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Karyological Analysis of Some Species of Aphanius (Osteichthyes: Cyprinodontidae) From Anatolia.

Byline: Muhammet Gaffaroglu Muradiye Karasu Ayata Sevgi Anal and Mustafa Ozkan


In this study chromosomal studies were carried out on four species of the genus Aphanius (Osteichthyes Cyprinodontidae). Metaphase chromosomes were obtained from kidney cells. The diploid chromosome numbers of Aphanius anatoliae A. danfordii A. splendens and A. villwocki were 2n=48 and consisted of three pairs of submetacentric and 21 pairs of subtelocentric chromosomes. The arm number (NF) was 54. Constitutive heterochromatin regions were observed on the centromeres of chromosomes in all four species by using C-banding. Nucleolus organizer regions (NOR) were observed on the short arms of two pairs of chromosomes. It is believed that the data obtained will make a contribution to fish cytogenetics.

Key words: Aphanius chromosome C-banding NOR


The family Cyprinodontidae is distributed throughout the United States Africa Southern Europe and Asia. It is represented by a genus in our country (Geldiay and Balik 2007). The genus Aphanius Nardo 1827 has 10 species in Anatolia. These species are Aphanius anatoliae A. asquamatus A. burdurensis A. danfordii A. fasciatus A. mento A. splendens A. sureyanus A. transgrediens and A. villwocki. All of these species are endemic to our country except for A. fasciatus and A. mento (Fricke et al. 2007).

Fish chromosome studies have been carried out for many years. Chromosome properties of 35 species from a total of 104 species (Fam: Cyprinodontidae) have been reported to date. The diploid chromosome numbers of these species were found to be 48 50 and 52 (Arai 2011).

The species of Aphanius living in Anatolia has been studied for molecular phylogeny and historical biogeography (Hrbek et al. 2002) genetic relationships (Bardakci et al. 2004) and population structure (GA1/4clA1/4 et al. 2007). Also molecular analysis of the genetic differentiation among Aphanius fasciatus populations has been studied in Tunis (Annabi et al. 2012).

There has been no a detailed study on the cytogenetic features of these species although only haploid chromosome number has been reported in A. anatoliae (Oztan 1954; Wildekamp 1993).

The purpose of this study is to reveal the chromosomal features (with Giemsa Ag-NOR staining and C-banding) of four species (A. anatoliae A. danfordii A. splendens and A. villwocki) from Anatolia.


Eighteen (10 female eight male) samples of Aphanius danfordii were collected from Sultansazligi Develi Kayseri (38 22' N 35 21' E) 17 (11 female six male) samples of A. anatoliae were collected from Egirdir Lake Isparta (37 51' N 30 50' E) 10 (four female six male) samples of A. splendens were collected from Salda Lake Burdur (37 31' N 29 43' E) and 15 (10 female five male) samples of A. villwocki were collected from Eminekin Eskisehir (39 22' N 31 06' E). These samples were transported live to the laboratory. Metaphase preparations were prepared according to Collares-Pereira (1992) from anterior kidney cells. The technique of Sumner (1972) was used for C-banding of sample preparations whereas the technique of Howell and Black (1980) was used for silver staining. The metaphase preparations were observed and photographed using a Leica DM3000 microscope. Chromosomes were classified according to Levan et al. (1964).


The diploid chromosome numbers of Aphanius anatoliae A. danfordii A. splendens and A. villwocki were determined to be 2n=48 and chromosome morphologies were three pairs of submetacentric (SM) and 21 pairs of subtelocentric (ST) chromosomes (Fig. 1). NF was found to be 54. The differentiations of sex chromosomes were not observed.

NOR was observed on the telomeric regions of the short arms of two large pairs of subtelocentric chromosomes in each species (Fig. 2a).

By using C-banding constitutive heterochromatin regions were observed on the centromeres of several chromosomes in all species. Additionally long arms of some chromosomes had telomeric heterochromatin bands (Fig. 2b).


According to Oztan (1954) the haploid chromosome number of Aphanius anatoliae was n=24. This study shares the results that we obtained from A. anatoliae. Except for this study no cytogenetic research has been reported on A. anatoliae A. danfordii A. splendens and A. villwocki.

A. anatoliae A. danfordii A. splendens and A. villwocki have the same diploid chromosome number as other species of Aphanius that were previously studied. However their chromosome morphologies are different from these species (Table I).

