The widely distributed ray-finned fish genus Carassius is very well known due to its unique biological characteristics such as polyploidy, clonality, and/or interspecies hybridization. These biological characteristics have enabled Carassius species to be successfully widespread over relatively short period of evolutionary time. Therefore, this fish model deserves to be the center of attention in the research field. Some studies have already described the Carassius karyotype, but results are inconsistent in the number of morphological categories for individual chromosomes. We investigated three focal species: Carassius auratus, C. carassius and C. gibelio with the aim to describe their standardized diploid karyotypes, and to study their evolutionary relationships using cytogenetic tools. We measured length (q+plength) of each chromosome and calculated centromeric index (i value). We found: (i) The relationship between q+plength and i value showed higher similarity of C. auratus and C. carassius. (ii) The variability of i value within each chromosome expressed by means of the first quartile (Q1) up to the third quartile (Q3) showed higher similarity of C. carassius and C. gibelio. (iii) The fluorescent in situ hybridization (FISH) analysis revealed higher similarity of C. auratus and C. gibelio. (iv) Standardized karyotype formula described using median value (Q2) showed differentiation among all investigated species: C. auratus had 24 metacentric (m), 40 submetacentric (sm), 2 subtelocentric (st), 2 acrocentric (a) and 32 telocentric (T) chromosomes (24m+40sm+2st+2a+32T); C. carassius: 16m+34sm+8st+42T; and C. gibelio: 16m+22sm+10st+2a+50T. (v) We developed R scripts applicable for the description of standardized karyotype for any other species. The diverse results indicated unprecedented complex genomic and chromosomal architecture in the genus Carassius probably influenced by its unique biological characteristics which make the study of evolutionary relationships more difficult than it has been originally postulated.

Department of Cell Biology Faculty of Science Charles University 12843 Prague Czech Republic

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Delaunay L. Comparative karyological study of species Muscari Mill. and Bellevalia Lapeyr. Bull. Tiflis Bot. Gard. 1922;2:1–32. (In Russian)

Levan A., Fredga K., Sandberg A.A. Nomenclature for centromeric position on chromosomes. Hereditas. 1964;52:201–220. doi: 10.1111/j.1601-5223.1964.tb01953.x. DOI

White M.J.D. Animal Cytology and Evolution. 3rd ed. Cambridge University Press; Cambridge, UK: 1973. p. 468.

White M.J.D. The Chromosomes. 6th ed. Chapman Hall; London, UK: 1973. p. 188.

Kretschmer R., Gunski R.J., Garnero A.D.V., de Freitas T.R.O., Toma G.A., Cioffi M.D.B., de Oliveira E.H.C., O’Connor R.E., Griffin D.K. Chromosomal Analysis in Crotophaga ani (Aves, Cuculiformes) Reveals Extensive Genomic Reorganization and an Unusual Z-Autosome Robertsonian Translocation. Cells. 2020;10:4. doi: 10.3390/cells10010004. PubMed DOI PMC

O’Connor R.E., Kiazim L.G., Rathje C.C., Jennings R.L., Griffin D.K. Rapid Multi-Hybridisation FISH Screening for Balanced Porcine Reciprocal Translocations Suggests a Much Higher Abnormality Rate Than Previously Appreciated. Cells. 2021;10:250. doi: 10.3390/cells10020250. PubMed DOI PMC

Cauret C.M., Gansauge M.T., Tupper A.S., Furman B.L., Knytl M., Song X.Y., Greenbaum E., Meyer M., Evans B.J., Wilson M. Developmental Systems Drift and the Drivers of Sex Chromosome Evolution. Mol. Biol. Evol. 2020;37:799–810. doi: 10.1093/molbev/msz268. PubMed DOI

Augstenová B., Pensabene E., Kratochvíl L., Rovatsos M. Cytogenetic Evidence for Sex Chromosomes and Karyotype Evolution in Anguimorphan Lizards. Cells. 2021;10:1612. doi: 10.3390/cells10071612. PubMed DOI PMC

