The genus Colomesus is the sole representative of the family Tetraodontidae in the Amazon region. Here, Colomesus asellus was analyzed using conventional and molecular cytogenetic protocols. Its diploid chromosome number is 2n = 46 with 12 meta-, 10 submeta-, 16 subtelo-, and 8 acrocentric chromosomes and a fundamental number of FN = 84. An XX/XY sex chromosome system was identified. Mapping of 18S rDNA correlated with the nucleolus organizer regions (Ag-NORs) in the short arms of the 2 X chromosomes in females and in the Y chromosome in males. C-banding revealed heterochromatin in the centromeric regions of all chromosomes, except for pair 3. Prominent sex chromosome-specific heterochromatin amplification was observed, covering the short arms of the Y chromosome almost entirely. FISH with telomeric and tropomyosin (tpm1) sequences, respectively, revealed terminal signals in all chromosomes. The analysis of extended DNA fibers confirmed the colocalization and the interspersed pattern of the telomeric and tpm1 sequences. Thus, this study highlights the remarkable evolutionary dynamism presented by the Amazonian puffer fish regarding the differentiation of a heteromorphic XY sex chromosome system and a particular sex-specific amplification of rDNA sites. This is the first record of such an association in the Tetraodontidae family.

Abramyan J, Ezaz T, Graves JAM, Koopman P: Z and W sex chromosomes in the cane toad (Bufo marinus). Chromosome Res 17:1015-1024 (2009).
Alves AL, Porto-Foresti F, Oliveira C, Foresti F: Supernumerary chromosomes in the pufferfish Sphoeroides spengleri: first occurrence in marine teleostean Tetraodontiformes fish. Genet Mol Biol 31:243-245 (2008).
Amaral CR, Brito PM, Silva DA, Carvalho EF: A new cryptic species of South American freshwater pufferfish of the genus Colomesus (Tetraodontidae), based on both morphology and DNA data. PLoS One 8:e74397 (2013).
Arai R: Karyological and osteological approach to phylogenetic systematics of tetraodontiform fishes. Bull Natl Sci Mus Tokyo 9:175-210 (1983).
Arai R, Nagaiwa K: Chromosomes of tetraodontiform fishes from Japan. Bull Natl Sci Mus Tokyo Ser A 2:59-72 (1976).
Arnason U, Allderdice PW, Lien J, Widegren B: Highly repetitive DNA in the baleen whale genera Balaenoptera and Megaptera. J Mol Evol 27:217-221 (1988).
Azzalin CM, Nergadze SG, Giulotto E: Human intrachromosomal telomeric-like repeats: sequence organization and mechanisms of origin. Chromosoma 110:75-82 (2001).
Bergero R, Charlesworth D: The evolution of restricted recombination in sex chromosomes. Trends Ecol Evol 24:94-102 (2009).
Bloch ME: Blochii Systema Ichthyologiae iconibus CX illustratum. Post obitum auctoris opus inchoatum absolvit, correxit, interpolavit JG Schneider (Berolini, Sumtibus Auctoris Impressum et Bibliopolio Sanderiano Commissum, Berlin 1801).
Born GG, Bertollo LAC: An XX/XY sex chromosome system in a fish species, Hoplias malabaricus, with a polymorphic NOR-bearing X chromosome. Chromosome Res 8:111-118 (2000).
Brainerd EL, Slutz SS, Hall EK, Phillis RW: Patterns of genome size evolution in tetraodontiform fishes. Evolution 55:2363-2368 (2001).
Brum MJI: Cytogenetic studies of Brazilian marine fish. Braz J Genet 19:421-427 (1996).
Brum MJI: Cytogenetic studies in tetraodontiforms Sphoeroides tyleri (Tetraodontidae) and Chilomycterus spinosus (Diodontidae) from Rio de Janeiro Coast, Brazil. Chromosome Sci 4:103-105 (2000).
