Efficiency of Two Enucleation Methods Connected to Handmade Cloning to Produce Transgenic Porcine Embryos

The purpose of our work was to establish an efficient-oriented enucleation method to produce transgenic embryos with handmade cloning (HMC). After 41–42 h oocytes maturation, the oocytes were further cultured with or without 0.4 μg/ml demecolcine for 45 min [chemically assisted handmade enucleation (CAHE) group vs polar body (PB) oriented handmade enucleation (OHE) group respectively]. After removal of the cumulus cells and partial digestion of the zona pellucida, oocytes with visible extrusion cones and/or polar bodies attached to the surface were subjected to oriented bisection. Putative cytoplasts without extrusion cones or PB were selected as recipients. Two cytoplasts were electrofused with one transgenic fibroblasts expressing green fluorescent protein (GFP), while non-transgenic fibroblasts were used as controls. Reconstructed embryos were cultured in Well of Wells (WOWs) with porcine zygote medium 3 (PZM-3) after activation. Cleavage and blastocyst rates were registered on day 2 and day 7 of in vitro culture respectively. Meanwhile, the total blastocyst cell number was counted on day 7. We found that the difference was only observed between blastocyst rates (38.6 ± 2% vs 48.1 ± 3%) of cloned embryos with GFP transgenic fibroblast cells after CAHE vs OHE. With adjusted time-lapse for zonae-free cloned embryos cultured in WOWs with PZM-3, it was obvious that in vitro developmental competence after CAHE was compromised when compared with the OHE method. OHE enucleation method seems to be a potential superior alternative method used for somatic cell nuclear transfer (SCNT) with transgenic fibroblast cells.     

Systemic antibacterial drug use in dogs in Australia

A survey of veterinarians' use of antibacterial drugs was conducted by distributing a questionnaire to Australian practitioners. Respondents were asked to indicate their general patterns of use of various systemic antibacterial drugs and drug combinations in dogs and their approach to certain specified clinical disorders. Overall, antibacterials of the p-lactam type (penicillins and cephalosporins) were most commonly used. Other antibacterials with substantial use were doxycycline, sulphonamide-trimethoprim, metronidazole, fluoroquinolones and clindamycin. Drug selection for different disorders was generally appropriate when compared with recommendations in recent texts, although inappropriate use was evident in some circumstances.     

Morphological Changes of the Endometrial Epithelium in the Bitch during Metoestrus and Anoestrus

Although cyclic changes of the endometrium in dogs involving both stromal and glandular compartments have been described, the fate of the surface epithelium after progressive growth and secretion is still unclear. In the present study, uteri of 43 healthy bitches in metoestrus and anoestrus were examined macroscopically and histologically. Tissue biopsies were taken from three different locations (cranial and middle parts of uterine horns and bifurcation). The stage of the oestrous cycle was determined by evaluation of progesterone and oestradiol levels in plasma hormone and was also confirmed clinically. Crypts formed in the luteal phase were covered with a columnar epithelium which gradually underwent fatty degeneration. In addition, the stromal part of the crypts disappeared and finally, in early anoestrus, epithelial sheaths desquamated and shed off into the uterine lumen. The surface epithelium was replaced by new cuboidal cells proliferating and migrating from the glandular openings. These findings were confirmed by oil red O staining and immunohistochemical detection of proliferation with Ki‐67 marker.     

Molecular epidemiology of clinical isolates of Pseudomonas aeruginosa isolated from horses in Ireland

Clinical isolates (n = 63) of Pseudomonas aeruginosa obtained from various sites in 63 horses were compared using ERIC2 RAPD PCR to determine their genetic relatedness. Resulting banding patterns (n = 24 genotypes) showed a high degree of genetic heterogeneity amongst all isolates examined, indicating a relative non-clonal relationship between isolates from these patients, employing this genotyping technique. This study characterised 63 clinical isolates into 24 distinct genotypes, with the largest cluster (genotype E) accounting for 10/63 (15.9%) of the isolates. ERIC2 RAPD PCR proved to be a highly discriminatory molecular typing tool of P. aeruginosa in isolates recovered from horses. With the adoption of several controls to aid reproducibility, this technique may be useful as an alternative to PFGE, particularly in epidemiological investigations of outbreaks where speed may be a significant parameter. This is the first report of clonal heterogeneity amongst P. aeruginosa from horses and demonstrated that ERIC RAPD PCR is a rapid method for the examination of this species in horses, which may be useful in outbreak analysis.     

Heliotropium europaeum poisoning of sheep with low liver copper concentrations and the preventive efficacy of cobalt and antimethanogen

In a field experiment in the Mallee district of Victoria, Merlno xBorder Leicester ewes and wethers grazed Heliotropium europaeum (heliotrope) over periods of 3 to 4 months in 4 successive years. By the end of the second year 12% (14 of 120) of the sheep had died; after 4 years the loss attributable to heliotrope was between 18% and 35%. Mortality was not affected by intraruminal treatment with cobalt or antimethanogen. At the end of the experiment the highest concentration of copper in the liver was 1.95 mmol/kg wet weight (approximately 413 micrograms/g dry weight). The relatively low mortality from primary heliotrope poisoning and the low concentration of copper in the liver of sheep grazing the plant are discussed in relation to the contrasting situation that prevails in the Riverina area of New South Wales. The importance of local environmental factors in the management of heliotrope grazing by sheep is emphasised, particularly in relation to the number of seasons in which the plant may be a major component of the diet.     

