cells Article Revisiting the Karyotypes of Alligators and Caimans (Crocodylia, Alligatoridae) after a Half-Century Delay: Bridging the Gap in the Chromosomal Evolution of Reptiles Vanessa C. S. Oliveira 1 , Marie Altmanová 2,3 , Patrik F. Viana 4 , Tariq Ezaz 5 , Luiz A. C. Bertollo 1 , Petr Ráb 3 , Thomas Liehr 6, * , Ahmed Al-Rikabi 6 , Eliana Feldberg 4 , Terumi Hatanaka 1 , Sebastian Scholz 7 , Alexander Meurer 8 and Marcelo de Bello Cioffi 1   Citation: Oliveira, V.C.S.; Altmanová,M.; Viana, P.F.; Ezaz, T.; Bertollo, L.A.C.; Ráb, P.; Liehr, T.; Al-Rikabi, A.; Feldberg, E.; Hatanaka, T.; et al. Revisiting the Karyotypes of Alligators and Caimans (Crocodylia, Alligatoridae) after a Half-Century Delay: Bridging the Gap in the Chromosomal Evolution of Reptiles. Cells 2021, 10, 1397. https://doi.org/ 10.3390/cells10061397 Academic Editor: Terje Raudsepp Received: 11 May 2021 Accepted: 2 June 2021 Published: 5 June 2021 Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affil- iations. Copyright: © 2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/). 1 Laboratório de Citogenética de Peixes, Departamento de Genética e Evolução, Universidade Federal de São Carlos, São Carlos 13565-905, Brazil; vanessacristina.sales@gmail.com (V.C.S.O.); bertollo@ufscar.br (L.A.C.B.); hterumi@yahoo.com.br (T.H.); mbcioffi@ufscar.br (M.d.B.C.) 2 Department of Ecology, Faculty of Science, Charles University, 12844 Prague, Czech Republic; altmanova.m@gmail.com 3 Laboratory of Fish Genetics, Institute of Animal Physiology and Genetics, Czech Academy of Sciences, 27721 Libˇ echov, Czech Republic; rab@iapg.cas.cz 4 Laboratório de Genética Animal, Coordenação de Biodiversidade, Instituto Nacional de Pesquisas da Amazônia, Manaus 69083-000, Brazil; patrik.biologia@gmail.com (P.F.V.); feldberg@inpa.gov.br (E.F.) 5 Institute for Applied Ecology, Faculty of Science and Technology, University of Canberra, Bruce, ACT 2617, Australia; tariq.ezaz@canberra.edu.au 6 Institute of Human Genetics, Jena University Hospital, Am Klinikum 1, 07747 Jena, Germany; ahmedgenetic@hotmail.com 7 An der Nachtweide 16, 60433 Frankfurt, Germany; chinemys@web.de 8 Alfred Nobel Strasse 1e, 55411 Bingen am Rhein, Germany; ameurer@online.de * Correspondence: Thomas.Liehr@med.uni-jena.de; Tel.: +49-36-41-939-68-50; Fax: +49-3641-93-96-852 Abstract: Although crocodilians have attracted enormous attention in other research fields, from the cytogenetic point of view, this group remains understudied. Here, we analyzed the karyotypes of eight species formally described from the Alligatoridae family using differential staining, flu- orescence in situ hybridization with rDNA and repetitive motifs as a probe, whole chromosome painting (WCP), and comparative genome hybridization. All Caimaninae species have a diploid chromosome number (2n) 42 and karyotypes dominated by acrocentric chromosomes, in contrast to both species of Alligatorinae, which have 2n = 32 and karyotypes that are predominantly metacentric, suggesting fusion/fission rearrangements. Our WCP results supported this scenario by revealing the homeology of the largest metacentric pair present in both Alligator spp. with two smaller pairs of acrocentrics in Caimaninae species. The clusters of 18S rDNA were found on one chromosome pair in all species, except for Paleosuchus spp., which possessed three chromosome pairs bearing these sites. Similarly, comparative genomic hybridization demonstrated an advanced stage of sequence divergence among the caiman genomes, with Paleosuchus standing out as the most divergent. Thus, although Alligatoridae exhibited rather low species diversity and some level of karyotype stasis, their genomic content indicates that they are not as conserved as previously thought. These new data deepen the discussion of cytotaxonomy in this family. Keywords: Alligatoridae; cytogenomics; chromosome; molecular cytogenetics 1. Introduction Unlike other vertebrates that underwent substantial diversification, extant crocodilian species have maintained morphological and ecological similarities for almost 100 mil- lion years (Myr) [1–4]. Crocodilians along with dinosaurs, pterosaurs, and birds form a monophyletic clade known as archosaurs. They represent a bridge between recent birds Cells 2021, 10, 1397. https://doi.org/10.3390/cells10061397 https://www.mdpi.com/journal/cells