Brum MJI, Mota LCG: C-banding and nucleolar organizer regions of Sphoeroides greeleyi (Tetraodontidae, Tetraodontiformes). Caryologia 55:171-174 (2002).
Choudhury RC, Prasad R, Das CC: Karyological studies in five tetraodontiform fishes from the Indian Ocean. Copeia 1982:728-732 (1982).
Cioffi MB, Martins C, Vicari MR, Rebordinos L, Bertollo LAC: Differentiation of the XY sex chromosomes in the fish Hoplias malabaricus (Characiformes, Erythrinidae): unusual accumulation of repetitive sequences on the X chromosome. Sex Dev 4:176-185 (2010).
Cioffi MB, Sánchez A, Marchal JA, Kosyakova N, Liehr T, et al: Whole chromosome painting reveals independent origin of sex chromosomes in closely related forms of a fish species. Genetica 139:1065-1072 (2011).
de Barros AV, Sczepanski TS, Cabrero J, Camacho JPM, Vicari MR, Artoni RF: Fiber FISH reveals different patterns of high-resolution physical mapping for repetitive DNA in fish. Aquaculture 322-323:47-50 (2011).
de Lima LCB, Martinez PA, Molina WF: Cytogenetic characterization of three Balistoidea fish species from the Atlantic with inferences on chromosomal evolution in the families Monacanthidae and Balistidae. Comp Cytogenet 5:61-69 (2011).
Elmerot C, Arnason U, Gojobori T, Janke A: The mitochondrial genome of the pufferfish, Fugu rubripes, and ordinal teleostean relationships. Gene 295:163-172 (2002).
Ezaz T, Sarre SD, O'Meally D, Graves JAM, Georges A: Sex chromosome evolution in lizards: independent origins and rapid transitions. Cytogenet Genome Res 127:249-260 (2009).
Ferreira M, Garcia C, Matoso DA, de Jesus IS, Feldberg E: A new multiple sex chromosome system X1X1X2X2/X1Y1X2Y2 in Siluriformes: cytogenetic characterization of Bunocephalus coracoideus (Aspredinidae). Genetica 144:591-599 (2016).
Fischer C, Ozouf-Costaz C, Crollius H, Dasilva C, Jaillon O, et al: Karyotype and chromosome location of characteristic tandem repeats in the pufferfish Tetraodon nigroviridis. Cytogenet Genome Res 88:50-55 (2000).
Friesen VL, Congdon BC, Kidd MG, Birt TP: Polymerase chain reaction (PCR) primers for the amplification of five nuclear introns in vertebrates. Mol Ecol 8:2147-2149 (1999).
Gagat M, Grzanka D, Izdebska M, Sroka WD, Hałas-Wiśniewska M, Grzanka A: Tropomyosin-1 protects transformed alveolar epithelial cells against cigaret smoke extract through the stabilization of F-actin-dependent cell-cell junctions. Acta Histochem 118:225-235 (2016).
Galetti PM Jr, Molina WF, Affonso PR, Aguilar CT: Assessing genetic diversity of Brazilian reef fishes by chromosomal and DNA markers. Genetica 126:161-177 (2006).
Galvis G, Mojica JI, Duque SR, Castellanos C, Sánchez-Duarte P, et al: Peces del Medio Amazonas, Región de Leticia. Serie de Guías Tropicales de campo, No. 5. Conservación Internacional (Editorial Panamericana, Formas e Impresos, Bogotá 2006).
Garrido-Ramos MA, de la Herrán R, Ruiz-Rejón C, Ruiz-Rejón M: A satellite DNA of the Sparidae family (Pisces, Perciformes) associated with telomeric sequences. Cytogenet Cell Genet 83:3-9 (1998).
Getlekha N, Molina WF, Cioffi MB, Yano CF, Maneechot N, et al: Repetitive DNAs highlight the role of chromosomal fusions in the karyotype evolution of Dascyllus species (Pomacentridae, Perciformes). Genetica 144:203-211 (2016).
