Phosphofructokinase and Malate Dehydrogenase Participate in the In Vitro Maturation of Porcine Oocytes

Oocyte maturation depends on the metabolic activity of cumulus–oocyte complex (COC) that performs nutritive and regulatory functions during this process. In this work, the enzymes [phosphofructokinase (PFK) and malate dehydrogenase (MDH)] were tested to elucidate the metabolic profile of porcine COCs during the in vitro maturation (IVM). Enzymatic activity was expressed in U/COC and U/mg protein (specific activity) as mean ± SEM. In vitro maturation was performed with 2‐oxoglutarate (5, 10 and 20 mm ) or hydroxymalonate (30, 60 and 100 mm ) inhibitors of PFK and MDH, respectively. The PFK and MDH activities (U) remained constant during maturation. For PFK, the U were (2.48 ± 0.23) 10^?5 and (2.54 ± 0.32) 10^?5, and for MDH, the U were (4.72 ± 0.42) 10^?5 and (4.38 ± 0.25) 10^?5 for immature and in vitro matured COCs, respectively. The specific activities were significantly lower after IVM, for PFK (4.29 ± 0.48) 10^?3 and (0.94 ± 0.12) 10^?3, and for MDH (9.08 ± 0.93) 10^?3 and (1.89 ± 0.10) 10^?3 for immature and in vitro matured COCs, respectively. In vitro maturation percentages and enzymatic activity diminished with 20 mm 2‐oxoglutarate or 60 mm hydroxymalonate (p < 0.05). Viability was not affected by any concentration of the inhibitors evaluated. The U remained unchanged during IVM; however, the increase in the total protein content per COC provoked a decrease in the specific activity of both enzymes. Phosphofructokinase and MDH necessary for oocyte IVM would be already present in the immature oocyte. The presence of inhibitors of these enzymes impairs the meiotic maturation. Therefore, the participation of these enzymes in the energy metabolism of the porcine oocyte during IVM is confirmed in this study.     

Cattle selected for lower residual feed intake have reduced daily methane production

Seventy-six Angus steers chosen from breeding lines divergently selected for residual feed intake (RFI) were studied to quantify the relationship between RFI and the daily rate of methane production (MPR). A 70-d feeding test using a barley-based ration was conducted in which the voluntary DMI, feeding characteristics, and BW of steers were monitored. The estimated breeding value (EBV) for RFI (RFI(EBV)) for each steer had been calculated from 70-d RFI tests conducted on its parents. Methane production rate (g/d) was measured on each steer using SF(6) as a tracer gas in a series of 10-d measurement periods. Daily DMI of steers was lower during the methane measurement period than when methane was not being measured (11.18 vs. 11.88 kg; P = 0.001). A significant relationship existed between MPR and RFI when RFI (RFI(15d)) was estimated over the 15 d when steers were harnessed for methane collection (MPR = 13.3 x RFI(15d) + 179; r(2) = 0.12; P = 0.01). Animals expressing lower RFI had lower daily MPR. The relationship established between MPR and RFI(15d) was used to calculate a reduction in daily methane emission of 13.38 g accompanied a 1 kg/d reduction in RFI(EBV) in cattle consuming ad libitum a diet of 12.1 MJ of ME/kg. The magnitude of this emission reduction was between that predicted on the basis of intake reduction alone (18 g x d(-1) x kg of DMI(-1)) and that predicted by a model incorporating steer midtest BW and level of intake relative to maintenance (5 g x d(-1) x kg of DMI(-1)). Comparison of data from steers exhibiting the greatest (n = 10) and lowest (n = 10) RFI(15d) showed the low RFI(15d) group to not only have lower MPR (P = 0.017) but also reduced methane cost of growth (by 41.2 g of CH(4)/kg of ADG; P = 0.09). Although the opportunity to abate livestock MPR by selection against RFI seems great, RFI explained only a small proportion of the observed variation in MPR. A genotype x nutrition interaction can be anticipated, and the MPR:RFI(EBV) relationship will need to be defined over a range of diet types to account for this.     

Infectivity and virulence of Australian strains of Moraxella bovis for the murine and bovine eye in relation to pilus serogroup sub-unit size and degree of piliation

The degree of piliation of 29 haemolytic and 4 non-haemolytic Australian strains of Moraxella bovis representing 7 different pilus antigen groups was determined. The infectivity and virulence for the eye was measured in steroid-treated mice and in cattle. Non-piliated strains failed to infect the murine eye. Most moderately or heavily piliated strains reproducibly produced the highest infectivity and virulence scores in mice when compared with lightly or very lightly piliated strains (p less than 0.05). Non-haemolytic, piliated strains were infective and in one instance virulent for mice. Almost similar levels of infectivity and virulence were observed for 7 representative haemolytic strains tested in both cattle and mice. The relative molecular weight of pilin sub-units was compared using sodium dodecyl-sulphate polyacrylamide gel electrophoresis. Three classes of pili, alpha, beta and gamma of ascending sub-unit size were identified among the 7 pilus antigen serogroups. Pilin sub-unit size bore no relationship to the degree of piliation but most strains that were highly virulent in mice and cattle expressed alpha and gamma sub-units. Some strains appeared capable of switching from alpha to beta or form beta to gamma sub-unit production.     

