Cytogenetics of the small-sized fish, Copeina guttata (Characiformes, Lebiasinidae): Novel insights into the karyotype differentiation of the family

Autoři: Gustavo Akira Toma aff001;  Renata Luiza Rosa de Moraes aff001;  Francisco de Menezes Cavalcante Sassi aff001;  Luiz Antonio Carlos Bertollo aff001;  Ezequiel Aguiar de Oliveira aff001;  Petr Rab aff003;  Alexandr Sember aff003;  Thomas Liehr aff004;  Terumi Hatanaka aff001;  Patrik Ferreira Viana aff005;  Manoela Maria Ferreira Marinho aff006;  Eliana Feldberg aff005;  Marcelo de Bello Cioffi aff001
Působiště autorů: Laboratório de Citogenética de Peixes, Departamento de Genética e Evolução, Universidade Federal de São Carlos, São Carlos, São Paulo, Brazil aff001;  Secretaria de Estado de Educação de Mato Grosso, Cuiabá, Mato Grosso, Brazil aff002;  Laboratory of Fish Genetics, Institute of Animal Physiology and Genetics, Czech Academy of Sciences, Liběchov, Czech Republic aff003;  Institute of Human Genetics, University Hospital Jena, Jena, Germany aff004;  Instituto Nacional de Pesquisas da Amazônia, Manaus, Amazonas, Brazil aff005;  Museu de Zoologia da Universidade de São Paulo, São Paulo, São Paulo, Brazil aff006
Vyšlo v časopise: PLoS ONE 14(12)
Kategorie: Research Article


Lebiasinidae is a small fish family composed by miniature to small-sized fishes with few cytogenetic data (most of them limited to descriptions of diploid chromosome numbers), thus preventing any evolutionary comparative studies at the chromosomal level. In the present study, we are providing, the first cytogenetic data for the red spotted tetra, Copeina guttata, including the standard karyotype, C-banding, repetitive DNA mapping by fluorescence in situ hybridization (FISH) and comparative genomic hybridization (CGH), providing chromosomal patterns and novel insights into the karyotype differentiation of the family. Males and females share diploid chromosome number 2n = 42 and karyotype composed of 2 metacentric (m), 4 submetacentric (sm) and 36 subtelocentric to acrocentric (st-a) chromosomes. Blocks of constitutive heterochromatin were observed in the centromeric and interstitial regions of several chromosomes, in addition to a remarkably large distal block, heteromorphic in size, which fully corresponded with the 18S rDNA sites in the fourth chromosomal pair. This overlap was confirmed by 5S/18S rDNA dual-color FISH. On the other hand, 5S rDNA clusters were situated in the long and short arms of the 2nd and 15th pairs, respectively. No sex-linked karyotype differences were revealed by male/female CGH experiments. The genomic probes from other two lebiasinid species, Lebiasina melanoguttata and Pyrrhulina brevis, showed positive hybridization signals only in the NOR region in the genome of C. guttata. We demonstrated that karyotype diversification in lebiasinids was accompanied by a series of structural and numeric chromosome rearrangements of different types, including particularly fusions and fissions.

Klíčová slova:

Comparative genomics – Cytogenetics – Fluorescent in situ hybridization – Hybridization – Chromosome mapping – Chromosome pairs – Karyotypes – Sex chromosomes


1. Albert JS, Reis RE. Historical Biogeography of Neotropical Freshwather Fishes. Berkeley: University of California Press; 2011. doi: 10.1525/california/9780520268685.001.0001

2. Reis RE, Albert JS, Di Dario F, Mincarone MM, Petry P, Rocha LA. Fish biodiversity and conservation in South America. J Fish Biol. 2016;89: 12–47. doi: 10.1111/jfb.13016 27312713

3. Schaefer SA. Conflict and resolution: impact of new taxa on phylogenetic studies of the Neotropical cascudinhos (Siluroidei: Loricariidae). In: Malabarba LR, Reis RE, Vari RP, Lucena ZMS, Lucena CAS, editors. Phylogeny and Classification of Neotropical fishes. Porto Alegre: Edipucrs; 1998. Pp. 375–400.

