Metagenomic identification of KPC-Klebsiella pneumoniae, and vancomycin resistant Enterococcus faecium, for surveillance and outbreak investigation. Clinical metagenomics of patient faecal sample

Culture-independent methods have shown promise in identifying pathogens, but high-level reconstruction of microbial genomes from microbiologically complex samples for more in-depth analyses remains a challenge. Here, using short-read metagenomic sequencing of a human faecal sample and analysis by tetranucleotide frequency profiling projected onto emergent self-organising maps, we were able to identify the presence, and then reconstruct near-complete Klebsiella pneumoniae and previously undetected Enterococcus faecium genomes, assign genotypes, identify the genomic context of key resistance elements, and identify highly discriminatory mutations in the metagenome to distinguish closely related strains, and infer transmission. These proof-of-principle results demonstrate the utility of clinical sample metagenomics to recover and discover sequences of important drug-resistant bacteria and application of the approach in outbreak investigations, independent of the need to culture the organisms

非培养依赖型方法在病原体鉴定领域已展现出应用前景，但从微生物群落组成复杂的样本中高完整度重建微生物基因组以开展更深入的分析，仍是一项亟待攻克的挑战。本研究针对一份人类粪便样本开展短读长宏基因组测序（short-read metagenomic sequencing），并结合投影于涌现自组织映射（emergent self-organising maps）的四核苷酸频率谱分析（tetranucleotide frequency profiling），成功完成以下工作：首先鉴定病原体的存在，随后重建出接近完整的肺炎克雷伯菌（Klebsiella pneumoniae）与此前未被检出的屎肠球菌（Enterococcus faecium）基因组；完成菌株基因型分型；明确关键耐药基因元件的基因组上下文；在宏基因组中识别出可区分近缘菌株的高区分度突变位点；并推断菌株的传播路径。本研究的原理验证性结果表明，无需对病原菌进行人工培养，临床样本宏基因组学技术即可实现重要耐药细菌基因组序列的获取与发现，且该方法可应用于暴发疫情调查工作。