Gross MC, Schneider CH, Valente GT, Porto JIR, Martins C, Feldberg E: Comparative cytogenetic analysis of the genus Symphysodon (Discus fishes, Cichlidae): chromosomal characteristics of retrotransposons and minor ribosomal DNA. Cytogenet Genome Res 127:43-53 (2009).
Gross MC, Schneider CH, Valente GT, Porto JIR, Martins C, Feldberg E: Variability of 18S rDNA locus among Symphysodon fishes: chromosomal rearrangements. J Fish Biol 76:1117-1127 (2010).
Guimarães EMC, Carvalho NDM, Schneider CH, Feldberg E, Gross MC: Karyotypic comparison of Hoplias malabaricus (Bloch, 1794) (Characiformes, Erythrinidae) in Central Amazon. Zebrafish 14:80-89 (2017).
Henning F, Trifonov V, Ferguson-Smith MA, de Almeida-Toledo LF: Non-homologous sex chromosomes in two species of the genus Eigenmannia (Teleostei: Gymnotiformes). Cytogenet Genome Res 121:55-58 (2008).
Holcroft NI: A molecular analysis of the interrelationships of tetraodontiform fishes (Acanthomorpha: Tetraodontiformes). Mol Phylogenet Evol 34:525-544 (2005).
Howell WM, Black DA: Controlled silver-staining of nucleolus organizer regions with a protective colloidal developer: a 1-step method. Experientia 36:1014-1015 (1980).
Ijdo JW, Wells RA, Baldini A, Reeders ST: Improved telomere detection using a telomere repeat probe (TTAGGG)n generated by PCR. Nucleic Acids Res 19:4780 (1991).
Janco M, Suphamungmee W, Li X, Lehman W, Lehrer S, Geeves M: Polymorphism in tropomyosin structure and function. J Muscle Res Cell Motil 34:177-187 (2013).
Kamiya T, Kai W, Tasumi S, Oka A, Matsunaga T, et al: A trans-species missense SNP in Amhr2 is associated with sex determination in the tiger pufferfish, Takifugu rubripes (fugu). PLoS Genet 8:e1002798 (2012).
Kawagoshi T, Uno Y, Matsubara K, Matsuda Y, Nishida C: The ZW micro-sex chromosomes of the Chinese soft-shelled turtle (Pelodiscus sinensis, Trionychidae, Testudines) have the same origin as chicken chromosome 15. Cytogenet Genome Res 125:125-131 (2009).
Kawagoshi T, Uno Y, Nishida C, Matsuda Y: The Staurotypus turtles and Aves share the same origin of sex chromosomes but evolved different types of heterogametic sex determination. PLoS One 9:e105315 (2014).
Kawai A, Nishida-Umehara C, Ishijima J, Tsuda Y, Ota H, Matsuda Y: Different origins of bird and reptile sex chromosomes inferred from comparative mapping of chicken Z-linked genes. Cytogenet Genome Res 117:92-102 (2007).
Kikuchi K, Kai W, Hosokawa A, Mizuno N, Suetake H, et al: The sex-determining locus in the tiger pufferfish, Takifugu rubripes. Genetics 175:2039-2042 (2007).
Kullander SO: Family Tetraodontidae, in Reis RE, Kullander SO, Ferraris CJ Jr (eds): Check List of the Freshwater Fishes of South and Central America, pp 670-670 (Edipucrs, Porto Alegre 2003).
Lauder GV, Liem KF: The evolution and interrelationships of the actinopterygian fishes. Bull Mus Comp Zool 150:95-197 (1983).
Lee MR, Elder FFB: Yeast stimulation of bone marrow mitosis for cytogenetic investigations. Cytogenet Cell Genet 26:36-40 (1980).
