Enzyme Immunoassays for the Determination of Ovine LH and FSH

The development of competitive enzyme immunoassays for ovine plasma LH (oLH) and FSH (oFSH) is described. Standards and plasma samples were preincubated with diluted antiserum to oLH or oFSH and the reacted solution (100 μl per well) was transferred to plates previously coated with oLH or oFSH, respectively. The second antibody used was anti‐rabbit IgG horseradish peroxidase. The measuring range was 0.39–50 ng/ml for each hormone and the 50% relative binding sensitivity was 9 ng/ml for oLH. The respective value for oFSH was 3.5 or 34 ng/ml with different hormone and antibody preparations used for the assay. The enzyme immunoassays were used to determine oLH and oFSH levels in plasma from ewes of two breeds during the oestrous cycle. The assays detected the first FSH surge coincident with the LH surge, the second FSH surge about 24 h later and the periodic fluctuations of FSH concentrations during the luteal phase of the oestrous cycle. These enzyme immunoassays are an efficient and economic alternative to the established radioimmunoassays (RIA) for oLH and oFSH.     

The combined effect of fenbendazole treatment and a move to aftermath 7 or 9 weeks after turnout on Dictyocaulus viviparus infections in calves

A grazing study was performed with the main objective of examining the effect of fenbendazole (FBZ) in a ‘dose and move’ system on nematode infections in calves with special emphasis on Dictyocaulus viviparus.

Three groups of six calves were grazed from May to October 1993. One group (DM7) was treated with FBZ and moved to aftermath (pasture which had only been mown) 7 weeks after turnout. The second group (DM9) was similarly treated and moved 9 weeks after turnout and the third group served as untreated pasture control group (PC) and was moved to aftermath 9 weeks after turnout.

FBZ treatment removed adult lungworms from DM7 and DM9. Tracer calves grazed during the first 7 or the first 9 weeks after turnout acquired mean burdens of 18 and 125 lungworms, respectively. In PC faecal larval counts increased until the end of August. Most of the animals in this group were then suffering from lungworm disease and emergency treatment with ivermectin was given. In both FBZ-treated groups, larvae reappeared in the faeces of some of the calves 4–5 weeks after treatment. Subsequent reinfection resulted in higher mean faecal larval counts in both groups 2 months after treatment, although variation in faecal larval counts was high. In DM7 values tended to be higher than in DM9. These higher larval counts were associated with mild signs of parasitic bronchitis in some calves of DM7, whereas no signs were seen in DM9.

At the end of the experiment, all calves, and also a group of six permanently housed non-infected control calves (HC), were infected experimentally with 5000 D. viviparus larvae to evaluate development of immunity. The worm counts at necropsy showed that all calves on pasture had developed immunity.     

Cooperia punctata trickle infections: parasitological parameters and evaluation of a Cooperia recombinant 14.2 kDa protein ELISA for estimating cumulative exposure of calves

We examined the effects of isoquinoline alkaloids in vitro in an effort to identify a treatment for Strongyloides stercoralis larva migrans in humans. Infective third-stage larvae of S. ratti and S. venezuelensis were used as model nematodes for S. stercoralis. Nematocidal activity was evaluated by the 50% paralysis concentration (PC(50)). Most of the tested isoquinoline alkaloids had activity for S. ratti and S. venezuelensis. We then evaluated in vitro cytotoxicity, which was the 50% inhibition concentration (IC(50)) of the compounds using HL60 tissue-culture cells. Three of the compounds (protopine, D-corydaline, and L-stylopine) which exhibited strong nematocidal activity, showed little cytotoxicity. In addition, we examined the relationship between nematocidal activity and cytotoxicity using the PC(50)/IC(50) ratio. A ratio equivalent to or lower than that calculated for the currently prescribed strongyloidosis treatments, ivermectin, albendazole and thiabendazole, was observed for allocryptopine, protopine, dehydrocorydaline, D-corydaline, L-stylopine, and papaverine. In contrast, the PC(50)/IC(50) ratios for protopine, D-corydaline, and L-stylopine were substantially more favorable. Therefore, protopine, D-corydaline, and L-stylopine were identified as potential effective treatments for strongyloidosis.     

