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. 2023 Dec 5;17:273–281. doi: 10.3897/compcytogen.17.108265

Metaphase chromosomes of five Neotropical species of the genus Drosophila (Diptera, Drosophilidae)

Doris Vela 1,✉, Erika Villavicencio 1
PMCID: PMC10752201  PMID: 38152389

Abstract

The mitotic metaphases of five Andean species of genus Drosophila are described for the first time. The evolutionary and interspecific genetic relationships within three Neotropical Drosophila species groups are analyzed. The diploid chromosome number for each species is as follows: D.cashapamba Céspedes et Rafael, 2012 2n = 6 (2V, 1J) (X = J, Y = R), D.ecuatoriana Vela et Rafael, 2004 2n = 10 (3R, 2V) (X = V, Y = R), D.ninarumi Vela et Rafael, 2005 2n = 10 (3R, 1V, 1D) (X = V, Y = R), D.urcu Vela et Rafael, 2005 2n = 12 (4R, 2V) (X = V, Y = R), D.valenteae Llangarí-Arizo et Rafael, 2018 2n = 8 (3R, 1J) (X = J, Y = R).

Keywords: Andean, Drosophila chromosomes, guarani , mesophragmatica , metaphase, tripunctata

Introduction

The ancestral karyotype for the genus Drosophila Fallén, 1823 (Diptera, Drosophilidae) consists of five pairs of large chromosomes (V shape or J shape) and one pair of dots (Sturtevant and Novitski 1941). This Drosophila metaphase chromosome configuration has been commonly observed, for instance, in some species of Neotropical groups of the type subgenus Drosophila: D.guarani group (King 1947), D.mesophragmatica group (Brncic and Koref 1957; Hunter and Hunter 1964), D.repleta group (Wasserman 1960) and D.tripunctata group (Pipkin and Heed 1964). The species of the type subgenus present a chromosome configuration ranging from three to six pairs of chromosomes. Cytogenetics studies demonstrated that in the genus Drosophila karyotypes of species may differ from the ancestral karyotype by the number of chromosomes and the chromosomal configuration, but chromosomal rearrangements do not break the integrity of Muller elements (chromosome arms and associated linkage groups) (Schaeffer 2018).

By means of the karyotypes, it is possible to observe the chromosomal rearrangements (inversions, translocations, duplications etc.) in species, and how they can limit the genetic exchange and potentially drive speciation (Noor et al. 2001). In addition, it is possible to detect interspecific and intraspecific polymorphism in species of Drosophila (Deng et al. 2007). Therefore, karyotypes are an important tool for understanding the evolutionary history of the Drosophila species, to conduct comparative genomics studies and to allow genome assembly at the chromosome level (Schaeffer 2018).

Most of the available cytological data about Neotropical species of Drosophila were reported in the past century (Metz and Moses 1923; Patterson and Wheeler 1942; Wharton 1943; Burla et al. 1949; Clayton and Wasserman 1957; Clayton and Wheeler 1975). In the most recent cytological studies of Neotropical species of Drosophila karyotypes of ten species from four sibling species groups have been described: D.chorlavi Céspedes et Rafael, 2012, D.mesophragmatica Duda, 1927 and D.rucux Céspedes et Rafael, 2012 from the D.mesophragmatica group (Mafla 2012), D.butantan Ratcov, Vilela et Goñi, 2017, D.sachapuyu Peñafiel-Vinueza et Rafael, 2018, and D.zamorana Peñafiel-Vinueza et Rafael, 2018 from the D.guarani group (Ratcov et al. 2017; Vela and Villavicencio 2021), D.huancavilcae Rafael et Arcos, 1989, D.inca Dobzhansky et Pavan, 1943, and D.yangana Rafael et Vela,;2003 from the D.repleta group (Mafla 2005, 2008), and D.montevidensis Goñi et Vilela, 2016 from the D.tripunctata group (Goñi and Vilela 2016).

