Research Article |
Corresponding author: Carlos Eduardo Faresin e Silva ( carlosfaresin@gmail.com ) Academic editor: T Chassovnikarova
© 2017 Carlos Eduardo Faresin e Silva, Rodrigo Amaral de Andrade, Érica Martinha Silva de Souza, Eduardo Schmidt Eler, Maria Nazareth da Silva, Eliana Feldberg.
This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Citation:
Silva CEF, Andrade RA, de Souza EMS, Eler ES, da Silva MNF, Feldberg E (2017) Comparative cytogenetics of some marsupial species (Didelphimorphia, Didelphidae) from the Amazon basin. Comparative Cytogenetics 11(4): 703-725. https://doi.org/10.3897/compcytogen.v11i4.13962
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We investigated the karyotype of 18 didelphid species captured at 13 localities in the Brazilian Amazon, after conventional staining, C-banding, Ag-NOR and fluorescent in situ hybridization (FISH) using the 18S rDNA probe. Variations were found in the X chromosome, heterochromatin distribution and the 18S rDNA sequence. The main variation observed was in the position of the centromere in the X chromosome of Caluromys philander Linnaeus, 1758 and Marmosa murina Linnaeus, 1758. For both species, the X chromosome showed a geographical segregation in the pattern of variation between eastern and western Brazil, with a possible contact area in the central Amazon. C-banding on the X chromosome revealed two patterns for the species of Marmosops Matschie, 1916, apparently without geographic or specific relationships. The nucleolus organizer region (NOR) of all species was confirmed with the 18S rDNA probe, except on the Y chromosome of Monodelphis touan Shaw, 1800. The distribution of this marker varied only in the genus Marmosa Gray, 1821 [M. murina Thomas, 1905 and M. demerarae Thomas, 1905]. Considering that simple NORs are seen as a plesiomorphic character, we conclude that the species Marmosa spp. and Didelphis marsupialis Linnaeus, 1758 evolved independently to the multiple condition. By increasing the sample, using chromosomal banding, and FISH, we verified that marsupials present intra- and interspecific chromosomal variations, which suggests the occurrence of frequent chromosomal rearrangements in the evolution of this group. This observation contrasts with the chromosomal conservatism expected for didelphids.
Marsupials, Amazon basin, C-band, NORs, 18S rDNA, Chromosomal rearrangements
In the Americas, subclass Metatheria Huxley, 1880 is represented by the three marsupial orders: Didelphimorphia Gill, 1872, Paucituberculata Ameghino, 1894 and Microbiotheria Ameghino, 1889. The largest of the three American orders is Didelphimorphia, which is represented by the family Didelphidae Gray, 1821, whose species are widely distributed throughout the continent. Didelphidae is the only marsupial group present in Brazil. Together with rodents, they make up an important part of the mammalian fauna of the Amazon region (
Historically, the first cytogenetic data on American marsupials were recorded by Jordan (1911; cited in
Unlike other mammal orders, such as Rodentia Bowdich, 1821, marsupials show relatively little chromosomal variation (
Among all the metatherian families, Macropodidae Gray, 1821 (order Diprotodontia Owen, 1866) is the most diverse in diploid number, varying from 2n=10 to 32. While the American Didelphidae has only the three main diploid numbers, with the most frequent being 2n=14 (
Limited sampling effort has hampered the estimation of species richness in the Amazon, leaving large gaps in our knowledge of the mammalian fauna (
The number of taxa analyzed to date is also limited, and existing cytogenetic analyses have been usually restricted only to the diploid and fundamental numbers (
In this study, we analyze the main morphological differences in the sex chromosomes and the C-band pattern of 18 didelphid species from the Brazilian Amazon. In addition, we describe for the first time karyotype for six species (Monodelphis touan, Monodelphis aff. adusta, Monodelphis sp., Marmosops impavidus, Marmosops bishopi and Marmosops pinheiroi) and discuss these patterns in a broader geographical context, including other regions of Brazil and South America.
