Development of a multilocus STR panel and assessment of the reliability of the origin of the aquaculture sterlet (Acipenser ruthenus)

Introduction. In modern practice of artificial breeding of sturgeon fish, obtaining viable offspring is of particular importance. In this context, the use of microsatellite markers to assess the relatedness and genetic diversity of sterlet in commercial aquaculture becomes relevant. This will make it possible to more accurately determine the origin of individuals and significantly increase the efficiency of breeding work at fisheries enterprises.
Object. The object of the study is samples of fin sections of aquaculture sterlet of various origins.
Materials and methods. The development of a microsatellite panel including 12 loci (AfuG 63, AfuG 112, Afu 68 b, LS-39, Spl-163, An20, AfuG 51, Aru13, AoxD161, AfuG 41, Aru18, LS-68) was carried out on the basis of the Federal State Budgetary Institution Federal Research Center VIZH them. OK. Ernst. Testing of the developed test system, as well as its information content, was carried out on a sample of hatchery sterlet (n=149) from three populations: Sukhonovskaya, among which there were two parental pairs with 30 known offspring (SX, n=65), Okaskaya (OK, n=50 ) and Lower Volga (NV, n=34).
Results and conclusions. Based on the results of using microsatellite markers, the reliability of the origin of three sterlet populations was assessed. Based on the polymorphism of 12 microsatellite loci for the three studied groups of sterlet, classical population genetic indicators were calculated: the average number of alleles per locus (NA = 7.528 ± 0.717), the number of effective alleles (NE = 4.098 ± 0.437), observed (HO = 0.571 ± 0.049) and expected (HE = 0.611 ± 0.049) heterozygosity, as well as the inbreeding coefficient (FIS = 0.046 ± 0.028). The probability of matching genotypes (PI) for the developed STR panel ranged from 3.7*10^-13 for OK to 8.1*10^-10 for SX, i.e. was practically excluded. The exclusion probability values (P1, P2, P3), calculated based on the results of the developed test system, were minimum for SX and maximum for OK and ranged from 99.88 to 99.99% for P1. The P2 parameter was 98.62% for the NV and 99.87% for the OK population. For P3 it was 99.99% for all studied groups. These data confirm the high information content of DNA analysis of 149 individuals to establish the authenticity of the origin of hatchery sterlet and the functionality of the developed STR panel. As a result of the research, a multiplex test system was created that allows analysis of twelve STR loci, and its high functional efficiency was demonstrated.

1. Volosnikov G. Review of data on biology of sterlet Acipenser ruthenus (Linnaeus, 1758). Vestnik of Astrakhan State Technical University. 2017. No. 2. Pp. 67-72.

2. Gessner J., Freyhof J., Kottelat M., Friedrich T. Acipenser ruthenus. The IUCN Red List of Threatened Species. 2022. e.T227A135062526.

3. Bykov A. D., Brazhnik S. Y. Current State of Sterlet Stocks and Artificial Reproduction in Russia. Fisheries issues. 2022. V. 23. № 3. Pp. 5-30.

4. Vasil'eva L. M. Modern problems of sturgeon breeding in Russia and the world. Technologies of the food and processing industry of the agro-industrial complex - healthy food products. 2015. № 2 (6). Pp. 30-36.

5. Abdelmanova A. S., Volkova V. V., Dotsev A. V., Zinovieva N. A. Characteristics of Genetic Diversity of Modern and Archival Populations of Black-and-White Cattle Using Microsatellite Markers. Achievements of science and technology of the agro-industrial complex. 2020. V. 34. № 2. Pp. 34-38.

6. Nikipelov V. I., Bardukov N. V., Kharzinova V. R., Grozescu Y. N., Zinovieva N. A. Characteristics of Microsatellite Loci and Their Polymorphism in Aquaculture Sterlet (Acipencer ruthenus). Genetics and animal breeding. 2023. No 5 (2).13 p.

7. Kharzinova V. R., Zinovieva N. A. Pattern of Genetic Diversity in Local and Commercial Pig Breeds Based on Microsatellite Analysis. Vavilov Journal of Genetics and Breeding. 2020. V. 24. № 7. Pp. 747-754.