In this study, the karyotypes of five Andean species of Drosophila from three sibling species groups are described for the first time: D.ecuatoriana Vela et Rafael, 2004 and D.valenteae Llangarí-Arizo et Rafael, 2018 from the D.guarani group, D.cashapamba Céspedes et Rafael, 2012 from the D.mesophragmatica group, D.ninarumi Vela et Rafael, 2005 and D.urcu Vela et Rafael, 2005 from the D.tripunctata group.

Methods

Species stock

The species analysed correspond to natural populations of: D.cashapamba (QCAZ-I 2349), Sangolquí Canton (location 0°19'59.3"S, 78°25'51"W DMS); D.ecuatoriana (QCAZ-I 1609), Yanacocha Forest (location 0°7'3.8"S, 78°35'9.4"W DMS); D.ninarumi (QCAZ-I 1765), Cruz Loma Forest (location 0°11'22"S, 78°31'17.2"W DMS); D.urcu (QCAZ-I 1755), Cruz Loma Forest (location 0°11'22"S, 78°31'17.2"W DMS) and D.valenteae (QCAZ-I 3142), Sangolquí Canton (location 0°19'59.3"S, 78°25'51"W DMS).

All species were provided by the Evolutionary Genetics Laboratory of Pontificia Universidad Católica del Ecuador. The flies were maintained in banana culture medium supplemented with fresh fruit, in a temperate room at 17 °C, with a 12 h light/dark cycle.

Chromosome plates

The metaphase nuclei of cerebral ganglia were obtained from third-instar larvae (ten males, ten females) of each species. Chromosomal plates were prepared by the cell suspension method (Cardoso and Dutra 1979) and thermic shock (Holmquist 1975) and stained with Giemsa. Ten metaphase nuclei were observed for each sex and species. A Ziess Axioskop 2 plus – HAL 100 microscope and a Cannon PowerShot A640 camera (100× objectives lens and optovar 2×) were used to observe and take the pictures of the mitotic chromosome cells. The modal number was considered the chromosome number of each species.

Mitotic chromosome analysis

For each species, the total length (TL), relative length (RL) and centromeric index (CI) of the chromosomes were estimated using the Axio Vision 4.4. Standard deviation of relative length was analysed using the SPSS statistical package 26.0v (Table 1).

Table 1.

Measurement of metaphase chromosomes of five Andean Drosophila species.

Species Chromosome TL (μm) RL (%) CI SD (n = 10) Morphology
D.ecuatoriana X 2,49 24,22 0,47 0,27 metacentric
2n = 10 Y 1,85 17,99 0,05 0,03 telocentric
2 1,65 16,05 0,49 0,12 metacentric
3 1,54 14,98 0,06 0,19 telocentric
4 1,42 13,81 0,07 0,21 telocentric
5 1,33 12,93 0,08 0,16 telocentric
D.valenteae X 2,09 27,42 0,37 0,23 submetacentric
2n = 8 Y 1,73 22,7 0,06 0,31 telocentric
2 1,4 18,37 0,07 0,21 telocentric
3 1,26 16,53 0,08 0,23 telocentric
4 1,14 14,96 0,09 0,14 telocentric
D.cashapamba X 2,88 26,2 0,38 0,12 submetacentric
2n = 6 Y 1,94 17,65 0,05 0,04 telocentric
2 3,21 29,2 0,47 0,11 metacentric
3 2,96 26,93 0,49 0,12 metacentric
D.ninarumi X 1,71 27,49 0,46 0,25 metacentric
2n = 10 Y 1,59 25,56 0,06 0,04 telocentric
2 1,12 18 0,09 0,26 telocentric
3 0,95 15,27 0,11 0,18 telocentric
4 0,83 13,34 0,12 0,2 telocentric
5 0,02 0,32 0,05 0,01 dot
D.urcu X 3,09 24,75 0,48 0,23 metacentric
2n = 12 Y 2,65 21,23 0,04 0,07 telocentric
2 1,62 12,98 0,49 0,17 metacentric
3 1,58 12,66 0,06 0,27 telocentric
4 1,45 11,61 0,07 0,21 telocentric
5 1,21 9,69 0,08 0,14 telocentric
6 0,88 7,05 0,11 0,29 telocentric
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TL: Total Length, RL: Relative Length, CI: Centromeric Index, SD: Standard deviation.