Levan A, Fredga K, Sandberg AA: Nomenclature for centromeric position on chromosomes. Hereditas 52:201-220 (1964).
Mandrioli M: Mariner-like transposable elements are interspersed within the rDNA-associated heterochromatin of the pufferfish Tetraodon fluviatilis. Chromosome Res 8:177-179 (2000).
Mandrioli M, Manicardi GC: Cytogenetics and molecular analysis of the pufferfish Tetraodon fluviatilis (Osteichthyes). Genetica 111:433-438 (2001).
Mank JE: The W, X, Y and Z of sex-chromosome dosage compensation. Trends Genet 25:226-233 (2009).
Mank JE, Ellegren H: Parallel divergence and degradation of the avian W sex chromosome. Trends Ecol Evol 22:389-391 (2007).
Mank JE, Promislow DEL, Avise JC: Evolution of alternative sex-determining mechanisms in teleost fishes. Biol J Linn Soc 87:83-93 (2006).
Marston S, Gautel M: Introducing a special edition of the Journal of Muscle Research and Cell Motility on tropomyosin: form and function. J Muscle Res Cell Motil 34:151-153 (2013).
Martinez PA: Processos carioevolutivos na Ordem Tetraodontiformes: uma visão através de suas diferentes linhagens. Dissertação, Universidade Federal do Rio Grande do Norte, Centro de Biociências, Programa de Pós-Graduação em Ecologia (2010).
Martinez PA, de Araujo WC, Molina WF: Derived cytogenetic traits, multiple NORs and B chromosomes in the compact karyotype of Canthigaster figueiredoi (Tetraodontiformes). Mar Genomics 3:85-89 (2010).
Matsubara K, Tarui H, Toriba M, Yamada K, Nishida-Umehara C, et al: Evidence for different origin of sex chromosomes in snakes, birds, and mammals and step-wise differentiation of snake sex chromosomes. Proc Natl Acad Sci USA 103:18190-18195 (2006).
Matsubara K, Sarre SD, Georges A, Matsuda Y, Marshall Graves JA, Ezaz T: Highly differentiated ZW sex microchromosomes in the Australian Varanus species evolved through rapid amplification of repetitive sequences. PLoS One 9:e95226 (2014).
Matsubara K, Uno Y, Srikulnath K, Matsuda Y, Miller E, Olsson M: No interstitial telomeres on autosomes but remarkable amplification of telomeric repeats on the W sex chromosome in the sand lizard (Lacerta agilis). J Hered 106:753-757 (2015).
Matsuura K: Taxonomy and systematics of tetraodontiform fishes: a review focusing primarily on progress in the period from 1980 to 2014. Ichthyol Res 62:72-113 (2015).
Monsch KA: Miocene fish faunas from the northwestern Amazonia basin (Colombia, Peru, Brazil) with evidence of marine incursions. Palaeogeogr Palaeoclimatol Palaeoecol 143:31-50 (1998).
Muller HJ: A gene for the fourth chromosome of Drosophila. J Exp Zool 17:325-336 (1914).
Müller J, Troschel FH: Fische, in Schomburgk R (ed): Reisen in Britisch-Guiana in den Jahren 1840-1844. Im Auftrag Sr. Mäjestat des Königs von Preussen ausgeführt von Richard Schomburgk, pp 618-644 (Verlagsbuchhandlung J.J. Weber, Leipzig 1849).
Neafsey DE, Palumbi SR: Genome size evolution in pufferfish: a comparative analysis of diodontid and tetraodontid pufferfish genomes. Genome Res 13:821-830 (2003).
Nelson JS, Grande TC, Wilson MV: Fishes of the World (John Wiley & Sons, Hoboken 2016).
Nergadze SG, Santagostino MA, Salzano A, Mondello C, Giulotto E: Contribution of telomerase RNA retrotranscription to DNA double-strand break repair during mammalian genome evolution. Genome Biol 8:R260 (2007).