Comparison of oestrus detection methods in dairy cattle

Sixty-seven Holstein-Friesian cows, from 20 days postpartum, were recruited into the study and fitted with both a pedometer (SAE Afikim) and a Heatime neck collar (SCR Engineers) and allocated a heat mount detector (either scratchcard [Dairymac] or KaMaR [KaMaR]) or left with none, relying only on farm staff observation. Common production stressors and other factors were assessed to determine their impact on the ability of each method to accurately detect oestrus and to investigate effects on the frequency of false-positive detections. Only 74 per cent of all potential oestrus periods (episodes of low progesterone) were identified by combining information from all methods. There was no difference between the methods in terms of sensitivity for detecting 'true oestrus events' (approximately 60 per cent), with the exception of scratchcards, which were less efficient (36 per cent). Pedometers and KaMaRs had higher numbers of false-positive identifications. No production stressors had any consequence on false-positives. The positive predictive values for neck collars or observation by farm staff were higher than those of other methods, and combining these two methods yielded the best results. Neck collars did not detect any of the nine oestrus events occurring in three cows with a body condition score (BCS) of less than 2, and the efficiency of correctly identifying oestrus was also reduced by high milk yield (odds ratio [OR]=0.34). Pedometer efficiency was reduced by lameness, low BCS or high milk yield (OR=0.42, 0.15 or 0.30, respectively).     

Retrospective study of 108 foals with septic osteomyelitis

Objective   To determine the clinical characteristics, short-term outcome and future athletic performance of foals with septic osteomyelitis.
Design   Retrospective clinical study of 108 Thoroughbred foals with radiographic evidence of bone infection that were presented at the Scone Veterinary Hospital between August 1995 and December 2001. Medical records were reviewed and information concerning signalment, the clinical, laboratory and radiographic findings, treatment and outcome was obtained. Racing records were obtained and evaluated for surviving foals that had reached racing age.
Results   Mean age of foals at initial evaluation was 39 days (range 1–180 days); 21 foals had multiple radiographic bone lesions (19.4%), and 76 had concurrent septic arthritis (70.4%). The most frequently affected bones were the femur, tibia and distal phalanx. In total, 87 foals were discharged from the hospital (80.6%), 79 survived long-term to reach racing age and 52 raced (65.8%). Overall, 48% (52/108) of the foals treated for osteomyelitis raced. Foals less than 30 days of age at the time of diagnosis, critically ill foals and those with multiple bones or joints affected were significantly less likely to be discharged from hospital. Multiple septic joints, but not multiple bone involvement, had an unfavourable prognosis for racing.
Conclusions   The prognosis for survival of foals with septic osteomyelitis or osteitis is favourable. Multiple bone or joint involvement is an important short-term prognostic indicator; however, the involvement of multiple joints, but not multiple infected bones, is associated with an unfavourable prognosis for racing.     

Hemodynamic effects of intravenous lidocaine in isoflurane anesthetized cats

Lidocaine dose‐dependently decreases the minimum alveolar concentration (MAC) of isoflurane in cats. The purpose of this study was to determine the hemodynamic effects of six lidocaine plasma concentrations in isoflurane anesthetized cats. Six cats were studied. After instrumentation, end‐tidal isoflurane concentration was set at 1.25 times the individual minimum alveolar concentration (MAC), which was determined in a previous study. Lidocaine was administered intravenously to target pseudo‐steady state plasma concentrations of 0, 3, 5, 7, 9, and 11 μg ml^–1, and isoflurane concentration was reduced to an equipotent concentration, determined in a previous study. Cardiovascular variables; blood gases; PCV; total protein and lactate concentrations; and lidocaine and monoethylglycinexylidide concentrations were measured at each lidocaine target concentration, before and during noxious stimulation. Derived variables were calculated. Data were analyzed using a repeated measures anova , followed by a Tukey test for pairwise comparisons where appropriate. One cat was excluded from analysis because the study was aborted at 7 μg ml^–1 due to severe cardiorespiratory depression. Heart rate, cardiac index, stroke index, right ventricular stroke work index, total protein concentration, mixed‐venous PO₂ and hemoglobin oxygen saturation, arterial and mixed‐venous bicarbonate concentrations, and oxygen delivery were significantly lower during lidocaine administration than when no lidocaine was administered. Mean arterial pressure, central venous pressure, pulmonary artery pressure, systemic and pulmonary vascular resistance indices, PCV, arterial and mixed‐venous hemoglobin concentrations, lactate concentration, arterial oxygen concentration, and oxygen extraction ratio were significantly higher during administration of lidocaine than when no lidocaine was administered. Most changes were significant at lidocaine target plasma concentrations of 7 μg ml^–1 and above. Noxious stimulation did not significantly affect most variables. Despite significantly decreasing in inhalant requirements, when combined with isoflurane, lidocaine produces greater cardiovascular depression than an equipotent dose of isoflurane alone. The use of lidocaine to reduce isoflurane requirements is not recommended in cats.