Prognostic significance of surgical margin, Ki-67 and cyclin D1 protein expression in grade II canine cutaneous mast cell tumor

The prognosis for canine cutaneous mast cell tumor (CCMT) is thought to be correlated with histopathological grading. However, the wide variety of histopathologic types of grade II is one of the most troublesome and difficult points for prognosis. The objective of this study is to determine the prognostic value of surgical margin, ki-67 and cyclin D1 protein expression in grade II tumor. Surgically resected specimens of solitary grade II CCMT from 48 dogs with follow-up periods over 360 days (median was 1080 days) were used in this study. The expression of cyclin D1 and ki-67 proteins was determined by morphometrically using slides stained immunocytochemically, and the correlations among the results, survival rate, and recurrence and/or metastasis rate of each dog were analyzed statistically. The recurrence and/or metastasis and mortality rate in the incomplete surgical excision group within 30 months postoperatively were higher than that of the complete surgical excision group. In the incomplete surgical excision group, dogs with low positive staining of ki-67 had a significantly better survival, but the recurrence and metastasis rate and ki-67 positivity failed to show a significant correlation. Only a small number of cases showed cyclin D1-positive tumor cells, but most of them had a poor outcome with a high recurrence rate. In grade II CCMT, incomplete excision induced a relatively high metastasis rate and poor prognosis. Ki-67 positivity is a marker for the estimation of overall survival in incomplete surgical excision cases. Cyclin D1 positivity was low and may not have a prognostic role.     

Antihyaluronidase action of ellagic acid effectively prevents polyspermy as a result of suppression of the acrosome reaction induced by sperm-zona interaction during in vitro fertilization of porcine oocytes

The present study was conducted to examine the effects of three tannin relatives (tannic acid, TA; gallic acid, GA; and ellagic acid, EA) on antihyaluronidase and reactive oxygen species (ROS) scavenging activity, in vitro fertilization (IVF) parameters, and the acrosome reaction (AR) induced by sperm-zona interaction. Among the three tannin relatives, TA and EA showed the strongest potency for blocking the hyaluronidase activity of boar sperm, with concentration-dependent inhibition over the range of 2-10 microg/ml. In contrast, ROSs were effectively scavenged by TA and GA, but not EA. When cumulus-free oocytes were inseminated in IVF medium containing 5 microg/ml of the tannin relatives, polyspermy was significantly reduced by TA and EA (32 and 29%, respectively) compared with oocytes treated with or without GA (51 and 69%, respectively) under conditions that maintained a high sperm penetration rate (P<0.05). Interestingly, induction of the AR by treatment of preincubated sperm with progesterone was blocked by TA and GA as a result of their higher levels of ROS scavenging activity, while EA, which possessed weak ROS scavenging activity, did not disturb induction of the AR with progesterone. However, the incidence of AR induced by sperm-zona interaction was significantly decreased by the strong antihyaluronidase actions of TA and EA compared with that in the absence of these compounds. Treatment with the compounds caused neither a protective proteolytic modification of the zona pellucida matrix before fertilization nor a reduction in acrosomal proteolytic activity or the number of zona-bound sperm. These findings suggest that the antihyaluronidase action of EA effectively prevents polyspermy by suppression of AR functionality induced by sperm-zona interaction and that hyaluronidase intervention is therefore required during porcine IVF.     

Spindle formation and microtubule organization during first division in reconstructed rat embryos produced by somatic cell nuclear transfer

The present study was conducted to demonstrate the spindle formation and behavior of chromosomes and microtubules during first division in reconstructed rat embryos produced by somatic cell nuclear transfer (SCNT) with cumulus cell nuclei. To demonstrate the effect of oocyte aging after ovulation on the cleavage of SCNT embryos, micromanipulation was carried out 11, 15 and 18 h after injection of hCG. SCNT oocytes were activated by incubation in culture medium supplemented with 5 microM ionomycin for 5 min followed by treatment with 2 mM 6-dimethylaminopurine (6-DMAP) in mR1ECM for 2-3 h. For immunocytochemical observation, the SCNT embryos were incubated with monoclonal anti-alpha-tubulin antibody and then fluorescein isothiocyanate (FITC)-conjugated goat anti-mouse IgG. Cleavage rates were significantly higher for oocytes collected after 15 and 18 h rather than for those collected 11 h after injection of hCG (56 and 53%, respectively vs. 28%; P<0.05). Premature chromosome condensation occurred before activation of the SCNT oocytes, but adequate spindle formation was only rarely observed. The distribution of microtubules in SCNT embryos after activation was different from those of fertilized and parthenogenic oocytes, i.e., a dense microtubule organization shaped like a ring was observed. Eighteen to 20 h post-activation, most SCNT embryos were in the 2-cell stage, but no nucleoli were clearly visible, which was quite different from the fertilized oocytes. In addition, first division with and without small cellular bodies containing DNA was observed in the rat SCNT embryos in some cases. The present study suggests that reorganization of transferred nuclei in rat SCNT embryos may be inadequate in terms of formation of the mitotic assembly and nucleolar reorganization.     

