title: Karyotype differentiation in 19 species of river loach fishes (Nemacheilidae, Teleostei): extensive variability associated with rDNA and heterochromatin distribution and its phylogenetic and ecological interpretation.
abstract: BACKGROUND
  Loaches of the family Nemacheilidae are one of the most speciose elements of Palearctic freshwater ichthyofauna and have undergone rapid ecological adaptations and colonizations. Their cytotaxonomy is largely unexplored; with the impact of cytogenetical changes on this evolutionary diversification still unknown. An extensive cytogenetical survey was performed in 19 nemacheilid species using both conventional (Giemsa staining, C- banding, Ag- and Chromomycin A3/DAPI stainings) and molecular (fluorescence in situ hybridization with 5S rDNA, 45S rDNA, and telomeric (TTAGGG)n probes) methods. A phylogenetic tree of the analysed specimens was constructed based on one mitochondrial (cytochrome b) and two nuclear (RAG1, IRBP) genes.
  
  
  RESULTS
  Seventeen species showed karyotypes composed of 2n = 50 chromosomes but differentiated by fundamental chromosome number (NF = 68-90). Nemachilichthys ruppelli (2n = 38) and Schistura notostigma (2n = 44-48) displayed reduced 2n with an elevated number of large metacentric chromosomes. Only Schistura fasciolata showed morphologically differentiated sex chromosomes with a multiple system of the XY1Y2 type. Chromomycin A3 (CMA3)- fluorescence revealed interspecific heterogeneity in the distribution of GC-rich heterochromatin including its otherwise very rare association with 5S rDNA sites. The 45S rDNA sites were mostly located on a single chromosome pair contrasting markedly with a pattern of two (Barbatula barbatula, Nemacheilus binotatus, N. ruppelli) to 20 sites (Physoschistura sp.) of 5S rDNA. The cytogenetic changes did not follow the phylogenetic relationships between the samples. A high number of 5S rDNA sites was present in species with small effective population sizes.
  
  
  CONCLUSIONS
  Despite a prevailing conservatism of 2n, Nemacheilidae exhibited a remarkable cytogenetic variability on microstructural level. We suggest an important role for pericentric inversions, tandem and centric fusions in nemacheilid karyotype differentiation. Short repetitive sequences, genetic drift, founder effect, as well as the involvement of transposable elements in the dispersion of ribosomal DNA sites, might also have played a role in evolutionary processes such as reproductive isolation. These remarkable dynamics of their genomes qualify river loaches as a model for the study of the cytogenetic background of major evolutionary processes such as radiation, endemism and colonization of a wide range of habitats.
authors.affiliation: Laboratory of Fish Genetics, Institute of Animal Physiology and Genetics, Czech Academy of Sciences, Rumburská 89, Liběchov, 277 21, Czech Republic. alexandr.sember@seznam.cz.|Department of Genetics and Microbiology, Faculty of Science, Charles University in Prague, Viničná 5, 128 44, Prague 2, Czech Republic. alexandr.sember@seznam.cz.
authors.forename: Alexandr
authors.initials: A
authors.lastname: Sember
authors.affiliation: Laboratory of Fish Genetics, Institute of Animal Physiology and Genetics, Czech Academy of Sciences, Rumburská 89, Liběchov, 277 21, Czech Republic. bohlen@iapg.cas.cz.
authors.forename: Jörg
authors.initials: J
authors.lastname: Bohlen
authors.affiliation: Laboratory of Fish Genetics, Institute of Animal Physiology and Genetics, Czech Academy of Sciences, Rumburská 89, Liběchov, 277 21, Czech Republic. v.slechtova@iapg.cas.cz.
authors.forename: Vendula
authors.initials: V
authors.lastname: Šlechtová
authors.affiliation: Laboratory of Fish Genetics, Institute of Animal Physiology and Genetics, Czech Academy of Sciences, Rumburská 89, Liběchov, 277 21, Czech Republic. altmanova.m@gmail.com.|Department of Ecology, Faculty of Science, Charles University in Prague, Viničná 7, 128 44, Prague 2, Czech Republic. altmanova.m@gmail.com.
authors.forename: Marie
authors.initials: M
authors.lastname: Altmanová
authors.affiliation: Laboratory of Fish Genetics, Institute of Animal Physiology and Genetics, Czech Academy of Sciences, Rumburská 89, Liběchov, 277 21, Czech Republic. radka.symonova@uibk.ac.at.|Research Institute for Limnology, University of Innsbruck, Mondseestraße 9, A-5310, Mondsee, Austria. radka.symonova@uibk.ac.at.
authors.forename: Radka
authors.initials: R
authors.lastname: Symonová
authors.affiliation: Laboratory of Fish Genetics, Institute of Animal Physiology and Genetics, Czech Academy of Sciences, Rumburská 89, Liběchov, 277 21, Czech Republic. rab@iapg.cas.cz.
authors.forename: Petr
authors.initials: P
authors.lastname: Ráb
mesh_terms: D000818:Animals; D003531:Cypriniformes / Q000145:classification* / Q000235:genetics*; D004251:DNA Transposable Elements; D004275:DNA, Ribosomal / Q000235:genetics; D005260:Female; D006570:Heterochromatin; D017404:In Situ Hybridization, Fluorescence; D007621:Karyotyping / Q000662:veterinary; D008297:Male; D010802:Phylogeny; D045483:Rivers
chemical_list: D004251:DNA Transposable Elements; D004275:DNA, Ribosomal; D006570:Heterochromatin