Karyotype description of three species of Loricariidae ( Siluriformes ) and occurrence of the ZZ / ZW sexual system in Hemiancistrus spilomma Cardoso & Lucinda , 2003

The neotropical freshwater systems have a high number of catfish species (Siluriformes), and many of those are denominated “cascudos” in Brazil. Cytogenetic data about three “cascudos” species fished in the rio Araguaia are described in the present study. The Pterygoplichthys joselimaianus showed 2n=52, with 28 metacentrics (M) chromosomes, 16 submetacentrics (SM) and 8 subtelocentrics/acrocentrics (ST/A) in both sexes. Hemiancistrus spinosissimus showed 2n=52, with karyotype formulae 26M+22SM+4ST, in both sexes. Hemiancistrus spilomma also showed 2n=52, but in this species a ZZ/ZW sex chromosome system (25M+21SM+6ST in females and 24M+22SM+6ST in males) was observed. The cells from H. spinosissimus and P. joselimaianus showed one chromosome pair bearing Ag-NORs, while in the H. spilomma three chromosome pairs bearing AgNORs were detected. The data showed in this work reveal particular chromosomal characteristics, important for a good recognition of both Hemincistrus species, and also show the importance of the insertion of cytogenetic data on taxonomic phylogenetic studies.

In the present work, the karyotypes and active rDNA nucleolar location of the three species of Loricariidae are presented, as well as the description of ZZ/ZW sexual chromosome system in Hemiancistrus spilomma.

Material and methods
In the present study, the taxonomy proposed by Armbruster (2004) who considers the genus Glyptoperichthys as synonyms with Pterygoplichthys was applied.In this way, P. joselimaianus (Weber, 1991) (three females, four males and one unidentified sex) were sampled in the lago Quatro Bocas (15º23'20.1''S,51º42'45.9''W),near the city of Araguaiana, Mato Grosso, Brazil.Voucher specimens were deposited in the fish collection of the Instituto de Ciências e Letras do Médio Araguaia, Universidade Federal de Mato Grosso, Brazil (ICLMA434).Hemiancistrus spinosissimus Cardoso & Lucinda, 2003 (two female and two males) and H. spilomma Cardoso & Lucinda, 2003 (eight females and two males) were catched in the rio Araguaia (15 o 50'15"S -051 o 58'43"W), near the city of Pontal do Araguaia, Mato Grosso, Brazil.Voucher specimens were deposited in the Museu de Ciências e Tecnologia of Pontifícia Universidade Católica do Rio Grande do Sul (MCP36689, MCP36686, MCP36685).

Results
In both sexes of Pterygoplichthys joselimaianus, the diploid number was 2n=52 and the karyotypic formulae 28M+16SM+8ST/A (Figure 1).In both sexes of Hemiancistrus spinosissimus the diploid number was 2n=52 and the karyotypic formulae 26M+22SM+4ST (Figure 2a).H. spilomma also showed 2n=52 but, a ZZ/ZW sex chromosome mechanism was observed, with a karyotypic formulae of 25M+21SM+6ST in females and 24M+22SM+6ST in males (Figure 2b).The ZZ chromosomes are represented by the first submetacentric pair whereas the W chromosome is morphologically similar to the third metacentric pair.
In the present study, it was observed that P. joselimaianus and H. spinosissimus cells showed one Ag-NOR-bearing chromosome pair (inset of Figure 1 and 2a, respectively).The Ag-NOR signals were subterminally located on the short arm of pair 9 in P. joselimaianus (Figure 1, inset) and terminally on the long arm of the pair 17 in H. spinosissimus (Figure 2a, inset).Three Ag-NOR-bearing pairs were detected in H. spilomma, one located in the terminal regions of the long arm in pairs 2 and 18 and other at the terminal end of the short arm of pair 3 (Figure 2b, inset).

Discussion
In Ancistrini, pericentric and paracentric inversions probably were the most important chromosome rearrangements ferentiated sex chromosomes, 64% showed female heterogamety and 36% male heterogamety.Eighty percent of the sex chromosomes corresponds to simple systems (77% ZZ/ZW and 23% XX/XY) and twenty percent correspond to multiple sex chromosome mechanisms (Centofante et al., 2002).The presence of a ZZ/ZW sex chromosome system in H. spilomma (present study) and in Ancistrus cf.dubius (Mariotto et al., 2004) suggest that this system may has a wide distribution in Ancistrini.Some fish groups have a predominance of one chromosome pair in their karyotype bearing just one active NOR, as some species of Pimelodidae (Dias & Foresti, 1993) and Loricariidae (Alves et al., 2003).On the other hand, occurrence of multiple NORs can be observed in some species of Hypostominae (Artoni & Bertollo, 1996).Among the species examined, Hemiancistrus spilomma is the only species thus far to show multiple NORs, as described in Table 1 and confirmed by the nucleolus analysis (Figure 2b).
It is early to draw conclusions on the significance of the ZZ/ZW occurrence associated with the multiplicity of Ag-NORs in Hemiancistrus spilomma, as well the maintenance of 2n=52 and single Ag-NOR in the majority of the Ancistrini species because this tribe is very specious (Fisch-Muller, 2003), and only a few species from five genus have been studied.that occurred in the karyotypic evolution of the group (Alves et al., 2003).The present data reinforce this hypothesis since all species analyzed also have 2n=52 chromosomes.These rearrangements keep the diploid number equal to 2n=52 chromosomes, metacentric and submetacentric, in most of the studied species (Table 1).Ancistrini has a lower diploid number when compared with Loricariinae (Alves et al., 2003), Hypostominae (Artoni &Bertollo, 1996), andHypoptopomatinae (Andreata et al., 1994).However, these conclusions are preliminary because Ancistrini is a very specious subfamily and these data are only related to a few species from five genera.The species P. joselimaianus also has 2n=52, the same diploid number found in Liposarcus anisitsi (Artoni & Bertollo, 1996), other species of the tribe Pterygoplichthini.According to Armbruster (2004), the tribes Ancistrini and Pterygoplichthini are sister groups and compose a derived clade of the Hypostominae.The present cytogenetic data reinforce the putative relationship between these tribes suggesting that 2n=52 could be a primitive diploid number for them.
A few years ago the presence of a sexual differentiation at a chromosome level was considered an uncommon occurrence in fishes.Advances in the studies of Neotropical fishes, however, have shown a great number of new occurrences in different species and families.Among those species with dif- Nevertheless, our research contributes to new karyological information that can aid in understanding of the evolutionary history of this group of fish.Hemiancistrus spilomma and Hemiancistrus spinosissimus are morphologically very similar and can easily be confused.Since these species have particular chromosomal characteristics the present cytogenetic data may be used as markers for differentiating the two species in the middle rio Araguaia.

Fig. 1 .
Fig. 1.Karyotypes of female and male of Pterygoplichthys joselimaianus with conventional Giemsa staining.The chromosome pair bearing Ag-NORs are in the inset.

Fig. 2 .
Fig. 2. Karyotypes of female and male of Hemiancistrus spinosissimus (a) and Hemiancistrus spilomma (b), with conventional Giemsa staining.The chromosome pair bearing Ag-NORs are in the inset.