Effect of Oil Overlay on Inhibition Potential of Roscovitine in Sheep Cumulus‐Oocyte Complexes

Inhibitors of cyclin‐dependent kinases, as roscovitine, have been used to prevent the spontaneous resumption of meiosis in vitro and to improve the oocyte developmental competence. In this study, the interference of oil overlay on the reversible arrest capacity of roscovitine in sheep oocytes as well as its effects on cumulus expansion was evaluated. For this, cumulus‐oocyte complexes (COCs) were cultured for 20 h in TCM 199 with 10% foetal bovine serum (Control) containing 75 μm roscovitine (Rosco). Subsequently, they were in vitro matured (IVM) for further 18 h in inhibitor‐free medium with LH and FSH. The culture was performed in Petri dishes under mineral oil (+) or in 96 well plates without oil overlay (?) at 38.5°C and 5% CO₂. At 20 and 38 h, the cumulus expansion and nuclear maturation were evaluated under stereomicroscope and by Hoechst 33342 staining, respectively. No group presented cumulus expansion at 20 h. After additional culture with gonadotrophins, a significant rate of COCs from both Control groups (+/?) exhibited total expansion while in both Rosco groups (+/?) the partial expansion prevailed. Among the oocytes treated with roscovitine, 65.2% were kept at GV in the absence of oil overlay while 40.6% of them reached MII under oil cover (p < 0.05). This meiotic arrest was reversible, and proper meiosis progression also occurred in the Control groups (+/?). So, the culture system without oil overlay improved the meiotic inhibition promoted by roscovitine without affecting the cumulus expansion rate or the subsequent meiosis progression.     

The influence of compost addition on heavy metal distribution between operationally defined geochemical fractions and on metal accumulation in plant

Purpose

Metal distribution patterns among geochemical fractions are informative for metal phytoavailability. Compost added to polluted soils may adsorb metals on the less phytoavailable fractions. A bioassay experiment was conducted to establish possible correlations between metal concentrations in different soil fractions and metal contents in edible plant parts and to investigate the influence of different compost loads on heavy metal availability to plants.

Materials and methods

Chinese cabbage plants were grown in pots with sandy and clayey soils and soils mixed with different doses of biosolid compost spiked with soluble heavy metal salts (Cd, Cu, and Pb). The metals’ distribution pattern in the soil and mixed samples was determined by sequential extraction procedure (modified BCR protocol). The studied fractions, from most to least bioavailable, were water-extractable (WE), exchangeable-adsorbed (EXC), associated with carbonates and acetic acid-soluble forms (CARB), occluded by reducible (hydro)oxides of Fe and Mn (RO), and associated with organic matter (OM) and a residual fraction (RES). Metal concentrations in soil extracts and in the digested plant tissue were measured by ICP-AES.

Results and discussion

The highest compost doses (72 and 115 Mg ha^?1) enhanced cabbage yield significantly. No excessive phytoaccumulation of metals was observed in plants grown in the clayey soil or its mixtures with compost. The compost dose of 72 Mg ha^?1 was optimal in decreasing Cu accumulation by plants grown in sandy soil, and 28.8 Mg ha^?1 was found to be effective in reducing Cd and Pb uptake. Metals were accumulated in plants primarily from the WE, EXC, and CARB fractions, whereas other fractions decreased phytoaccumulation. Compost addition suppressed heavy metal mobility, but different fractions were active in pollutant sorption, depending on soil type and metal.

Conclusions

Compost addition increased metal proportions in the RO and OM fractions, reducing metal phytoavailability. This is especially important for sandy soils with low adsorption ability and higher vulnerability to metal pollution than clayey soils. A compost dose of 20% v/v (or 28.8 Mg ha^?1) effectively reduced plant accumulation of Cd and Pb. We propose using the first three steps of the modified BCR protocol as a three-step sequential-extraction procedure for the most phytoavailable fractions of heavy metal: WE, EXC, and CARB.     

Potassium uptake characteristics of prunus rootstocks: Influence of solution Ca/Mg ratios and solution nickel

Abstract

Nutrient solution experiments were conducted in the growth chamber to study the influence of rootstock, solution Ca/Mg ratios and solution nickel on K uptake. The experimental plants were one‐year‐old prune trees: ‘French’ prune (Prunus domestica L.) scions grafted on Myrobalan 29C (P. cerasifera Ehrh.), Marianna 2624 (P. cerasifera x P. munsoniana?) or Nemaguard (P. persica x P. davidiana) rootstocks. Ion uptake parameters Imax, Km, and Cmin were calculated from ion depletion measurements over a 6 to 10‐hr period.