Noleto RB, Vicari MR, Cipriano RR, Artoni RF, Cestari MM: Physical mapping of 5S and 45S rDNA loci in pufferfishes (Tetraodontiformes). Genetica 130:133-138 (2007).
Noleto RB, Guimarães FSF, Paludo KS, Vicari MR, Artoni RF, Cestari MM: Genome size evaluation in tetraodontiform fishes from the Neotropical region. Mar Biotechnol 11:680-685 (2009).
Noleto RB, Vicari MR, Cestari MM, Artoni RF: Variable B chromosomes frequencies between males and females of two species of pufferfishes (Tetraodontiformes). Rev Fish Biol Fisheries 22:343-349 (2012).
Ohno S: Sex Chromosomes and Sex-Linked Genes (Springer-Verlag, Berlin 1967).
Oliveira JS, Fernandes SCR, Schwartz CA, Bloch C, Melo JAT, et al: Toxicity and toxin identification in Colomesus asellus, an Amazonian (Brazil) freshwater puffer fish. Toxicon 48:55-63 (2006).
O'Meally D, Patel HR, Stiglec R, Sarre SD, Georges A, et al: Non-homologous sex chromosomes of birds and snakes share repetitive sequences. Chromosome Res 18:787-800 (2010).
Perry SV: Vertebrate tropomyosin: distribution, properties and function. J Muscle Res Cell Motil 22:5-49 (2001).
Pinheiro VSDS, Carvalho NDM, do Carmo EJ, Schneider CH, Feldberg E, Gross MC: Karyoevolution in Potamorhina (Cope, 1878) (Ostariophysi, Curimatidae): using repetitive DNA for the elucidation of genome organization. Zebrafish 13:118-131 (2016).
Pinkel D, Straume T, Gray JW: Cytogenetic analysis using quantitative, high-sensitivity, fluorescence hybridization. Proc Natl Acad Sci USA 83:2934-2938 (1986).
Pokorná M, Kratochvíl L, Kejnovský E: Microsatellite distribution on sex chromosomes at different stages of heteromorphism and heterochromatinization in two lizard species (Squamata: Eublepharidae: Coleonyx elegans and Lacertidae: Eremias velox). BMC Genet 12:90 (2011).
Reed KM, Phillips RB: Polymorphism of the nucleolus organizer region (NOR) on the putative sex chromosomes of Arctic char (Salvelinus alpinus) is not sex related. Chromosome Res 5:221-227 (1997).
Ruiz-Herrera A, Nergadze SG, Santagostino M, Giulotto E: Telomeric repeats far from the ends: mechanisms of origin and role in evolution. Cytogenet Genome Res 122:219-228 (2008).
Sá-Gabriel LG, Molina WF: Sex chromosomes in Stephanolepis hispidus (Monacanthidae, Tetraodontiformes). Cytologia 69:447-452 (2004).
Sá-Gabriel LG, Molina WF: Karyotype diversification in fishes of the Balistidae, Diodontidae and Tetraodontidae (Tetraodontiformes). Caryologia 58:229-237 (2005).
Sambrook J, Russell DW: Molecular Cloning: A Laboratory Manual, ed 3 (Cold Spring Harbor Laboratory Press, Cold Spring Harbor 2001).
Santini F, Tyler JC: A phylogeny of the families of fossil and extant tetraodontiform fishes (Acanthomorpha, Tetraodontiformes), Upper Cretaceous to recent. Zool J Linn Soc 139:565-617 (2003).
Santini F, Nguyen NTT, Sorenson L, Waltzek TB, Lynch Alfaro JW, et al: Do habitat shifts drive diversification in teleost fishes? An example from the pufferfishes (Tetraodontidae). J Evol Biol 26:1003-1018 (2013).
Santos FA, Marques DF, Terencio ML, Feldberg F, Rodrigues LRR: Cytogenetic variation of repetitive DNA elements in Hoplias malabaricus (Characiformes - Erythrinidae) from white, black and clear water rivers of the Amazon basin. Genet Mol Biol 39:40-48 (2016).
