Functional Role of Feline Zona Pellucida Protein 4 Trefoil Domain: A Sperm Receptor or Structural Component of the Domestic Cat Zona Pellucida?

The trefoil domain (TFD) is a protein structure characterized by six cysteines, which form a typical three-loop structure by three disulphide bridges. It is assumed that two of these loops generate a hydrophobic groove, which could be a binding site for carbohydrate residues or proteins. The zona pellucida (ZP) protein, ZP4, contains such a TFD. The carbohydrate-/protein-binding property of TFD allows us to assume a potential sperm receptor function of this domain. Additionally, gastrointestinal trefoil peptides are stable against proteases; therefore, a structural role of TFD within the ZP might also be possible. We were able to show that the synthesized and natural folded feline TFD (fTFD) expresses the typical protease resistance that vanished under reducing conditions and after substitution of cysteine residues within the peptide. Furthermore, an antibody directed against the first loop of fTFD was almost unable to bind to intact in vitro mature cat oocytes. Pre-incubation of oocytes in the reducing agent (DDT), however, improved antibody binding substantially. Therefore, we suggest structural masking of the fTFD domain within the intact ZP. An interaction between fTFD and feline sperm cells was examined using several methods, including immunocytochemistry, immunoelectron microscopy, co-immunoprecipitation and far western blot, but we found no indication for an involvement of TFD in the primary sperm binding to the ZP. To summarize, there is increasing evidence that the TFD of fZP4 has a structural rather than a sperm-binding function.     

Pharmacokinetics of pradofloxacin and doxycycline in serum, saliva, and tear fluid of cats after oral administration

The pharmacokinetic properties of pradofloxacin and doxycycline were investigated in serum, saliva, and tear fluid of cats. In a crossover study design, six cats were treated orally with a single dose of pradofloxacin (Veraflox^® Oral Suspension 2.5%) and doxycycline (Ronaxan^® 100 mg) at 5 mg/kg body weight. Following administration, samples of serum, saliva, and tear fluid were taken in regular intervals over a period of 24 h and analysed by turbulent flow chromatography/tandem mass spectrometry. All values are given as mean ± SD. Pradofloxacin reached a mean maximum serum concentration ( C _max) of 1.1 ± 0.5 μg/mL after 1.8 ± 1.3 h ( t _max). In saliva and tear fluid, mean C _max was 6.3 ± 7.0 and 13.4 ± 20.9 μg/mL, respectively, and mean t _max was 0.5 ± 0 and 0.8 ± 0.3 h, respectively. Doxycycline reached a mean C _max in serum of 4.0 ± 0.8 μg/mL after 4.3 ± 3.2 h. Whilst only at two time-points doxycycline concentrations close to the limit of quantification were determined in tear fluid, no detectable levels were found in saliva. The high concentrations of pradofloxacin in saliva and tear fluid are promising to apply pradofloxacin for the treatment of conjunctivitis and upper respiratory tract infections in cats. As doxycycline is barely secreted into these fluids after oral application the mechanisms of its clinical efficacy remain unclear.     

Carbon in Amazon forests: unexpected seasonal fluxes and disturbance-induced losses

The net ecosystem exchange of carbon dioxide was measured by eddy covariance methods for 3 years in two old-growth forest sites near Santarém, Brazil. Carbon was lost in the wet season and gained in the dry season, which was opposite to the seasonal cycles of both tree growth and model predictions. The 3-year average carbon loss was 1.3 (confidence interval: 0.0 to 2.0) megagrams of carbon per hectare per year. Biometric observations confirmed the net loss but imply that it is a transient effect of recent disturbance superimposed on long-term balance. Given that episodic disturbances are characteristic of old-growth forests, it is likely that carbon sequestration is lower than has been inferred from recent eddy covariance studies at undisturbed sites.     

