The present study was designed to evaluate sperm phenotypic variables during in vivo and in vitro storage following multiple sperm stripping in common carp (Cyprinus carpio L.). Each male was injected 3 times with carp pituitary 3 days apart. Sperm was stored in vivo in the body cavity for 0.5 days (Fresh sperm) and 3 days (Old sperm) after hormonal stimulation. Then sperm was collected and diluted with a carp extender at a ratio of 1:1, and stored in vitro on ice for 0, 3, and 6 days. The phenotypic parameters, including the number of total motile spermatozoa, number of fast motile spermatozoa, number of motile spermatozoa, percentage of fast motile spermatozoa, and percentage of spermatozoa motility were the major components of principal component analysis (PCA). In general, Fresh sperm from the first stripping showed slightly better quality than Old sperm from the second and third stripping, especially in the phenotypic parameters of a number of total spermatozoa and a number of total motile spermatozoa (P < 0.05). The highest kinetic and quantitative spermatozoa variables were obtained in Fresh and Old sperm just after sperm collection (0-day storage in vitro), and then they were decreased during the period of in vitro storage up to 6 days (P < 0.05). However, the fertilization, hatching, and malformation rates from Fresh sperm were similar compared with the Old sperm. Sperm could be stripped 0.5 days post hormonal treatment and stored in vitro up to 6 days with good fertilization performance (fertility, hatching, and malformation rates were 92.5%, 91.5%, and 1.3%, respectively). Therefore, our results suggested that multiple hormonal treatments with multiple stripping could be used in artificial reproduction in common carp.

• Klíčová slova
• Common carp, In vitro, In vivo, Sperm storage, Spermatozoa aging,
• MeSH
• kapři * MeSH
• kryoprezervace metody   MeSH
• led MeSH
• motilita spermií fyziologie   MeSH
• sperma fyziologie   MeSH
• spermie fyziologie   MeSH
• stárnutí MeSH
• uchování spermatu * metody   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• led MeSH

The purpose of the current study was to analyze phenotypic and functional characteristics of common carp (Cyprinus carpio) spermatozoa during in vitro aging and to investigate whether global DNA methylation is affected by sperm aging. Milt was collected from five individual males, stored in vitro on ice in a refrigerator for up to 96 h post stripping (HPS) and used to fertilize eggs with intervals of 1, 24 and 96 h. Computer-assisted sperm analysis and a S3e Cell Sorter was employed to determine the spermatozoa phenotypic characteristics (motility, velocity, concentration and viability). In addition, pH and osmolality of the seminal fluid and the capacity of the spermatozoa to fertilize, hatching rate and health of the resulting embryos were examined at different aging times. Whole-genome bisulfite sequencing was used to compare the global and gene-specific DNA methylation in fresh and aged spermatozoa. The results demonstrated that spermatozoa aging in common carp significantly affects their performance and thus the success of artificial fertilization. The methylation level at the cytosine-phosphate-guanine (CpG) sites increased significantly with 24 HPS spermatozoa compared to the fresh group at 1 HPS and then decreased significantly at 96 HPS. A more detailed investigation of gene specific differences in the DNA methylation was hindered by incomplete annotation of the C. carpio genome in the public databases.

• Klíčová slova
• DNA methylation, common carp, epigenetics, fertilization, fish, milt, sperm aging, sperm quality, sperm storage,
• MeSH
• kapři genetika   růst a vývoj   MeSH
• metylace DNA genetika   MeSH
• spermie metabolismus   patologie   MeSH
• stárnutí genetika   patologie   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

The lipid composition of sperm membranes is crucial for fertilization and differs among species. As the evolution of internal fertilization modes in fishes is not understood, a comparative study of the sperm lipid composition in freshwater representatives of externally and internally fertilizing fishes is needed for a better understanding of taxa-specific relationships between the lipid composition of the sperm membrane and the sperm physiology. The lipidomes of spermatozoa from stingray, a representative of cartilaginous fishes possessing internal fertilization, and sterlet, a representative of chondrostean fishes with external fertilization, have been studied by means of nuclear magnetic resonance (NMR), matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), electrospray MS, gas chromatography-(GC) MS, and thin-layer chromatography (TLC). NMR experiments revealed higher cholesterol content and the presence of phosphatidylserine in stingray compared to sterlet sperm. Unknown MS signals could be assigned to different glycosphingolipids in sterlet (neutral glycosphingolipid Gal-Cer(d18:1/16:0)) and stingray (acidic glycosphingolipid sulpho-Gal-Cer(d18:1/16:0)). Free fatty acids in sterlet sperm indicate internal energy storage. GC-MS experiments indicated a significant amount of adrenic acid, but only a low amount of docosahexaenoic acid in stingray sperm. In a nutshell, this study provides novel data on sperm lipid composition for freshwater stingray and sterlet possessing different modes of fertilization.

• Klíčová slova
• fertilization mode, freshwater fish, lipidomics, mass spectrometry, sperm, thin-layer chromatography,
• MeSH
• chromatografie na tenké vrstvě MeSH
• druhová specificita MeSH
• fertilizace fyziologie   MeSH
• glykosfingolipidy chemie   MeSH
• hmotnostní spektrometrie s elektrosprejovou ionizací MeSH
• kyseliny dokosahexaenové chemie   MeSH
• lipidomika MeSH
• lipidy chemie   MeSH
• magnetická rezonanční spektroskopie MeSH
• plynová chromatografie s hmotnostně spektrometrickou detekcí MeSH
• ryby fyziologie   MeSH
• spektrometrie hmotnostní - ionizace laserem za účasti matrice MeSH
• spermie chemie   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• glykosfingolipidy MeSH
• kyseliny dokosahexaenové MeSH
• lipidy MeSH