4. Pereira LHG, Hanner R, Foresti F, Oliveira C. Can DNA barcoding accurately discriminate megadiverse Neotropical freshwater fish fauna? BMC Genet. 2013;14: 1–14. doi: 10.1186/1471-2156-14-1

5. Pires AA, Ramirez JL, Galetti PM, Troy WP, Freitas PD. Molecular analysis reveals hidden diversity in Zungaro (Siluriformes: Pimelodidade): a genus of giant South American catfish. Genetica. 2017;145: 335–340. doi: 10.1007/s10709-017-9968-8 28501957

6. Prizon AC, Bruschi DP, Borin-Carvalho LA, Cius A, Barbosa LM, Ruiz HB, et al. Hidden diversity in the populations of the armored catfish Ancistrus Kner, 1854 (Loricariidae, Hypostominae) from the Paraná River Basin revealed by molecular and cytogenetic data. Front Genet. 2017;8: 185. doi: 10.3389/fgene.2017.00185 29225612

7. Ramirez JL, Birindelli JL, Carvalho DC, Affonso PRAM, Venere PC, Ortega H, et al. Revealing hidden diversity of the underestimated neotropical ichthyofauna: DNA barcoding in the recently described genus Megaleporinus (Characiformes: Anostomidae). Front Genet. 2017;8: 149. doi: 10.3389/fgene.2017.00149 29075287

8. Cioffi MB, Moreira-Filho O, Ráb P, Sember A, Molina WF, Bertollo LAC. Conventional Cytogenetic Approaches—Useful and Indispensable Tools in Discovering Fish Biodiversity. Curr Genet Med Rep. 2018;6: 176–186.

9. Cioffi MB, Bertollo LAC. Chromosomal distribution and evolution of repetitive DNAs in fish. In: Garrido-Ramos MA, editor. Repetitive DNA. Basel: Karger Publishers; 2012. pp. 197–221.

10. Blanco D, Vicari M, Lui R, Artoni R, Almeida M, Traldi J, et al. Origin of the X1X1X2X2/X1X2Y sex chromosome system of Harttia punctata (Siluriformes, Loricariidae) inferred from chromosome painting and FISH with ribosomal DNA markers. Genetica. 2014;142.

11. Schemberger MO, Nascimento VD, Coan R, Ramos É, Nogaroto V, Ziemniczak K, et al. DNA transposon invasion and microsatellite accumulation guide W chromosome differentiation in a Neotropical fish genome. Chromosoma. 2019; 1–14. doi: 10.1007/s00412-018-0679-4

12. Utsunomia R, de Andrade Silva DMZ, Ruiz-Ruano FJ, Goes CAG, Melo S, Ramos LP, et al. Satellitome landscape analysis of Megaleporinus macrocephalus (Teleostei, Anostomidae) reveals intense accumulation of satellite sequences on the heteromorphic sex chromosome. Sci Rep. 2019;9: 5856. doi: 10.1038/s41598-019-42383-8 30971780

13. Artoni RF, Castro JP, Jacobina UP, Lima-Filho PA, da Costa F, Werneck GW, et al. Inferring diversity and evolution in fish by means of integrative molecular cytogenetics. Sci World J. 2015; 365787.

14. Barbosa P, de Oliveira LA, Pucci MB, Santos MH, Moreira-Filho O, Vicari MR, et al. Identification and chromosome mapping of repetitive elements in the Astyanax scabripinnis (Teleostei: Characidae) species complex. Genetica. 2015;143: 55–62. doi: 10.1007/s10709-014-9813-2 25549800

15. Schemberger MO, Nogaroto V, Almeida MC, Artoni RF, Valente GT, Martins C, et al. Sequence analyses and chromosomal distribution of the Tc1/Mariner element in Parodontidae fish (Teleostei: Characiformes). Gene. 2016;593: 308–314. doi: 10.1016/j.gene.2016.08.034 27562083