Surface Deformation and Lower Crustal Flow in Eastern Tibet

Field observations and satellite geodesy indicate that little crustal shortening has occurred along the central to southern margin of the eastern Tibetan plateau since about 4 million years ago. Instead, central eastern Tibet has been nearly stationary relative to southeastern China, southeastern Tibet has rotated clockwise without major crustal shortening, and the crust along portions of the eastern plateau margin has been extended. Modeling suggests that these phenomena are the result of continental convergence where the lower crust is so weak that upper crustal deformation is decoupled from the motion of the underlying mantle. This model also predicts east-west extension on the high plateau without convective removal of Tibetan lithosphere and without eastward movement of the crust east of the plateau.     

Managing anthelmintic resistance: Is it feasible in New Zealand to delay the emergence of resistance to a new anthelmintic class?

The recent registration in New Zealand of the first new class of broad-spectrum anthelmintic, for use against nematode parasites of ruminants, in nearly three decades has raised the possibility that parasite management practices could be improved to minimise the emergence of resistance to the new drug. A review of knowledge pertaining to the selection of anthelmintic resistance in nematode parasites of sheep highlights a number of management practices which could be altered to achieve this.

A number of previously common practices such as whole-flock treatment of adult ewes around lambing, and treatment of lambs as they are moved onto pastures with low parasite contamination have been clearly identified as high risk for selecting resistant parasites. Once high-risk practices have been identified steps can be taken to either eliminate their use or mitigate the associated risk. Much of the focus on the management of resistance around the world is on the retention of susceptible genotypes in refugia. While approaches to retaining unselected parasites are likely to vary around the world, empirical studies indicate that the practice is likely to be effective at slowing the development of resistance. The challenge for farmers and advisors will be to strike a balance between retaining sufficient susceptible para-sites to usefully delay the development of resistance while not unduly compromising animal performance and farm profitability. The merits of combining different classes of anthelmintic in order to slow the development of resistance remains somewhat contentious in some countries. However, the attributes of oral anthelmintics are such that they seem likely to meet most, if not all, of the criteria for combinations to be highly effective at slowing the build-up of resistance in nematode parasites.

It is evident that considerable progress has been made in understanding the factors involved in selecting anthelmintic-resistant nematodes since the last broad-spectrum anthelmintic class was released in the early 1980s. Therefore, it should be possible to manage a new class of anthelmintic in such a way as to significantly extend its effective life. The challenge is likely to be in convincing farmers of the merits of adopting such pro-active strategies.     

Post-vaccinal distemper encephalitis in two Border Collie cross littermates

Abstract

CASE HISTORY: One 4.5-month-old male Border Collie cross presented with aggression and seizures in October 2006. A 16-month-old, female, spayed Border Collie cross presented with hypersalivation and a dropped jaw and rapidly became stuporous in September 2007. The dogs were littermates and developed acute neurological signs 5 and 27 days, respectively, after vaccination with different modified live vaccines containing canine distemper virus.

HISTOPATHOLOGICAL FINDINGS: Sections of brain in both dogs showed evidence of encephalitis mainly centred on the grey matter of brainstem nuclei, where there was extensive and intense parenchymal and perivascular infiltration of histiocytes and lymphocytes. Intra-nuclear and intra-cytoplasmic inclusions typical of distemper were plentiful and there was abundant labelling for canine distemper virus using immunohistochemistry.

DIAGNOSIS: Post-vaccinal canine distemper.

CLINCIAL RELEVANCE: Post-vaccinal canine distemper has mainly been attributed to virulent vaccine virus, but it may also occur in dogs whose immunologic nature makes them susceptible to disease induced by a modified-live vaccine virus that is safe and protective for most dogs.     