We cytogenetically analyzed 111 individuals in 18 species and 8 didelphid genera, collected in 13 localities in the Amazon (Table
Didelphid species and their respective localities. Species analyzed in the current study were collected at localities 1 to 13, with number of individuals of males (M) and females (F) indicated. Geographic references for the current project were collected in a decimal degree projection using the WGS 84 reference. For literature data we insert converted geographical references where available. Localities with coordinates are presented only the first time they are cited in the table.
Species | Locality | Locality Number | Coordinates† | M | F | Total | Reference |
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Caluromys philander | Trombetas River, Pará State | 10 | 1.48163888889°S, 56.4573333333°W | 9 | 5 | 14 | Present work |
Tapajós River, Pará State | 11 | 3.35486111111°S, 55.2031666667°W | 1 | 1 | 2 | Present work | |
Purus River, Amazonas State | 4 | 4.98066666667°S, 62.9770000000°W | 1 | 1 | Present work | ||
Manaus, Amazonas State | 6 | 3.100548°S, 59.974595°W | 1 | 1 | Present work | ||
Aragua, Venezuela | 14 | – |
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Manaus, Amazonas State | 15 | 3.13333333333°S, 59.9500000000°W |
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Jari, River, Pará State, Brazil | 12 | 0.7000000000°S, 52.6666666667°W | 1 | 1 |
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Pernambuco state | 16 | – |
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São Paulo state | 17 |
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Caluromys lanatus | Japurá River, Amazonas State | 1 | 1.84341666667°S, 69.0264722222°W | 1 | Present work | ||
Iquitos, Peru | – | – |
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Manaus, Amazonas State | – | – |
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Rondônia, Brasil | 13 | – |
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Marmosa demerarae | Aripuanã River, Amazonas State | 7 | 6.00000000000°S, 60.1666666667°W | 4 | 4 | 8 | Present work |
Manaus, Amazonas State | 6 | 3.13333333333°S, 59.9500000000°W | 7 | 11 | 18 | Present work | |
Cuieiras River, Amazonas State | 5 | 2.70708611111°S, 60.3738388889°W | 4 | 2 | 6 | Present work | |
Purus River, Amazonas State | 4 | 0.57725000000°S, 64.8976944444°W | 3 | 4 | 7 | Present work | |
Negro River, Amazonas State | 3 | 0.57725000000°S, 64.8976944444°W | 1 | 5 | 7 | Present work | |
Tapajós River, Pará State | 11 | 3.35486111111°S, 55.2031666667°W | 3 | 5 | 9 | Present work | |
Trombetas River, Pará State | 10 | 1.48163888889°S, 56.4573333333°W | 9 | 5 | 14 | Present work | |
Jari River, Pará State | 12 | 0.7000000000°S, 52.6666666667°W | 9 | 2 | 11 | Present work | |
Juruá River, Amazonas State | 2 | 3.64151111111°S, 66.1006916667°W | 1 | 1 | Present work | ||
Jatapú River, Amazonas State | 9 | 2.017940°S, 58.203228°W | 1 | 1 | Present work | ||
Jari River, Pará State | 12 | 0.7000000000°S, 52.6666666667°W | 1 | 1 | Present work | ||
Uatumã River, Amazonas State | 8 | 1.84998888889°S, 59.4402000000°W | 5 | 3 | 9 | Present work | |
Trombetas River, Pará sate | 10 | 1.48163888889°S, 56.4573333333°W | 1 | 1 | Present work | ||
Negro River, Amazonas State | 3 | 0.57725000000°S, 64.8976944444°W | 1 | 1 | 2 | Present work | |
Juruá River | 2 | 3.64151111111°S, 66.1006916667°W | 1 | 1 | Present work | ||
