NEW DATA ON THE POLYMORPHISM OF THE VOLGA POPULATION OF THE IMPERIAL EAGLE (AQUILA HELIACA, FALCONIFORMES, ACCIPITRIDAE) BASED ON THE RESULTS OF THE ANALYSIS OF MITOCHONDRIAL AND MICROSATELLITE DNA

Abstract

Background. The analysis of the available data on the study of the polymorphism of the mtDNA control region in the Eurasian populations of burial eagles (1996-2020) indicates their genetic differentiation by geographical distribution. The periodic detection of new haplotypes indicates the relevance of such studies and insufficient knowledge of this issue. The aim of the work is to clarify the level of polymorphism and identify specific genetic features of the Volga population of the imperial eagle (A. heliaca) according to the analysis of the variability of mitochondrial and microsatellite DNA. Materials and methods. The material for genetic research was new collections (n = 22) of linn feathers of burial eagles collected in the Ulyanovsk region (n = 15), the Republic of Tatarstan (n = 4), the Republic Mordovia (n = 2) and the Chuvash Republic (n = 1). DNA was isolated from feathers of adult birds and the pulp and blood of chicks according to the standard method of phenol-chloroform extraction. A fragment (345 pn) of the mtDNA control region (D-loop) and three tetranucleotide microsatellite repeats (IEAAAG09, IEAAAG11, IEAAAG15) were used for genetic analysis. Fragments of the mtDNA control region (n = 22) and microsatellite DNA (n = 171) were sequenced using an ABI 3500 sequencer (Applied Biosystems). Haplotypic and nucleotide diversity were studied using the DnaSP 5.10.01 program. To reconstruct phylogenetic relationships, Maximum Likelihood (ML) and Neighbor-Joining (NJ) methods were used in the MEGA X program. Microsatellite genetic data was analyzed using the Arlequin 3.5.2 and Structure 2.3.4 programs. The obtained nucleotide sequences of newly identified mtDNA haplotypes of the imperial eagle were deposited in GenBank NCBI under the numbers OK504653 (haplotype RE) and OK504654 (haplotype RF). For statistical processing of the results, a significance level of p < 0.05 was set for all tests. Results. Analysis of the distribution of mtDNA control region haplotypes across 5 geographically isolated Imperial eagle breeding groups in the Volga region (ULR-NW – northwest of the Ulyanovsk region, UlR-NE – northeast of the Ulyanovsk region, UlR–C – center of the Ulyanovsk region, UlR-S – south of the Ulyanovsk region, RTat – Republic of Tatarstan) revealed their subdivision. When analyzing the average numbers of nucleotide differences, the isolation of the breeding groups from the Republic of Tatarstan and the Ulyanovsk region is noticeable. The average number of nucleotide differences (k) between all nest groups was 1.606 and was lower than this indicator when comparing the group from Tatarstan with the Ulyanovsk groups (1.739). In the phylogenetic analysis, two haplogroups of the imperial eagle were identified. The first haplogroup (AQH_1) is formed by mtDNA haplotypes characterized by one or two nucleotide substitutions. The second haplogroup (AQH_2) consists of haplotypes having from 3 to 5 nucleotide substitutions. The results of the median test also indicate the existence of 2 genetically close groups of sequences of a fragment of the mtDNA control region corresponding to more or less altered haplotypes. The analysis of microsatellite DNA of geographically isolated breeding groups of imperial eagles in the Ulyanovsk region revealed a low level of internal polymorphism. Conclusion. The conducted studies have shown that the Volga population of the imperial eagle is genetically differentiated. It consists of two subpopulations – Ulyanovsk and Tatarstan, characterized by specific mtDNA haplotypes. These subpopulations are confined to the Volga and Bugulminsko-Belebeevskaya uplands. The analysis of microsatellite DNA, as well as the reconstruction of evolutionary events related to the variability of mtDNA haplotypes show that the level of polymorphism of the Ulyanovsk subpopulation of imperial eagles is low, and the population itself is fairly homogeneous.

Keywords

• aquila heliaca
• control region
• mtdna
• haplotype diversity
• volga region population