Heat Shock Protein 70 and Sex Steroid Receptors in the Follicular Structures of Induced Ovarian Cysts

The purpose of this study was to estimate the expression and relative amounts of estrogen (ER) and progesterone receptors (PR) and their isoforms as well as heat shock protein 70 (HSP70) in ovaries of rats with induced cystic ovarian disease (COD). Primary, secondary, tertiary, atretic and cystic follicles were evaluated by immunohistochemistry and total ovarian proteins were analyzed by Western blot. In the granulosa layer, growing and cystic follicles in the treated group have a higher expression of ERα than growing follicles of control individuals. In the theca interna layer, tertiary follicles presented a significantly higher expression of ERα in the treated group. An increase in total ERα protein was detected in the treated group. Granulosa cells of all growing, atretic and cystic follicles show a lower expression of ERβ in animals with COD, and the total protein expression of ERβ was lower in this group. The expression of PR was lower in the granulosa cell layer of tertiary and cystic follicles in treated animals, and theca interna layer had less intense immunostaining in this group. Although there were no differences in the expression of PR-B by Western blotting, the expression of PR-A was higher and the expression of PR-C was smaller in the treated group. An intense HSP70 immunostaining was observed in the cells of cystic follicles. By Western blotting, higher protein expression of HSP70 was detected in the ovarian samples of the control group than those of the treated ones. Ovaries of animals with COD exhibited an altered steroid receptor expression and subtype balance as compared with control animals, and an increase in HSP70 immunoexpression.     

Leaching of dissolved organic carbon and nitrogen under cotton farming systems in a Vertisol

Leaching with deep drainage is one of the loss pathways of carbon (C) and nitrogen (N) in cropping fields. However, field studies in irrigated row cropping systems are sparse. A 3‐year investigation on C and N leaching associated with deep drainage was overlaid on a long‐term experiment on tillage practices and crop rotations in Australia. The treatments included cotton (Gossypium hirsutum L.) monoculture and cotton–wheat (Triticum aestivum L.) or maize (Zea maize L.) rotations with maximum or minimum tillage. The deep drainage C and N concentrations at 0.6 and 1.2 m depth were measured after furrow irrigation with ceramic cup samplers during the 2014–15, 2015–16 and 2016–17 cotton seasons. Pre‐planting dissolved organic carbon (DOC) concentration in soil at 0.6–1.2 m depth during 2016–17 was 64 mg kg^?1 for maximum tilled cotton monoculture, 36 mg kg^?1 for minimum tilled cotton monoculture and 39 mg kg^?1 for cotton–wheat, and in maize and cotton subplots 51 and 41 mg kg^?1, respectively. Post‐harvest DOC values in soil were similar in all treatments (average of 32 mg DOC kg^?1). Total organic carbon (TOC) losses in deep drainage were equal to 2%–30% of TOC gained in irrigation water. Oxidized N losses in deep drainage ranged from 0.7% to 12% of applied N (260 kg ha^?1). NOx‐N concentrations in leachate under maize systems (20 mg L^?1) were up to 73% lower than those in cotton systems (75 mg L^?1). Maize sown in rotation with cotton can improve cotton yield, reduce N leaching and improve N use efficiency of subsequent cotton.     