Marmosa murina | Purus River, Amazonas State | 4 | 0.57725000000°S, 64.8976944444°W | 2 | 2 | Present work | |
Pernambuco State | 16 | – |
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Villa Vivencio, Colombia | 18 | – |
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Bolivar, Venezuela | 19 | – |
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Tartarugalzinho, Amapá State | 21 |
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Loreto, Peru | 20 | – |
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Vila Rica, Mata Grosso State | 22 | 10°01'S, 51°07'W |
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UHE Peixe Angical, Tocantins State | 23 | 12°01'30”S, 48°32'21"W |
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Porto Nacional, Tocantins state | 24 | 10°42'S, 48°25'W |
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Uruaçú, Goiás state | 25 | 14°31'S, 49°08'W |
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Colinas do Sul, Goiás state | 26 | 14°09'S, 48°04'W |
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UHE Corumbá IV Luziania, Goiás state | 27 | 16°15'09"S, 47°57'01"W |
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Pacoti, Ceará state | 28 | 4°13'S, 38°55'W |
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Reserva Biológica Duas Bocas, Espírito Santo state | 29 | 20°16'S, 40°28'W |
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Gracilinanus emiliae | Tapajós River, Pará state | 11 | 3.35486111111°S, 55.2031666667°W | 3 | 1 | 4 | Present work |
Serra da Mesa, Colinas do Sul, Goiás state | 26 | 14°09'S 48°04'W |
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UHE Corumbá IV, Luziania, | 27 | 16°15'09"S, 47°57'01"W |
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Metachirus nudicaudatus | Trombetas River, Pará state | 10 | 1.48163888889°S, 56.4573333333°W | 1 | 1 | Present work | |
Jari River, Pará state | 12 | 0.7000000000°S, 52.6666666667°W | 1 | 1 | Present work | ||
Cuieiras River, Amazonas state | 5 | 2.70708611111°S, 60.3738388889°W | 1 | 1 | Present work | ||
Juruá River, Amazonas state | 2 | 3.64151111111°S, 66.1006916667°W | 1 | 1 | Present work | ||
Tapajós River, Pará state | 11 | 3.5486111111°S, 55.2031666667°W | 2 | 2 | 4 | Present work | |
Glironia venusta | Porto Velho, Rondônia State | 13 | 8.87416666667°S, 64.0077777778°W | 1 | 1 | Present work | |
Monodelphis touan | Jari River, Pará state | 12 | 0.7000000000°S, 52.6666666667°W | 3 | 3 | Present work | |
Monodelphis sp. | Purus River, Amazonas state | 4 | 0.57725000000°S, 64.8976944444°W | 1 | 1 | Present work | |
Monodelphis aff. adusta | Aripuanã River, Amazonas state | 7 | 6.00000000000°S, 60.1666666667°W | 1 | 1 | Present work | |
Monodelphis emiliae | Aripuanã River, Amazonas state | 7 | 6.00000000000°S, 60.1666666667°W | 1 | 1 | Present work | |
Juruá River, Acre state | 8°40'S 72°47'W |
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Monodelphis brevicaudata | Negro River state | 3 | 0.57725000000°S, 64.8976944444°W | 1 | 1 | Present work | |
Marmosops bishop | Aripuanã River, Amazonas state | 7 | 6.00000000000°S, 60.1666666667°W | 5 | 6 | 11 | Present work |
Purus River, Amazonas state | 4 | 0.57725000000°S, 64.8976944444°W | 2 | 1 | 3 | Present work | |
Negro River, Amazonas state | 3 | 0.57725000000°S, 64.8976944444°W | 1 | 1 | Present work | ||
Marmosops pinheiroi | Tapajós River, Pará state | 11 | 3.5486111111°S, 55.2031666667°W | 4 | 2 | 6 | Present work |
