Production of Lambs After the Transfer of Fresh and Cryopreserved in vitro Produced Embryos from Prepubertal Lamb Oocytes and Unsorted and Sex-sorted Frozen-thawed Spermatozoa

Cumulus-oocyte complexes from hormone-stimulated 3-4-week-old (n=43) and 6-7-week-old (n=12) prepubertal lambs were matured in vitro and incubated with unsorted, or X- or Y-spermatozoa separated with a high-speed cell sorter (SX MoFlo)frozen-thawed. Presumptive zygotes were then cultured to the blastocyst stage, and transferred to recipients fresh or after cryopreservation (frozen). Oocyte cleavage was higher (p <0.05) with unsorted (515/926, 55.6%) than X- or Y-spermatozoa (261/672, 38.8% and 229/651, 35.2%, respectively) and blastocyst formation (% zygotes) by Day 9 of in vitro culture was lower (p <0.05) for X- (102/261, 39.1%) than unsorted spermatozoa (249/515, 48.3%), but did not differ between Y-spermatozoa (103/229, 45.0%) and unsorted spermatozoa, or between X- and Y-spermatozoa (p >0.05). For fresh embryos, survival to term was 50.0% (3/6) for unsorted, 0.0% (0/6) for X- and 16.7% (1/6) for Y-spermatozoa-derived embryos (p >0.05), and for frozen embryos was 4.0% (2/50) for unsorted, 9.1% (2/22) for X- and 2.9% (1/34) Y-spermatozoa-derived embryos (p >0.05). Of the two lambs born from X-spermatozoa-derived embryos, one was female (50%), and from the two Y-spermatozoa-derived lambs, both were male (100%), demonstrating that lambs can be produced after the transfer of fresh and cryopreserved IVP embryos derived from prepubertal lamb oocytes and frozen-thawed sex-sorted sperm.     

Expression of the porcine beta2-adrenergic receptor in Chinese hamster ovary cells

The gene for the porcine beta2-adrenergic receptor (pbeta2AR) was transfected into Chinese hamster ovary (CHO) cells for expression. Fourteen stable cell lines were obtained and exhibited receptor densities ranging from 12 to 2,371 fmol/mg membrane protein. The receptor density was not correlated with estimates of gene copy number obtained by Southern hybridization. The pbeta2AR in CHO cells exhibited saturable binding of [125I]CYP (Kd = 14.5 pM) and stereospecificity for (-)- and (+)-isoproterenol. The relative affinities for (-)-isoproterenol (ISO), (-)-epinephrine (EPI), and (-)-norepinephrine (NEPI) were ISO > EPI > NEPI, which are characteristic of beta2AR. The affinity values for these ligands were similar to those in other species. Binding of ISO, EPI, and NE revealed two affinity states of the betaAR; the high-affinity state was eliminated by adding Gpp(NH)p, a nonhydrolyzable GTP analogue. Binding of the antagonist propranolol modeled to only one affinity state, and Gpp(NH)p did not affect binding. Multiple affinity states are characteristic of agonist-induced coupling of betaAR with G-proteins, and the data suggest that the cloned pbetaAR is functionally competent. Data confirm that the pbeta2AR is the pig version of beta2AR. Stable CHO cell lines will be useful for characterization of pbeta2AR and screening and designing potential drugs that may be used to enhance pig production.     

Development and evaluation of a rapid absorbed enzyme immunoassay test for the diagnosis of Johne''s disease in cattle

An absorbed enzyme immunoassay (EIA) test for Johne's disease in cattle was developed in which absorption of cross-reacting antibodies occurred as a rapid reaction in solution rather than overnight with whole organisms and a subsequent centrifugation step. Total test time was reduced to less than 2 h with a minimum of manipulations. The test was evaluated in cattle herds from Johne's disease-endemic and Johne's disease-free regions of Australia. Specificity was 99.8%. Calculations of sensitivity were affected by the history of the herd under test. However, the EIA detected in excess of 80% of animals before onset of clinical disease and 65% of faecal shedders were EIA positive on, or before, first detection of Mycobacterium paratuberculosis in their faeces. The test should aid epidemiological studies and be a useful tool in the management and control of Johne's disease.     

