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Genotyping of European Toxoplasma gondii strains by a new high-resolution next-generation sequencing-based method
M. Joeres, P. Maksimov, D. Höper, S. Calvelage, R. Calero-Bernal, M. Fernández-Escobar, B. Koudela, R. Blaga, MG. Vrhovec, K. Stollberg, N. Bier, S. Sotiraki, J. Sroka, W. Piotrowska, P. Kodym, W. Basso, FJ. Conraths, A. Mercier, L. Galal, ML....
Language English Country Germany
Document type Journal Article
NLK
ProQuest Central
from 1997-01-01 to 1 year ago
Medline Complete (EBSCOhost)
from 2000-01-01 to 1 year ago
Health & Medicine (ProQuest)
from 1997-01-01 to 1 year ago
- MeSH
- Genetic Variation MeSH
- Genotype MeSH
- Humans MeSH
- Multiplex Polymerase Chain Reaction MeSH
- Polymorphism, Restriction Fragment Length MeSH
- DNA, Protozoan genetics MeSH
- Pregnancy MeSH
- Toxoplasma * genetics MeSH
- High-Throughput Nucleotide Sequencing MeSH
- Check Tag
- Humans MeSH
- Pregnancy MeSH
- Female MeSH
- Publication type
- Journal Article MeSH
PURPOSE: A new high-resolution next-generation sequencing (NGS)-based method was established to type closely related European type II Toxoplasma gondii strains. METHODS: T. gondii field isolates were collected from different parts of Europe and assessed by whole genome sequencing (WGS). In comparison to ME49 (a type II reference strain), highly polymorphic regions (HPRs) were identified, showing a considerable number of single nucleotide polymorphisms (SNPs). After confirmation by Sanger sequencing, 18 HPRs were used to design a primer panel for multiplex PCR to establish a multilocus Ion AmpliSeq typing method. Toxoplasma gondii isolates and T. gondii present in clinical samples were typed with the new method. The sensitivity of the method was tested with serially diluted reference DNA samples. RESULTS: Among type II specimens, the method could differentiate the same number of haplotypes as the reference standard, microsatellite (MS) typing. Passages of the same isolates and specimens originating from abortion outbreaks were identified as identical. In addition, seven different genotypes, two atypical and two recombinant specimens were clearly distinguished from each other by the method. Furthermore, almost all SNPs detected by the Ion AmpliSeq method corresponded to those expected based on WGS. By testing serially diluted DNA samples, the method exhibited a similar analytical sensitivity as MS typing. CONCLUSION: The new method can distinguish different T. gondii genotypes and detect intra-genotype variability among European type II T. gondii strains. Furthermore, with WGS data additional target regions can be added to the method to potentially increase typing resolution.
Bavarian Health and Food Safety Authority Erlangen Germany
Central European Institute of Technology University of Veterinary Sciences Brno Brno Czech Republic
Centre National de Référence Toxoplasmose Centre Hospitalier Universitaire Dupuytren Limoges France
Centre of Epidemiology and Microbiology National Institute of Public Health Prague Czech Republic
Chemisches und Veterinäruntersuchungsamt Westfalen Standort Arnsberg Arnsberg Germany
Department of Analysis and Diagnostics Norwegian Veterinary Institute Ås Norway
Department of Animal Health Welfare and Food Safety Norwegian Veterinary Institute Tromsø Norway
Department of Microbiology National Veterinary Institute Uppsala Sweden
Faculty of Veterinary Medicine University of Veterinary Sciences Brno Brno Czech Republic
German Federal Institute for Risk Assessment Department for Biological Safety Berlin Germany
IDEXX Laboratories Kornwestheim Germany
Infectious Disease Preparedness Statens Serum Institut Copenhagen Denmark
Institute of Parasitology Vetsuisse Faculty University of Bern Bern Switzerland
Italian National Institute of Health Rome Italy
Landeslabor Berlin Brandenburg Frankfurt Germany
Parasitology Laboratory Instituto Nacional de Investigação Agrária e Veterinária Oeiras Portugal
University of Agricultural Sciences and Veterinary Medicine Cluj Napoca Romania
Veterinary Research Institute Hellenic Agricultural Organisation DIMITRA Thessaloniki Greece
References provided by Crossref.org
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