Effect of Point Mutation in the Growth Differentiation Factor 9 Gene of Oocytes on the Sterility and Fertility of Awassi Sheep

Document Type: Original Articles

Authors

1 College of Veterinary Medicine, University of Baghdad, Bagdad, Iraq

2 College of Veterinary Medicine, University of Tikrit, Tikrit, Iraq

Abstract

Growth differentiation factor 9 (GDF9) plays a critical role in ovarian follicular development and ovulation rate. The present study aimed to investigate the correlation between the single-nucleotide polymorphism (SNP) of the GDF9 gene and reproductive performance variables, such as fertility and sterility in Awassi sheep. Forty pairs of ovaries from a total of 40 slaughtered Iraqi Awassi ewes were used in this study. Twenty of the ovaries were collected from sterile ewes and the other 20 ovaries were taken from fertile ewes for genomic DNA extraction, polymerase chain reaction, and sequencing to detect GDF9 gene polymorphism. Follicles and oocytes of all the 40 ovaries were evaluated and compared with the results of genotyping. Furthermore, histopathological and microscopic evaluations were performed for 40 ovarian tissues of the two groups. The sequence analysis revealed that exon I had three SNPs, including T(114)C, G(129)R, and G(199)A. The first two SNPs were silent mutations and the last mutation was missense responsible for the substitution of glutamic acid with lysine at position 67. The current study showed a significant increase (P≤0.01) in GG, AA, CC, GA, and GG genotypes at G(129)R,  G(199)A, T(114)C, G(129)R, and  G(199)A loci, respectively. Moreover, the TT genotype in locus T(114)C was recorded to significantly augment (P≤0.05) in the fertile ewes. Mutant GA genotype of the G(129)R locus led to a significant (P≤0.05) increase in the percentage of follicles (4-8 mm) and oocytes number, compared to wild GG. On the other hand, a significant decrease was recorded in the mutant AA genotype in G(199)A, compared to wild GG. Differences between CC and TT genotypes at T(114)C locus were not significant. Histopathological examination revealed hypoplasia in the ovarian tissue of sterile ewes accompanied by fibrous connective tissue invasion and follicles degeneration. However, in the fertile ewes, the ovarian tissues were normal with the presence of numerous corpus albicans and degenerative corpus luteum. According to the findings of this study, the homozygote mutation in fertile ewes minimized the number of follicles and oocytes leading to sterility, while the heterozygote mutation was reported in the fertile Awassi ewes.

Keywords


Article Title [French]

Effet de la Mutation Ponctuelle dans le Gène du Facteur de Différenciation de Croissance 9 des Ovocytes sur la Stérilité et la Fertilité des Moutons Awassi

Abstract [French]

: Le facteur de différenciation de croissance 9 (GDF9) joue un rôle essentiel dans le développement folliculaire ovarien et le taux d'ovulation. Cette étude visait à étudier la corrélation entre le polymorphisme mononucléotidique (SNP) du gène GDF9 et les variables de performance de reproduction, telles que la fertilité et la stérilité chez les moutons Awassi. A cet effet, 40 paires d'ovaires ont été prélevées à partir de 40 brebis Awassi irakiennes abattues. Au total, 20 ovaires ont été prélevés sur des brebis stériles et les 20 autres sur des brebis fertiles. Les échantillons prélevés ont été ensuite soumis à une extraction d'ADN génomique suivie d’une réaction en chaîne par polymérase et d’un séquençage pour détecter le polymorphisme du gène GDF9.Les follicules et les ovocytes des 40 ovaires ont été ensuite évalués et comparés aux résultats du génotypage. De plus, des évaluations histopathologiques et microscopiques ont été réalisées pour les 40 tissus ovariens des deux groupes. L'analyse des séquences a révélé que l'exon I avait trois SNPs, en l’occurence T (114) C, G (129) R et G (199) A. Les deux premiers SNP étaient des mutations silencieuses et la dernière était une mutation faux-sens, responsable de la substitution de l'acide glutamique par la lysine en position 67. L'étude actuelle a montré une augmentation significative (P≤0.01) des génotypes GG, AA, CC, GA et GG à G (129) R, G (199) A, T (114) C, G (129) R, et G (199) A loci, respectivement. De plus, le génotype TT dans le locus T (114) C, était significativement augmenté (P≤0.05) chez les brebis fertiles. Le génotype mutant GA du locus G (129) R a conduit à une augmentation significative (P≤0.05) du pourcentage de follicules (4-8 mm) et du nombre d'ovocytes, par rapport au GG sauvage. En revanche, une diminution significative a été enregistrée dans le génotype AA mutant dans G (199) A, comparé au GG sauvage. Il n'y avait aucune différence significative entre les génotypes CC et TT au locus T (114) C. L'examen histopathologique a révélé une hypoplasie dans le tissu ovarien des brebis stériles accompagnée d'une invasion du tissu conjonctif fibreux et d'une dégénérescence des follicules. Cependant, les tissus ovariens étaient normaux avec la présence de nombreux corps albicans et de corps jaune dégénératif chez les brebis fertile. Selon les résultats de cette étude, la mutation homozygote chez les brebis fertiles a diminué le nombre de follicules et d'ovocytes qui ont conduit à la stérilité, tandis que la mutation hétérozygote a été rapportée chez les brebis fertiles Awassi.

Keywords [French]

  • Fertilité
  • Hétérozygote
  • Homozygote
  • Infertilité
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