Marmosops parvidens | Trombetas River, Pará state | 10 | 1.48163888889°S, 56.4573333333°W | 8 | 1 | 9 | Present work |
Cuieiras River, Amazonas state | 5 | 2.70708611111°S, 60.3738388889°W | 3 | 2 | 5 | Present work | |
Jari River, Pará state | 12 | 0.7000000000°S, 52.6666666667°W | 2 | 2 | Present work | ||
Jatapú River, Amazonas state | 9 | 2.017940°S, 58.203228°W | 4 | 3 | 7 | Present work | |
La Paz, Bolívia | – | – |
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Serra da Mesa, Colinas do Sul, Goiás state | 26 | 14°09'S, 48°04'W |
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Apiacás, Mato Grosso state | 9°34'S, 57°23'W |
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Marmosops impavidus | Juruá River, Amazonas state | 2 | 3.64151111111°S, 66.1006916667°W | 2 | 1 | 3 | Present work |
Marmosops pakaraimae | Japurá River, Amazonas state | 1 | 1.84341666667°S, 69.0264722222°W | 1 | 3 | Present work | |
Didelphis marsupialis | Tapajós River, Pará state | 11 | 3.5486111111°S, 55.2031666667°W | 1 | 3 | 4 | Present work |
Trombetas River, Pará state | 10 | 1.48163888889°S, 56.4573333333°W | 1 | 2 | 3 | Present work | |
Manaus, Amazonas state | 6 | 3.13333333333°S, 59.9500000000°W | 8 | 4 | 12 | Present work | |
Uatumã River, Amazonas stateM | 9 | 2.017940°S, 58.203228°W | 1 | 1 | 2 | Present work | |
Cuieiras River, Amazonas state | 5 | 2.70708611111°S, 60.3738388889°W | 2 | 2 | 4 | Present work |
Sampling sites plotted on the Amazon basin map, Amazonas State: 1 Japurá River, Japurá city 2 Juruá River, Juruá city 3 Negro River, Santa Isabel do Rio Negro city 4 Purus River, Tapauá city 5 Cuieiras River, Manaus city 6 Manaus city, urban área: Federal University of Amazonas’s campus (UFAM) and Isaac Sabá Oil Refinery) 7 Aripuanã River, Novo Aripuanã city 8 Uatumã River, Presidente Figueiredo city 9 Jatapú River, São Sebastião do Uatumã city; Pará State: 10 Trombetas River, Oriximiná city 11 Tapajós River, Aveiro and Santarém cities 12 Jari River, Almeirim city; Rondônia State: 13 Madeira River, Porto Velho city. Geographic coordinates at the Table
Karyotypes under conventional staining (I), C-band (II), 18S rDNA and Ag-NOR (III), sex chromosomes in the boxes: a Marmosa demerarae b Metachirus nudicaudatus c Gracilinanus emiliae d Marmosa murina, (IV) variations on the 18S sites found in the individuals from Purus River, Tapauá city, Amazonas State e Caluromys philander f Caluromys lanatus g Marmosops pinheiroi.
Karyotypes under conventional staining (I), C-band (II), 18S rDNA and Ag-NOR (III), sex chromosomes in the boxes: a Glironia venusta bMonodelphis aff. adustac Monodelphis touan d Monodelphis emiliae e Monodelphis brevicaudata f Monodelphis sp. g Didelphis marsupialis.
All voucher specimens: Glironia venusta Thomas, 1912: (BAC 80) – Caluromys philander Linnaeus, 1758: Tapajós River (male: SISTAP-M-297; SISTAP-M-305; SISTAP-M-318; SISTAP-M-382; female: SISTAP-M-244); Trombetas River (female: CTGA-M-652); Purus River (female: CAN 34); Manaus (female: MSN 01); (female: BAC 102) – Caluromys lanatus Olfers, 1818: Japurá River (female: CTGA-M-701) – Marmosops sp. Matschie, 1916: Aripuanã River (female: MCA 3; MCA 7; MCA 8; MCA 26; MCA 31; MCA 35; male: MCA 4; MCA 16; MCA 38; MCA 39); Jari River (female: TAG 3459; RNL 70); Juruá River (male: EE 107; EE 139; female: EE135); Cuieiras River (female: EE 198; EE 211; male: EE 192; EE 201; EE216) – Marmosops bishopi Pine, 1981: Negro