Comparison of efficiency of in vitro cloned sheep embryo production by conventional somatic cell nuclear transfer and handmade cloning technique

Conventional somatic cell nuclear transfer (SCNT) technique of in vitro production of cloned embryos involves use of costly and complicated micromanipulators. Handmade cloning (HMC) technique has been applied as efficient and cost‐effective alternative in many livestock species. The aim of the present study was to compare the efficiency of in vitro production and in vitro development of cloned sheep embryos by the two techniques. Cloned embryos were produced by conventional SCNT using micromanipulator apparatus and by HMC technique. Enucleation efficiency and efficiency of fusion with somatic cell (nucleus donor) were compared. Cleavage percentage was observed on day 2 of in vitro culture (IVC), and morula and blastocyst percentages were calculated on day 7 of IVC. Higher enucleation efficiency (96.98 ± 1.01 vs. 93.62 ± 1.03; p > .05) as well as fusion efficiency was obtained with HMC technique than with conventional SCNT (96.26 ± 1.34 vs. 92.63 ± 0.70, p < .05); 181 cloned sheep embryos were produced in vitro by conventional SCNT and 92 by HMC. Cleavage percentage observed on day 2 of in vitro culture was higher in HMC than SCNT (66.92 ± 3.72 vs. 55.97 ± 2.5, respectively, p < .05). Morula percentage obtained was higher in SCNT than HMC (44.12 ± 2.93 vs. 30.43 ± 6.79, respectively, p < .05), whereas blastocyst percentage obtained by HMC was higher (12.46 ± 4.96) than SCNT (5.31 ± 2.25; p > .05). It was inferred that HMC technique provides a cost‐effective and efficient method of in vitro production of cloned sheep embryos with a comparatively simpler technique with a possibility of automation. Efficiency of cloned embryo production could be improved further by propagating and standardizing this technique.     

Changes in plasma concentrations of S100A7 and S100A8 in dairy cows during pregnancy

This study was carried out to examine the changes in plasma concentrations of the Ca‐binding antimicrobial proteins S100A7 and S100A8 during pregnancy in dairy cows. Holstein Friesian cows (n = 19) were inseminated with Holstein Friesian semen. Blood was collected at days 30, 60, 90, 120, 150, 180, 210, 240 and 270 after insemination. Plasma was used for measuring the concentrations of S100A7 and S100A8. Both S100A7 and S100A8 concentrations showed similar patterns during gestation; they increased during the midgestation, between days 90 and 180, and then declined before calving. The findings indicated that plasma concentrations of S100A7 and S100A8 did not change significantly during pregnancy in cows. Further studies are required to determine the roles of S100A7 and S100A8 in physiological function during pregnancy in dairy cows.     

Resynchronising returns to service in anoestrous dairy cows in the South Island of New Zealand

AIMS: To investigate the effect of targeted resynchronisation of cows treated for non-observed oestrus before the planned start of mating (PSM), that were not detected in oestrus or pregnant 23 days after treatment (phantom cows), on the proportion pregnant at 42 days after PSM and the end of mating.

METHODS: Farm staff from eight herds in two regions of the South Island of New Zealand identified 1,819 cows not showing oestrus by 10 days before PSM. These cows were treated with intravaginal progesterone for 7 days, and I/M gonadorelin 10 days and 1 day before PSM. Three days before PSM they were injected with cloprostenol and equine chorionic gonadotrophin, with fixed time artificial insemination (FTAI) at PSM. By 23 days after PSM, 1,218 cows had not returned to oestrus. Of these, 161 cows confirmed not pregnant by transrectal ultrasonography were randomly assigned to no treatment (control group; n=74) or were resynchronised 25 days after PSM using the same treatment programme as above, with FTAI 35 days after PSM (n=87). All cows that returned to oestrus were artificially inseminated until 42 days after PSM, when natural mating was used. All cows were examined using transrectal ultrasonography 80 to 90 days after PSM to confirm conception dates.

RESULTS: Of the 1,819 anoestrous cows treated before PSM, 526 (29 (95% CI=23.1–34.0)%) had not been observed in oestrus by 23 days after PSM and had not conceived, so were diagnosed as phantoms cows. For resynchronised cows, 42/87 (48 (95% CI=37.8–58.8)%) were pregnant by 42 days after PSM compared to 21/74 (28 (95% CI=18.1–38.7)%) control cows (p=0.009). At the end of mating 58/87 (67 (95% CI=56.6–76.7)%) cows in the resynchronised group were pregnant and 46/74 (62 (95% CI=50.9–73.2)%) in the control group (p=0.554). The hazard of conception from 21 to 42 days after PSM was 1.9 (95% CI=1.07–3.12) times greater for resynchronised than control cows (p=0.026).

CONCLUSION: In cows not observed in oestrus and treated before PSM, resynchronisation increased the proportion pregnant by 42 days after PSM.

CLINICAL RELEVANCE: The benefit of resynchronisation depends on the number of anoestrous cows before PSM and the number of phantom cows after PSM. However at the herd-level it is likely that providing advice to reduce the known risk factors for cows not being observed in oestrus before the PSM may well be more cost effective than identifying and treating a sub-population of phantom cows.     

Comparison of two topical treatments on wound healing 7 days after disbudding of calves using thermocautery

ABSTRACT

Aims To compare wound healing 7 days after cautery disbudding of dairy calves treated immediately after disbudding with a traditional antimicrobial spray (OXY) or with a topical anaesthetic gel containing cetrimide, adrenaline and two local anaesthetics, lignocaine and bupivacaine (TA).

