Dynamics in the membrane organization of the mammalian sperm cell and functionality in fertilization

The capacitation process of sperm cells involves complex changes in the composition and orientation of molecules at the surface of the sperm cell. Here we focus on the lipid architecture in the sperm plasma membrane and demonstrate that the sperm plasma membrane is not static but is an extremely dynamic structure. Advanced fluoroscopic techniques enabled continuous monitoring of lipid organization in living cells and extremely rapid lipid movements were observed. The orientation of lipids in the sperm plasma membrane changed under capacitative treatments, was found to be sensitive for temperature and also changed upon binding of sperm cells to the zona pellucida. The changes in membrane properties coincided with an activation of protein kinases resulting in tyrosine phosphorylation of specific plasma membrane proteins. The detected membrane changes relate to intrinsic membrane properties such as fluidity, permeability, adhesiveness and fusibility. We think that these results may provide a physiological basis for new assays, able to discriminate between functional and non-physiological sperm cells.     

Rickets caused by excessive renal phosphate loss and apparent abnormal vitamin D metabolism in a cat

Rickets was diagnosed in a 1-year-old cat with a history of weakness, osteopenia, and recurrent fractures. Processes causing rickets include vitamin D deficiency caused by inadequate, nutrition, lack of exposure to sunlight, defective metabolism of parent vitamin D to active metabolites, inherited vitamin D receptor defects, hypoparathyroidism, chronic renal failure, renal loss of phosphate, or malabsorptive states resulting from gastrointestinal or hepatic diseases. On the basis of analysis of serum 25-hydroxyvitamin D3 and 1,25-dihydroxyvitamin D2 and D3 concentrations, serum biochemical analysis, and urinary fractional clearance of electrolytes, the causes of rickets in our cat, were most compatible with a combination of excessive loss of phosphorus via the kidneys and deficient or abnormal hepatic 25-hydroxylation of vitamin D. Calcifediol treatment and twice daily administration of phosphate salts resulted in clinical improvement and increases in mineralization of the skeleton, as evidenced on radiographic evaluation.     

Hemorrhagic septicemia in fallow deer (Dama dama) caused by Pasteurella multocida multocida.

Four outbreaks of hemorrhagic septicemia caused by Pasteurella multocida multocida occurred in a population of 1,800 fallow deer (Dama dama) during 1992-1996. A total of 340 fallow deer were submitted for postmortem examination. Pasteurellosis was diagnosed in 273 of 312 deer suspected of having septicemia. Pasteurella multocida was isolated from 257 animals, and the diagnosis was based on typical pathologic changes alone in the other 16 animals. Pasteurella multocida was isolated in pure culture from 219 of 248 samples of cerebrospinal fluid. Eighteen animals were observed moribund with severe depression, foamy nasal discharge, and respiratory distress, and 257 were found dead. Major clinical signs and pathologic changes included extensive swelling of the head and the neck and peracute or acute septic pneumonia, petechial and ecchymotic hemorrhages on serous membranes, and severely hemorrhagic adrenal glands and abomasum. Rhinitis and necrotic pharyngeal mucosae were common. Histologically, the most advanced lesions were in the nasal mucosa and pharynx. The swelling of the head and the neck arose from a diffuse cellulitis in the subcutaneous and intermuscular tissues. The earliest lesions in the lungs included large numbers of bacteria in the pulmonary capillaries, but various degrees of fibrinous exudation to the alveoli and infiltration with heterophils usually were observed.     

Fecal steroid analysis for evaluating ovarian function in the greater kudu (Tragelaphus strepsiceros) and lesser kudu (Tragelaphus imberbis)

Seasonal reproductive-endocrine norms have not been described for the genus Tragelaphus, which consists of seven species of African antelope. Longitudinal patterns of progesterone metabolite excretion were assessed by radioimmunoassays in fecal samples collected noninvasively (three to seven samples per week) from greater kudu (Tragelaphus strepsiceros, n = 4) and lesser kudu (Tragelaphus imberbis, n = 4). Progesterone metabolite excretion patterns revealed seasonal estrous cycles in both species, and discrimination of pregnant versus nonpregnant females was achieved in lesser kudu. These data reveal the value of fecal progesterone metabolites for establishing reproductive-endocrine norms in both zoo-maintained and free-living antelopes of the genus Tragelaphus.     

