Preservation of the genetic diversity of the sevryuga (Acipenser stellatus Pall) in conditions of artificial reproduction

. According to the lactadehydrogenase locus, the genetic diversity of 9 samples of the Volga sevryuga of different origin was studied. In total, 733 pieces of young sevryuga, 99 pieces of producers and 107 pieces were studied. four-year-olds of sevryuga with MPERZ. To assess the genetic variability of lactate dehyrogenase allozymes, the method of polyacrylamide gel electrophoresis was used, as the most appropriate for assessing the influence of the environment in natural and domesticated populations. It is shown that the young sevryuga, grown in the conditions of 6 Astrakhan sturgeon hatcheries, differs significantly from the control group of natural producers. The LDH-3 lactate dehydrogenase locus is represented by 6 genotypes. 5 genotypes were found in producers, in the absence of only genotype 100/112. Genotypes 70/112 and 112/122 are missing in the samples from all plants and MPERZ, while genotype 70/70, which was found in excess at MPERZ, is also missing in three plants. However, in general, only 2 genotypes out of 6 are missing for all ARI. A study of the dynamics of the frequency of alleles in this set of samples showed that in 4 samples with ARI and on MPRZ there is no allele 112. However, in general, the ARI allele persists. Thus, it is shown that due to the different genetic variability of LDH-3 in juveniles from different acute respiratory infections, the genetic diversity in the samples as a whole approaches the indicators of producers.


Introduction
Sevryuga is a semi-passable fish of the sturgeon family, which feeds in the marine environment, and reproduces in rivers.In the past, it was the most numerous species of sturgeon.Its maximum catch in the Volga River in 1986 was 5.2 thousand tons [1].Currently, the catch of the sevryuga is carried out only for scientific research and artificial reproduction.However, the number of sturgeon fish in the sea is estimated in tens of thousands of pieces, mature producers do not enter the river, so the catch according to quotas for reproduction is carried out in the pre-estuary zone of the Volga River and in the sea [2].
Sevryuga in the Astrakhan region is reproduced mainly by artificial method at 6 sturgeon hatcheries (ARI).The total area of the outgrowth ponds of fish hatcheries is 982.4 hectares, the potential total capacity is up to 57.522 million pieces of juvenile sturgeon annually.
Due to the poor survival rate of sturgeon juveniles released into the river and the low commercial return of factory fish, the idea of transferring the reproduction of sturgeon fish completely into artificial conditions of industrial systems has been actively promoted in recent decades.It is assumed that the cultivation of large juveniles will increase its resilience.
The distinctive features of artificial reproduction from natural reproduction are the following: hormonal stimulation, natural or domesticated producers; lifetime production of mature eggs by straining, fertilization and de-gluing; incubation at high density of bookmarks in Sturgeon-type apparatuses; keeping larvae in pools, before switching to active nutrition; transplanting larvae into gardens installed in ponds, for transition to active nutrition; transfer of larvae to ponds for further cultivation.At the same time, at each stage of the production process, eggs and larvae fall into conditions different from natural ones and selection takes place in favor of the fittest individuals [3,4,5,6].
However, in order to preserve the genetic diversity of a species, an indispensable condition is the preservation of the habitual environment in which it has developed phylogenetically and to which it is ontogenetically adapted.
In this regard, the aim of the study was to study the genetic diversity of the lactate dehydrogenase locus in the young sevryuga, grown in various conditions of the Astrakhan ARI.

Materials and methods
The material of juveniles and producers was collected at the BIOS Research and Production Center (BIOS) and 6 operating ARI of the Astrakhan region: Kizansky, Bertyulsky Sergievsky, Alexandrovsky, Zhitninsky, Lebyazhye.Growing ponds and production facilities at all plants have their own characteristics, so the young are grown in non-standard conditions.The material from four-year-olds of the Volga sevryuga, grown in industrial conditions in a closed water supply installation from caviar, was collected at the Mozhaisk industrial and experimental fish hatchery of FSUE Glavrybvod.
Biological material (tissues and heart) was taken from the young sevryuga when it was released into the river or during the last bonitation in ponds in 2014.Biological samples were collected from producers and four-year-olds of sevryuga in the form of pieces of fins.The bioassays were frozen and stored in the refrigerator at a temperature no higher than minus 18 ° C. The juveniles of the sevryuga and 99 pieces of producers were examined, as well as 107 pieces.four-year-olds of sevryuga with MPERZ.
To assess the genetic variability of lactate dehyrogenase allozymes, the method of polyacrylamide gel electrophoresis was used, as the most appropriate for assessing the influence of the environment in natural and domesticated populations.As practice shows, the use of this method allows us to characterize both individual fish genotypes and the level of heterogeneity of the reproductive herd as a whole [7,8].
The selective load of protein markers is recognized as very informative when analyzing the relationship between the fitness of the genome to environmental conditions, while DNA markers that have adaptive significance for sevryuga have not yet been studied [9].In addition, the technique of electrophoretic detection of allozymes is easy to implement and much cheaper compared to the analysis of DNA markers.
Genetic and biochemical studies were carried out by polyacrylamide gel electrophoresis in the institute laboratory.The distribution of allelic variants of the enzymes lactate dehydrogenase (LDH-3 locus) and esterase (EST-2 locus) in stellate sturgeon was studied.The genetic heterogeneity of the samples was assessed according to the following parameters: the frequency of alleles and genotypes (p); levels of observed (Hob) and expected (He) heterozygosity; deviation of heterozygosity from the expected (D).The correspondence between the observed and theoretical expected frequencies was estimated according to the Hardy-Weinberg principle using the χ2 test.The zymograms were interpreted according to the Atlas supplement.[8].

