Genes of the major histocompatibility complex (MHC) code for cell surface proteins essential for adaptive immunity. They show the most outstanding genetic diversity in vertebrates, which has been connected with various fitness traits and thus with the long-term persistence of populations. In this study, polymorphism of the MHC class II DRB locus was investigated in chamois with Single-Strand Conformation Polymorphism (SSCP)/Sanger genotyping and Ion Torrent S5 next-generation sequencing (NGS). From eight identified DRB variants in 28 individuals, five had already been described, and three were new, undescribed alleles. With conventional SSCP/Sanger sequencing, we were able to detect seven alleles, all of which were also detected with NGS. We found inconsistencies in the individual genotypes between the two methods, which were mainly caused by allelic dropout in the SSCP/Sanger method. Six out of 28 individuals were falsely classified as homozygous with SSCP/Sanger analysis. Overall, 25% of the individuals were identified as genotyping discrepancies between the two methods. Our results show that NGS technologies are better performing in sequencing highly variable regions such as the MHC, and they also have a higher detection capacity, thus allowing a more accurate description of the genetic composition, which is crucial for evolutionary and population genetic studies.

• Klíčová slova
• Ion Torrent, Rupicapra rupicapra, major histocompatibility complex, next-generation sequencing,
• Publikační typ
• časopisecké články MeSH

In Europe the Borrelia burgdorferi sensu lato complex is represented by five distinct genospecies: Borrelia burgdorferi sensu stricto, Borrelia afzelii, Borrelia garinii, Borrelia valaisiana, and Borrelia lusitaniae. These taxonomic entities are known to differ in their specific associations with vertebrate hosts and to provoke distinct clinical manifestations in human patients. However, exceptions to these rules have often been observed, indicating that strains belonging to a single genospecies may be more heterogeneous than expected. It is, therefore, important to develop alternative identification tools which are able to distinguish Borrelia strains not only at the specific level but also at the intraspecific level. DNA from a sample of 370 Ixodes ricinus ticks collected in the Czech Republic was analyzed by PCR for the presence of a approximately 230-bp fragment of the rrfA-rrlB intergenic spacer of Borrelia spp. A total of 20.5% of the ticks were found to be positive. The infecting genospecies were identified by analyzing the amplified products by the restriction fragment length polymorphism (RFLP) method with restriction enzyme MseI and by single-strand conformation polymorphism (SSCP) analysis. The two methods were compared, and PCR-SSCP analysis appeared to be a valuable tool for rapid identification of spirochetes at the intraspecific level, particularly when large samples are examined. Furthermore, by using PCR-SSCP analysis we identified a previously unknown Borrelia genotype, genotype I-77, which would have gone unnoticed if RFLP analysis alone had been used.

• MeSH
• Borrelia burgdorferi komplex klasifikace   genetika   MeSH
• DNA bakterií genetika   izolace a purifikace   MeSH
• fylogeneze MeSH
• genetická variace * MeSH
• genotyp MeSH
• klíště mikrobiologie   MeSH
• lidé MeSH
• mezerníky ribozomální DNA genetika   MeSH
• molekulární sekvence - údaje MeSH
• polymerázová řetězová reakce metody   MeSH
• polymorfismus délky restrikčních fragmentů MeSH
• polymorfismus konformace jednovláknové DNA * MeSH
• sekvenční analýza DNA MeSH
• techniky typizace bakterií MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• hodnotící studie MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Česká republika MeSH
• Názvy látek
• DNA bakterií MeSH
• mezerníky ribozomální DNA MeSH