Assessment of diclofenac LC50 reference values in juvenile and embryonic stages of the zebrafish (Danio rerio)

The aim of the study was to compare the acute toxicity of diclofenac to juvenile and embryonic stages of the zebrafish (Danio rerio). Acute toxicity tests were performed on the aquarium fish Danio rerio, which is one of the model organisms most commonly used in toxicity testing. The tests were performed using a semi-static method according to OECD guideline No. 203 (Fish, acute toxicity test). Embryo toxicity tests were performed in zebrafish embryos (Danio rerio) in compliance with OECD No. 212 methodology (Fish, short-term toxicity test on embryo and sac-fry stages). The results were subjected to a probit analysis using the EKO-TOX 5.2 programme to determine 96hLC50 and 144hLC50 (median lethal concentration, 50% mortality after a 96 h or 144 h interval, respectively) values of diclofenac. The statistical significance of the difference between LC50 values in juvenile and embryonic stages of Danio rerio was tested using the Mann-Whitney non-parametric test implemented in the Unistat 5.1 programme. The LC50 mean value of diclofenac was 166.6 +/- 9.8 mg/L in juvenile Danio rerio, and 6.11 +/- 2.48 mg/L in embryonic stages of Danio rerio. The study demonstrated a statistically higher sensitivity to diclofenac (P < 0.05) in embryonic stages compared to the juvenile fish.     

Serum chemistry comparisons between captive and free-ranging giraffes (Giraffa camelopardalis)

Serum chemistry analyses were compared between captive and free-ranging giraffes (Giraffa camelopardalis) in an attempt to better understand some of the medical issues seen with captive giraffes. Illnesses, including peracute mortality, energy malnutrition, pancreatic disease, urolithiasis, hoof disease, and severe intestinal parasitism, may be related to zoo nutrition and management issues. Serum samples were collected from 20 captive giraffes at 10 United States institutions. Thirteen of the captive animal samples were collected from animals trained for blood collection; seven were banked samples obtained from a previous serum collection. These samples were compared with serum samples collected from 24 free-ranging giraffes in South Africa. Differences between captive and free-ranging giraffes, males and females, and adults and subadults were analyzed by using a 2 x 2 x 2 factorial and Fisher's least significant difference for mean separation; when necessary variables were ranked and analyzed via analysis of variance. Potassium and bilirubin concentrations and alanine aminotransferase (ALT) activities were different between captive and free-ranging giraffes, but all fell within normal bovid reference ranges. The average glucose concentration was significantly elevated in free-ranging giraffes (161 mg/dl) compared with captive giraffes (113 mg/dl). All giraffes in this study had glucose concentrations higher than bovine (42-75 mg/ dl) and caprine (48-76 mg/dl) reference ranges. Differences were also seen in lipase, chloride, and magnesium though these findings are likely not clinically significant. There were no differences detected between sexes. Adults had higher concentrations of potassium, total protein, globulins, and chloride and higher gamma glutamyltransferase activities, whereas subadults had higher concentrations of phosphorus. Within the captive group, nonimmobilized animals had higher concentrations of total protein and globulins. Captive giraffe diets need further investigation to determine if the differences seen in this study, especially glucose and bilirubin concentrations and ALT activities, may result in some health problems often seen in captive giraffes.     

Additivity of effects from dietary copper and zinc on growth performance and fecal microbiota of pigs after weaning

Four experiments were conducted to determine the interactive effects of pharmacological amounts of Zn from ZnO and Cu from organic (Cu-AA complex; Cu-AA) or inorganic (CuSO(4)) sources on growth performance of weanling pigs. The Cu was fed for 4 (Exp. 1) or 6 (Exp. 2, 3, and 4) wk after weaning, and Zn was fed for 4 (Exp. 1) or 2 (Exp. 2, 3, and 4) wk after weaning. Treatments were replicated with 7 pens of 5 or 6 pigs per pen (19.0 ± 1.4 d of age and 5.8 ± 0.4 kg of BW, Exp. 1), 12 pens of 21 pigs per pen (about 21 d of age and 5.3 kg of BW, Exp. 2), 5 pens of 4 pigs per pen (20.3 ± 0.5 d of age and 7.0 ± 0.5 kg of BW, Exp. 3), and 16 pens of 21 pigs per pen (about 21 d of age and 5.7 kg of BW, Exp. 4). In Exp. 1 and 2, Cu-AA (0 vs. 100 mg/kg of Cu) and ZnO (0 vs. 3,000 mg/kg of Zn) were used in a 2 × 2 factorial arrangement. Only Exp. 1 used in-feed antibiotic (165 mg of oxytetracycline and 116 mg of neomycin per kilogram feed), and Exp. 2 was conducted at a commercial farm. In Exp. 3, sources of Cu (none; CuSO(4) at 250 mg/kg of Cu; and Cu-AA at 100 mg/kg of Cu) and ZnO (0 vs. 3,000 mg/kg of Zn) were used in a 3 × 2 factorial arrangement. In Exp. 4, treatments were no additional Cu, CuSO(4) at 315 mg/kg of Cu, or Cu-AA at 100 mg/kg of Cu to a diet supplemented with 3,000 mg/kg of Zn from ZnO and in-feed antibiotic (55 mg of carbadox per kilogram of feed). In Exp. 1 and 2, both Zn and Cu-AA improved (P < 0.001 to P = 0.03) ADG and ADFI. No interactions were observed, except in wk 1 of Exp. 2, where Zn increased the G:F only in the absence of Cu-AA (Cu-AA × Zn, P = 0.04). A naturally occurring colibacillosis diarrhea outbreak occurred during this experiment. The ZnO addition reduced (P < 0.001) the number of pigs removed and pig-days on antibiotic therapy. In Exp 3, ADFI in wk 2 was improved by Zn and Cu (P < 0.001 and P = 0.09, respectively) with no interactions. In wk 1, G:F was reduced by ZnO only in the absence of Cu (Cu × Zn, P = 0.03). Feeding Zn decreased fecal microbiota diversity in the presence of CuSO(4) but increased it in the presence of Cu-AA (Cu source × Zn, P = 0.06). In Exp. 4, Cu supplementation improved the overall ADG (P = 0.002) and G:F (P < 0.001). The CuSO(4) effect on G:F was greater (P < 0.001) than the Cu-AA effect. Our results indicate that pharmacological amounts of ZnO and Cu (Cu-AA or CuSO(4)) are additive in promoting growth of pigs after weaning.     

