Cytogenetic analysis of three sea catfish species (Teleostei, Siluriformes, Ariidae) with the first report of Ag-NOR in this fish family

Despite their ecological and economical importance, fishes of the family Ariidae are still genetically and cytogenetically poorly studied. Among the 133 known species of ariids, only eight have been karyotyped. Cytogenetic analyses performed on Genidens barbus and Sciades herzbergii revealed that both species have 2n = 56 chromosomes and Cathorops aff. mapale has 2n = 52 chromosomes: Genidens barbus has 10 Metacentrics (M), 14 Submetacentrics (SM), 26 Subtelocentrics (ST), and 6 Acrocentrics (A), Sciades herzbergii has 14M, 20SM, 18ST and 4A, whereas Cathorops aff. mapale has 14M, 20SM, and 18ST. The nucleolus organizer regions (NORs) were found in a single chromosome pair on the short arm of a large-sized ST pair in Genidens barbus and on the short arm of a middle-size SM pair in Cathorops aff. mapale. Multiple NORs on the short arms of two large-sized ST pairs were found in Sciades herzbergii. The occurrence of diploid numbers ranging from 2n = 52 through 56 chromosomes and the presence of different karyotypic compositions, besides the number and position of NORs suggest that several numeric and structural chromosome rearrangements were fixed during the evolutionary history of this fish family.

The order Siluriformes (catfishes) has 3,088 species, divided into 36 families and 477 genera distributed worldwide, except for the coldest areas in the Southern and Northern hemispheres (Ferraris, 2007). There are only two predominantly marine families: Plotosidae and Ariidae. The family Ariidae, know as sea catfishes, includes 133 species distributed in 26 genera. It presents a worldwide distribution with species that live in marine and brackish water (Ferraris, 2007). Recent phylogenetic studies showed that the family Ariidae is monophyletic, but its relationships with other siluriforms remain unclear. Some authors suggest a close relationship with Pangasiidae, Ictaluridae, and mainly Anchariidae (de Pinna, 1998;Kailola, 2004;Hardman, 2005;Sullivan et al., 2006;Betancur-R et al., 2007).
Karyotypes of only eight ariid species have been described so far and showed diploid numbers ranging from 2n = 54 to 2n = 56 and complements mainly constituted of biarmed chromosomes (Table 1). Considering the worldwide distribution of Ariidae and its commercial importance, the main objective of the present study was to describe the karyotypes of Cathorops aff. mapale, Genidens barbus, and Sciades herzbergii for a better understanding of the karyotypic evolution and relationships among genera and species in the family.
Mitotic chromosome preparations were performed according to the technique described by Foresti et al. (1993). Nucleolar organizer regions (Ag-NORs) were revealed by the silver-staining method (Howell and Black, 1980). The chromosome morphology was determined based on arm ratios, as proposed by Levan et al. (1964), and the chromosomes were classified according to their morphology as Metacentrics (M), Submetacentrics (SM), Subtelocentrics (ST), and Acrocentrics (A).

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Cytogenetic analyses showed that Genidens barbus has 2n = 56 (10M+14SM+ 26ST+ 6A - Figure 1), Cathorops aff. mapale has 2n = 52 (14M+20SM+18ST - Figure 2a), and Sciades herzbergii has 2n = 56 (14M+20SM+18ST+4A - Figure 2b). Early karyotypic studies of G. barbus by Gomes et al. (1994, cited as Netuma barba) and S. herzbergii by Molina et al. (2004, cited as Hexanematichthys herzbergii) showed the same diploid numbers found in the present study. However, the karyotypic formulae previously described for these species were different from those found here (Table 1). These differences may be due to technical artifacts, such as differences in chromosome condensation, or may be real differences that should be checked in further studies covering the distribution area of these species.
Up to now, only two species of Cathorops were cytogenetically investigated: C. aff. mapale (2n = 52 -present study) and one unidentified species, Cathorops sp., that has 2n = 54 (Gomes et al., 1992). This difference in diploid numbers between species of a single genus was also found in Bagre (Fitzsimons et al., 1988;Gomes et al., 1990) (Table 1), suggesting that this is not a rare phenomenon among ariids. On the other hand, the two Genidens species analyzed (Gomes et al., 1994) presented the same diploid number (Table 1).
The karyotypes of ariids are composed of all morphological types of chromosomes (Table 1). However, in some species such as those of the genus Bagre and Sciades herzbergii, a large number of metacentric and submetacentric chromosomes is observed, while in other species, such as Bagre marinus, a large number of subtelocentric and acrocentric chromosomes is observed (Table 1). This variation allows to hypothesize that many structural chromosome rearrangements were fixed during the karyotypic evolution among species of this family. Oliveira and Gosztonyi (2000) studied the karyological evolution of the order Siluriformes, particularly of the family Diplomystidae, and reached the conclusion that the ancestral diploid number for this order is 2n = 56. Among the families closely related to Ariidae, species of Pangasiidae have about 2n = 60 chromosomes (Magtoon and Donsakul, 1987;Manosroi et al. 2003), and among Ictaluridae, the diploid numbers range from 2n = 40 to 72 (LeGrande and Cavender, 1980;LeGrande, 1981;Clark and Mathis, 1982;LeGrande et al., 1984;Amemiya, 1986). This diploid number variation, as well as that observed among ariids, suggests that numeric chromosome rearrangements (fusions and fissions) may have had an important role in the karyotypic evolution of this group.
The present results on Ag-NORs location are the first described for ariids. The ariid species studied have single or multiple Ag-NORs (Table 1). The NORs were found on the short arm of a biggest-size ST pair in Genidens barbus, and on the short arm of a middle-size SM pair in Cathorops aff. mapale. Among the Ictaluridae species, only single NORbearing chromosomes have been observed (Amemiya et al., 1986). This is also the most common condition in Siluriformes (Oliveira and Gosztonyi, 2000;Oliveira et al., 2006) and even in Teleostei (Klinkhardt, 1998). On the other hand, multiple NORs on the short arm of two large-Nirchio et al. 263  sized ST pairs were found in Sciades herzbergii, which reinforces the hypothesis that structural chromosome rearrangements were also fixed in the karyotypic evolution of ariids. Chromosomal rearrangements, indicated by differences in diploid numbers and karyotypes among species and by morphological changes in the position of the Ag-NORs, are widespread among fishes of the family Ariidae (Table 1). These chromosome rearrangements may have played an important role in the karyotypic evolution of the family, as already suggested by LeGrande (1980) and Fitzsimons et al. (1988). However, additional data should be added to clarify the importance of these chromosomal rearrangements in the evolution of the species and genera of ariids.