Rainbow trout (Oncorhynchus mykiss Walbaum) and sea trout (Salmo trutta Linnaeus, 1758) show large karyotypic differences and their hybrid offspring is not viable due to unstable karyotype and chromosome fragmentation. However, gametes from these two species were used to induce gynogenetic development. Rainbow trout eggs activated by UV-irradiated sea trout sperm were subjected to high hydrostatic pressure (HHP) shock to prevent release of the 2nd polar body (early shock) or to inhibit the first cleavage (late shock) in order to produce diploid meiotic gynogenotes and gynogenetic doubled haploids (DHs), respectively. Cytogenetic analysis proved fish that development was induced by the sea trout spermatozoa were rainbow trout. In turn, molecular examination confirmed homozygosity of the gynogenetic DHs. Presumed appearance of the recessive alleles resulted in lower survival of the gynogenetic DH larvae (~25%) when compared to survival of the heterozygous (meiotic) gynogenotes (c. 50%). Our results proved that genomic incompatibilities between studied trout species result in the hybrid unviability. However, artificial gynogenesis including activation of rainbow trout eggs with UV-irradiated sea trout spermatozoa was successfully induced. As both species are unable to cross, application of the UV-irradiated sea trout spermatozoa to activate rainbow trout development assures only maternal inheritance with no contamination by the residues of the paternal chromosomes.

Poor quality eggs produced by the fully homozygous doubled haploids (DHs) may impair generation of clonal lines in fish species. In the present research, gynogenetic development of rainbow trout (Oncorhynchus mykiss) was induced in eggs originated from the DH females. Eggs were activated with the UV-irradiated grayling (Thymallus thymallus) spermatozoa and subjected to the high hydrostatic pressure (HHP) shock to provide diploid clonal individuals. Only two of four DH females produced eggs that were successfully activated by the irradiated spermatozoa and subsequently developed into the gynogenetic embryos. Survival rates of rainbow trout from the clonal lines equalled 21.5% and 19.8% during embryogenesis and decreased after hatching to 18.6% and 14.9%, respectively. Some of the dead rainbow trout clones collected between hatching and swim-up stage were emaciated and exhibited spinal deformities including scoliosis. Provided results confirmed limited developmental competences of eggs produced by rainbow trout DH females. Clonal rainbow trout developing in such eggs exhibited reduced survival and increased frequency of the body abnormalities.

In the present research, the eggs from four rainbow trout females were used to provide four groups of gynogenetic doubled haploids (DHs). The quality of the eggs from different clutches was comparable, however, interclutch differences were observed in the gynogenetic variants of the experiment and the survival of DH specimens from different groups varied from 3% to 57% during embryogenesis. Transcriptome analysis of the eggs from different females exhibited inter-individual differences in the maternal genes’ expression. Eggs originating from females whose gynogenetic offspring had the highest survival showed an increased expression of 46 genes when compared to the eggs from three other females. Eggs with the highest survival of gynogenetic embryos showed an up-regulation of genes that are associated with cell survival, migration and differentiation (tyrosine-protein kinase receptor TYRO3-like gene), triglyceride metabolism (carnitine O-palmitoyltransferase 1 gene), biosynthesis of polyunsaturated fat (3-oxoacyl-acyl-carrier-protein reductase gene), early embryogenic development (protein argonaute-3 gene, leucine-rich repeat-containing protein 3-like gene), 5S RNA binding (ribosome biogenesis regulatory protein homolog) as well as senescence and aging (telomerase reverse transcriptase, TERT gene), among others. Positive correlation between the genotypic efficiency and egg transcriptome profiles indicated that at least some of the differentially expressed genes should be considered as potential candidate genes for the efficiency of gynogenesis in rainbow trout.

ContentsIonizing radiation (IR) is applied to inactivate nuclear genome in the salmonid eggs to induce androgenetic development. However, it has been considered that doses of IR used to damage maternal chromosomes may also affect morphology of the eggs and decrease their developmental potential. Thus, the main goal of the present research was to assess alterations in the rainbow trout (Oncorhynchus mykiss) eggs caused by the high dose of IR administered during androgenesis. In the present research, rainbow trout eggs were irradiated with 350 Gy of X-rays, inseminated and exposed to the high hydrostatic pressure (HHP) shock to develop as androgenetic doubled haploids (DHs). The distribution of lipid droplets in the irradiated and non-irradiated rainbow trout eggs, survival rates and morphology of larvae from androgenetic and control groups were compared. It has been observed that non-irradiated and irradiated eggs exhibited altered distribution of lipid droplets. Most of the eggs before IR treatment displayed rather equal distribution of the oil droplets. In turn, majority of eggs studied after irradiation had coalesced lipid droplets, a pattern found in eggs with reduced quality. Incidences of abnormally developed larvae were more frequently observed among fish that hatched from the irradiated eggs. Observed changes suggest X-rays applied for the genetic inactivation of rainbow trout eggs may lead to decrease of their developmental competence. K E Y W O R D Sandrogenesis, body malformations, egg quality, lipid droplets, reactive oxygen species | 713 POLONIS et aL. SUPPORTING INFORMATIONAdditional supporting information may be found online in the Supporting Information section at the end of the article.How to cite this article: Polonis M, Jagiełło K, Dobosz S, et al.Alterations in the rainbow trout (Oncorhynchus mykiss) eggs exposed to ionizing radiation during induced androgenesis.

Ionizing radiation (IR) is applied to inactivate the nuclear genome in rainbow trout eggs during induced androgenetic development. However, IR-generated reactive oxygen species (ROS) may affect developmental potential of eggs and reduce the effectiveness of androgenesis. To verify this assumption, androgenetic development of rainbow trout was induced in eggs irradiated with 350 Gy of X-rays. Survival rates, pH of the ovarian fluid and activity of antioxidant enzymes, including SOD, CAT and GPx, were examined in non-irradiated and irradiated eggs originating from four females. Survival rates of androgenetic embryos developing in eggs produced by different females varied from 1% to 57% and these inter-clutch differences were significant. Eggs from female F4, which showed the highest developmental competence for androgenesis, also showed increased activities of SOD, CAT and GPx enzymes. The pH value of the ovarian fluid of each female was over 8 before and after irradiation, therefore it seems that radiation did not affect the ovarian fluid pH. Considering the above-mentioned inter-clutch differences, a strong maternal effect on the effectiveness of androgenesis can be assumed. Eggs with increased activity of antioxidant enzymes before irradiation should be expected to show increased developmental competence for androgenesis.