Department of Microbiology, Austin Health, Melbourne, Australia.
J Clin Microbiol. 2013 May;51(5):1396-401. doi: 10.1128/JCM.03332-12. Epub 2013 Feb 13.
Next-generation sequencing (NGS) of bacterial genomes has recently become more accessible and is now available to the routine diagnostic microbiology laboratory. However, questions remain regarding its feasibility, particularly with respect to data analysis in nonspecialist centers. To test the applicability of NGS to outbreak investigations, Ion Torrent sequencing was used to investigate a putative multidrug-resistant Escherichia coli outbreak in the neonatal unit of the Mercy Hospital for Women, Melbourne, Australia. Four suspected outbreak strains and a comparator strain were sequenced. Genome-wide single nucleotide polymorphism (SNP) analysis demonstrated that the four neonatal intensive care unit (NICU) strains were identical and easily differentiated from the comparator strain. Genome sequence data also determined that the NICU strains belonged to multilocus sequence type 131 and carried the bla(CTX-M-15) extended-spectrum beta-lactamase. Comparison of the outbreak strains to all publicly available complete E. coli genome sequences showed that they clustered with neonatal meningitis and uropathogenic isolates. The turnaround time from a positive culture to the completion of sequencing (prior to data analysis) was 5 days, and the cost was approximately $300 per strain (for the reagents only). The main obstacles to a mainstream adoption of NGS technologies in diagnostic microbiology laboratories are currently cost (although this is decreasing), a paucity of user-friendly and clinically focused bioinformatics platforms, and a lack of genomics expertise outside the research environment. Despite these hurdles, NGS technologies provide unparalleled high-resolution genotyping in a short time frame and are likely to be widely implemented in the field of diagnostic microbiology in the next few years, particularly for epidemiological investigations (replacing current typing methods) and the characterization of resistance determinants. Clinical microbiologists need to familiarize themselves with these technologies and their applications.
新一代测序（NGS）的细菌基因组最近变得更加容易获得，现在已经可用于常规的诊断微生物学实验室。然而，关于其可行性仍存在一些问题，特别是在非专业中心进行数据分析方面。为了测试 NGS 在暴发调查中的适用性，我们使用 Ion Torrent 测序法调查了澳大利亚墨尔本 Mercy 妇女医院新生儿科的一个推定的多药耐药大肠杆菌暴发。对四个疑似暴发菌株和一个对照菌株进行了测序。全基因组单核苷酸多态性（SNP）分析表明，四个新生儿重症监护病房（NICU）菌株完全相同，与对照菌株容易区分。基因组序列数据还确定，NICU 菌株属于多位点序列类型 131，携带 bla(CTX-M-15)超广谱β-内酰胺酶。将暴发菌株与所有公开的完整大肠杆菌基因组序列进行比较表明，它们与新生儿脑膜炎和泌尿道致病性分离株聚类。从阳性培养物到测序完成（数据分析之前）的周转时间为 5 天，每个菌株的成本约为 300 美元（仅试剂）。诊断微生物学实验室采用 NGS 技术的主要障碍目前是成本（尽管这在降低）、缺乏用户友好和以临床为重点的生物信息学平台，以及研究环境之外缺乏基因组学专业知识。尽管存在这些障碍，NGS 技术在短时间内提供无与伦比的高分辨率基因分型，并且可能在未来几年内广泛应用于诊断微生物学领域，特别是用于流行病学调查（取代当前的分型方法）和耐药决定因素的表征。临床微生物学家需要熟悉这些技术及其应用。