Table I.- Karyotype studies in the genus Aphanius.



###number (2n)

A.###48###48 ST-A###48###Vitturi et al.


A.###48###22 SM+6 ST###70###Esmaeili et al.


###48###28 SM+20 ST###76###Esmaeili et al.

A. sophiae###(2007)

A.###48###14 SM+34 ST###31###Esmaeili et al.


###48###16 SM+32 ST###32###Esmaeili et al.

A. dispar###(2008a)

A.###48###12 SM+39 ST###30###Esmaeili et al.


A.###48###8 SM+40 ST###28###Esmaeili et al.


A.###48###6 SM+42 ST###54###In this study


A.###48###6 SM+42 ST###54###In this study


A.###48###6 SM+42 ST###54###In this study


A.###48###6 SM+42 ST###54###In this study


The karyotype of the Aphanius species that has been studied in this research is very similar to A. vladykovi which has eight SM and 40 ST chromosomes (Esmaeili et al. 2009).

It has been reported that diploid chromosome numbers of A. asquamatus and A. mento collected from inland waters of Turkey was 2n=48. However there was no information about the chromosome morphologies of these species (Arai 2011). The similarity of the chromosome number between Aphanius species shows that the chromosome number is conservative in this genus. A similar situation has also been reported by other authors (Esmaeili et al. 2009).

It has been shown that some species from the genus Cyprinodon Garmanella Jordanella Megupsilon and Orestias (Fam: Cyprinodontidae) have 2n=48 (Arai 2011). On the other hand some species of the genus Fundulus Gambusia and Poecilia (Order: Cyprinodontiformes) also have 2n=48 (Arai 2011). The findings from A. anatoliae A. danfordii A. splendens and A. villwocki are similar to these studies.

According to our observations there was no sex chromosome differentiation in A. anatoliae A. danfordii A. splendens and A. villwocki. In this regard these species are similar to A. persicus A. sophiae A. ginaonis A. dispar A. isfahanensis and A. vladykovi in which sex chromosomes were unreported (Esmaeili et al. 2007 2008ab 2009).

C-band patterns of fish chromosomes have been studied for many years. Constitutive heterochromatin regions are found at or around centromeres and telomeres using the C-banding method. These regions may also be found within chromosomal arms and sometimes the short arms of acrocentric chromosomes may be entirely heterochromatic (Gaffaroglu and YA1/4ksel 2009).

There is no difference in the patterns of heterochromatin regions among the species in this study. The findings that C-bands localize in the centromere and telomere regions of the chromosomes in A. fasciatus (Vitturi et al. 1995). A. anatoliae A. danfordii A. splendens and A. villwocki are similar to this study in terms of including centromeric and telomeric C-bands. However inter-individual polymorphism of telomeric C-bands that have been reported in A. fasciatus (Vitturi et al. 1995) were not observed in this study. Moreover A. anatoliae A. danfordii A. splendens and A. villwocki are similar to Gambusia holbrooki which has centromeric C-bands (Russo et al. 1999). Similar results have also been obtained by other researchers in cyprinid taxa (Gaffaroglu and YA1/4ksel 2009).

NOR numbers and locations have proven useful in fish cytotaxonomy (Amemiya and Gold 1988). In addition to the diploid chromosome numbers these properties have also been examined in many fish species (Arai 2011). In this study no significant differences in the number and location of the NOR have been observed between the species.

It was concluded that the NOR number of A. fasciatus varied between one and eight (Vitturi et al. 1995). While polymorphism of NOR number and localization has been reported in A. fasciatus no polymorphism was observed in the Aphanius species in this study. They are similar to G. holbrooki regarding the existence of NOR in the short arms of two pairs of chromosomes (Russo et al. 1999).

Chromosome banding studies (especially AgNOR and C-banding) have not been found on other Aphanius species (except A. fasciatus) to date. For this reason a comparison in this regard cannot be made in detail within the genus.

This study may contribute to cytogenetics of the Aphanius species.


This study was supported by TABITAK (No. 110T054 project). The authors thank to Salim Serkan GACLA (SA1/4leyman Demirel University Turkey) for helping in the field and identification of the samples.


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Author:Gaffaroglu, Muhammet; Karasu, Muradiye; Anal, Ayata Sevgi; Ozkan, Mustafa
Publication:Pakistan Journal of Zoology
Article Type:Report
Geographic Code:90ASI
Date:Oct 31, 2014
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