With K solution concentrations initially adjusted to 100 μM, K uptake rates of Prunus rootstocks were constant down to approximately 20–30 μM, then declined. Rootstocks were able to deplete solution K to concentrations less than 1 μM. There were no significant differences in K uptake parameters among the rootstocks tested.

Varying solution Ca/Mg ratio from 2.75/1 to 1/4 (Ca + Mg = 3.75 mM) had no effect on K uptake. Potassium uptake rates of Myrobalan 29C rootstocks in the presence of 100 μM nickel were not significantly different from those in the absence of nickel. Rates of nickel uptake were significantly lower than those of K. After eight days of pretreatment in solutions adjusted daily to 100 μM Ni(NO₃)₂, prune leaves began to show signs of interveinal chlorosis. Potassium uptake by nickel pretreated trees was not significantly different from that by control trees. Results are discussed in relation to field observations of K deficiency in prune orchards.     

The Effect of Calcium Sprays and Fruit Thinning on Bitter Pit Incidence and Calcium Content in ‘Honeycrisp’ Apple

‘Honeycrisp’ is a recently released cultivar of apple (Malus x domestica) developed by the University of Minnesota, now in high demand by growers and consumers. This cultivar is susceptible to bitter pit, especially when the trees are young and the fruit load is low. The effects of calcium (Ca) sprays, fruit thinning, vegetative growth, fruit and leaf nutrient concentration on bitter pit incidence, and two sampling methods for nutrient analysis were studied. Bitter pit incidence was associated with several variables, including Ca and phosphorus (P) concentrations in leaves and fruits, magnesium (Mg) concentration in leaves, and yield parameters. Fruit Ca concentration and content for the growing season were modeled, which allowed the determinations of Ca levels associated with bitter pit incidence. Our data suggest that plug analysis was less sensitive than fruit-segment analysis for finding differences in Ca concentrations.     

Plasma lipopolysaccharide elevations in cattle associated with early-stage infection by Fasciola hepatica

Fasciolosis is an endemic zoonotic parasitic disease with significant impacts on human health and both animal health and production. Early post-infection impacts on the host remain unclear. The objective of this study was to determine the changes, if any, to levels of endotoxin in cattle plasma in response to early-stage infection with Fasciola hepatica. Thirty-six (36) commercial bred cattle were experimentally infected with approximately 400 viable metacercariae. Plasma lipopolysaccharide (endotoxin) levels were examined on 24 occasions from 0 h before infection to 336 h after infection using the Limulus Amoebocyte Lysate chromogenic end point assay and compared with that of six (6) uninfected control animals. Peak lipopolysaccharide levels in infected animals were reached at 52 h after infection and returned to pre-infection levels at time 144 h after infection. Infected animals had significantly elevated lipopolysaccharide levels between 24 and 120 h after infection when compared to uninfected animals. The mean change in endotoxin units (EU)/mL over time after infection was statistically significant in infected animals. Elevations of lipopolysaccharide occurred in all infected animals suggesting a possible repeatable and titratable endotoxemia conducive to therapeutic agent model development.     

Segmental axonopathy of Merino sheep in New Zealand

AIM: To investigate an axonopathy of Merino sheep that caused progressive hindlimb ataxia and slight to moderate paresis, with the purpose of understanding its pathogenesis.

METHODS: Tissues were fixed in buffered paraformaldehyde or paraformaldehyde and glutaraldehyde, processed into wax and epoxy resin, respectively, and examined by light and electron microscopy. Fresh frozen spinal cord and trigeminal nerve roots were subjected to homogenisation, centrifugation and two-dimensional electrophoresis. Selected protein spots were identified using matrix-assisted laser desorption ionisation (MALDI) mass spectrometry.

RESULTS. By light microscopy, there were large pale foamy spheroidal axonal swellings affecting peripheral as well as central axons. By electron microscopy, these were shown to contain many membrane-bound vesicles. The main abnormalities in expressed proteins involved cytoskeletal elements and myosin heavy chain, the latter interpreted as associated with the molecular motor myosin Va.

CONCLUSIONS: The disorder is the same as that described in Merinos in Australia as segmental axonopathy, and believed to have an inherited aetiology. The lesions and protein changes indicate abnormalities of the cytoskeleton, its relationship with the myelin sheath, and myosin Va molecular motor. The consequence appears to be abnormal axonal transport and inability to maintain the integrity of axons and their myelin sheaths.     

Automated sample cleanup for pesticide multiresidue analysis. Part II--Design and evaluation of column chromatography module

An automated, continuous flow system is described for Florisil column chromatography of pesticide residues from food extracts. Evaluation of the system using 5 common organochlorine and organophosphorus pesticides in 2 crop matrices demonstrates essentially no difference in recovery or precision between automated and currently used manual analyses. The automated procedure uses only 20% of the solvents and adsorbents used in the manual procedure and is 3 times faster.