16. Utsunomia R, Silva DMZ de A, Ruiz-Ruano FJ, Araya-Jaime C, Pansonato-Alves JC, Scacchetti PC, et al. Uncovering the ancestry of B chromosomes in Moenkhausia sanctaefilomenae (Teleostei, Characidae). PLoS One. 2016;11: e0150573. doi: 10.1371/journal.pone.0150573 26934481

17. Barros AV, Wolski MAV, Nogaroto V, Almeida MC, Moreira-Filho O, Vicari MR. Fragile sites, dysfunctional telomere and chromosome fusions: what is 5S rDNA role? Gene. 2017;608: 20–27. doi: 10.1016/j.gene.2017.01.013 28111257

18. Yano CF, Bertollo LAC, Rebordinos L, Merlo MA, Liehr T, Portela-Bens S, et al. Evolutionary dynamics of rDNAs and U2 small nuclear DNAs in Triportheus (Characiformes, Triportheidae): high variability and particular syntenic organization. Zebrafish. 2017; 14: 146–154. doi: 10.1089/zeb.2016.1351 28051362

19. de Oliveira EA, Sember A, Bertollo LAC, Yano CF, Ezaz T, Moreira-Filho O, et al. Tracking the evolutionary pathway of sex chromosomes among fishes: characterizing the unique XX/XY1Y2 system in Hoplias malabaricus (Teleostei, Characiformes). Chromosoma. 2018;127: 115–128. doi: 10.1007/s00412-017-0648-3 29124392

20. Borges AT, Cioffi MB, Bertollo LAC, Soares RX, Costa GWWF, Molina WF. Paracentric inversions differentiate the conservative karyotypes in two Centropomus species (Teleostei: Centropomidae). Cytogenet Genome Res. 2019;157:239–248. doi: 10.1159/000499748 30991393

21. Nakayama C, Jégu M, Porto JIR, Feldberg E. Karyological evidence for a cryptic species of piranha within Serrasalmus rhombeus (Characidae, Serrasalminae) in the Amazon. Copeia. 2001; 3: 866–869.

22. Milhomem SSR, Pieczarka JC, Crampton WGR, Silva DS, De Souza ACP, Carvalho JR, et al. Chromosomal evidence for a putative cryptic species in the Gymnotus carapo species-complex (Gymnotiformes, Gymnotidae). BMC Genet. 2008;9: 75. doi: 10.1186/1471-2156-9-75 19025667

23. Ferreira-Neto M, Artoni RF, Vicari MR, Moreira-Filho O, Camacho JPM, Bakkali M, et al. Three sympatric karyomorphs in the fish Astyanax fasciatus (Teleostei, Characidae) do not seem to hybridize in natural populations. Comp Cytogenet. 2012;6: 29–40. doi: 10.3897/CompCytogen.v6i1.2151 24260650

24. Ferreira M, Kavalco KF, de Almeida-Toledo LF, Garcia C. Cryptic diversity between two imparfinis species (Siluriformes, Heptapteridae) by cytogenetic analysis and DNA barcoding. Zebrafish. 2014;11: 306–317. doi: 10.1089/zeb.2014.0981 24937469

25. Ferreira M, Garcia C, Matoso DA, de Jesus IS, Cioffi M de B, Bertollo LAC, et al. The Bunocephalus coracoideus species complex (Siluriformes, Aspredinidae). Signs of a speciation process through chromosomal, genetic and ecological diversity. Front Genet. 2017;8: 120. doi: 10.3389/fgene.2017.00120 28983316

26. do Nascimento VD, Coelho KA, Nogaroto V, de Almeida RB, Ziemniczak K, Centofante L, et al. Do multiple karyomorphs and population genetics of freshwater darter characines (Apareiodon affinis) indicate chromosomal speciation? Zool Anz. 2018;272: 93–103. doi: 10.1016/j.jcz.2017.12.006

