Abstract
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.
Highlights
On a global scale human brucellosis is one of the most common bacterial zoonotic diseases [1]
All available strains from culture-positive cases of human brucellosis that had been processed by the National Consultant Laboratory for Brucella and that had been identified as B. melitensis by real-time and conventional PCR methods between January 2014 and June 2016 (n = 57) were subjected to whole genome sequencing (WGS)
The decline can be attributed to the implementation of public health measures including testing and slaughtering of cattle, livestock vaccination, border control and surveillance programmes that largely eradicated the causative agent from the animal reservoir and minimized the risk of transmission to humans
Summary
On a global scale human brucellosis is one of the most common bacterial zoonotic diseases [1]. The disease has its highest incidence and prevalence in countries of the Mediterranean basin, Middle East, some parts of Central and South America, Africa and Asia [2]. The highest annual incidences of human brucellosis per million of the population are observed in Syria and in Mongolia [3]. Each year a relatively small number of cases are reported in Germany, with most of them having a history of travelling to or immigrating from endemic regions like the Mediterranean basin. Within the last two years an increase of cases was observed. In 2014 and 2015, the number of annual cases nearly doubled to 47 and 44, respectively
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