Evaluation of epididymis storage temperature and cryopreservation conditions for improved mitochondrial membrane potential,membrane integrity,sperm motility and in vitro fertilization in bovine epididymal sperm

The maintaining of the epididymis at lower temperatures during storage and transport improves sperm quality. Our study aimed to test whether epididymis storage temperature (post‐mortem) and sperm cryopreservation affect sperm kinetics, membrane integrity, mitochondrial potential and fertility capacity. Thirty‐six epididymides were collected from 18 bulls after slaughter and divided into two groups: at 4 or 34°C for 2–3 hr. The sperm was collected from the epididymis cauda. The evaluation consisted of computer‐assisted sperm analysis (CASA), SYBR14/PI/JC1 to evaluate membrane integrity, mitochondrial membrane potential (MMP) and measurement of lipid peroxidation (TBARS). The sperm was then frozen using an automatic device. After thawing, sperm samples were evaluated by the same variables and further in vitro fertilization rates. Cryopreservation negatively affected sperm motility in samples stored at 4 and 34°C. Nevertheless, the 4°C samples yielded higher rates of blastocyst formation. Pre‐freeze sperm motility, progressive motility and velocity were higher in sperm from epididymis stored at 4°C while post‐thaw sperm motility, progressive motility and velocity remained the same among samples from epididymis stored at 4 or 34°C. However, with regard to the kinetic patterns, samples collected from epididymis stored at 34°C had lower values when compared to those stored at 4°C prior the cryopreservation process. Our results indicate that epididymis handling conditions after cryopreservation may affect sperm quality after thawing, especially due to compromised MMP in sperm collected from epididymis stored at higher temperatures.     

Induction of fertilizable eggs by conspecific vitellogenin implantation in captive female walking catfish,Clarias batrachus (Linn.)

The synthesis of vitellogenin (Vg) is induced by conspecific Vg (Vg1 and Vg2) and estradiol‐17β (E₂) as demonstrated by the pattern of ³H‐serine incorporation in the liver and plasma proteins. The incorporation studies indicated that the label was first incorporated into the liver after which it appeared in the blood in both E₂‐ and Vg‐treated male catfish. Since Vg was capable of inducing its own synthesis, experiments were conducted in females during preparatory–prespawning period (March–May) to make them gravid by implanting Vg pellets. Two implantations of 4 mg Vg1 pellets into female catfish with an interval of 15 days, followed by laboratory maintenance for 45 days of initial implantation showed a significant increment in ovarian weight with concomitant formation of yolky oocytes through synthesis and incorporation of Vg, whereas Vg2 implantation was not effective in this regard. Histological observation of yolky oocytes in Vg1‐treated group showed the peripheral migration of germinal vesicle (eccentric germinal vesicle), which indicates the onset of maturation. On 45th day, third implantation with 2 mg Vg pellets was performed and after 15 days, fish were hormonally induced with a single injection of hCG (2,000 IU/kg fish). Six groups were considered such as initial control, BSA‐implanted control, Vg1‐implanted, Vg2‐implanted, catfish collected from the field on the last day of the experiment and catfish collected during spawning period in this experiment with 3–7 fish in each group. Each of the experimental fish was sexually mature and the body weight was between 100 and 125 g. The percentage of ovulation and fertilization in the eggs of Vg1‐implanted group was 91% and 78%, respectively, which was almost similar to that of gravid female catfish collected during breeding period (July). The breeding performance in BSA‐ and Vg2‐treated females was very poor. The fertilized eggs were hatched in the laboratory conditions. Thus, in the female catfish, Vg1 not only induces vitellogenesis but also makes the oocytes viable for fertilization.     

Air Contact Influences the pH of Extended Porcine Semen

In this study, the pH-rise during storage of extended porcine semen was examined. This pH-rise was found to be caused by CO(2)-loss from the buffering system in the extender and was more pronounced with increasing air volume in the recipient. An influence on sperm motility parameters was observed between semen samples stored in the presence of different amounts of ambient air in the recipient. Velocity parameters and percentage motile spermatozoa were significantly lower for semen stored in recipients with higher air volume and elevated pH. Adjusting extender preparation by avoiding air contact in commercial AI-centres may minimize the pH-rise and its influence on sperm motility.     

