Effect of Overnight Staining on the Quality of Flow Cytometric Sorted Stallion Sperm: Comparison with Tradtitional Protocols

Flow cytometry is considered the only reliable method for the separation of X and Y chromosome bearing spermatozoa in equines. The MoFlo SX DP sorter is highly efficient, allowing the production of foals of the desired sex. However, to achieve acceptable pregnancy rates the currently used protocol requires working with fresh semen obtained close to, or at, the sorting facility. An alternative protocol was tested during two consecutive breeding seasons. Fresh stallion semen was cooled for 20 h, during which staining with Hoechst 33342 took place. On the following day, this sample was flow sorted and compared with spermatozoa from the same ejaculate that had been sexed on the previous day. All sperm parameters evaluated remained unchanged when fresh sorted and refrigerated sorted semen were compared. Pre‐sorting storage at 5°C did not alter sperm velocities nor kinetics, viability or membrane permeability, production of reactive oxygen species, mitochondrial membrane potential or DNA fragmentation index of the sorted sample. The findings open for the possibility of using semen from stallions housed far from the sorting facilities. Processed and stained sperm could be shipped refrigerated on the previous day, sorted and inseminated on the next day.     

An ethogram of post-anesthetic recovery behaviors in horses: comparison of pre- and post-anesthetic behaviors

Pain management is an important post-operative concern. Pain scales may rely on the observer's subjective assessment of the level of discomfort and may not correlate with physiologic or pharmacologic measures of pain. The purpose of this study was to develop an objective measure of behavior in healthy pain-free horses recovering from anesthesia that could be used for comparison with the behavior of horses recovering from a surgical procedure.
Focal sampling with videotape and observation was done on five healthy horses before anesthesia to establish baselines. Behavioral measures included head turns, tail swishes, eyelid aperture/size, ear position, angle of neck, weight shifts, and ambulation. Physiologic measures included heart rate, respiratory rate, and temperature. The horses were anesthetized for 2 hours with isoflurane, and in the recovery stall, data were collected continuously on videotape from the time of extubation to standing. Focal sampling of 15 minutes was repeated at 1, 2, 4, and 24 hours after the horses returned to their stalls. Video data were analyzed using the Noldus Observer Video Analysis System.
A wide variation in behavior was observed between horses in the recovery stall. Observations at 1, 2, and 4 hours revealed change from the up/forward ear position to the down ear position, and a decrease in weight bearing of the hind limb from a baseline of 11–12 minutes out of 15 minutes to 7–9 minutes with an increase in toe pointing. Time spent standing was similar to baseline of 93.8%, and the neck angle was not changed from baseline of 173.5°. Values had returned to baseline at 24 hours.
Changes in behavior were induced by anesthesia alone and must be taken into consideration when evaluating analgesic treatments.     

Quantification of transmission of foot-and-mouth disease virus caused by an environment contaminated with secretions and excretions from infected calves

Foot-and-mouth disease virus (FMDV) infected animals can contaminate the environment with their secretions and excretions. To quantify the contribution of a contaminated environment to the transmission of FMDV, this study used calves that were not vaccinated and calves that were vaccinated 1 week prior to inoculation with the virus in direct and indirect contact experiments. In direct contact experiments, contact calves were exposed to inoculated calves in the same room. In indirect contact experiments, contact calves were housed in rooms that previously had held inoculated calves for three days (either from 0 to 3 or from 3 to 6 days post inoculation). Secretions and excretions from all calves were tested for the presence of FMDV by virus isolation; the results were used to quantify FMDV transmission. This was done using a generalized linear model based on a 2 route (2R, i.e. direct contact and environment) SIR model that included information on FMDV survival in the environment. The study shows that roughly 44% of transmission occurs via the environment, as indicated by the reproduction ratio R^02Renvironment that equalled 2.0, whereas the sum of R^02Rcontact and R^02Renvironment equalled 4.6. Because vaccination 1 week prior to inoculation of the calves conferred protective immunity against FMDV infection, no transmission rate parameters could be estimated from the experiments with vaccinated calves. We conclude that a contaminated environment contributes considerably to the transmission of FMDV therefore that hygiene measures can play a crucial role in FMD control.

Electronic supplementary material

The online version of this article (doi:10.1186/s13567-015-0156-5) contains supplementary material, which is available to authorized users.     

Cholelithiasis and hyperthyroidism in a cat

A 14-year-old domestic short-hair cat presented with a history of intermittent malaise and increased drinking. A diagnosis of hyperthyroidism and cholelithiasis was made by a combination of blood testing, radiography and ultrasonography. After medical management of hyperthyroidism, thyroidectomy and cholecystectomy were successfully performed. Removed choleliths were comprised of calcium carbonate and bilirubinate. Histopathological analysis of tissue suggested low grade pancreatic and hepatobiliary disease, as well as hyperthyroidism, might have contributed to stone formation.     