River (male: SISIS-M-127); Purus River (male: SISPUR-M-135; SISPUR-M-157; SISPUR-M-160; SISPUR-M-164; SISPUR-M-135; CAN 30; CAN 51; female: CAN 48) – Marmosops pinheiroi Pine, 1981: Tapajós River (male: SISTAP-M-237; SISTAP-M-278; female: SISTAP-M-268; SISTAP-M-277) – Marmosops parvidens Tate, 1931: Trombetas River (male: CTGA-M-501; CTGA-M-516; CTGA-M-531; CTGA-M-532; CTGA-M-551; CTGA-M-555; CTGA-M-581; CTGA-M-600; female: CTGA-M-533) – Marmosops impavidus Tschudi, 1845: Purus River (male: SISPUR-M-149) – Marmosops cf. pakaraimae Voss, Lim, Díaz-Nieto et Jansa 2013: Japurá River (male: SISJAP-M-705) – Marmosa murina Linnaeus, 1758: Jari River (male: RNL 45); Uatumã River (male: CEF 4; CEF 8; CEF 18; CEF 27; CEF 28; CEF 32; female: CEF 16; CEF 34; CTGA-M-8; CTGA-M-22; CTGA-M-41;), Negro River (male: SISIS-M-57; SISIS-M-63); Trombetas River (female: CTGA-M—519); Purus River (male: CAN 43); Japurá River (male: CTGA-M-708) – Marmosa murina Linnaeus, 1758: Aripuanã River (female: MCA12, Japurá River (male: SISJAP-M-764)- Gracilinanus emiliae Thomas, 1909: Tapajós River: (male: SISTAP- M-245; SISTAP- M-343; SISTAP- M-344; SISTAP- M-345) – Marmosa demerarae Thomas, 1905: Aripuanã River (female: MCA 27; MCA 36; MCA 58; MCA 65; male: MCA 21; MCA59); Jari River (female: RNL 31; RNL 48; male: RNL 30; MCA 32; MCA 46; MCA 49; MCA 58; MCA 61; MCA 64; MCA 66; MCA 67) Juruá River (female: EE136; male: EE 143); Manaus (female: EE 149: EE 150; EE 151; EE 154; EE 158; EE 159; EE 169; EE 222; EE 228; 229; EE 234; male: EE 157; EE 167; EE 170; EE 176; EE 189; EE 194; EE 196; EE 202; EE 215; EE 220; EE 235); Cuieiras River (female: EE 193; EE 219); Tapajós River (female: SISTAP-M-229; SISTAP-M-241; SISTAP-M-321; SISTAP-M-333; SISTAP-M-369; male: SISTAP-M-267; SISTAP-M-279; SISTAP-M-322); Trombetas River (female: CTGA-M-579; CTGA-M-590; CTGA-M-622; CTGA-M-667; CTGA-M-672; male: CTGA-M-535; CTGA-M-539; CTGA-M-557; CTGA-M-558; CTGA-M-572; CTGA-M-573; CTGA-M-578; CTGA-M-580; CTGA-M-613); Negro River (female: SISIS-M-85; SISIS-M-110; SISIS-M-117; SISIS-M-128; male SISIS-M- 86); Purus River (female: SISPUR-M-145; CAN 25; CAN 31; CAN 50: male: SISPUR-M-144; SISPUR-M-147; SISPUR-M-148) – Monodelphis aff. adusta Thomas, 1897: Madeira River (male: MCA 15) – Monodelphis touan: Jari River (male: TAG 2731; RNL 68) – Monodelphis sp. Burnett, 1830: Purus River: (male: CAN 44) – Monodelphis emiliae Thomas, 1912: Aripuanã River (female: MCA 31) – Metachirus nudicaudatus Geoffroy et Saint-Hilaire, 1803: Jari: River (RNL 47); Cuieiras River: (female: EE 200); Tapajós River (female: SISTAP-M-230; SISTAP-M-230; male: SISTAP-M-251; SISTAP-M-269); Trombetas River: (female: CTGA-M-655); Jatapú River: (female: CTGA-M-52; CTGA-M-58); Negro River: (female: SISIS-M-64; SISIS-M-78; male: SISIS-M-84; SISIS-M-116); Purus River: (male: CAN 33) – Didelphis marsupialis Linnaeus 1758: Jari River: (female: RNL 44; RNL 53; RNL 59; male: RNL 52; RNL 55; RNL 62; RNL 63); Manaus: (female EE 174; EE 197; EE 204; EE 224; EE 227; EE 246; EE 250; EE 155; EE 155; EE 173; EE 183; EE 190; EE 203; EE 205; EE 206; EE 223; EE 232; EE 233; EE 237; EE 247;EE 248; EE249; EE 190); Uatumã River (female: CEF 5; male: CEF 13); Trombetas River (female: CTGA-M-594; CTGA-M-606; male: CTGA-M-607); Purus River (male: SISPUR-M-185); Negro River (male: SISIS-M-73):Tapajós River (female: SISTAP-M-324; SISTAP-M-346; SISTAP–M-347;male: SISTAP-M-243); Japurá River: (male: CTGA-M-732).
