Georgi Enrico, Walter Mathias C, Pfalzgraf Marie-Theres, Northoff Bernd H, Holdt Lesca M, Scholz Holger C, Zoeller Lothar, Zange Sabine, Antwerpen Markus H
Bundeswehr Institute of Microbiology, Munich, Germany.
Institute of Laboratory Medicine, Ludwig-Maximilians University, Munich, Germany.
PLoS One. 2017 Apr 7;12(4):e0175425. doi: 10.1371/journal.pone.0175425. eCollection 2017.
Brucellosis, a worldwide common bacterial zoonotic disease, has become quite rare in Northern and Western Europe. However, since 2014 a significant increase of imported infections caused by Brucella (B.) melitensis has been noticed in Germany. Patients predominantly originated from Middle East including Turkey and Syria. These circumstances afforded an opportunity to gain insights into the population structure of Brucella strains. Brucella-isolates from 57 patients were recovered between January 2014 and June 2016 with culture confirmed brucellosis by the National Consultant Laboratory for Brucella. Their whole genome sequences were generated using the Illumina MiSeq platform. A whole genome-based SNP typing assay was developed in order to resolve geographically attributed genetic clusters. Results were compared to MLVA typing results, the current gold-standard of Brucella typing. In addition, sequences were examined for possible genetic variation within target regions of molecular diagnostic assays. Phylogenetic analyses revealed spatial clustering and distinguished strains from different patients in either case, whereas multiple isolates from a single patient or technical replicates showed identical SNP and MLVA profiles. By including WGS data from the NCBI database, five major genotypes were identified. Notably, strains originating from Turkey showed a high diversity and grouped into seven subclusters of genotype II. MLVA analysis congruently clustered all isolates and predominantly matched the East Mediterranean genetic clade. This study confirms whole-genome based SNP-analysis as a powerful tool for accurate typing of B. melitensis. Furthermore it allows special allocation and therefore provides useful information on the geographic origin for trace-back analysis. However, the lack of reliable metadata in public databases often prevents a resolution below geographic regions or country levels and corresponding precise trace-back analysis. Once this obstacle is resolved, WGS-derived bacterial typing adds an important method to complement epidemiological surveys during outbreak investigations. This is the first report of a detailed genetic investigation of an extensive collection of B. melitensis strains isolated from human cases in Germany.
布鲁氏菌病是一种全球常见的细菌性人畜共患病，在北欧和西欧已变得相当罕见。然而，自2014年以来，德国注意到由羊种布鲁氏菌（B. melitensis）引起的输入性感染显著增加。患者主要来自包括土耳其和叙利亚在内的中东地区。这些情况为深入了解布鲁氏菌菌株的种群结构提供了机会。2014年1月至2016年6月期间，从57名患者中分离出布鲁氏菌，经国家布鲁氏菌病咨询实验室培养确诊为布鲁氏菌病。使用Illumina MiSeq平台生成了它们的全基因组序列。为了解析地理归因的遗传簇，开发了一种基于全基因组的单核苷酸多态性（SNP）分型检测方法。将结果与布鲁氏菌分型的当前金标准——多位点可变数目串联重复序列分析（MLVA）分型结果进行比较。此外，检查序列在分子诊断检测靶区域内是否存在可能的遗传变异。系统发育分析揭示了空间聚类，并在两种情况下区分了不同患者的菌株，而来自单个患者的多个分离株或技术重复显示出相同的SNP和MLVA谱型。通过纳入来自NCBI数据库的全基因组测序（WGS）数据，鉴定出五种主要基因型。值得注意的是，源自土耳其的菌株表现出高度多样性，并分为基因型II的七个亚簇。MLVA分析一致地将所有分离株聚类，并且主要与东地中海遗传分支匹配。本研究证实基于全基因组的SNP分析是准确分型羊种布鲁氏菌的有力工具。此外，它允许进行特殊分配，因此为溯源分析提供了有关地理来源的有用信息。然而，公共数据库中缺乏可靠的元数据通常会妨碍在地理区域或国家层面以下进行解析以及相应的精确溯源分析。一旦解决了这一障碍，WGS衍生的细菌分型将为疫情调查期间的流行病学调查增添一种重要的补充方法。这是对从德国人类病例中分离出的大量羊种布鲁氏菌菌株进行详细基因研究的首份报告。