Scacchetti PC, Utsunomia R, Pansonato-Alves JC, da Costa Silva GJ, Vicari MR, et al: Repetitive DNA sequences and evolution of ZZ/ZW sex chromosomes in Characidium (Teleostei: Characiformes). PLoS One 10:e0137231 (2015).
Schneider CH, Gross MC, Terencio ML, Artoni RF, Vicari MR, et al: Chromosomal evolution of neotropical cichlids: the role of repetitive DNA sequences in the organization and structure of karyotype. Rev Fish Biol Fisheries 23:201-214 (2013a).
Schneider CH, Gross MC, Terencio ML, Carmo EJ, Martins C, Feldberg E: Evolutionary dynamics of retrotransposable elements Rex1, Rex3 and Rex6 in neotropical cichlid genomes. BMC Evol Biol 13:152 (2013b).
Sumner AT: A simple technique for demonstrating centromeric heterochromatin. Exp Cell Res 75:304-306 (1972).
Tyler JC: A diagnosis of the two species of South American puffer fishes (Tetraodontidae, Plectognathi) of the genus Colomesus. Proc Acad Nat Sci Philadelphia 116:119-148 (1964).
Viana PF, Ribeiro LB, Souza GM, Chalkidis HM, Gross MC, Feldberg E: Is the karyotype of neotropical boid snakes really conserved? Cytotaxonomy, chromosomal rearrangements and karyotype organization in the Boidae family. PLoS One 11:e0160274 (2016).
Vieira FG, Souza DS, Freitas MHM, Mota ACF: Tetraodontidae. Peixes do rio Madeira, vol 3, ed 1, pp 403-405 (Dialeto, São Paulo 2013).
Wright AE, Dean R, Zimmer F, Mank JE: How to make a sex chromosome. Nat Commun 7:12087 (2016).
Yamanoue Y, Miya M, Doi H, Mabuchi K, Sakai H, Nishida M: Multiple invasions into freshwater by pufferfishes (Teleostei: Tetraodontidae): a mitogenomic perspective. PLoS One 6:e17410 (2011).
Yano CF, Bertollo LAC, Liehr T, Troy WP, Cioffi MB: W chromosome dynamics in Triportheus species (Characiformes, Triportheidae): an ongoing process narrated by repetitive sequences. J Hered 107:342-348 (2016a).
Yano CF, Bertollo LAC, Ezaz T, Trifonov V, Sember A, et al: Highly conserved Z and molecularly diverged W chromosomes in the fish genus Triportheus (Characiformes, Triportheidae). Heredity 118:276-283 (2016b).
Copyright / Drug Dosage / Disclaimer
Copyright: All rights reserved. No part of this publication may be translated into other languages, reproduced or utilized in any form or by any means, electronic or mechanical, including photocopying, recording, microcopying, or by any information storage and retrieval system, without permission in writing from the publisher.
Drug Dosage: The authors and the publisher have exerted every effort to ensure that drug selection and dosage set forth in this text are in accord with current recommendations and practice at the time of publication. However, in view of ongoing research, changes in government regulations, and the constant flow of information relating to drug therapy and drug reactions, the reader is urged to check the package insert for each drug for any changes in indications and dosage and for added warnings and precautions. This is particularly important when the recommended agent is a new and/or infrequently employed drug.
Disclaimer: The statements, opinions and data contained in this publication are solely those of the individual authors and contributors and not of the publishers and the editor(s). The appearance of advertisements or/and product references in the publication is not a warranty, endorsement, or approval of the products or services advertised or of their effectiveness, quality or safety. The publisher and the editor(s) disclaim responsibility for any injury to persons or property resulting from any ideas, methods, instructions or products referred to in the content or advertisements.
You do not currently have access to this content.