The metaphases were obtained from bone marrow by in vivo method according to Ford and Harmerton (1956). Each animal received 1mL/100g weight of a 0,0125% colchicine solution for 30 minutes, the cells were exposed for 20 minutes to a 0,075M KCl solution, fixed 3:1 in methanol and acetic acid and stored at -20 °C. The C-band and Nucleolus Organizing Regions (NORs) patterns were determined according to the techniques described by
Among the 18 species analyzed, 11 showed 2n=14; six 2n=18 and one 2n=22 chromosomes (Table
In the species with 2n=14, we observed a very similar structure among the autosomes. These karyotypes include six autosome pairs (Fig.
We observed three different morphologies for X chromosome: metacentric in G. emiliae and Marmosops spp. (Fig.
The bare-tailed woolly opossum (C. philander) has X chromosome either acrocentric or submetacentric, with females being either homozygous or heterozygous carriers of the heteromorphic X (Fig.
Geographic locations of Caluromys philander individuals and its sexual chromosomes morphology data in South America. Literature data represented by empty circles and present work represented by full circles: (● 14) Venezuela,
Geographic locations of Marmosa murina individuals and its X chromosome morphology data in South America. Literature data represented by empty circles and present work represented by full circles: (● 18) Villa Vivêncio, Colômbia,
The Y chromosome was acrocentric in G. emiliae, Marmosops spp., M. demerarae and M. nudicaudatus (Fig.
Among the species with 2n=18 chromosomes, FNa=20 was recorded in Glironia venusta Thomas, 1912, with formula 2m+2sm+2st+10a+XX/XY (Fig.
Didelphis marsupialis was the only species that presented 2n=22 chromosomes and FNa=20, with formula 20a+XX/XY (Fig.
The position of the heterochromatin on the 2n=14 species was centromeric, being conspicuous in M. demerarae (Fig.
Two C-band patterns were present in the X chromosome for species of Marmosops. In pattern 1, X was entirely heterochromatic except for a proximal band in the long arms (Fig.
Comparative cytogenetic data of the didelphid species analyzed in the present study and those from the literature. In Locality, numbers indicate sampling sites as in the maps of Figures
Species | Locality number | 2n | FNa | NORs Pair/arm | 18S rDNA | X/Y | X chromosome C–band | Source |
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Caluromys philander | 10; 11; 15; 16 | 14 | 22 | 6p | 6p | a/d | B |
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4; 6; 14; 15; 17 | 14 | 22 | 6p | 6p | sm/d | B | São Paulo State, |
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12 | 14 | 22 | 6p | 6p | sm/a | B |
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Caluromys lanatus | 1 | 14 | 22 | 6p | 6p | sm/– | A. | Present work |
Marmosa murina | 2; 3; 8; 10; 12; 18; 19; 20; 25; 26 | 14 | 22 | 5q;6p | 5q;6p | (m)sm/ d | A |
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16; 22; 25; 24; 26; 27; 28; 29 | 14 | 22 | 5q;6p | 5q;6p | a/ d | A |
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4 | 14 | 22 | 5q;6p | 1p; 3p; 5q; 6p | a/d | A | Present work | |
Marmosa demerarae | 2; 3; 4; 5; 6; 7; 9; 10; 11; 12 25, 26 | 14 | 20 | 5q; 6p | 5q;6p | a/a | C |
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La Paz, Bolívia | 14 | 20 | – | – | a/a | – |
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16 | 14 | 24 | 5q; 6p | a/a |
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– | 14 | 24 | 5pq; 6p | 5pq; 6p | a/a |
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Rio Grande do Sul | 14 | 24 | 5pq; 6p | – | a/a |
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Marmosops bishopi | 4; 7; | 14 | 24 | 6p | m/a | C; D | Present work | |
3 | 14 | 24 | 6p | m/a | C | Present work | ||
Marmosops pinheiroi | 11 | 14 | 24 | 6p | 6p | m/a | C | Present work |
Marmosops parvidens | 5; 10; 12 | 14 | 24 | 6p | m/a | C; D | Present work | |
9 | 14 | 24 | 6p | m/a | D | Present work | ||
Marmosops impavidus | 2 | 14 | 24 | 6p | m/a | C | Present work | |