Ciclosporin A therapy is associated with disturbances in glucose metabolism in dogs with atopic dermatitis

The effect of ciclosporin A (CsA) on glucose homeostasis was investigated in 16 dogs with atopic dermatitis by determining plasma glucose, serum fructosamine and insulin concentrations, and serial insulin and glucose concentrations following a glucagon stimulation test, before and 6 weeks after CsA therapy at 5 mg/kg once daily. All dogs completed the study. Following CsA treatment, the median serum fructosamine concentrations were significantly higher (pretreatment 227.5 μmol/L; post-treatment 246.5 μmol/L; P = 0.001; reference range 162-310 μmol/L). Based on analyses of the areas under concentration-time curves (AUC) pre- and post-CsA treatment, plasma glucose concentrations were significantly higher (AUC without baseline correction 31.0 mmol/L/min greater; P = 0.021) and serum insulin concentrations were significantly lower (AUC without baseline correction 217.1 μIU/mL/min lower; P = 0.044) following CsA treatment. Peak glucose concentrations after glucagon stimulation test were significantly higher following CsA treatment (10.75 versus 12.05 mmol/L; P = 0.021), but there was no significant difference in peak serum insulin (52.0 versus 35.0 μIU/mL; P = 0.052). There was a negative correlation between baseline uncorrected insulin AUC and trough serum log CsA concentrations (r = -0.70, P = 0.005). The administration of CsA to dogs with atopic dermatitis leads to disturbances in glucose homeostasis. The clinical significance of this is unclear, but it should be taken into account when considering CsA treatment in dogs that already have such impairments.     

Imputation of missing single nucleotide polymorphism genotypes using a multivariate mixed model framework

The objective of this paper was to investigate, for various scenarios at low and high marker density, the accuracy of imputing genotypes when using a multivariate mixed model framework using information from 2, 4, or 10 surrounding markers. This model predicts genotypes at a locus, using genotypes at nearby loci as correlated traits, and the additive genetic relationship matrix to use information from genotyped relatives. For 2 scenarios this method was compared with the population-based imputation algorithms FastPHASE and Beagle. Accuracies of imputation were obtained with Monte Carlo simulation and predicted with selection index theory, using input from the simulated data. Five different scenarios of missing genotypes were considered: 1) genotypes of some loci are missing due to genotyping errors, 2) juvenile selection candidates are genotyped using a smaller SNP panel, 3) some animals in the pedigree of a breeding population are not genotyped, 4) juvenile selection candidates are not genotyped, and 5) 1 generation of animals in the top of the pedigree are not genotyped. Surrounding marker information did not improve accuracy of imputation when animals whose genotypes were imputed were not genotyped for those surrounding markers. When those animals were genotyped for surrounding markers, results indicated a limited gain when linkage disequilibrium (LD) between SNP was low, but a substantial increase in accuracy when LD between SNP was high. For scenario 1, using 1 vs. 11 SNP, accuracy was respectively 0.75 and 0.81 at low, and 0.75 and 0.93 at high density. For scenario 2, using 1 vs. 11 SNP, accuracy was, respectively, 0.70 and 0.73 at low, and 0.71 and 0.84 at high density. Beagle outperformed the other methods at high SNP density, whereas the multivariate mixed model was clearly superior when SNP density was low and animals where genotyped with a reduced SNP panel. The results showed that extending the univariate gene content method to a multivariate BLUP model with inclusion of surrounding marker information only yields greater imputation accuracy when the animals with imputed loci are at least genotyped for some SNP that are in LD with the SNP to be imputed. The equation derived from selection index theory accurately predicted the accuracy of imputation using the multivariate mixed model framework.