Real-Time Whole-Genome Sequencing for Routine Typing, Surveillance, and Outbreak Detection of Verotoxigenic Escherichia coli
Real-Time Whole-Genome Sequencing for Routine Typing, Surveillance, and Outbreak Detection of Verotoxigenic Escherichia coli
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DOI：
10.1128/jcm.03617-13
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发表时间：
2014-05-01
影响因子：
9.4
通讯作者：
Aarestrup, Frank M.
中科院分区：
文献类型：
--
作者：
Joensen, Katrine Grimstrup;Scheutz, Flemming;Aarestrup, Frank M.
病原体的快速准确鉴定和分型对于有效监测和暴发检测至关重要。目前的常规程序是基于各种技术，使程序费力，耗时，昂贵。随着全基因组测序（WGS）变得越来越便宜，它在诊断和常规监测方面都有巨大的潜力。本研究的目的是进行实时评估的WGS的常规分型和监测的大肠埃希菌（VTEC）产生的细胞毒素。在丹麦，Statens血清研究所（SSI）常规接收所有疑似VTEC分离株。在2012年秋季的7周期间，使用IonTorrent PGM同时对所有传入分离株进行WGS。利用网络工具（www.genomicepidemiology.org）进行实时生物信息学分析，用于种鉴定、多位点序列类型（MLST）分型和系统发育关系的确定，以及用于检测E.大肠杆菌毒力基因作为本研究的一部分开发。在研究期间，共对46株疑似VTEC分离株进行了平行表征。毒力检测成功地检测了常规分型中的毒力基因，明确的是verocytotoxin 1（vtx1）、verocytotoxin 2（vtx2）和intimin（eae），也检测到了其他毒力基因。毒力测定法也是一种确定verocytotoxin（vtx）亚型的可靠方法。从WGS建立了与流行病学一致的分离株实时聚类，从而能够区分散发和爆发分离株。总的来说，WGS分型比目前的常规方法更快，成本更低。因此，WGS分型是常规分型策略的上级替代方案。这种方法也可以应用于其他病原体的分型和监测。
Fast and accurate identification and typing of pathogens are essential for effective surveillance and outbreak detection. The current routine procedure is based on a variety of techniques, making the procedure laborious, time-consuming, and expensive. With whole-genome sequencing (WGS) becoming cheaper, it has huge potential in both diagnostics and routine surveillance. The aim of this study was to perform a real-time evaluation of WGS for routine typing and surveillance of verocytotoxin-producing Escherichia coli (VTEC). In Denmark, the Statens Serum Institut (SSI) routinely receives all suspected VTEC isolates. During a 7-week period in the fall of 2012, all incoming isolates were concurrently subjected to WGS using IonTorrent PGM. Real-time bioinformatics analysis was performed using web-tools (www.genomicepidemiology.org) for species determination, multilocus sequence type (MLST) typing, and determination of phylogenetic relationship, and a specific VirulenceFinder for detection of E. coli virulence genes was developed as part of this study. In total, 46 suspected VTEC isolates were characterized in parallel during the study. VirulenceFinder proved successful in detecting virulence genes included in routine typing, explicitly verocytotoxin 1 (vtx1), verocytotoxin 2 (vtx2), and intimin (eae), and also detected additional virulence genes. VirulenceFinder is also a robust method for assigning verocytotoxin (vtx) subtypes. A real-time clustering of isolates in agreement with the epidemiology was established from WGS, enabling discrimination between sporadic and outbreak isolates. Overall, WGS typing produced results faster and at a lower cost than the current routine. Therefore, WGS typing is a superior alternative to conventional typing strategies. This approach may also be applied to typing and surveillance of other pathogens.