Knytl M., Tlapakova T., Vankova T., Krylov V. Silurana Chromosomal Evolution: A New Piece to the Puzzle. Cytogenet. Genome Res. 2018;156:223–228. doi: 10.1159/000494708. PubMed DOI

Gillberg C. Chromosomal disorders and autism. J. Autism Dev. Disord. 1998;28:415–425. doi: 10.1023/A:1026004505764. PubMed DOI

Erkilic N., Gatinois V., Torriano S., Bouret P., Sanjurjo-Soriano C., Luca V.D., Damodar K., Cereso N., Puechberty J., Sanchez-Alcudia R., et al. A Novel Chromosomal Translocation Identified due to Complex Genetic Instability in iPSC Generated for Choroideremia. Cells. 2019;8:1068. doi: 10.3390/cells8091068. PubMed DOI PMC

Sember A., Bertollo L.A.C., Ráb P., Yano C.F., Hatanaka T., de Oliveira E.A., Cioffi M.D.B. Sex Chromosome Evolution and Genomic Divergence in the Fish Hoplias malabaricus (Characiformes, Erythrinidae) Front. Genet. 2018;9:71. doi: 10.3389/fgene.2018.00071. PubMed DOI PMC

Miura I., Shams F., Lin S.M., Cioffi M.D.B., Liehr T., Al-Rikabi A., Kuwana C., Srikulnath K., Higaki Y., Ezaz T. Evolution of a Multiple Sex-Chromosome System by Three-Sequential Translocations among Potential Sex-Chromosomes in the Taiwanese Frog Odorrana swinhoana. Cells. 2021;10:661. doi: 10.3390/cells10030661. PubMed DOI PMC

Seroussi E., Knytl M., Pitel F., Elleder D., Krylov V., Leroux S., Morisson M., Yosefi S., Miyara S., Ganesan S., et al. Avian Expression Patterns and Genomic Mapping Implicate Leptin in Digestion and TNF in Immunity, Suggesting That Their Interacting Adipokine Role Has Been Acquired Only in Mammals. Int. J. Mol. Sci. 2019;20:4489. doi: 10.3390/ijms20184489. PubMed DOI PMC

Tymowska J. Polyploidy and cytogenetic variation in frogs of the genus Xenopus. In: Green D.M., Sessions S.K., editors. Amphibian Cytogenetics and Evolution. Academic Press; San Diego, CA, USA: 1991. pp. 259–297.

Knytl M., Kalous L., Symonová R., Rylková K., Ráb P. Chromosome studies of European cyprinid fishes: Cross-species painting reveals natural allotetraploid origin of a Carassius female with 206 chromosomes. Cytogenet. Genome Res. 2013;139:276–283. doi: 10.1159/000350689. PubMed DOI

Kobayasi H., Kawashima Y., Takeuchi N. Comparative Chromosome Studies in the Genus Carassius, Especially with a Finding of Polyploidy in the Ginbuna (C. auratus langsdorfii) Jpn. J. Ichthyol. 1970;17:153–160. doi: 10.11369/JJI1950.17.153. DOI

Ojima Y., Takai A. Further cytogenetical studies on the origin of the gold-fish. Proc. Jpn. Academy. Ser. B Phys. Biol. Sci. 1979;55:346–350. doi: 10.2183/pjab.55.346. DOI

Yang L., Sado T., Vincent Hirt M., Pasco-Viel E., Arunachalam M., Li J., Wang X., Freyhof J., Saitoh K., Simons A.M., et al. Phylogeny and polyploidy: Resolving the classification of cyprinine fishes (Teleostei: Cypriniformes) Mol. Phylogenet. Evol. 2015;85:97–116. doi: 10.1016/j.ympev.2015.01.014. PubMed DOI

Lusk S., Hanel L., Lojkásek B., Lusková V., Muška M. The Red List of Lampreys and Fishes of the Czech Republic. In: Němec M., Chobot K., editors. Red List of Threatened Species of the Czech Republic, Vertebrates. Příroda; Prague, Czech Republic: 2017. pp. 51–82. Chapter 34.

