Motility Characteristics and Fertilizing Capacity of Boar Spermatozoa Stained with Hoechst 33342

Flow cytometry sorting of spermatozoa using fluorescence dye Hoechst 33342 is the only effective sex selection methodology validated in numerous laboratories. This study was carried out to determine the effect of Hoechst 33342 on the motility and fertility of stained boar spermatozoa. Experiment 1 evaluated motility parameters (percentage of motile spermatozoa, velocity, angularity and oscillation) of boar spermatozoa stained with Hoechst 33342 by a computer‐aided sperm analysis (CASA) instrument. Spermatozoa (30 million/ml) were divided into five treatment groups and stained during 1 h at 35°C with 9, 18, 27, 60 and 90 μM of H33342. There were no differences in sperm motility patterns nor percentages of motile spermatozoa incubated in the presence of 9, 18 or 27 μM. Percentage of motile spermatozoa and motility parameters decreased significantly (p < 0.05) at 60 μM of Hoechst 33342. Spermatozoa were immotile at concentration of 90 μM. In experiment 2, pregnancy rates, farrowing rates and litter size from sows (n = 275) artificially inseminated (AI) with either Hoechst 33342 stained (27 μM) or unstained (control) spermatozoa were determined. Sows inseminated with stained spermatozoa had no significant lower pregnancy rate (88.33%) as compared with controls (90.32%). Staining neither affected farrowing rates (85.0 vs 87.7%) nor total number of piglets born (10.56 ± 0.32 vs 10.47 ± 0.24, stained and controls, respectively). No phenotypical abnormalities were registered among the newborn piglets. The data suggest that incubating spermatozoa with Hoechst 33342 at levels required for X‐ and Y‐bearing chromosome sperm sorting, does not impair sperm viability or their fertility after AI.     

An overview of the topical management of wounds

Wounds in animals are a common and frequent reason for seeking veterinary attention. The way in which wounds are managed affect the rate of healing, the time to return to normal function, the final cosmetic appearance, and hence the satisfaction of customers. The management of wounds depends on the stage of wound healing and can include irrigation, mechanical and chemical debridement, the use of antiseptics and antimicrobials, adherent and nonadherent dressings, and miscellaneous topical applications such as aloe vera, honey and live yeast cell derivative. The advantages, disadvantages and indications for initial wound management, topical applicants and dressings are discussed.     

Application of high-resolution melt curve analysis for classification of infectious bronchitis viruses in field specimens

Objective A real-time polymerase chain reaction (PCR)/high-resolution melt (HRM) curve analysis protocol was developed in our laboratory to differentiate infectious bronchitis (IB) virus reference strains. In the current study, this method was used to detect and classify IB viruses in field submissions. Procedure Over an 11-month period samples from 40 cases of suspected IB virus were received and 17 submissions were positive for IB virus by polymerase chain reaction. HRM curve analysis classified each strain as subgroup 1, 2 or 3 strain (12 submissions) or a strain that was unable to be classified (5 submissions). The 3′ untranslated region (UTR) and partial S1 gene nucleotide sequences for the 17 IB virus strains were determined and their identity with those of the relative reference strains compared to confirm the classifications generated using the HRM curve analysis. Results Of the 12 IB field viruses classified as subgroup 1, 2, or 3 using HRM curve analysis, the 3′UTR and S1 gene nucleotide sequences had identities ≥99% with the respective subgroup reference strain. Analysis of the 3′ UTR and S1 gene nucleotide sequences for the five IB virus strains that could not be classified indicated that four belonged to one of the subgroups, and one was a potential recombinant strain (between strains from subgroups 2 and 3). A novel recombinant strain was also detected. Conclusion HRM curve analysis can rapidly assign the majority of IB viruses present in field submissions to known subgroups. Importantly, HRM curve analysis also identified variant genotypes that require further investigation.     

Disease signs reported in south-eastern Australian dairy cattle while grazing Brassica species