27. Gavazzoni M, Paiz LM, Oliveira CAM, Pavanelli CS, Graça WJ, Margarido VP. Morphologically cryptic species of the Astyanax bimaculatus “caudal peduncle spot” subgroup diagnosed through cytogenetic characters. Zebrafish. 2018;15: 382–388. doi: 10.1089/zeb.2018.1574 29634423

28. Nirchio M, Paim FG, Milana V, Rossi AR, Oliveira C. Identification of a new mullet species complex based on an integrative molecular and cytogenetic investigation of Mugil hospes (Mugilidae: Mugiliformes). Front Genet. 2018;9: 1–9. doi: 10.3389/fgene.2018.00001

29. Santos EO dos Deon GA, Almeida RB de, Oliveira EA de, Nogaroto V, Silva HP da, et al. Cytogenetics and DNA barcode reveal an undescribed Apareiodon species (Characiformes: Parodontidae). Genet Mol Biol. 2019; 42:365–373. doi: 10.1590/1678-4685-GMB-2018-0066 31259363

30. Neto CCM, Lima-Filho PA, Araújo WC, Bertollo LAC, Molina WF. Differentiated evolutionary pathways in Haemulidae (Perciformes): karyotype stasis versus morphological differentiation. Rev Fish Biol Fish. 2012;22: 457–465.

31. Barby F, Rab P, Lavoue S, Ezaz T, Bertollo LAC, Kilian A, et al. From chromosomes to genome: insights into the evolutionary relationships and biogeography of Old World knifefishes (Notopteridae; Osteoglossiformes). Genes. 2018; 9(6). pii: E306. doi: 10.3390/genes9060306 29921830

32. da Silva FA, Feldberg E, Carvalho NDM, Rangel SMH, Schneider CH, Carvalho-Zilse GA, et al. Effects of environmental pollution on the rDNAomics of Amazonian fish. Environ Pollut. 2019;252: 180–187. doi: 10.1016/j.envpol.2019.05.112 31146233

33. Soto MÁ, Castro JP, Walker LI, Malabarba LR, Santos MH, de Almeida MC, et al. Evolution of trans-Andean endemic fishes of the genus Cheirodon (Teleostei: Characidae) are associated with chromosomal rearrangements. Rev Chil Hist Nat. 2018;91: 8.

34. Weitzman M, Weitzman SH. Family Lebiasinidae. In: Reis RE Kullander SO, Ferraris CJ Jr, editors. Check List of the Freshwater fishes of South and Central America. Porto Alegre: Edipucrs. 2003; pp. 241–250.

35. Eschmeyer WN, Fricke R, van der Laan R. Catalog of fishes: Genera, species, references. California Academy of Sciences, San Francisco, USA. 2019. Available from:

36. Netto-Ferreira AL, Marinho MMF. New species of Pyrrhulina (Ostariophysi: Characiformes: Lebiasinidae) from the brazilian shield, with comments on a putative monophyletic group of species in the genus. Zootaxa. 2013;3664: 369–376. doi: 10.11646/zootaxa.3664.3.7 26266308

37. Scheel JJ. Fish chromosomes and their evolution. Intern Rep Danmarks Akvar. 1973;22.

38. Arefjev VA. Karyotypic diversity of characidae families (Pisces, characidae). Caryologia. 1990;43: 291–304. doi: 10.1080/00087114.1990.10797008

39. Oliveira C, Andreata AA, Toledo LFA, Toledo SA. Karyotype and nucleolus organizer regions of Pyrrhulina cf australis (Pisces, Characiformes, Lebiasinidae). Rev Bras Genética. 1991; 685–690.

40. Oliveira MIB, Sanguino ECB, Falcão JN. Estudos citogenéticos em Pyrrhulina sp. Teleostei, Characiformes, Lebiasinidae) IV Simpósio de Citogenética Evolutiva e Aplicada de Peixes Neotropicais. 1992;13.