Testing Usability of Butylated Hydroxytoluene in Conservation of Goat Semen

The objective of this study was to investigate whether butylated hydroxytoluene (BHT) could be used as a suitable supporter or alternative of egg yolk during preservation of goat spermatozoa. Three in vitro experiments and a fertility test were conducted to evaluate the effect of BHT on viability of chilled‐stored semen as well as motility and kidding rate of frozen‐thawed spermatozoa. In the first two experiments, ejaculates (n = 30/experiment) were collected from 10 bucks, split, diluted with egg yolk‐based and egg yolk‐free extenders supplemented with or without 0.3, 0.6, 2, 5 and 8 mm BHT and stored at 5°C for 168 h. In the third experiment, 30 ejaculates were collected from the above‐mentioned bucks, split and diluted with egg yolk‐free extenders supplemented with or without 0.3, 0.6 and 0.9 mm BHT and egg yolk‐based extenders supplemented with or without 5 mm BHT. Diluted semen was cooled to 5°C over a period of 4 h, frozen and thawed in the form of 0.3‐ml pellets. In the fertility test, 75 ejaculates were collected from two proven fertile bucks, split, diluted with egg yolk‐free extenders containing 0.6 mm BHT and egg yolk‐based extenders supplemented with or without 5 mm BHT, frozen and thawed as described above. An insemination volume of 0.6 ml containing 120–140 × 10⁶ progressively motile spermatozoa was used for a single cervical insemination of cloprostenol‐synchronized does (n = 230). The results showed that addition of 5 mm BHT to egg yolk‐deficient (2.5%) extenders significantly improved viability of chilled‐stored semen together with motility (48.5%) and fertility (62.5%) of frozen‐thawed spermatozoa. Replacement of egg yolk in semen extenders by 0.6 mm BHT could sustain not only viability of chilled‐stored semen but also post‐thaw motility (47.5%) and fertility (53.75%) of frozen‐thawed spermatozoa. In conclusion, supplementation of semen diluents with BHT can ameliorate preservability of goat sperm.     

A "Schrodinger Cat" Superposition State of an Atom

A "Schrodinger cat"-like state of matter was generated at the single atom level. A trapped 9Be+ ion was laser-cooled to the zero-point energy and then prepared in a superposition of spatially separated coherent harmonic oscillator states. This state was created by application of a sequence of laser pulses, which entangles internal (electronic) and external (motional) states of the ion. The Schrodinger cat superposition was verified by detection of the quantum mechanical interference between the localized wave packets. This mesoscopic system may provide insight into the fuzzy boundary between the classical and quantum worlds by allowing controlled studies of quantum measurement and quantum decoherence.     

Soil properties in furrows of an irrigated Vertisol sown with continuous cotton (Gossypium hirsutum L.)

Average in-field water application efficiency in furrow-irrigated cotton (Gossypium hirsutum L.) in Australia is less than optimal, and The underlying reasons may include surface sealing, exposure of sodic soil by laser levelling, and soil compaction due to wheel-trafficking. The objective of this study was to quantify the effects of reducing traffic and tillage intensity on furrow soil properties in an irrigated Vertisol. Soil was sampled during the growing seasons of 2001–02, 2003–04 and 2005–06 from the surface 50-mm of adjacent wheel-tracked and non-wheel-tracked furrows in an experiment in north-western New South Wales, Australia. The treatments were: cotton sown either after conventional tillage (disc-ploughing and incorporating cotton stalks to 0.2 m, chisel ploughing to 0.3 m followed by bed construction) or on “permanent beds” (slashing cotton plants after harvest, followed by root cutting and bed renovation with a disc-hiller). Irrigation water was alkaline but had low salinity and sodium adsorption ratio. Soil properties measured were pH, EC_1:5, geometric mean diameter (GMD) and specific volume (SV) of dry soil aggregates, exchangeable cations and plastic limit. Permanent bed systems had generally lower pH and higher SOC than conventionally tilled furrows, although differences were small. Soil pH and SOC averaged over the three growing seasons was 6.9 and 0.89 g/100 g, respectively, in permanent bed furrows, and 7.1 and 0.84 g/100 g, respectively, in conventionally tilled furrows. Compared to non-wheel-tracked furrows, plastic limit was lower (0.24 vs. 0.25 g/g), and EC_1:5 (0.24 vs. 0.20 dS/m) and GMD (2.6 vs. 2.1 mm) higher in wheel-tracked furrows. Intra-seasonal changes in soil properties of furrows were also small, and are unlikely to significantly affect any hydrological processes. Inter-seasonal differences were, however, significant, and could affect hydrological processes in this soil.     

NFAT binding and regulation of T cell activation by the cytoplasmic scaffolding Homer proteins

T cell receptor (TCR) and costimulatory receptor (CD28) signals cooperate in activating T cells, although understanding of how these pathways are themselves regulated is incomplete. We found that Homer2 and Homer3, members of the Homer family of cytoplasmic scaffolding proteins, are negative regulators of T cell activation. This is achieved through binding of nuclear factor of activated T cells (NFAT) and by competing with calcineurin. Homer-NFAT binding was also antagonized by active serine-threonine kinase AKT, thereby enhancing TCR signaling via calcineurin-dependent dephosphorylation of NFAT. This corresponded with changes in cytokine expression and an increase in effector-memory T cell populations in Homer-deficient mice, which also developed autoimmune-like pathology. These results demonstrate a further means by which costimulatory signals are regulated to control self-reactivity.