Method Eighty-one female dairy calves between 6–8 weeks of age were disbudded using a standard cautery disbudding protocol (sedation, cornual block, and analgesia), with complete removal of the horn bud. After disbudding, the wound on the right and left horn buds within each animal were randomly allocated to receive either OXY or TA. One week after disbudding, wounds were visually assessed for the presence of exudate, necrotic tissue, crust, or granulation tissue which were each assigned a grade from 1–3, where 1?=?no evidence, 2?=?moderate presence; and 3?=?marked presence.

Results At 7 days after disbudding, the prevalence of wounds with exudate or necrotic tissue was very low, and independent of treatment. The odds of a disbudding wound treated with TA being scored as having granulation tissue and was 5.2 (95% CI?=?1.72–15.7) times that of a wound treated with OXY. Conversely, the odds of wounds treated with TA being scored as having crusts was 0.18 (95% CI?=?0.06–0.57) that of a wound treated with OXY. No sign of infection was seen in any calves.

Conclusion and clinical relevance The greater prevalence of granulation tissue and reduction in crusting in disbudding wounds treated with topical anaesthetic gel compared to those treated with the antimicrobial spray suggests that use of the topical anaesthetic gel may speed wound healing compared to using antimicrobial spray. This should be further tested in a larger study undertaken over a longer period.     

Suspected inherited rickets in Corriedale sheep

A suspected inherited form of rickets was diagnosed in Corriedale sheep on a commercial sheep farm in Marlborough. Affected lambs were normal at birth but showed a reduced growth rate and a range of skeletal abnormalities, including both varus and valgus deformities and a characteristic dip in the shoulder region. Up to 20 affected lambs had been born on the property over the last two seasons. Skeletal lesions in affected lambs included swollen costochondral junctions, moderate to marked thickening of physes in long bones, and collapse of weakened subchondral bone, particularly in the proximal humerus. These lesions, together with the microscopic changes, strongly suggested a diagnosis of rickets. Affected lambs were hypocalcaemic and hypophosphataemic, consistent with a deficiency of vitamin D.

Vitamin D assays revealed significantly higher 1,25(OH) vitamin D concentrations in affected lambs than in controls, suggesting a defect in the vitamin D receptor as the likely mechanism. DNA testing revealed that 4/10 Corriedale rams used during the previous season had sired affected lambs. Studies aimed at confirming the genetic nature of the disease and the mechanism are continuing at Massey University.     

Copper deficiency in sheep: An assessment of the relationship between concentrations of copper in serum and plasma

AIM: To assess the relationship between concentrations of copper in serum and plasma in sheep.

METHODS: Concentrations of Cu were measured in paired serum and heparinised plasma samples collected from 110 sheep in nine flocks. Linear regression was used to evaluate whether flock or gender had a significant effect on the association between concentrations of Cu in serum and plasma. The individual results for concentrations of Cu in serum were then compared with those from plasma, using correlation and limits of agreement plotting.

RESULTS: Concentrations of Cu in serum ranged from 7.3 to 22 (mean 14.0) µmol/L, while concentrations in plasma ranged from 9 to 27 (mean 16.3) µmol/L. On average, concentrations of Cu in serum were 2.3 µmol/L lower than in plasma. Over the range of values seen in this study, concentrations of Cu in plasma and serum were significantly correlated (r=0.89), and mean concentrations in serum were 87% of those in plasma. There was no effect of flock or gender on the relationship between concentrations of Cu in serum and plasma. Despite the significant correlation, there was marked variability between individual samples in the proportion of Cu that was lost during clotting, with the 95% limits of agreement for serum Cu ranging from 70 to 104% of the plasma concentration.

CONCLUSIONS: As in cattle, the individual variability in the loss of Cu during clotting in sheep is too great for concentration of Cu in serum to be used as a substitute for that in plasma.

CLINICAL RELEVANCE: When assessing the blood Cu pool as part of the diagnosis of Cu-responsive disease in sheep, the concentration of Cu in plasma should be measured in preference to that of serum. We suggest that a range of 4.5 to 9 µmol/L in plasma be used to define marginal Cu status in sheep.     

Direct Radiative Forcing by Smoke from Biomass Burning

Airborne measurements in smoke from biomass burning in Brazil have yielded optical parameters that permit an improved assessment of the effects of smoke on Earth's radiation balance. The global-mean direct radiative forcing due to smoke from biomass burning worldwide is estimated to be no more than about -0.3 watt per square meter (cooling), compared with +2.45 watts per square meter (warming) due to anthropogenic greenhouse gases. On regional scales, direct radiative forcing due to smoke can be large and might indirectly affect global climate.     

Simultaneous Measurement of Local Gain and Electron Density in X-ray Lasers

X-ray lasers (XRLs) have experimental average gains that are significantly less than calculated values and a persistently low level of spatial coherence. An XRL has been used both as an injected signal to a short XRL amplifier and as an interferometer beam to measure two-dimensional local gain and density profiles of the XRL plasma with a resolution near 1 micrometer. The measured local gain is in agreement with atomic models but is unexpectedly spatially inhomogeneous. This inhomogeneity is responsible for the low level of spatial coherence observed and helps explain the disparity between observed and simulated gains.