Replacement of by-catch fishmeal with dried chicken viscera meal in extruded feeds: effect on growth,nutrient utilisation and carcass composition of catfish <Emphasis Type="Italic">Clarias batrachus</Emphasis> (Linn.) fingerlings

An 84-day feeding trial was conducted to study the effect of replacing dietary fishmeal with dried chicken viscera meal (CVM) on the growth (net biomass gain, specific growth rate, SGR), feed acceptability, feed conversion ratio (FCR), protein efficiency ratio (PER) and carcass composition of Clarias batrachus fingerlings. Triplicate groups of fingerlings with mean initial body weight of 13.35 g were fed on six iso-nitrogenous and iso-lipidic diets. The control diet (CVM0) used marine by-catch fishmeal as the sole source of animal protein. In the other five diets (CVM100–CVM500), 20–100% of fishmeal was substituted by dried CVM at 20% increments. The highest body weight gain, SGR and PER, and the lowest FCR were observed in fish fed a diet containing 300–500 g CVM kg⁻¹. The fish accumulated increasing quantities of lipids and decreasing levels of ash in their carcasses with increasing levels of dietary CVM.     

Purification and development of ELISAs for two forms of vitellogenin in Indian walking catfish, <Emphasis Type="Italic">Clarias batrachus</Emphasis> (L.)

Two forms of vitellogenin (Vg: Vg1 and Vg2) were purified from the plasma of estradiol-17β (E₂)-treated Indian walking catfish, Clarias batrachus, by gel filtration and adsorption chromatography. Native Vg1 and Vg2 had apparent molecular masses of 375 and 450 kDa, respectively, and both Vgs resolved into two similar major bands (95 and 67 kDa) in SDS-PAGE under reducing condition. Polyclonal antisera raised against each form of Vg were absorbed with a combination of hypophysectomized male catfish serum proteins and alternate Vg to ensure specificity. Immunological analyses verified the presence of Vg1 and Vg2 in the plasma of female catfish. Homologous ELISAs were developed for Vg1 and Vg2 using their respective harvested antisera, which exhibited the detection limit of 100 ng ml^?1 for Vg1 and 40 ng ml^?1 for Vg2, and low level of cross-reactivity (not parallel to the standard) was found with alternate Vg in each assay. Treatment of male catfish with E₂ induced both Vgs showing a proportionate ratio of Vg1 to Vg2 at 5.6:1. Plasma concentrations of both Vgs measured by ELISAs at different reproductive phases of field collected female catfish increased in accordance with the ovarian development, keeping the proportionate ratio of Vg1 to Vg2 at about 2:1 in fish undergoing vitellogenesis during prespawning period and 1:20 during spawning period, suggesting that Vg1 may be the major Vg to contribute in yolk formation, whereas Vg2, besides its role in yolk formation, may facilitate other physiological functions. The present study, thus, demonstrates the occurrence of two unequally synthesized Vgs in the catfish.     

SPATIAL DISTRIBUTION AND ACCUMULATION OF CALCIUM IN DIFFERENT TISSUES,DEVELOPING SPIKES AND SEEDS OF FINGER MILLET GENOTYPES

□ Total calcium (Ca) content in different tissues, developing spikes (S1, S2, S3 and S4 stages), and distribution sites in seeds of five finger millet genotypes, which were differed in total calcium content, were analyzed. Atomic absorption spectroscopy (AAS) revealed stage-dependent quantitative changes in calcium accumulation from low to high during different stages of developing spikes and also in different tissues and grains. Results of scanning electron microscopic (SEM) energy dispersive X-ray (EDX) analysis showed differential distributions of the calcium in seed components viz. seed coat, aleurone layer and endosperm. Highest calcium content was observed in aleurone layer followed by seed coat while lowest calcium was found in endosperm of all genotypes. Major differences were found amongst genotypes with regard to the calcium distribution in seed components. The differential regulation of transport machinery might be responsible for differential calcium ion delivery and spatial distribution in the seed. A clear understanding of differential accumulation, spatial distribution and the variation of calcium within finger millet genotypes/seeds could be exploited for breeding for their bio-fortification and development of calcium rich functional foods.     

Relationship between apparent electrical conductivity and soil physical properties in a Malaysian paddy field

Site-specific crop management, well-established in some developed countries, is now being considered in developing countries such as Malaysia. The apparent electrical conductivity (EC_a) of the soil can be used as an indirect indicator of a number of soil physical properties and even crop yield. Commercially available EC_a sensors can efficiently develop the spatially dense data sets desirable in describing within-field spatial soil variability for precision farming. The main purpose of this study was to generate a variability map of soil EC_a within a Malaysian paddy field using a VerisEC sensor. The EC_a values were then compared with some soil variables within classes after delineation. Measured parameters were mapped using the kriging technique and their correlation with soil EC_a was determined. The study showed that the VerisEC can determine soil spatial variability, and can acquire soil EC_a information quickly. Spatial variability of shallow and deep EC_a showed the same patterns. Estimation of soil properties based on EC_a varied from one soil parameter to another and all could be estimated better by deep EC_a. Cross-validation results showed that shallow and deep EC_a, and also bulk density, gave more accurate estimates compared with other variables.