CYTOGENETIC CHARACTERIZATION OF Ameivula ocellifera (SPIX, 1825) (SQUAMATA, TEIIDAE) FROM THE BRAZILIAN NORTHEAST CARACTERIZAÇÃO CITOGENÉTICA DE Ameivula ocellifera (SPIX, 1825) (SQUAMATA, TEIIDAE) DO NORDESTE BRASILEIRO

Ameivula is as a new genus of Teiidae family that emerged after extensive revision of species that comprised the former complex of species called Cnemidophorus group. Its species has a wide distribution from the northeast of Brazil to northern Argentina. Cytogenetic studies in the Teiidae family have shown that karyotypical data are important tools in phylogenetic and systematic studies within this group allowing to determine the position of species in the family. Thus, this study aimed to describe the karyotype of Ameivula ocellifera (Spix, 1825) from Picos, Piauí state in the Brazilian Northeast. Specimens were collected from August 2014 to October 2015 using interception traps and pitfalls, mounted randomly along the Caatinga area. The animals were collected and transported to Federal Institute of Piauí, campus Picos, where was carried out all laboratory procedures. Individuals analyzed showed a diploid number of 2n = 50 for both sexes, with karyotype composed by 30 macrochromosomes and 20 microchromosomes of telocentric and subtelocentric types. There were no heteromorphic sex chromosomes in the studied specimens. C-band technique evidenced the heterochromatic blocks in pericentromeric and telomeric regions of chromosomes. The nucleolar organizing regions appeared as a simple unit located at the terminal portion of the long arm of chromosomal pair number 5. The chromosomal characteristics of A. ocellifera analyzed do not show divergences regarding individuals from other regions. However, the nucleolar organizing regions seems to be a good chromosomal marker that permits to distinguish the species already studied.


INTRDUCTION
The Teiidae family is a group of lizards comprising 160 species distributed in three subfamilies: Teiinae integrates the genera Ameiva The Ameivula genus emerged from extensive systematic revisions in Teiidae family totaling 11 valid species of small animals (UETZ, 2019). Its species are widely distributed throughout Brazil and can be found from the Brazilian Northeast to the north of Argentina (VANZOLINI; RAMOS-COSTA;VITT, 1980). In Brazil, this lizard occurs in several environments, such as the Caatinga (VITT, 1983;MENEZES et al. 2011), Cerrado (MESQUITA;COLLI, 2003), and restinga of Atlantic Forest (SANTANA et al. 2010). Ameivula ocellifera (Spix, 1825) is a species of active foraging habit, with ecological and morphological characteristics identical to other species of the genus (SILVA; ARAÚJO, 2005).
In the '70s Teiidae was divided into three groups based on the chromosomal characteristics of its members: (1) a group consisting of the genera Dracaena, Callopistes, Crocodilurus and Tupinambis, with a diploid number of 2n = 34-38, divided into five to six pairs of metacentric macrochromosomes and one satellite in pair 2; (2) a group formed by the genera Ameiva, Dicrodon, Kentropyx, Teius, and Cnemidophorus of which of the latter, some species became Ameivula, with diploid number 2n = 46-56 with many acrocentric chromosomes; and (3) a group with the microteiids with a diploid number of 2n = 46 (GORMAN, 1970).
The ancient karyotype of Teiidae would have originated from centric fission of macrochromosomes of an ancestral karyotype of Tupinambinae with 2n = 50 (GORMAN, 1973). Additionally, pericentric inversions, fusions and fissions may have been responsible for the karyotypic diversification of some of the species, e.g., those belonging Cnemidophorus genus, in which chromosome numbers in diploid species vary between 2n = 46 and 52. In triploid species, they vary from 3n = 69 to 71, and it is of 4n = 92 chromosomes in tetraploid species (GORMAN, 1970;PECCININI-SEALE, 1981, 1989.
Thus, considering the karyotype variation already defined for different genera and species of the Teiidae family, this study aimed to describe the karyotype of a population of Ameivula ocellifera, looking for genetic patterns that can be used as a tool to establish differences between species.