41. Arai R. Fish karyotype a check list. Japan: Springer press; 2011. doi: 10.1007/978-4-431-53877-6

42. Sassi F de MC, Oliveira EA de, Bertollo LAC, Nirchio M, Hatanaka T, Marinho MMF, et al. Chromosomal evolution and evolutionary relationships of Lebiasina species (Characiformes, Lebiasinidae). Int J Mol Sci. 2019;20: 2944.

43. Moraes RLR, Bertollo LAC, Marinho MMF, Yano CF, Hatanaka T, Barby FF, et al. Evolutionary relationships and cytotaxonomy cin the genus Pyrrhulina (Characiformes, Lebiasinidae). Zebrafish. 2017;00: zeb.2017.1465. doi: 10.1089/zeb.2017.1465 28767325

44. Moraes RLR, Sember A, Bertollo LAC, De Oliveira EA, Ráb P, Hatanaka T, et al. Comparative cytogenetics and neo-Y formation in small-sized fish species of the genus Pyrrhulina (Characiformes, Lebiasinidae). Front Genet. 2019;10: 678. doi: 10.3389/fgene.2019.00678 31428127

45. Bertollo LAC, Cioffi MB, Moreira-Filho O. Direct chromosome preparation from freshwater teleost fishes. In: Ozouf-Costaz C, Pisano E, Foresti F, Almeida Toledo LF, editors. Fish cytogenetic techniques (Chondrichthyans and Teleosts). Enfield USA: CRC Press; 2015. pp. 21–26. doi: 10.1201/b18534-4

46. Sumner AT. A simple technique for demonstrating centromeric heterochromatin. Exp Cell Res. 1972;75: 304–306. doi: 10.1016/0014-4827(72)90558-7 4117921

47. Pendás AM, Morán P, Garcia-Vázquez E. Ribosomal RNA genes are interspersed throughout a heterochromatic chromosome arm in Atlantic salmon. Cytogenet Genome Res. 1993;63: 128–130.

48. Cioffi MB, Martins C, Centofante L, Jacobina U, Bertollo LAC. Chromosomal Variability among Allopatric Populations of Erythrinidae Fish Hoplias malabaricus: Mapping of three classes of repetitive DNAs. Cytogenet Genome Res. 2009;125: 132–141. Available from: doi: 10.1159/000227838 19729917

49. Sambrook J, Russell DW. Molecular cloning: a laboratory manual. 3rd ed. New York, USA: Cold Spring Harbor Laboratory Press; 2001.

50. Zwick MS, Hanson RE, Mcknight TD, Islam-Faridi MH, Stelly DM, Wing RA, et al. A rapid procedure for the isolation of C 0 t-1 DNA from plants. Genome. 1997;40: 138–142. doi: 10.1139/g97-020 18464813

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

52. Naorem S, Bhagirath T. Chromosomal differentiations in the evolution of channid fishes–molecular genetic perspective. Caryologia. 2006;59:235–40.

53. Cioffi MB, Bertollo LAC, Villa MA, Oliveira EA, Tanomtong A, Yano CF. Genomic organization of repetitive DNA elements and its implications for the chromosomal evolution of channid fishes (Actinopterygii, Perciformes). PLoS One. 2015;10(6):e0130199. doi: 10.1371/journal.pone.0130199 26067030

54. Lowry DB, Willis JH. A widespread chromosomal inversion polymorphism contributes to a major life-history transition, local adaptation, and reproductive isolation. PLoS Biol. 2010;8: e1000500. doi: 10.1371/journal.pbio.1000500 20927411

55. Ortiz-Barrientos D, Engelstädter J, Rieseberg LH. Recombination rate evolution and the origin of species. Trends Ecol Evol. 2016;31: 226–236. doi: 10.1016/j.tree.2015.12.016 26831635

56. Kirkpatrick M. The evolution of genome structure by natural and sexual selection. J Hered. 2017;108: 3–11. doi: 10.1093/jhered/esw041 27388336