Sayer C.D., Copp G.H., Emson D., Godard M.J., Ziȩba G., Wesley K.J. Towards the conservation of crucian carp Carassius carassius: Understanding the extent and causes of decline within part of its native English range. J. Fish Biol. 2011;79:1608–1624. doi: 10.1111/j.1095-8649.2011.03059.x. PubMed DOI

Ohno S., Muramoto J., Christian L., Atkin N.B. Diploid-tetraploid relationship among old-world members of the fish family Cyprinidae. Chromosoma. 1967;23:1–9. doi: 10.1007/BF00293307. DOI

Balon E.K. About the oldest domesticates among fishes. J. Fish Biol. 2004;65:1–27. doi: 10.1111/j.0022-1112.2004.00563.x. DOI

Lusková V., Lusk S., Halačka K., Vetešník L. Carassius auratus gibelio—The most successful invasive fish in waters of the Czech Republic. Russ. J. Biol. Invasions. 2010;1:176–180. doi: 10.1134/S2075111710030069. DOI

Tarkan A.S., Güler Ekmekçi F., Vilizzi L., Copp G.H. Risk screening of non-native freshwater fishes at the frontier between Asia and Europe: First application in Turkey of the fish invasiveness screening kit. J. Appl. Ichthyol. 2014;30:392–398. doi: 10.1111/jai.12389. DOI

Card J.T., Hasler C.T., Ruppert J.L., Donadt C., Poesch M.S. A Three-pass electrofishing removal strategy is not effective for eradication of prussian Carp in a North American stream network. J. Fish Wildl. Manag. 2020;11:485–493. doi: 10.3996/JFWM-20-031. DOI

Haynes G.D., Gongora J., Gilligan D.M., Grewe P., Moran C., Nicholas F.W. Cryptic hybridization and introgression between invasive Cyprinid species Cyprinus carpio and Carassius auratus in Australia: Implications for invasive species management. Anim. Conserv. 2012;15:83–94. doi: 10.1111/j.1469-1795.2011.00490.x. DOI

Kalous L., Knytl M. Karyotype diversity of the offspring resulting from reproduction experiment between diploid male and triploid female of silver Prussian carp, Carassius gibelio (Cyprinidae, Actinopterygii) Folia Zool. 2011;60:115–121. doi: 10.25225/fozo.v60.i2.a5.2011. DOI

Xiao J., Zou T., Chen Y., Chen L., Liu S., Tao M., Zhang C., Zhao R., Zhou Y., Long Y., et al. Coexistence of diploid, triploid and tetraploid crucian carp (Carassius auratus) in natural waters. BMC Genet. 2011;12:20. doi: 10.1186/1471-2156-12-20. PubMed DOI PMC

Jiang F.F., Wang Z.W., Zhou L., Jiang L., Zhang X.J., Apalikova O.V., Brykov V.A., Gui J.F. High male incidence and evolutionary implications of triploid form in northeast Asia Carassius auratus complex. Mol. Phylogenet. Evol. 2013;66:350–359. doi: 10.1016/j.ympev.2012.10.006. PubMed DOI

Knytl M., Kalous L., Rab P. Karyotype and chromosome banding of endangered crucian carp, Carassius carassius (Linnaeus, 1758) (Teleostei, Cyprinidae) Comp. Cytogenet. 2013;7:205–213. doi: 10.3897/compcytogen.v7i3.5411. PubMed DOI PMC

Knytl M., Kalous L., Rylková K., Choleva L., Merilä J., Ráb P. Morphologically indistinguishable hybrid Carassius female with 156 chromosomes: A threat for the threatened crucian carp, C. carassius, L. PLoS ONE. 2018;13:e0190924. doi: 10.1371/journal.pone.0190924. PubMed DOI PMC

Bertollo L., Cioffi M. Direct chromosome preparation from freshwater teleost fishes. In: Ozouf-Costaz C., Pisano E., Foresti F., Foresti L.D.A.T., editors. Fish Cytogenetic Techniques: Ray-Fin Fishes and Chondrichthyans. CRC Press; Enfield, CT, USA: 2015. pp. 21–26.