Marmosops pakaraimae | 1 | 14 | 24 | 6p | m/a | C | Present work | |
Gracilinanus emiliae | 11; 25; 26 | 14 | 22 | 6p | 6p | m/a | A |
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Metachirus nudicaudatus | 2; 5 10; 11; 12 | 14 | 20 | 6p | 6p | a/a | A | Present work |
Glironia venusta | 13 | 18 | 20 | 6p | 6p | a/– | A | Fantin e da Silva 2011, Present work |
Monodelphis touan | 12 | 18 | 28 | Xp | Xp | a/a | A | Present work |
Monodelphis sp. | 4 | 18 | 32 | 7p | 7p | sm/a | – | Present work |
Monodelphis aff.adusta | 7 | 18 | 30 | 7p | a/d | A | Present work | |
Monodelphis emiliae | 7, Juruá River, Acre | 18 | 30 | 7p | sm/– | A |
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Monodelphis brevicaudata | 3; | 18 | 30 | 7q, Xp, Yq | 7q, Xp, Yq | sm/a | – | Present work |
Roraima and Pará states | 18 | 30 | Xp | a/d |
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Didelphis marsupialis | 5; 6; 9; 10; 11 | 22 | 20 | 5q;7pq;8q | 5q;7pq;8q | a/a | A | Present work |
In the species with 2n=18 chromosomes, the heterochromatin was centromeric in G. venusta (Fig.
NORs confirmed by FISH using the 18S rDNA probe were present in the short arms of pair 6 in all 2n=14 species and G. venusta (2n=18). In M. demerarae and M. murina sites were also detected in the terminal position of the long arms of pair 5 (Fig.
In D. marsupialis, both the 18S rDNA probe and silver were detected in three chromosome pairs. In two pairs, the sites were located in the terminal region of the long arms, while in one pair they were bitelomeric (Fig.
In the last decade, advances in systematic and taxonomic studies of the family Didelphidae introduced changes in the taxonomy and nomenclature of several of its taxa (
The autosome structure observed here corroborates karyotypic conservation in the diploid number and chromosomal formula (NFa) as previously described in the didelphid species Didelphis marsupialis, Marmosa demerarae, Metachirus nudicaudatus, Monodelphis touan (previously named M. brevicaudata), Monodelphis aff. adusta (previously named as M. cf. emiliae) and for species of Marmosops (
Although didelphids are generally considered to have a conserved karyotype, by comparing the karyotypes among different genera, it was possible to associate them with certain species due to the presence of diagnostic characters. For example, M. demerarae and M. murina differ in their FNa, morphology, and sex chromosome size. In species of the genus Monodelphis, morphological variation in chromosomes was restricted to pair 6, which grants an FNa varying between 30 (as observed in M. aff. adusta, M. touan and, M. brevicaudata) and 32 arms (Monodelphis sp.). However, the same does not occur for the genus Marmosops, in which the five species analysed, present a very similar chromosome macrostructure.
The genus Marmosa has a complex taxonomy and recently underwent great taxonomic changes, with all species of Micoureus, formerly treated as a separate genus, now considered as a subgenus of Marmosa. Considering the taxonomic instability in Didelphidae, with individuals being reclassified, and some complex of species being divided into two or more valid taxa, even purportedly karyotyped species may in fact have their karyotypes still unknown. Thus, our knowledge as to how many and which species among didelphids were karyotyped remains unstable. A revision of the literature for species with reported karyotypes is required.
Apparently, there is a likely geographical structure in the distribution of the morphological forms of the X chromosome in Marmosa murina, with the metacentric X so far found in the northern and western parts of its distribution, the submetacentric X prevailing in the Amazon basin of Brazil and the acrocentric forms prevailing in the other known localities in central and eastern Brazil (Fig.
Among the Amazonian marsupials analyzed here, the variation in centromere position and heterochromatin patterns of the X chromosome is noteworthy.
