了其能够识别与感染的组织学证据一致的细菌种类的能力，即使无法使用常规的广泛细菌检测方法进行诊断也是如此。16S Capture为有效，灵敏地检测嵌入人体组织中的细菌以及含有高度片段化模板DNA的标本提供了一种新颖的方法。