Schneider C.A., Rasband W.S., Eliceiri K.W. NIH Image to ImageJ: 25 years of image analysis. Nat. Methods. 2012;9:671–675. doi: 10.1038/nmeth.2089. PubMed DOI PMC

R Core Team . R: A Language and Environment for Statistical Computing. R Core Team; Vienna, Austria: 2020. [(accessed on 20 August 2021)]. Available online: https://www.R-project.org.

RStudio Team . RStudio: Integrated Development Environment for R. RStudio, PBC.; Boston, MA, USA: 2019. [(accessed on 20 August 2021)]. Available online: http://www.rstudio.com.

Naito E., Dewa K., Ymanouchi H., Kominami R. Ribosomal ribonucleic acid (rRNA) gene typing for species identification. J. Forensic Sci. 1992;37:396–403. doi: 10.1520/JFS13249J. PubMed DOI

Sember A., Bohlen J., Šlechtová V., Altmanová M., Symonová R., Ráb P. 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. BMC Evol. Biol. 2015;15:1–22. doi: 10.1186/s12862-015-0532-9. PubMed DOI PMC

Knytl M., Smolík O., Kubíčková S., Tlapáková T., Evans B.J., Krylov V. Chromosome divergence during evolution of the tetraploid clawed frogs, Xenopus mellotropicalis and Xenopus epitropicalis as revealed by Zoo-FISH. PLoS ONE. 2017;12:e0177087. doi: 10.1371/journal.pone.0177087. PubMed DOI PMC

Spoz A., Boron A., Porycka K., Karolewska M., Ito D., Abe S., Kirtiklis L., Juchno D. Molecular cytogenetic analysis of the crucian carp, Carassius carassius (Linnaeus, 1758) (Teleostei, Cyprinidae), using chromosome staining and fluorescence in situ hybridisation with rDNA probes. Comp. Cytogenet. 2014;8:233–248. doi: 10.3897/compcytogen.v8i3.7718. PubMed DOI PMC

Sember A., Pelikánová Š., de Bello Cioffi M., Šlechtová V., Hatanaka T., Do Doan H., Knytl M., Ráb P. Taxonomic Diversity Not Associated with Gross Karyotype Differentiation: The Case of Bighead Carps, Genus Hypophthalmichthys (Teleostei, Cypriniformes, Xenocyprididae) Genes. 2020;11:479. doi: 10.3390/genes11050479. PubMed DOI PMC

Makino S. Notes on the chromosomes of some fresh-water Teleosts. Jpn. J. Genet. 1934;9:100–103.

Cherfas N.B. Natural triploidy in females of the unisexual form of silver crucian carp (Carassius auratus gibelio Bloch) Genetika. 1966;2:16–24.

Chiarelli B., Ferrantelli O., Cucchi C. The caryotype of some teleostea fish obtained by tissue culture in vitro. Experientia. 1969;25:426–427. doi: 10.1007/BF01899963. PubMed DOI

Raicu P., Taisescu E., Banarescu P. Carassius carassius and Carassius auratus, a pair of diploid and tetraploid representative species (Pices, Cyprinidae) Cytologia. 1981;46:233–240. doi: 10.1508/cytologia.46.233. DOI

Fister S., Soldatovic B. Karyotype analysis of male and diploid female Carassius auratus gibelio, Bloch (Pisces, Cyprinidae) caught in the Danube at Belgrade, evidence for the existence of a bisexual population. Acta Vet. Beogr. 1991;41:81–90.

Makino S. A Karyological Study of Gold-fish of Japan. Cytologia. 1941;12:96–111. doi: 10.1508/cytologia.12.96. DOI

Ohno S., Atkin N.B. Comparative DNA values and chromosome complements of eight species of fishes. Chromosoma. 1966;18:455–466. doi: 10.1007/BF00332549. PubMed DOI

Ojima Y., Hitotsumachi S., Makino S. Cytogenetic Studies in Lower Vertebrates. I A Preliminary Report on the Chromosomes of the Funa (Carassius auratus) and Gold-fish (A Revised Study) Proc. Jpn. Acad. 1966;42:62–66. doi: 10.2183/pjab1945.42.62. DOI

Ojima Y., Hitotsumachi S. Cytogenetic studies in lower vertebrates. IV. a note on the chromosomes of the carp (cyprinus carpio) in comparison with those of the funa and the goldfish (Carassius auratus) Jpn. J. Genet. 1967;42:163–167. doi: 10.1266/jjg.42.163. DOI

Arai R., Fujiki A. Chromosomes of three races of Goldfish, Kuro-demekin, Sanshiki- demekin and Ranchu. Bull. Natl. Sci. Mus. Ser. A Zool. 1977;3:187–192.