Objective
To estimate the relative frequency of various disease signs in dairy cattle fed brassicas in south-eastern Australia, to estimate the incidence of disease signs within affected groups and to identify risk factors for commonly reported signs in brassica-fed dairy cows.
Design
A case series study and a case-control study.
Procedures
Case data was collected using an incident reporting system. Herd managers and veterinarians recorded details about disease incidents in brassica-fed dairy cattle in summer and autumn 1995 using a standard questionnaire. Potential risk factors for photosensitisation were assessed using a case-control study.
Results
Disease signs were reported in 66 groups of brassica-fed dairy cattle. Photosensitisation and bloat were the more frequently reported signs. While high incidences were reported in some groups, the incidence of disease signs was low within most affected groups. Photosensitisation occurred more frequently among groups of cows fed brassica crops treated with nitrogenous fertilisers or which were low yielding. Risks of other disease signs were greatest while the first one quarter of the crop was grazed.
Conclusions
In the study population, most disease outbreaks occurring in brassica-fed cattle were of low incidence. However, some high incidence outbreaks occurred. Results from this study suggest that important risk factors exist for disease signs in brassica-fed dairy cattle. Further studies are required if these risk factors are to be fully identified. This would allow the development of preventative strategies for high incidence disease outbreaks while feeding brassicas to dairy cattle.     

Reference ranges for biochemical and haematological values in farmed saltwater crocodile (Crocodylus porosus) yearlings

Objective To determine reference ranges for healthy yearling farmed saltwater crocodiles by performing routine biochemical and haematological laboratory tests on blood samples.
Design A clinico-pathological study.
Procedure Blood samples were collected from 120 healthy yearlings from four Northern Territory crocodile farms and body weight and length were measured. All animals had been fasted for 2 days before sample collection. Routine biochemical analytes were determined on 120 samples and haematological values determined on 30 samples (from one farm).
Results Reference ranges for biochemical and haemato-logical values were determined for farmed yearling saltwater crocodiles in the Northern Territory.
Conclusion The results were comparable with published reference ranges for other crocodilian species. Other published results of haematological values from saltwater crocodiles were from very young (6-week-old) hatchlings and older (2- to 4-year-old) crocodiles. Differences in values were presumed to be caused by age-related factors.     

A screening test for subclinical liver disease in horses affected by pyrrolizidine alkaloid toxicosis

Objective: To evaluate various biochemical tests as indicators of subclinical liver disease in horses exposed to pyrrolizidine alkaloid toxicosis.
Design: A clinical pathology field study.
Animals: Twenty-two clinically normal horses from four properties in the Kimberley region of Western Australia.
Procedure: Serum samples from each horse were assayed for gamma glutamyltransferase, alkaline phosphatase and aspartate aminotransferase activities, and for serum bile acid concentration, albumin and total protein. Serum protein electrophoresis was performed and their amino acid profiles determined. Bromosulphophihalein halfclearance times were measured. Horses were then subjected to a single liver biopsy. Results were analysed by, variance of group means, the Fisher-Irwin exact test, and by sensitivity and specificity calculation.
Results: Horses were classified into 2 groups, of 10 unaffected and 12 subclinically affected, on the basis of liver histology. Significant differences between the unaffected and subclinical groups were observed for gamma glutamyltransferase and alkaline phosphatase activities (P < 0.01). Gamma glutamyltransferase had sufficient sensitivity (75%) and specificity (90%) to function as a primary screening test for subclinical liver disease in horses exposed to pyrrolizidine alkaloids. Alkaline phosphatase was useful, but with lower sensitivity (58%).
Conclusion: Serum gamma glutamyltransferase activity is a useful screening test for detecting subclinical liver disease in horses exposed to pyrrolizidine alkaloids under field conditions in northern Australia.     

Early Development and Putative Primordial Germ Cells Characterization in Dogs

Previously, three distinct populations of putative primordial germ cells (PGCs), namely gonocytes, intermediate cells and pre‐spermatogonia, have been described in the human foetal testis. According to our knowledge, these PGCs have not been studied in any other species. The aim of our study was to identify similar PGC populations in canine embryos. First, we develop a protocol for canine embryo isolation. Following our protocol, 15 canine embryos at 21–25 days of pregnancy were isolated by ovaryhysterectomy surgery. Our data indicate that dramatic changes occur in canine embryo development and PGCs specification between 21 to 25 days of gestation. At that moment, only two PGC populations with distinct morphology can be identified by histological analyses. Cell population 1 presented round nuclei with prominent nucleolus and a high nuclear to cytoplasm ratio, showing gonocyte morphology. Cell population 2 was often localized at the periphery of the testicular cords and presented typical features of PGC. Both germ cell populations were positively immunostained with anti‐human OCT‐4 antibody. However, at day 25, all cells of population 1 reacted positively with OCT‐4, whereas in population 2, fewer cells were positive for this marker. These two PGCs populations present morphological features similar to gonocytes and intermediate cells from human foetal testis. It is expected that a population of pre‐spermatogonia would be observed at later stages of canine foetus development. We also showed that anti‐human OCT‐4 antibody can be useful to identify canine PGC in vivo.