Seminal plasma proteins of Atlantic halibut (Hippoglossus hippoglossus L.).

Protein content and properties in the seminal plasma of Atlantic halibut (Hippoglossus hippoglossus) were assayed using spectrophotometric and electrophoretic methods. The protein concentration ranged from 6.4 +/- 3.1 to 19.4 +/- 3.4 mg ml(-1) and anti-proteolytic activity from 585.2 +/- 104.6 to 2912.4 +/- 367.4 U l(-l). A high correlation between anti-proteolytic activity and protein concentration (r = 0.95), and between sperm concentration and osmolality was found (r = 0.92). There was a significant decrease in anti-proteolytic activity from the first to the second sampling, but not in protein concentration. Anti-proteolytic activity and protein concentration were significantly affected by variations in individual males. Electrophoresis revealed four anti-proteolytic bands and individual differences in bands of proteolytic activity, which were subsequently characterized as metalloproteases and serine proteases.

Acrosome staining and motility characteristics of sterlet spermatozoa after cryopreservation with use of methanol and DMSO.

In this study we describe acrosome staining and motility characteristics of fresh and cryopreserved sterlet (Acipenser ruthenus L.) spermatozoa using soybean trypsin inhibitor-Alexa conjugate fluorescent staining and computer-aided sperm analysis (CASA), respectively. Methanol or dimethylsulfoxide (DMSO) were used as cryoprotectants. After cryopreservation a decline in sperm motility characteristics occurred, but no differential effect between cryoprotectant was observed. Cryopreservation caused a significant increase in the percentage of spermatozoa with acrosome stained by SBTI-Alexa for samples cryopreserved using DMSO compared to methanol. These data suggest that the low usefulness of DMSO for cryopreservation of sturgeon spermatozoa is related to its harmful specific effect towards the acrosome, probably by causing its precocious triggering, much before any egg contact.

Isolation, characterization and cDNA sequencing of a Kazal family proteinase inhibitor from seminal plasma of turkey (Meleagris gallopavo).

The turkey reproductive tract and seminal plasma contain a serine proteinase inhibitor that seems to be unique for the reproductive tract. Our experimental objective was to isolate, characterize and cDNA sequence the Kazal family proteinase inhibitor from turkey seminal plasma and testis. Seminal plasma contains two forms of a Kazal family inhibitor: virgin (Ia) represented by an inhibitor of moderate electrophoretic migration rate (present also in the testis) and modified (Ib, a split peptide bond) represented by an inhibitor with a fast migration rate. The inhibitor from the seminal plasma was purified by affinity, ion-exchange and reverse phase chromatography. The testis inhibitor was purified by affinity and ion-exchange chromatography. N-terminal Edman sequencing of the two seminal plasma inhibitors and testis inhibitor were identical. This sequence was used to construct primers and obtain a cDNA sequence from the testis. Analysis of a cDNA sequence indicated that turkey proteinase inhibitor belongs to Kazal family inhibitors (pancreatic secretory trypsin inhibitors, mammalian acrosin inhibitors) and caltrin. The turkey seminal plasma Kazal inhibitor belongs to low molecular mass inhibitors and is characterized by a high value of the equilibrium association constant for inhibitor/trypsin complexes.

Effects of different surfactants on motility and DNA integrity of brown trout (Salmo trutta fario) and common carp (Cyprinus carpio) spermatozoa.

In this study we evaluated effects of surfactants on motility parameters and DNA integrity of spermatozoa of freshwater teleost fish. Common carp (Cyprinus carpio) and brown trout (Salmo trutta fario) spermatozoa were exposed to either sodium dodecyl sulphate (SDS, anionic surfactant) or octoxynol 9 ( Triton X-100, nonionic surfactant). Both surfactants added at activation caused a decrease in sperm motility characteristics measured by computer-assisted sperm analysis (CASA). Intraspecific differences in speed and trajectory of movement were detected. Triton X-100 and SDS when added to non activated sperm were also effective in the decrease of sperm motility and caused an increase of DNA fragmentation. Our results suggest that not only sperm motility apparatus but also DNA are targets for surfactant action. Therefore any exposure of spermatozoa to surfactants, in aquaculture conditions or natural environment, would have a negative impact on fish reproduction.

Polymorphism of transferrin of carp seminal plasma: relationship to blood transferrin and sperm motility characteristics.

Transferrin (Tf) is a major protein of carp (Cyprinus carpio) seminal plasma. Its relationship with milt quality is unknown. In this study, we sought to determine if Tf is polymorphic in carp seminal plasma and if this polymorphism is related to sperm motility characteristics. We screened males of purebred common carp line (Polish line R6) for Tf polymorphism in blood plasma. The majority of Tf genotypes represented only DD and DG variants. We then collected milt from preselected DD and DG genotypes and tested their sperm motility characteristics using computer-aided sperm analysis (CASA). Tf polymorphism in seminal plasma was found to be identical with that of blood. However, the relationships between Tf polymorphism and iron metabolic parameters were different for blood and semen. These data suggest different regulation of Tf in liver and testis. We found substantial differences in sperm motility characteristics between both genotypes. Spermatozoa of DG males were characterized by lower curvilinear velocity (VCL), amplitude of lateral head displacement (ALH), higher linearity (LIN) and straightness (STR) of movement as compared to DD males. No differences were found in other sperm characteristics such as sperm concentration and percentage of sperm motility. Our results suggest that sperm motility parameters are related to Tf polymorphism and therefore this polymorphism may be related to sperm competitive ability.

Application of biochemical markers for identification of biological properties of animal semen.

The use of biochemical markers for identification of biological properties of semen will help to develop new criteria that are accurate and objective in predicting and improving male fertility. Understanding and controlling the mechanisms involved in fertility is a key challenge, which is of fundamental importance in successful animal reproductive performance. Moreover, unraveling the unique molecular mechanism associated with sperm function might have considerable diagnostic value in the evaluation of male infertility. This review offered insights into some recent achievements and provided perspectives for possible applications of the biochemical markers of semen.

Improvement of electrophoretic detection of antitrypsin activity in fish blood and seminal plasma.

The original method of Uriel and Berges for detection of trypsin inhibitors lacks specificity due to masking effects of nonspecific esterases. We report a modification of this method based on inhibition of esterases in samples by phenylmethylsulfonyl fluoride (PMSF). This method can be particularly useful for characterization profiles of antitrypsin activity in seminal plasma of salmonid fish where esterases and inhibitors migrate at the same mobility.