Ojima Y., Ueda T., Narikawa T. A Cytogenetic Assessment on the Origin of the Gold-fish. Proc. Jpn. Acad. Ser. B. 1979;55:58–63. doi: 10.2183/pjab.55.58. DOI

Ojima Y., Yamano T. The assignment of the nucleolar organizer in the chromosomes of the funa (Carassius, cyprinidae, pisces) Proc. Jpn. Acad. Ser. B. 1980;56:551–556. doi: 10.2183/pjab.56.551. DOI

Zan R.G., Song Z. Analysis and comparison between the karyotypes of Cyprinus carpio and Carassius auratus as well as Aristichthys nobilis and Hypophthalmichthys molitrix. Acta Genet. Sin. 1980;7:72–77.

Zan R.G. Studies of sex chromosomes and C-banding karyotypes of two forms of Carassius auratus in Kunming Lake. Acta Genet. Sin. 1982;9:32–39.

Wang R.F., Shi L.M., He W.S. A Comparative Study of the Ag-NORS of Carassius auratus from Different Geographic Districts. Zool. Res. 1988;9:165–169.

Kasama M., Kobayasi H. Hybridization Experiment Between Female Crucian Carp and Male Grass Carp. Nippon. Suisan Gakkaishi Jpn. Ed. 1989;55:1001–1006. doi: 10.2331/suisan.55.1001. DOI

Kasama M., Kobayasi H. Hybridization experiment between Carassius carassius female and Gnathopogon elongatus elongatus male. Jpn. J. Ichthyol. 1990;36:419–426. doi: 10.1007/BF02905461. DOI

Kasama M., Kobayashi H. Hybridization experiment between Gnathopogon elongatus elongatus female and Carassius carassius male. Jpn. J. Ichthyol. 1991;38:295–300. doi: 10.1007/BF02905575. DOI

Hafez R., Labat R., Quillier R. Cytogenetic study of some species of Cyprinidae from the Midi-Pyrenees region. Bull. Soc. D’Hist. Nat. Toulouse. 1978;114:122–159.

Sofradžija A., Berberović L., Hadžiselimović R. Hromosomske garniture karaša (Carassius carassius) i babuške (Carassius auratus gibelio) Ichthyologia. 1978;10:135–148.

Mayr B., Ráb P., Kalat M. NORs and counterstain-enhanced fluorescence studies in Cyprinidae of different ploidy level. Genetica. 1986;69:111–118. doi: 10.1007/BF00115130. PubMed DOI

Kobayasi H., Ochi H., Takeuchi N. Chromosome studies in the genus Carassius: Comparison of C. auratus grandoculis, C. auratus buergeri, and C. auratus langsdorfii. Jpn. J. Ichthyol. 1973;20:6. doi: 10.11369/jji1950.20.7. DOI

Boron A. Karyotypes of diploid and triploid silver crucian carp Carassius auratus gibelio (Bloch) Cytobios. 1994;80:117–124.

Boron A., Szlachciak J., Juchno D., Grabowska A., Jagusztyn B., Porycka K. Karyotype, morphology, and reproduction ability of the Prussian carp, Carassius gibelio (Actinopterygii: Cypriniformes: Cyprinidae), from unisexual and bisexual populations in Poland. Acta Ichthyol. Piscat. 2011;41:19–28. doi: 10.3750/AIP2011.41.1.04. DOI

Kalous L., Bohlen J., Rylková K., Petrtýl M. Hidden diversity within the Prussian carp and designation of a neotype for Carassius gibelio (Teleostei: Cyprinidae) Ichthyol. Explor. Freshwaters. 2012;23:11–18.