Results

The description of new karyotypes of Drosophila species is presented below:

The Drosophilaguarani group

The karyotype of D.ecuatoriana is 2n = 10 (3R, 2V), comprising of four autosomes – a large V-shaped metacentric (pair 2) and three pairs of rod-shaped telocentric chromosomes (pairs 3, 4 and 5) – and the sexual pair (X = V, Y = R). The X chromosome is V-shaped metacentric and the Y chromosome is rod-shaped telocentric (Fig. 1A, B, Table 1).

Figure 1.

Figure 1.

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Metaphase karyotype of AD.ecuatoriana female BD.ecuatoriana male CD.valenteae female DD.valenteae male ED.cashapamba female FD.cashapamba male GD.ninarumi female HD.ninarumi male ID.urcu female JD.urcu male. Scale bar: 3 µm (A–J).

The karyotype of D.valenteae is 2n = 8 (3R, 1J), comprising of three rod-shaped telocentric autosomes (pairs 2, 3 and 4), and the sexual pair (X = J, Y = R). The X chromosome is J-shaped submetacentric, and the Y chromosome is rod-shaped telocentric (Fig. 1C, D, Table 1).

The Drosophilamesophragmatica group

The karyotype of D.cashapamba is 2n = 6 (2V, 1J) comprising of two V-shaped metacentric autosomes (pairs 2 and 3) and the sexual pair (X = J, Y = R). The X chromosome is J-shaped submetacentric and the Y chromosome is rod-shaped telocentric (Fig. 1E, F, Table 1).

The Drosophilatripunctata group

The karyotype of D.ninarumi is 2n = 10 (3R, 1V, 1D), comprising of four autosomes – three rod-shaped telocentric (pairs 2, 3 and 4) and one pair of dot-shaped chromosomes (pair 5), and the sexual pair (X = V, Y = R). The X chromosome is V-shaped metacentric and the Y chromosome is rod-shaped telocentric (Fig. 1G, H, Table 1).

The karyotype of D.urcu is 2n = 12 (4R, 2V) comprising of five autosomes – a pair of V-shaped metacentric (pair 2) and four pairs of rod-shaped telocentric chromosomes (pairs 3, 4, 5 and 6) – and the sexual pair (X = V, Y = R). The X chromosome is V-shaped metacentric and the Y chromosome is rod-shaped telocentric (Fig. 1I, J, Table 1).

Discussion

Considering the high diversity of Drosophila species in the Neotropical region little is known about diploid chromosome numbers of these species.

In the Drosophilaguarani group, the most common karyotype is 2n = 12. In the present study, the karyotype of D.ecuatoriana is 2n = 10 (Fig. 1A, B). A similar 2n = 10 karyotype was reported in other species of this group: D.guaraja King, 1947 (King 1947), D.butantan (Ratcov et al. 2017) and D.sachapuyu (Vela and Villavicencio 2021). The karyotype of D.valenteae is 2n = 8 (Fig. 1C, D) and is similar to D.alexandrei Cordeiro, 1951 (Cordeiro 1951), both species present the lowest diploid chromosome reported for the Drosophilaguarani species group.