Characterization of transgenic livestock production

The objective of transgenic livestock improvement projects is to develop and bring to market superior breeding stock, as well as germplasm for the artificial insemination and embryo transfer industries. Livestock animal biotechnology programs hold the promise of achieving, in a single generation, improvements in commercially important livestock species previously possible only through long-term traditional selective breeding practices or by chance mutation. Transgenic farm animals harboring growth hormone or metabolically related structural genes have been created. Studies of these animals demonstrate the effects of inadequate regulation of transgene expression. Research continues to explore the intricacies of developmental regulation of such genes and phenotypic consequences of mammalian gene transfer. Ultimately, genetically engineered livestock will provide producers with the benefit of increased production efficiencies while the consumer will have healthier animal food products. Conceivably, products will be produced with lower levels of fat, cholesterol, feed additives and pharmaceutical residues from animals with altered carcass composition that will result in greater nutritional benefit for the consumer.     

The Control of Reactive Oxygen Species Influences Porcine Oocyte In Vitro Maturation

The aim of this study was to examine the effect of varying intracellular reactive oxygen species (ROS) levels during oocyte in vitro maturation with enzymatic ROS production systems (xanthine + xanthine oxidase or xanthine + xanthine oxidase + catalase), scavenger systems (catalase or superoxide dismutase + catalase) or cysteine on porcine oocyte maturation. Oocyte ROS levels showed an increase when H₂O₂ or O₂?^‐ production systems were added to the culture medium (p < 0.05). On the other hand, the presence of ROS scavengers in the maturation medium did not modify oocyte ROS levels compared with the control after 48 h of maturation, but the addition of cysteine induced a decrease in oocyte ROS levels (p < 0.05). The ROS production systems used in this work did not modified the percentage of oocyte nuclear maturation, but increased the decondensation of sperm head (p < 0.05) and decreased the pronuclear formation (p < 0.05). In turn, the addition of O₂?^‐ and H₂O₂ scavenging systems during in vitro maturation did not modify the percentage of oocytes reaching metaphase II nor the oocytes with decondensed sperm head or pronuclei after fertilization. However, both parameters increased in the presence of cysteine (p < 0.05). The exogenous generation of O₂?^‐ and H₂O₂ during oocyte in vitro maturation would not affect nuclear maturation or later sperm penetration, but most of the spermatozoa cannot progress to form the pronuclei after fusion with the oocyte. The decrease in endogenous ROS levels by the addition of cysteine would improve pronuclear formation after sperm penetration.     

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.     

The resistance of meat chickens vaccinated by aerosol with a live V4 Newcastle disease virus vaccine in the field to challenge with a velogenic Newcastle disease virus

Meat chickens housed on a commercial broiler farm in Australia were vaccinated once at 10 to 11 days-of-age by aerosol with live V4 Newcastle disease virus (NDV) vaccine. Groups of vaccinated and unvaccinated birds were flown to Malaysia, where they were challenged with a virulent strain of NDV. Survival rates in vaccinated chickens challenged 7, 14, 21 or 31 d after vaccination were 0.47, 0.77, 0.97 and 0.92, respectively. All unvaccinated chickens died due to Newcastle disease (ND) following challenge. Chickens in Australia and Malaysia were bled and the serums tested for haemagglutination-inhibiting (HI) antibody to NDV. Many vaccinated birds with no detectable antibody, and all birds with a log2 titre of 2 or greater, survived challenge. The results showed that this V4 vaccine induced protective immunity in a significant proportion of chickens within 7 d of mass aerosol vaccination. This early immunity occurred in the absence of detectable circulating HI antibody. Non-HI antibody mediated immunity continued to provide protection up to 31 d after vaccination. Almost all vaccinated birds were protected within 3 w of vaccination. It is concluded that the V4 vaccine is efficacious and could be useful during an outbreak of virulent ND in Australia.     

Immunity of calves treated with an oxfendazole pulse release bolus to challenge with Dictyocaulus viviparus

Two experiments were performed to study the effect of an oxfendazole pulse release intraruminal device (OPRB) on the development of lungworm infection and immunity in calves. Infections were monitored in OPRB-treated and control calves by faecal and pastural larval counts. Levels of immunity of grazing groups and helminth-free housed groups were compared by challenge infection. Even though signs of parasitic bronchitis were seen only in control animals, sufficient immunity had developed in OPRB-treated calves to prevent disease after a substantial challenge infection. However, the degree of immunity was less than in untreated grazing control groups and there were indications that it was related to initial infection levels. These results imply that the OPRB may successfully control parasitic bronchitis, provided that pasture larval infections are sufficient to allow the development of immunity.