| Summary: | <p>Abstract</p> <p>Background</p> <p>Seven karyomorphs of the fish, <it>Hoplias malabaricus </it>(A-G) were previously included in two major groups, Group I (A, B, C, D) and Group II (E, F, G), based on their similar karyotype structure. In this paper, karyomorphs from Group I were analyzed by means of distinct chromosomal markers, including silver-stained nucleolar organizer regions (Ag-NORs) and chromosomal location of repetitive sequences (18S and 5S rDNA, and satellite 5S<it>Hind</it>III-DNA), through fluorescence in situ hybridization (FISH), in order to evaluate the evolutionary relationships among them.</p> <p>Results</p> <p>The results showed that several chromosomal markers had conserved location in the four karyomorphs. In addition, some other markers were only conserved in corresponding chromosomes of karyomorphs A-B and C-D. These data therefore reinforced and confirmed the proposed grouping of karyomorphs A-D in Group I and highlight a closer relationship between karyomorphs A-B and C-D. Moreover, the mapping pattern of some markers on some autosomes and on the chromosomes of the XY and X<sub>1</sub>X<sub>2</sub>Y systems provided new evidence concerning the possible origin of the sex chromosomes.</p> <p>Conclusion</p> <p>The <it>in situ </it>investigation of repetitive DNA sequences adds new informative characters useful in comparative genomics at chromosomal level and provides insights into the evolutionary relationships among <it>Hoplias malabaricus </it>karyomorphs.</p>
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