Several reports have shown that the karyotype of Drosophila species of the D.mesophragmatica group is highly conserved, 2n = 10, including a pair of rod-shaped or a dot-like fifth chromosomes (Brncic 1957). Additionally, paracentric inversions are the principal chromosomal rearrangements attributed to this species group (Brncic and Koref 1957). In our study, the chromosome number of D.cashapamba is 2n = 6, the chromosomes are large and present a small pericentromeric heterochromatin (Fig. 1E, F). It has been suggested that D.cashapamba is a junior synonym of D.dreyfusi Dobzhansky et Pavan, 1943 (Dr Carlos Vilela, pers. communication) due to the similarity of the male genitalia and the same chromosome number, 2n = 6 (Dobzhansky and Pavan 1943). However, in this study we maintain the current taxonomical classification until new taxonomic studies confirm the junior synonym status of D.cashapamba.

According to the information available in the Drosophila karyotype databases (Morelli et al. 2022), the chromosome number 2n = 6 is rarely reported in Drosophila subgenus. Only thirteen species of Drosophila subgenus present three pairs of chromosomes: D.canalinea Patterson et Mainland, 1944 from D.canalinea group, D.dreyfusi and D.wingei Cordeiro, 1964 from D.dreyfusi group, D.albomicans Duda, 1923, D.annulipes Duda, 1924, D.neohypocausta Lin et Wheeler, 1973 from D.immigrans group, D.atalaia Vilela et Sene, 1982 from D.peruensis group, D.pinicola Sturtevant, 1942 from D.pinicola group, D.quinaria Loew, 1866 from D.quinaria group; D.neoguaramunu Frydenberg, 1956 from D.tripunctata group, D.montana Patterson et Wheeler, 1942 from D.virilis group, D.aracea Heed et Wheeler, 1957 and D.tranquilla Spencer, 1942 (not grouped).

Most species of the D.tripunctata group have a karyotype 2n = 12, the sixth pair is a dot chromosome; some members of D.tripunctata group have a karyotype 2n = 10 (Morelli et al. 2022). In the karyotype of D.ninarumi, 2n = 10, it is present a dot-like fifth pair of chromosome (Fig. 1G, H) which is reported in the most species of Drosophilatripunctata group. This karyotype is similar to D.fairchaldi Pipkin et Heed, 1964 and D.unipunctata Patterson, 1943 (Wharton 1943; Pipkin and Heed 1964; Clayton and Wheeler 1975) but in these species the dot-like chromosome is absent. In the case of D.urcu, the karyotype is 2n = 12, all the chromosomes are large metacentric or telocentric (Fig. 1I, J). Our data show that the karyotype of D.ninarumi and D.urcu have a relevant similitud, the sexual chromosomes are the largest of the chromosome set, with a Y chromosome heteropycnotic (Fig. 1G, J).

Traditional studies like genetic crosses, in situ hybridization, polytene chromosomes maps or karyotype description are not commonly performed. However, for the genus Drosophila, the information provided by cytological studies is the initial tool in understanding the evolutionary history and the high radiation of the Drosophila species in the Neotropical region and also important in the beginning of genomic studies on these species.

Conclusions

This study reveals the first karyotype description of five Neotropical species of Drosophila. Only the karyotype of D.urcu, 2n = 12, is similar to the ancestral karyotype of Drosophila, but the sixth pair are large chromosomes. The karyotypes of D.ecuatoriana and D.ninarumi are 2n = 10, but only the last one has a dot-like chromosome. The karyotype of D.valenteae is 2n = 8; this is the second species of D.guarani group that have this chromosome number. The karyotype of D.cashapamba presents a low chromosome number, 2n = 6, which is only reported in other thirteen species of subgenus Drosophila.

Acknowledgements

The present research has been supported by the Pontificia Universidad Católica del Ecuador through the project QINV0320-IINV529010200.

Citation

Vela D, Villavicencio E (2023) Metaphase chromosomes of five Neotropical species of the genus Drosophila (Diptera, Drosophilidae). Comparative Cytogenetics 17: 273–281. https://doi.org/10.3897/compcytogen.17.108265

Funding Statement

The present research has been supported by the Pontificia Universidad Católica del Ecuador throught the project QINV0320-IINV529010200.

ORCID

Doris Vela https://orcid.org/0000-0001-7690-7758

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