Results and discussion
The study of the genetic variability of the sevryuga showed that LDH-3 should be represented by 6 genotypes (Table 1).The genotypes of 9 samples of juveniles and producers were analyzed.5 genotypes were found in producers, in the absence of only genotype 100/112.There are no genotypes 70/112 and 112/122 in the samples of juveniles from all plants and MPERZ, while the genotype 70/70 found in excess at MPERZ is also missing in three plants.
In general, it can be said that 3 genotypes out of 6 possible were detected in 3 samples with acute respiratory infections and MPERZ, only 2 genotypes were detected in 3 samples with acute respiratory infections and 4 genotypes were detected on the Swan ARI.Due to some discrepancy between the missing and present genotypes on different ARI, in general, only 2 genotypes out of 6 are missing for all ARI.Thus, we see that due to the different genetic variability of LDH-3 in juveniles from different ARI, the genetic diversity in the samples as a whole approaches the indicators of producers.
However, the absence of 2 genotypes out of 6 in the young sevryuga released into the river shows that changes in values exceed 10-20% of the population variability level.This makes it possible to characterize the gene pool of the population at the studied locus as unstable.A study of the dynamics of the frequency of alleles in this set of samples showed that in 4 samples with ARI and on MPRZ there is no allele 112 (Table 2).However, in general, the ARI allele remains.When growing sevryuga, MPERZ allele 112 is eliminated.Therefore, when creating reproductive herds in industrial and natural conditions, it should be borne in mind that the most rare alleles are eliminated and the gene pool is depleted.Only producers represented mainly by fish of their natural population and a combined sample of all ARI had all alleles marked.The observed heterozygosity at the plants ranges from 0.056 to 0323, and on average is 0.212, which is a normal value, relatively high heterozygosity at the MPERZ, reaching 0.383.
It is known that the genetic structure of any set of organisms is influenced by environmental conditions.The adaptive nature of protein polymorphism makes it possible to assess this effect, since individuals with certain enzyme genotypes gain an advantage in transformed environmental conditions.We have studied an enzyme included in the respiratory process, which is influenced by a combination of factors: temperature, flow rate, hydrochemical composition, water exchange rate, feeding intensity, stress, and more.In addition to technological factors, a number of abiotic and biotic factors affect the dynamics of the gene pool when growing juveniles in the outgrowth ponds of the Astrakhan ARI.First of all, this is the temperature and oxygen regime, the area and depth of ponds, the availability of food, the presence of predators and much more, which differ in different farms.Also, the farms have their own domesticated reproductive herds, which have a specific gene pool [14,15].
All factors together provide a variety of growing conditions and, accordingly, a variety of the gene pool in different ARI, not only by the LDH-3 locus, but also EST, according to which rare genotypes are also eliminated in juveniles raised on ARI [16].In general, a variety of conditions allows to preserve the genetic diversity of the young sevryuga released into the river.
Unification, industrialization of technologies and conditions for growing young sevryuga for release into the natural environment and the creation of reproductive herds will lead to depletion of the gene pool due to both the effect of the founders of reproductive herds and the selective influence of habitat conditions on certain genotypes.This is confirmed by our data on the elimination of 2 to 4 genotypes out of 6 in different ARI and the preservation of all genetic diversity in producers represented by fish from their natural population and a combined sample of fish from all ARI.

Conclusion
The study of the dynamics of genetic indicators in young sevryuga reared in different ARI showed that, compared with producers caught from natural populations, the gene pool is depleted in all ARI due to the elimination of the allele 112 LDH -3 and genotypes 70/112, 100/112, 112/112 with its participation.The multidirectional change in the frequency of occurrence of individual genotypes and alleles in different acute respiratory infections allows, in general, for acute respiratory infections to maintain genetic indicators at a relatively stable level.Therefore, the preservation of a variety of growing conditions for young sevryuga will contribute to the preservation of its genetic diversity.

Table 1 .
The frequency of occurrence of LDH-3 genotypes in the Volga sevryuga of different origin*.

Table 2 .
The frequency of occurrence of alleles at the LDH-3 locus and heterozygosity in samples of the Volga Sevryuga*.