8. Kharzinova V. R., Kudryavtsev A. V., Semerikova M. N., Zinovieva N. A. Study of the population structure and genetic diversity of the Chukchi reindeer breed based on the analysis of microsatellites. Achievements of science and technology of the agro-industrial complex. 2023. V. 37. № 9. Pp. 87-92.

9. https://www.syntol.ru/catalog/nabory-dlya-analiza-str-lokusov-zhivotnykh/nabor-reagentov-genekspertosyetr-dlya-geneticheskoy-pasportizatsii-i-opredeleniya-rodstva-osetrovykh.html.

10. Barmintseva A. E., Muge N. S. Use of Microsatellite Loci to Establish Sturgeon (Acipenseridae) Species and Identify Individuals of Hybrid Origin. Genetics. 2013. V. 49. № 9. Pp. 1093-1105.

11. Makarova E. G., Kozlova N. V. Genetic Monitoring of Sterlet Juveniles (Acipenser ruthenus) in the Volga River. Fish farming and fisheries. 2023. № 5.

12. Dudu A., Georgescu S., Burcea A., Florescu I., Costache M. Microsatellites Variation in Sterlet Sturgeon, Acipenser ruthenus from the Lower Danube Scientific Papers: Animal Science and Biotechnologies. 2013. V. 46. Pp. 90-96.

13. Pekarik L., et al. Current stocking program of the sterlet (Acipenser ruthenus, L.) can negatively shape its genetic variability in the Middle Danube. Zoology Lab, Plant Science and Biodiversity Center. 2019. V. 8. No 19. Pp. 2-8.

14. Welsh A., May B. Development and standardization of disomic microsatellite markers for lake sturgeon genetic studies. Journal of Applied Ichthyology. 2006. V. 22 (5). Pp. 337-344.

15. Yacheng Hu, et al. Development and characterization of novel cross–species tetranucleotide microsatellite markers for sterlet (Acipenser ruthenus) from Chinese sturgeon (Acipenser sinensis). Chinese Sturgeon Research Institute. Yichang, 2019. Pp. 310.

16. Peakall R., Ebert D., Cunningham R., Lindenmayer D. B. Mark–recapture by genetic tagging reveals restricted movements by bush rats, Rattus fuscipes, in a fragmented landscape. Journal of Zoology. 2006. No 268. Рp. 207-216.

17. Jamieson A. The effectiveness of using co-dominant polymorphic allelic series for (1) checking pedigrees and (2) distinguishing full-sib pair members. Animal Genetics. 1994. V. 25 (1). Pp. 37-44.

18. Jamieson A., Taylor S. C. S. Comparisons of three probability formulae for parentage exclusion. Animal Genetics. 1997. V. 28. Pp. 397-400.

19. Brown A. H. D., Weir B. S. Measuring genetic variability in plant populations. In: Isozymes in plant genetics and breeding, Part A. Elsevier Science Publisher, Amsterdam. 1983. Pp. 219-239.

20. Hartl D. L., Clark A. G. Principles of population genetics. Sinauer Associates. 1997. 542 p.

21. Peakall R., Smouse P. E. GenAlEx 6.5: genetic analysis in Excel. Population genetic software for teaching and research – an update. Bioinformatics. 2012. V. 28. Pp. 2537-2539.

22. Kohlmann K., Kersten P., Gessner J., Onara D., Taflan E., Radu S. New microsatellite multiplex PCR sets for genetic studies of the sterlet sturgeon, Acipenser ruthenus. Environmental Biotechnology. 2017. V. 13. Pp. 11-17.

23. Wang J., Sun Z., Jiang L., Hu Y. Developing microsatellite duplex PCR reactions for sterlet (Acipenser ruthenus) and their application in parentage identification. Scientific Reports. 2022. No. 12. P. 12036.

24. Maletsanake D., Nsoso S. J., Kgwatalala P. M. Genetic variation from 12 microsatellite makers in an indigenous Tswana goat flock in Southeastern Botswana. Livestock research for rural development. 2013. V. 25. P. 2.