| Authors |
Maxim D. Simakov, Research engineer of the laboratory of molecular ecology and systematics of animals, Penza State University (40 Krasnaya street, Penza, Russia), E-mail: maksimakov@bk.ru
Mikhail V. Korepov, Candidate of biological sciences, associate professor of the sub-department of biology and chemistry, Ulyanovsk State Pedagogical University named after I.N. Ulyanov (4/5 Lenina square, Ulyanovsk, Russia), E-mail: korepov@list.ru
Anton A. Kuzmin, Candidate of biological sciences, associate professor of the sub-department of biotechnology and technosphere security, Penza State Technological University (1A/11 Baidukova lane / Gagarina street, Penza, Russia), E-mail: kuzmin-puh@yandex.ru
Sergey V. Titov, Doctor of biological sciences, professor, dean of the faculty of physics, mathematics and natural sciences, Penza State University (40 Krasnaya street, Penza, Russia), E-mail: svtitov@yandex.ru
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| Abstract |
Background. Despite the study of the variability of the mtDNA control region of the esternimperial eagles in the Volga population, due attention was not paid to cyt b, the second mitochondrial marker. Therefore, studies of cyt b gene polymorphism in Volga population of Aquila heliaca, taking into account the available data on mtDNA variability, are relevant and useful. This is necessary to clarify the population differentiation of imperial eagles at the level of regional breeding groups. The purpose of the research is to study the level of polymorphism of the imperial eagle (A. heliaca) on the territory of the Ulyanovsk region according to the analysis of the variability of the cyt b gene of mitochondrial DNA. In addition, it was supposed to identify specific genetic and ecological traits of the Volga population. Materials and methods. The material for genetic studies was obtained from samples (n = 15) of linn feathers of imperial eagles collected in the Ulyanovsk region. DNA was isolated according to the standard procedure of phenol-chloroform extraction after treatment of the material with proteinase K and SDS. Original primers – AqH Cyt b D (5'–GAAAGTCCCACCCTCTGCT–3') and AqH Cyt b R (5'–GCTCCAATTAGGGAAGAG–3') were used for genetic analysis of polymorphism of the cyt b gene fragment (977 bp). The polymerase chain reaction (PCR) was carried out in 25 μl of a standard reaction mixture
at an annealing temperature of 60 ºC. Fragments of the mtDNA control region (n = 15) were sequenced using an ABI 3500 sequencer (Applied Biosystems). Haplotypic and nucleotide diversity were studied using the DnaSP 5.10.01 program. Maximum likelihood (ML) methods in the MEGA X program were used to reconstruct phylogenetic relationships. The Median network (Median Joining) of mitochondrial haplotypes was built in the PopART program using the TCS algorithm. The obtained nucleotide sequences of mtDNA haplotypes of the cyt b gene of the imperial eagle were deposited in GenBank NCBI – OL421575–OL421587. For statistical processing of the results, the significance level p ≤ 0.05 was set for all tests. Results. The data obtained in the course of the study on the latent polymorphism of the mitochondrial cyt b gene of geographical breeding groups of the imperial eagle on the territory of the Ulyanovsk region indicates the haplotypic diversity and specificity of the Volga population of A. heliaca. The maximum number of specific haplotypes is observed in the northeastern and southern breeding groups of the Volga population. They are confined to the most fragmented forest-steppe and steppe areas of the Ulyanovsk region (Hap 3, Hap 6, Hap 7 – 60 % and Hap 9, Hap 10, Hap 11 – 43 %, respectively). At the same time, the most common haplotype in the Volga population is Hap 4. It forms the basis of haplogroup AqH_1 (= G1) and each of the selected geographical breeding groups (NE – 40 %, C – 33 %, S – 57 %). Such pronounced differences in the shared haplotypic structure of the groups were insufficient to detect statistically significantly genetic differentiation of the Ulyanovsk subpopulation of the Volga population of the imperial eagle. Conclusions. The obtained data on the variability of the cyt b gene fragment indicate that the level of polymorphism of the imperial eagle is low, and the population recorded in the Ulyanovsk region is fairly homogeneous.
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| References |
1. Krasnaya kniga Penzenskoy oblasti: v 2 t. = The Red Book of Penza region: in 2 volumes. 2nd ed. Penza; Voronezh: Voronezhskaya oblastnaya tip. – Izd-vo im. E.A. Bolkhovitinova, 2019;2:264. (In Russ.)
2. Krasnaya kniga Respubliki Mordoviya: v 2 t. = The Red Book of the Republic of Mordovia: in 2 volumes. Saransk: Mordovskoe knizhnoe izd-vo, 2005;2:336. (In Russ.)
3. Krasnaya kniga Chuvashskoy Respubliki. T. 1. Ch. 2. Zhivotnye = The Red Book of the Chuvash Republic. Volume 1. Part 2. Animals. Cheboksary: IPK Chuvashiya, 2011:372. (In Russ.)
4. Krasnaya kniga Rossiyskoy Federatsii (zhivotnye) = The Red Book of the Russian Federation (animals). Moscow: Tovarishchestvo nauchnykh izdaniy KMK, 2008:855. (In Russ.)