We observed chromosomal conservatism in the heterochromatin pattern in eight didelphid species: (C. lanatus, G. venusta, D. marsupialis, M. touan, M. aff. adusta, M. emiliae, G. emiliae and M. nudicaudatus). C. philander presented heterochromatic pattern different from the heterochromatic distribution reported in the literature for this species (
Thus, heterochromatin distribution patterns can serve as a cytotaxonomic character, as well as shedding light on chromosomal evolution and regulation of gene expression. However, our results demonstrate that, except for Marmosops spp., the other species under study presented little heterochromatin intraspecific variation, including the X chromosome. Thus, this character alone does not allow for distinguishing among didelphid populations, although heterochromatin distribution may be an effective character for distinguishing between certain species pairs. This is the case for M. demerarae and M. murina, with the former presenting larger centromeric heterochromatic blocks than the latter, and between C. philander and C. lanatus, both with 2n=14 and NF=24, but with distinct heterochromatic patterns.
The NOR in Didelphidae can be simple or multiple. According to
According to the literature, in Monodelphis there are NOR sites on pair 7 and on the X chromosome of Monodelphis aff. adusta and Monodelphis sp. (
In the Y chromosome of M. touan, FISH did not confirm the marking. This situation was verified in other organisms, where precipitation in the heterochromatic regions took place but could lead to an erroneous interpretation of the distribution of this marker (
When mapping the NOR character on the phylogenetic tree of Jansa and Voss (2014, fig. 01) (not shown here), we verified that multiple NORs are distributed in two distinct lineages: the first in species of the genus Marmosa and the second in species with 22 chromosomes of the genera Didelphis and Philander Brisson, 1762. The mapping of the simple condition onto the phylogenetic tree depicts a wide distribution for this character, present at the base of the tree (Caluromys philander, C. lanatus, Glironia venusta) and in at least one or more species of the remaining major clades (Gracilinanus emiliae, Marmosops spp., Metachirus nudicaudatus, Monodelphis touan, Monodelphis kunsi, and Monodelphis dimidiata) (
When mapping the NOR character on the phylogenetic tree of
In M. murina, intraspecific geographic variation in NORs were detected. Specimens from the Purus River have multiple NORs, those collected in the state of Goiás have simple NOR in the short arms of pair 6 (
Our results indicate geographic variation in NORs for M. demerarae. Amazonian specimens analysed did not present ribosomal cistrons in the short arms of the fifth pair, as recorded for specimens from the Atlantic forest in the Rio Grande do Sul and São Paulo states of southern Brazil (
In Didelphis marsupialis from several Amazonian sites, only NOR activity varied, as was already reported in specimens from the Atlantic forest (
The cytogenetic data presented here shows that didelphid marsupial karyotypes present intraspecific variation in the morphology of sex chromosomes and in chromosomic markers (C-band and NOR) and present some geographic variation in the distribution of these features for several species. Furthermore, there are many areas in the Amazon, including the transition zone between the Amazon and the Cerrado biomes, which do not have cytogenetic records for any didelphid species. This situation seriously undermines our understanding of the significance of the recorded variation, whether it is part of a continuous gradient, or whether it represents intraspecific gradations, or whether it is related to new lineages or cryptic species still uncovered. Thereby, despite the chromosomal stability related to diploid numbers and chromosomal formula in marsupials across continents, didelphids present some intra- and interspecific chromosomal variations, probably related to frequent chromosomal rearrangements. Additional systematic sampling and analyses will be required for a better understanding of the karyotypic evolution of this group.
This work was supported by Fundação de Amparo a Pesquisas do Estado do Amazonas (FAPEAM), Coordenação de Aperfeiçoamento Pessoal (CAPES), the SISBIOTA/Rede BIOPHAM (CNPq); and Pró-Amazônia (CAPES). We thank Dra. MC Gross e Dr. CH Schneider who cooperated with research development and G.H. Shepard Jr. for reviewing the English of the manuscript.
Voucher specimens
Data type: Microsoft Word Document (.docx)
Explanation note: All analyzed specimens were deposited at Mammals Collection in the Instituto Nacional de Pesquisas da Amazônia (