Acceptance by stoats (Mustela erminea) of 1080 (sodium monofluoroacetate) in small-volume baits and its effect on behaviour and time to death

AIMS: To assess whether stoats (Mustela erminea) would eat small baits containing 0.1% sodium monofluoroacetate (1080); whether they would die from it; how long it would take to kill them; and to document the behaviour of 1080-intoxicated stoats.

METHODS: Stoats were offered 1-g baits of two semi-fluid formulations containing 0.1% 1080, presented in open dishes, and their subsequent behaviour was monitored by video and direct observation. Muscle samples from stoats that died were analysed for 1080 residues.

RESULTS: There was no significant difference between two types of bait with regard to acceptance, mortality, and time to death, and behavioural effects were similar; consequently, results from the two types of bait were combined. Twelve of 14 stoats offered the baits ate them voluntarily, and a 13th licked bait off its fur; all 13 died between 1 h 15 min and 4 h 7 min (mean 2 h 38 min) later. At first (range 29 min — 2 h 7 min, mean 1 h 1 min), their behaviour appeared to be normal. Ataxia and hyperactivity were the first behavioural signs of poisoning, and lasted 2 min — 1 h 40 min (mean 26 min). This was followed by recumbency with convulsions and rapid breathing (range 16 min to 2 h, mean 58 min), then recumbency with limited activity and progressively shallow breathing prior to death (range 1–51 min, mean 33 min). Stoats became non-responsive to a light being turned on, or to touch once recumbency became sustained.

Residues of 1080 were found in muscle tissue of all 13 dead stoats, at concentrations ranging from 0.075 μg/g in a 287-g male that died 4 h 7 min after eating only 0.74 g of bait, to 2.5 μg/g in a 254-g female that died 2 h 42 min after taking a whole 1-g bait.

CONCLUSION: Stoats will voluntarily take small (1-g) baits containing a lethal dose of 1080 at 0.1%, and die from it comparatively rapidly for a mammalian carnivore.     

The effect of different combinations of lignocaine,ketoprofen, xylazine and tolazoline on the acute cortisol response to dehorning in calves

AIMS: The aims of this study were (a) to evaluate the effect of xylazine and tolazoline, with and without lignocaine, on the cortisol response of calves following amputation dehorning and (b) to assess the effect of a non-steroidal anti-inflammatory drug (ketoprofen) and local anaesthesia on the cortisol response of calves to amputation dehorning.

METHODS: Plasma cortisol concentrations were measured in 100 dehorned or non-dehorned 3-month-old calves over an 8-h period following five different sedative/analgesic or control treatments. Sedative/analgesic treatments were: control (no anaesthesia); local anaesthesia and ketoprofen; local anaesthesia and xylazine; local anaesthesia, xylazine and tolazoline; and xylazine only. Within each sedative/analgesic treatment group, half the calves (n=10 per group) were amputation dehorned and half were not dehorned.

RESULTS: The change in plasma cortisol concentrations in calves dehorned after being given ketoprofen and local anaesthesia did not differ significantly from that of non-dehorned control calves for at least 8 h. In contrast, the cortisol response of dehorned calves not given analgesic drugs peaked 30 min after dehorning and lasted >4 h. Xylazine injected before dehorning significantly reduced but did not eliminate the peak of the cortisol response. When both xylazine and local anaesthesia were administered before dehorning the peak in the cortisol response was virtually eliminated. In the dehorned calves that received xylazine with or without local anaesthesia, cortisol concentration increased significantly 3 h after dehorning and did not return to baseline until at least 5 h later. When tolazoline was administered shortly after xylazine, it caused a marked cortisol response, higher than the response to any other treatment.

CONCLUSIONS: Combining ketoprofen and local anaesthesia minimised the cortisol response, and by inference the pain- induced distress, following amputation dehorning in calves. Xylazine reduced the initial cortisol response to dehorning but not as much as when local anaesthesia was also given. The increase in cortisol concentration from 3–8 h after dehorning in calves given xylazine alone or in combination with local anaesthesia suggests that calves experienced pain-induced distress during this time and that xylazine had no long-term analgesic effect. Tolazoline, used to reverse the sedative effects of xylazine, caused a marked cortisol response in calves via a mechanism which remains unclear.     

Systemic protozoal disease in hihi and saddlebacks

Extract

Juvenile hihi or stitchbirds (Notiomystis cincta) and saddlebacks (Philestumus carunculatus) are particularly prone to systemic protozoal infections when kept in high density populations. Granu-lomatous lesions in the intestine, liver and spleen are associated with proliferating intracellular schizonts. Affected birds become anorexic and lose weight or die suddenly without clinical signs. The protozoan in hihi appears most likely to be Atoxoplasma spp whereas that in saddlebacks is more like Plasmodium spp.