Factors affecting the success rate of treatment of recumbent dairy cows suffering from hypocalcaemia

We aimed to investigate the ratio of accompanying diseases in cows suffering from clinical hypocalcaemia and their influence on cure rate. In five veterinary practices in different regions of Germany, all recumbent cows around parturition were included in the study for a period of 1 year. After recording the case history a clinical examination was done and a serum sample was taken to measure the concentrations of calcium and phosphorus magnesium, beta-hydroxybutyrate, total bilirubin, cholesterol, urea, and the activities of ASAT, CK, and GLDH. Only cows with hypocalcaemia entered the statistical analysis. Hypocalcaemia was the major cause of recumbency in cows of the second lactation or elder. Muscle damage was the second frequent diagnose in recumbent cows and the major concomitant disease in hypocalcaemic cows. The overall cure rate was between 89.4% and 94.8%. Calcium and phosphorus concentrations did not have an influence on cure rate. Non-cured cows had higher serum activities of CK (p<0.043) and ASAT (p<0.006). Nevertheless, the activities of CK and ASAT were no good predictors of treatment failure because of their low specificity and the high cure rate of the cows in the five practices.     

Evaluation of an indirect enzyme-linked immunosorbent assay (ELISA) using recombinant toxin for detection of antibodies against Pasteurella multocida toxin

To facilitate the control of progressive atrophic rhinitis (PAR) of swine caused by toxigenic Pasteurella multocida, an enzyme-linked immunosorbent assay (ELISA) and a serum neutralization test (NT) have recently been developed to detect antibodies against the P. multocida dermonecrotic toxin (PmDNT). However, the NT is a cumbersome and time-consuming technique. To overcome these drawbacks, we developed an indirect ELISA, using recombinant PmDNT expressed in Escherichia coli, for the detection of antibodies to PmDNT in serum samples from pigs. The practical usefulness of this ELISA was compared with the NT using serum samples obtained from experimentally infected and naturally infected pigs. In the pigs experimentally inoculated with vaccine including PmDNT toxoid, the ELISA and neutralization antibodies were detected at almost the same time, and a good correlation was demonstrated between both tests (P<0.01, R(2)=0.807). Therefore, the ELISA can be used to evaluate the immune reaction of pigs after vaccination with P. multocida toxoid. In a survey conducted on a field herd with a history of clinical AR, the seropositivity by ELISA in pigs of age 4.5-6 months was increased even though the NT was negative, and the correlation was low between the results obtained with the two tests (P<0.01, R(2)=0.38). Therefore, the results indicated that this ELISA might be a useful alternative to the NT currently used to detect the antibody to PmDNT after vaccination or infection with P. multocida.     

Tsetse control in cattle from pyrethroid footbaths

In Burkina Faso, we assessed the efficacy of treating cattle with a footbath containing aqueous formulations of pyrethroids to control two tsetse-fly species, Glossina tachinoides Westwood, 1850 (Diptera, Glossinidae) and Glossina palpalis gambiensis Vanderplank 1949. Legs were the most targeted parts of the body for tsetse-fly blood meals: 81% (95% CI: 73, 89) for G. tachinoides and 88% (81, 95) for G. palpalis.

The in-stable efficacy of footbath treatments was compared with manual full spraying with a 0.005% alphacypermethrin (Dominex, FMC, Philadelphia, USA) formulation (250 mL versus 2 L). The proportions of knocked-down flies were the same with footbath and full spray but the latter was more protective against fly bites. In field use, the efficacy of both methods should be similar given the recommended treatment frequency: 3 days for footbath versus 7 days for full spray.

Among 96 cattle drinking at the same water point in Dafinso (Burkina Faso), 68 (71%) were treated with a footbath containing a 0.005% deltamethrin formulation (Vectocid, CEVA SA, Libourne, France). We observed the effect of this live-bait technique on the one hand on released cohorts of reared, irradiated flies, and on the other hand on wild tsetse flies. In both cases, the footbath treatment was associated with a reduction of the apparent fly density probably related to an increased mortality.     

Clinical, neurological and clinicopathological signs, treatment and outcome of metaldehyde intoxication in 18 dogs

OBJECTIVES: To describe the clinical signs, clinicopathological abnormalities and outcome of metaldehyde intoxication in dogs. METHODS: Medical records of dogs presenting between 1989 and 2005 with a diagnosis of metaldehyde toxicity were reviewed retrospectively. Data obtained from the medical record included signalment, history, clinical signs, laboratory tests results, hospitalisation period length, treatments and outcome. RESULTS: Eighteen dogs fulfilled the inclusion criteria. The most prevalent clinical signs were seizures, hyperthermia, tachycardia and muscle tremors. Serum biochemistry abnormalities included increased serum muscle enzymes activities, acidaemia (six dogs) and decreased blood bicarbonate (eight dogs). Treatment was symptomatic and supportive. Hyperbilirubinaemia was observed in two dogs. Diazepam was the most commonly used anticonvulsant followed by phenobarbitone and pentobarbital. General inhalant anaesthesia was required in nine of 18 dogs with seizures unresponsive to anticonvulsants. The survival was 83 per cent (15 of 18 dogs). CLINICAL SIGNIFICANCE: This clinical study recorded, for the first time in the veterinary literature, several clinicopathological abnormalities from severely intoxicated dogs. Metabolic acidosis was common, while acute or delayed hepatotoxicity was an uncommon complication.     

Culicoides biting midges at the National Zoological Gardens of South Africa

Culicoides biting midges (Diptera: Ceratopogonidae) are responsible for the transmission of a large number of pathogens to livestock and wild animals. In this study the presence of the genus, using light traps based at four different sites within the National Zoological Gardens of South Africa, was investigated during 2002-2004. In total, 37 species were recorded, including large numbers of Culicoides imicola Kieffer, 1913, which is responsible for the transmission of economically important arboviruses in South Africa, Europe, Middle and Far East. These results are discussed with reference to the wider Culicoides fauna in the Onderstepoort area of South Africa, their vector competence as well as biosecurity at the National Zoological Gardens.