5. Belik V.P., Galushin V.M. Population structure of the range of the Imperial Eagle in Northern Eurasia. Korolevskiy orel: Rasprostranenie, sostoyanie populyatsiy i perspektivy okhrany orla-mogil'nika (Aquila heliaca) v Rossii: sb. nauch. tr. = Royal Eagle: Distribution, population status and prospects for the conservation of the Eastern Imperial Eagle (Aquila heliaca) in Russia: collected papers. Moscow: Soyuz okhrany ptits Rossii, 1999:129–139. (In Russ.)
6. Korepov M.V. Distribution and abundance of the Eastern imperial eagle (Aquila heliaca, Falconiformes, Accipitridae) in the central part of the Volga Upland. Zoologicheskiy zhurnal = Zoological journal. 2012;91(2):190–201. (In Russ.)
7. Korepov M.V., Borodin O.V. Solnechnyy orel (Aquila heliaca) – prirodnyy simvol Ul'yanovskoy oblasti = The solar eagle (Aquila heliaca) is a natural symbol of the Ulyanovsk region. Ul'yanovsk: Povolzh'e, 2013:213. (In Russ.)
8. Seibold I., Helbig A.J., Meyburg B.-U. [et al.]. Genetic Differentiation and Molecular Phytogeny of European Aquila Eagles according to Cytochrome b Nucleotide Sequences. Eagle Studies. World Working Group on Birds of Prey (WWGBP). Berlin, London & Paris. 1996:1–15.
9. Korepov M.V., Stryukov S.A., Korepova D.A. [et al.]. Genetic differentiation and polymorphism of the Volga region population of the imperial eagle (Aquila heliaca, Falconiformes, Accipitridae) according to mitochondrial DNA analysis. Izvestiya vysshikh uchebnykh zavedeniy. Povolzhskiy region. Estestvennye nauki = University proceedings. Volga region. Natural sciences. 2017;(3):3–15. (In Russ.). doi:10.21685/2307-9150-2017-3-1
10. Simakov M.D., Korepov M.V., Stryukov S.A. [et al.]. New data on the polymorphism of the Volga region population of the imperial eagle (Aquila heliaca, Falconiformes, Accipitridae) based on the results of the analysis of mitochondrial and microsatellite DNA. Russian Journal of Ecosystem Ecology. 2021;6(3). (In Russ.). doi:10.21685/ 2500-0578-2021-3-2
11. Wink M., Elsayed A.-A., Gonzalez J., Olsen J. Phylogenetic relationships in the Bonelli’s eagle (Hieraaetus fasciatus) species complex based on nucleotide sequences of mitochondrial and nuclear marker genes. Heidelberg, Baden-Wuerttemberg, Germany, 2016.
12. Helbig A.J., Kocum A., Seibold I., Braun M.J. A multi-gene phylogeny of aquiline eagles (Aves: Accipitriformes) reveals extensive paraphyly at the genus level. Mol. Phylogenet. Evol. 2005;35(1):147–164.
13. Lerner H.R., Mindell D.P. Phylogeny of eagles, Old World vultures, and other Accipitridae based on nuclear and mitochondrial DNA. Mol. Phylogenet. Evol. 2005;37(2): 327–346.
14. Zhou L., Dong Y. Complete mitochondrial genome of Aquila heliaca. Hefei, Anhui, China, 2017.
15. Vili N., Nemesházi E., Kovács S. [et al.]. Factors affecting DNA quality in feathers used for non-invasive sampling. Journal of Ornithology. 2013;154:587–595.
16. Arrigi F.E., Bergendahl G., Mandel M. Isolation and characterization of DNA from fixed cells and tissues. Exp. Cell. Res. 1968;(50):47–53.
17. Sambrook J., Fritsch E.F., Maniatis T. Molecular cloning: a laboratory manual. New York: Cold Spring Harbor Laboratory Press, 1989. Available at: www.cshlpress.com
18. Hall T.A. BioEdit: a user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. Nucleic Acids Symposium Series. 1999;41:95–98.
19. BioEdit 7.2. Available at: https://bioedit.software.informer.com
20. Multiple Sequence Alignment. Available at: http://www.ebi.ac.uk/clustalw
21. Librado P., Rozas J. DnaSP v5: A software for comprehensive analysis of DNA polymorphism data. Bioinformatics. 2009;25:1451,1452.
22. Kumar S., Stecher G., Li M. [et al.]. MEGA X: Molecular Evolutionary Genetics Analysis across computing platforms. Molecular Biology and Evolution. 2018;35:
1547–1549.
23. Leigh J.W., Bryant D., Nakagawa S. POPART: full feature software for haplotype network construction. Methods in Ecology and Evolution. 2015;6(9):1110–1116. doi:10.1111/2041-210x.12410
24. Clement M., Posada D., Crandall K.A. TCS: a computer program to estimate gene genealogies. Molecular Ecology. 2000;9(10):1657–1659. doi:10.1046/j.1365-294x.2000. 01020.x
25. Borodin O.V., Smirnova S.L., Sviridova T.V. [et al.]. The current state of the imperial eagle in the Ulyanovsk region. Korolevskiy orel: Rasprostranenie, sostoyanie populyatsiy i perspektivy okhrany orla-mogil'nika (Aquila heliaca) v Rossii: sb. nauch. tr. = Royal Eagle: Distribution, population status and prospects for the conservation of the Imperial Eagle (Aquila heliaca) in Russia: collected papers. Moscow: Soyuz okhrany ptits Rossii, 1999:68–73. (In Russ.)
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