57. Jay P, Whibley A, Frézal L, Rodríguez de Cara MÁ, Nowell RW, Mallet J, et al. Supergene evolution triggered by the introgression of a chromosomal inversion. Curr Biol. 2018;28: 1839–1845.e3. doi: 10.1016/j.cub.2018.04.072 29804810

58. Mérot C, Berdan EL, Babin C, Normandeau E, Wellenreuther M, Bernatchez L. Intercontinental karyotype-environment parallelism supports a role for a chromosomal inversion in local adaptation in a seaweed fly. Proc R Soc B Biol Sci. 2018; 285: 20180519. doi: 10.1098/rspb.2018.0519 29925615

59. Supiwong W, Pinthong K, Seetapan K, Saenjundaeng P, Bertollo LAC, de Oliveira EA, et al. Karyotype diversity and evolutionary trends in the Asian swamp eel Monopterus albus (Synbranchiformes, Synbranchidae): a case of chromosomal speciation? BMC Evol Biol. 2019;19: 73. doi: 10.1186/s12862-019-1393-4 30849933

60. Roussel P, André C, Comai L, Hernandez-Verdun D. The rDNA transcription machinery is assembled during mitosis in active NORs and absent in inactive NORs. J Cell Biol. 1996;133: 235 LP– 246. doi: 10.1083/jcb.133.2.235 8609158

61. Collares-Pereira MJ, Ráb P. NOR polymorphism in the Iberian species Chondrostoma lusitanicum (Pisces: Cyprinidae)–re-examination by FISH. Genetica. 1999;105: 301–303. doi: 10.1023/a:1003885922023 10761113

62. Nirchio M, Róndon R, Oliveira C, Ferreira IA, Martins C, Pérez J, et al. Cytogenetic studies in three species of Lutjanus (Perciformes: Lutjanidae: Lutjaninae) from the Isla Margarita, Venezuela. Neotrop Ichthyol. 2008;6: 101–108.

63. Ghigliotti L, Near TJ, Ferrando S, Vacchi M, Pisano E. Cytogenetic diversity in the Antarctic plunderfishes (Notothenioidei: Artedidraconidae). Antarct Sci. 2010;22: 805–814. doi: 10.1017/S0954102010000660

64. Sochorová J, Garcia S, Gálvez F, Symonová R, Kovařík A. Evolutionary trends in animal ribosomal DNA loci: introduction to a new online database. Chromosoma. 2018;127: 141–150. doi: 10.1007/s00412-017-0651-8 29192338

65. Martins C, Galetti PM. Two 5S rDNA arrays in Neotropical fish species: is it a general rule for fishes? Genetica. 2001;111: 439–446. doi: 10.1023/a:1013799516717 11841188

66. Foster HA, Bridger JM. The genome and the nucleus: a marriage made by evolution. Chromosoma. 2005;114: 212–229. doi: 10.1007/s00412-005-0016-6 16133352

67. Garrido-Ramos MA. Satellite DNA: an evolving topic. Genes. 2017: 8:230. doi: 10.3390/genes8090230 28926993

68. Reichwald K, Petzold A, Koch P, Downie BR, Hartmann N, Pietsch S, et al. Insights into sex chromosome evolution and aging from the genome of a short-lived fish. Cell. 2015;163: 1527–1538. doi: 10.1016/j.cell.2015.10.071 26638077

69. Krysanov E, Demidova T. Extensive karyotype variability of African fish genus Nothobranchius (Cyprinodontiformes). Comp Cytogenet. 2018;12: 387–402. doi: 10.3897/CompCytogen.v12i3.25092 30338046

70. De Souza E Sousa JF, Viana PF, Bertollo LAC, Cioffi MB, Feldberg E. Evolutionary relationships among Boulengerella Species (Ctenoluciidae, Characiformes): genomic organization of repetitive DNAs and highly conserved karyotypes. Cytogenet Genome Res. 2017; 152: 194–203. doi: 10.1159/000480141 28942442

Článek vyšel v časopise


2019 Číslo 12
Nejčtenější tento týden