Papoušek I., Vetešník L., Halačka K., Lusková V., Humpl M., Mendel J. Identification of natural hybrids of gibel carp Carassius auratus gibelio (Bloch) and crucian carp Carassius carassius (L.) from lower Dyje River floodplain (Czech Republic) J. Fish Biol. 2008;72:1230–1235. doi: 10.1111/j.1095-8649.2007.01783.x. DOI

Wouters J., Janson S., Lusková V., Olsén K.H. Molecular identification of hybrids of the invasive gibel carp Carassius auratus gibelio and crucian carp Carassius carassius in Swedish waters. J. Fish Biol. 2012;80:2595–2604. doi: 10.1111/j.1095-8649.2012.03312.x. PubMed DOI

King M. Species Evolution: The Role of Chromosome Change. Cambridge University Press; Cambridge, UK: 1993. p. 336.

Cremer T., Cremer C. Chromosome territories, nuclear architecture and gene regulation in mammalian cells. Nat. Rev. Genet. 2001;2:292–301. doi: 10.1038/35066075. PubMed DOI

Chevret E., Volpi E., Sheer D. Mini review: Form and function in the human interphase chromosome. Cytogenet. Genome Res. 2000;90:13–21. doi: 10.1159/000015654. PubMed DOI

Claussen U., Michel S., Mhlig P., Westermann M., Grummt U.W., Kromeyer-Hauschild K., Liehr T. Demystifying chromosome preparation and the implications for the concept of chromosome condensation during mitosis. Cytogenet. Genome Res. 2002;98:136–146. doi: 10.1159/000069817. PubMed DOI

Tjio J.H., Levan A. The chromosome number of man. Hereditas. 1956;42:1–6. doi: 10.1111/j.1601-5223.1956.tb03010.x. DOI

Zhu H.P., Ma D.M., Gui J.F. Triploid origin of the gibel carp as revealed by 5S rDNA localization and chromosome painting. Chromosome Res. 2006;14:767–776. doi: 10.1007/s10577-006-1083-0. PubMed DOI

Mantovani M., Abel L.D.D.S., Moreira-Filho O. Conserved 5S and variable 45S rDNA chromosomal localisation revealed by FISH in Astyanax scabripinnis (Pisces, Characidae) Genetica. 2005;123:211–216. doi: 10.1007/s10709-004-2281-3. PubMed DOI

Gromicho M., Coutanceau J.P., Ozouf-Costaz C., Collares-Pereira M.J. Contrast between extensive variation of 28S rDNA and stability of 5S rDNA and telomeric repeats in the diploid-polyploid Squalius alburnoides complex and in its maternal ancestor Squalius pyrenaicus (Teleostei, Cyprinidae) Chromosome Res. 2006;14:297–306. doi: 10.1007/s10577-006-1047-4. PubMed DOI

Schmid M., Vitelli L., Batistoni R. Chromosome banding in amphibia. XI. Constitutive heterochromatin, nucleolus organizers, 18S + 28S and 5S ribosomal RNA genes in Ascaphidae, Pipidae, Discoglossidae and Pelobatidae. Chromosoma. 1987;95:271–284. doi: 10.1007/BF00294784. PubMed DOI

Zhou L., Wang Y., Gui J.F. Genetic Evidence for Gonochoristic Reproduction in Gynogenetic Silver Crucian Carp (Carassius auratus gibelio Bloch) as Revealed by RAPD Assays. J. Mol. Evol. 2000;51:498–506. doi: 10.1007/s002390010113. PubMed DOI

Daněk T., Kalous L., Veselý T., Krásová E., Reschová S., Rylková K., Kulich P., Petrtýl M., Pokorová D., Knytl M. Massive mortality of Prussian carp Carassius gibelio in the upper Elbe basin associated with herpesviral hematopoietic necrosis (CyHV-2) Dis. Aquat. Org. 2012;102:87–95. doi: 10.3354/dao02535. PubMed DOI

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