Ovulation,Pregnancy Rate and Early Embryonic Development in Vernal Transitional Mares Treated with Equine‐ or Porcine‐FSH

The objective of this study was to compare the efficacy of purified equine‐ and porcine‐FSH treatment regimes in mares in early vernal transition. Mares (n = 22) kept under ambient light were examined ultrasonographically per‐rectum, starting January 30th. They were assigned to one of two treatment groups using a sequential alternating treatment design when a follicle ≥ 25 mm was detected. In the eFSH group, mares were treated twice daily with equine‐FSH, and in the pFSH group mares were treated twice daily with porcine‐FSH; treatments were continued until follicle(s) ≥ 35 mm, and 24 h later hCG was administered. Oestrous mares were inseminated with fresh semen and examined for pregnancy on days 11–20 post‐ovulation. In the eFSH group, 11/11 (100%) mares developed follicle(s) ≥ 35 mm, 8/11 (73%) ovulated and 6/8 (75%) conceived. In the pFSH group, 5/11 (45%) developed follicle(s) ≥ 35 mm, 4/11 (36%) ovulated and 3/4 (75%) conceived. Treatment with eFSH resulted in a greater ovarian stimulation; higher number of pre‐ovulatory‐sized follicles, higher number of ovulations and higher number of embryos (p < 0.05). Following ovulation, serum progesterone concentrations were correlated with the number of CLs and supported early embryonic development; maternal recognition of pregnancy occurred in all pregnant mares. We concluded that eFSH can be used to effectively induce follicular growth and ovulation in vernal transitional mares; however, if bred, diagnosis and management of twins’ pregnancies would be required prior to day 16 because of the increased risk of multiple embryos per pregnancy. Conversely, the current pFSH treatment regime cannot be recommended.     

Effect of Roscovitine‐Treated Donor Cells on Development of Porcine Cloned Embryos

Synchronization of the donor cell cycle is an important factor for successful animal cloning by nuclear transfer. To improve the efficiency of porcine cloning, in the present report, we evaluated effects of contact inhibition, serum starvation and roscovitine treatment of donor cells on in vitro and in vivo developmental potency of cloned porcine embryos. Fibroblasts derived from a porcine foetus at day 30 of gestation were isolated and cultured to 70% confluency. Then, cells were either cultured to 100% confluency for contact inhibition, or cultured in 0.5% serum for 72 h for serum starvation or with 15 μm roscovitine for 24 h. Cells were most effectively synchronized at G0/G1 in the serum starvation group (87.5%) compared with the contact inhibition and roscovitine treatment groups (76.3% and 79.9% respectively p < 0.05). However, after somatic cell nuclear transfer followed by in vitro culture, the serum starvation group showed a significantly lower blastocyst formation rate (5.6%) compared with the contact inhibition and roscovitine treatment groups (11.6% and 20.0% respectively). Differential expression of apoptosis‐related genes and the level of apoptosis in each treatment group explain the variation in developmental competence among the groups. Significantly higher level of apoptosis was observed in the serum starvation group. On the other hand, the roscovitine treatment group shows the lowest level of apoptosis and the best in vitro development among the groups. Cloned embryos derived from roscovitine‐treated donor cells were transferred to surrogate pigs. Three healthy live piglets were produced. In conclusion, we suggest that roscovitine treatment of donor cells improves development of cloned porcine embryos and can raise the efficiency of cloned piglet production.     

Sensitivity of boreal forest carbon balance to soil thaw

We used eddy covariance; gas-exchange chambers; radiocarbon analysis; wood, moss, and soil inventories; and laboratory incubations to measure the carbon balance of a 120-year-old black spruce forest in Manitoba, Canada. The site lost 0.3 +/- 0.5 metric ton of carbon per hectare per year (ton C ha-1 year-1) from 1994 to 1997, with a gain of 0.6 +/- 0.2 ton C ha-1 year-1 in moss and wood offset by a loss of 0.8 +/- 0.5 ton C ha-1 year-1 from the soil. The soil remained frozen most of the year, and the decomposition of organic matter in the soil increased 10-fold upon thawing. The stability of the soil carbon pool ( approximately 150 tons C ha-1) appears sensitive to the depth and duration of thaw, and climatic changes that promote thaw are likely to cause a net efflux of carbon dioxide from the site.