MATERIAL AND METHODS
The specimens were collected from August 2014 to October 2015, in areas of Caatinga in the city of Picos, state of Piauí, Brazil. The collection used pitfalls mounted in places where the presence of lizards was observed. The traps were checked three times a day and the captured animals were collected to prevent them from dying due to excessive heat.
Thus, the collected animals were taken to the Biology Laboratory of the Federal Institute of Piauí, campus Picos, where all procedures were performed for the obtention of cytogenetic material, cataloguing, recording and subsequent fixation and preservation. All under license of collection and processing of the animals, issued by the Chico Mendes Institute for Biodiversity Conservation -ICMBIO through the SISBIO platform, number: 47710-1.
Chromosomal preparations were obtained through the air-drying method as described by Bertollo, Takahashi and Moreira-Filho (1978). Cbanding (Sumner, 1972) was used to visualize Heterochromatin regions and silver nitrate staining to detect NORs, according to the procedure described by Howel, Black, (1980).
Chromosomes were observed using a microscope coupled to a camera, and the best images captured. The chromosomal nomenclature was based on the system of Levan, Fredga and Sandberg (1964) taking into account only macrochromosomes (M) and microchromosomes (m).

RESULTS
Cytogenetic analyses were performed on three males and four females of Ameivula ocellifera. The individuals analyzed showed a diploid number of 2n = 50 for both sexes, with a karyotype composed of 30 macrochromosomes and 20 microchromosomes of telocentric and subtelocentric types ( Figure 1). There were no heteromorphic sex chromosomes observed in the specimens studied.

DISCUSSION
In recent years, the Teiidae family and its genera have undergone major changes that have led to the repositioning of species into complexes and the creation of new genera. Ameivula is a new genus that accommodates the species that belonged to the former group Cnemidophorus ocellifera. The complexity in the precise determination involving the species Cnemidophorus ocellifera arouse works based on phylogenetic evidence and molecular data. These works positioned these species in the new genus Ameivula, even though the latter having many features shared with the Ameiva species (GIUGLIANO; CONTEL; COLLI, 2006).
The diploid number of 2n = 50 that includes biarmed chromosomes observed in Ameivula ocellifera in this study remains unaltered despite its wide geographic distribution. This fact also corroborates the studies in individuals also attributed to the Northeast and Southeast of Brazil Signs of chromosome rearrangements in the species of Ameivula are evident when comparing the A. littoralis karyotype from the Brazilian Southeast, with other South American species (PECCININI-SEALE et al. 2004). According to the latter authors, the decrease in diploid number observed can be attributed to centric fusions of microchromosomes. In addition, pericentric inversions in the first and second pair of chromosomes occurred in A. littoralis, making these pairs of the submetacentric type.
The constitutive heterochromatin patterns observed in Ameivula ocellifera shows pericentromeric and telomeric regions with numerous blocks of heterochromatin in the two arms of several pairs of chromosomes (Table 1). This is very similar among the species of the Teiidae family such as Ameiva ameiva (LINNAEUS, 1758) and Kentropyx paulensis (BOETTGER, 1893) (SANTOS et al. 2007). The constitutive heterochromatin is restricted to the telomeric and centromeric region in Ameiva and Cnemidophorus species from other regions of the country. In specimens of these regions, the microchromosomes were almost entirely heterochromatic, and this fact appears being common among those related groups.  Rocha, Bergallo and Peccinini-Seale (1997) Regarding the NORs, the number and position seem remaining conserved in Ameivula ocellifera in accordance with other populations previously analyzed (Table 1). The exception is the absence of size polymorphism as those observed by Santos et al. (2007). It was also detected in the congeneric species Ameivula littoralis a single pair carrying the NOR (PECCININI-SEALE et al. 2004). However, in Ameivula nativo, multiple NOR were noticed in a different chromosome pair (ROCHA BERGALLO;PECCININI-SEALE, 1997). This differentiation maybe due rearrangements involving the nucleolar organizer regions among the Ameivula species from Brazil.
In this way, despite the stability in the diploid number proven with this study, the NORs pattern is a useful tool to establish differences among the Ameivula species, contributing to correct identification of regional forms of these lizards.

ACKNOWLEDGMENTS
The authors are grateful Instituto Federal do Piauí (PROAGRUPAR-INFRA 033/2014) for logistic and financial support and to the anonymous reviewers for suggestions to our manuscript.