The Effect of Temperature on First Feeding, Growth, and Survival of Larval Witch Flounder Glyptocephalus cynoglossus

Witch flounder Glyptocephalus cynoglossus has recently been identified as a candidate species for aquaculture in the northeastern United States and the Canadian Atlantic Provinces. This study investigated the optimal temperatures for witch flounder larval first feeding and for long term larval culture from hatching through metamorphosis. Maximum first feeding occurred between 15.0 and 16.2 C. Larvae did not survive beyond first feeding when reared at mean temperatures of 5.1, 10.4, or 19.5 C and were unable to initiate feeding at mean rearing temperatures below 6.0 C. At a rearing temperature of 15.0 C in 16-L tanks, mean larval survival to 60 days post hatch (dph) was 14.1%. Mean overall length-specific growth rate for larvae reared to 60 dph at 15.0 C was 3.5%/d and mean absolute growth was 0.62 mm/d. Subsequent larval growth at 15.6 C began to taper off towards 70 dph at the onset of weaning which overlapped with larval metamorphosis. Growth plateaued at 85 dph, followed by a rebound between 90 and 95 dph. Survival was 100% when weaning onto a dry, pelleted diet was initiated at 70 dph with a 10-d live diet co-feeding period. These results are favorable and encourage the further pursuit of commercial witch flounder culture.     

Investigations of an enteric infection of cockatoos caused by an enterovirus-like agent

An enteric infection in cockatoos associated with a 30 nm diameter enterovirus-like agent seen in faeces and intestinal epithelial cells is described. The disease is characterised by intractable, profuse, mucoid diarrhoea, weight loss, dehydration and death. Lesions in the intestine consist of villous atrophy, villous fusion, enterocyte hyperplasia and, in some cases, chronic inflammation. Affected birds so far examined have concurrent psittacine beak and feather disease.     

Conservation of Pattern and Process: Developing an Alternative Paradigm of Rangeland Management

This article examines the question of how well the rangeland management profession has served conservation of patterns and processes that support multiple ecosystem services. We examine the paradigms under which rangeland management operates and argue that our profession developed under the utilitarian paradigm with the primary goals of sustainable forage for livestock production. While optimization of multiple rangeland products and services has always been a consideration, a comprehensive set of principles have not be been developed to advance this concept. We argue that fire and grazing, often viewed as mere tools used for production goals, should rather be viewed as essential ecosystem processes. Rangeland management continues to operate under the utilitarian paradigm appropriate to societal values of the 20th century and by and large has failed to provide management guidance to reverse degradation of several highly valued ecosystem services. We support this argument with evidence that biodiversity has declined on rangelands in the past half century and that much of this decline is due to management goals that favor a narrow suite of species. The full suite of ecosystem services valued by society will only benefit by management for heterogeneity, which implies that there is no one goal for management and that landscape-level planning is crucial. Explicitly incorporating heterogeneity into state-and-transition models is an important advancement not yet achieved by our profession. We present new principles for rangeland management formed on the basis of conservation of pattern and process. While recognizing that many rangelands have significant deviations from historic plant communities and disturbance regimes, we suggest that management for conservation of pattern and process should focus on fire and grazing to the extent possible to promote a shifting mosaic across large landscapes that include patches that are highly variable in the amount of disturbance rather than the current goal of uniform moderate disturbance.     

Epitope-tagged insulin-like growth factor-I expression in muscle

Development of a recombinant insulin like growth factor I (IGF-I) that is distinguishable from its endogenous counterpart would provide a powerful tool for delineating the role of IGF in myogenesis. Therefore, the objective of this study was to create an epitope-tagged IGF-I that retains biological activity and determine whether expression of this construct is possible in muscle tissue following direct DNA injection. Expression vectors were created that encoded porcine IGF-I containing a T7 (11-amino acid) epitope-tag (TIGF). Immunoreactivity of the purified recombinant TIGF was confirmed using monoclonal antibodies. Biological activity was evaluated by examining differentiation of myoblasts cultured with TIGF or transfected with TIGF plasmid DNA. Addition of purified TIGF to myoblast cultures stimulated (P < 0.05) muscle creatine kinase levels similar to insulin (10(-5) M). Likewise, transfection of L6A1 with TIGF DNA hastened (P < 0.01) differentiation compared to control pcDNA-transfected myoblasts. The integrity of the recombinant protein was confirmed using a sandwich-configured enzyme linked immunosorbent assay. Finally, recombinant TIGF DNA was injected in porcine muscle and the ability to detect TIGF protein was evaluated. TIGF expression was detected in muscle fibers of injected porcine muscle. These data show that a T7 amino acid tag placed on the amino terminus of the IGF-I protein remains intact during processing and does not interfere with the biological activity of the molecule. Use of this DNA construct is an excellent tool for investigating the role of IGFs in control muscle development and provides a model to investigate other regulators of animal growth.