Vesicles Cytoplasmic Injection: An Efficient Technique to Produce Porcine Transgene‐Expressing Embryos

The use of vesicles co‐incubated with plasmids showed to improve the efficiency of cytoplasmic injection of transgenes in cattle. Here, this technique was tested as a simplified alternative for transgenes delivery in porcine zygotes. To this aim, cytoplasmic injection of the plasmid alone was compared to the injection with plasmids co‐incubated with vesicles both in diploid parthenogenic and IVF zygotes. The plasmid pcx‐egfp was injected circular (CP) at 3, 30 and 300 ng/μl and linear (LP) at 30 ng/μl. The experimental groups using parthenogenetic zygotes were as follows: CP naked at 3 ng/μl (N = 105), 30 ng/μl (N = 95) and 300 ng/μl (N = 65); Sham (N = 105); control not injected (N = 223); LP naked at 30 ng/μl (N = 78); LP vesicles (N = 115) and Sham vesicles (N = 59). For IVF zygotes: LP naked (N = 44) LP vesicles (N = 94), Sham (N = 59) and control (N = 79). Cleavage, blastocyst and GFP+ rates were analysed by Fisher's test (p < 0.05). The parthenogenic CP naked group showed lower cleavage respect to control (p < 0.05). The highest concentration of plasmids to allow development to blastocyst stage was 30 ng/μl. There were no differences in DNA fragmentation between groups. The parthenogenic LP naked group resulted in high GFP rates (46%) and also allowed the production of GFP blastocysts (33%). The cytoplasmic injection with LP vesicles into parthenogenic zygotes allowed 100% GFP blastocysts. Injected IVF showed higher cleavage rates than control (p < 0.05). In IVF zygotes, only the use of vesicles produced GFP blastocysts. The use of vesicles co‐incubated with plasmids improves the transgene expression efficiency for cytoplasmic injection in porcine zygotes and constitutes a simple technique for easy delivery of plasmids.     

Influence of Gonadotrophin‐Induced First Oestrus on Gilt Fertility

The aim of this study was to determine the association between the oestrous response of pre‐pubertal gilts to gonadotrophin injection or boar exposure and their subsequent farrowing rate and litter size. At 154 days of age, randomly selected pre‐pubertal gilts received an intramuscular injection of 400 IU equine chorionic gonadotrophin plus 200 IU human chorionic gonadotrophin (PG600^®; Merck Animal Health; n = 181). From the remaining pool of animals not treated with hormones, the first gilts showing signs of oestrus were selected to act as controls (n = 201). Boar exposure began at 155 days of age for both groups, and gilts were bred at a weight of approximately 130 kg. Comparisons were made between PG600^®‐treated gilts exhibiting oestrus or not within 7 days post‐injection (early and late responders, respectively) and control gilts exhibiting oestrus or not within 30 days after beginning of boar exposure (select and non‐select control gilts, respectively). By 162 days, oestrus was detected in 67.5% of PG600^®‐treated gilts compared with 5.7% of control gilts (p < 0.0001). The proportion of animals observed in oestrus at least three times before breeding was greater for select control gilts compared with early and late responder PG600^®‐treated gilts (p ≤ 0.001). There were no significant differences in farrowing rate and litter size between the four treatment groups. These data indicate that PG600^® is an effective tool to induce an earlier oestrus in gilts, that subsequent farrowing rate and born alive litter size compare favourably to that of select gilts and that gilts failing to respond promptly to hormonal stimulation do not exhibit compromised fertility.     

Off-Resonance Conduction Through Atomic Wires

The electrical resistance of wires consisting of either a single xenon atom or two xenon atoms in series was measured and calculated on the basis of an atom-jellium model. Both the measurement and the calculation yielded a resistance of 10(5) ohms for the single-xenon atom system and 10(7) ohms for the two-xenon atom system. These resistances greatly exceeded the 12,900-ohm resistance of an ideal one-dimensional conduction channel because conduction through the xenon atoms occurs through the tail of the xenon 6s resonance, which lies far above the Fermi level. This conduction process in an atom-sized system can now be understood in terms of the electronic states of individual atoms.     

Inherited chondrodysplasia in Texel sheep

CASE HISTORY: A skeletal disease characterised by dwarfism, limb deformity and sometimes sudden death occurred over a period of 5 years in lambs born on a commercial sheep farm in Southland. The disease showed variable expression and occurred in crossbred sheep. A genetic aetiology was supported by the birth of affected lambs over two seasons in a flock of putative carrier and affected sheep transported to Massey University.

CLINICAL FINDINGS: Affected lambs appeared normal at birth but showed evidence of dwarfism, wide-based stance and exercise intolerance as early as 1 week of age. Most died within the first 3 months of life, often after developing bilateral varus deformity of the forelimbs. Some severely-affected lambs died suddenly of respiratory embarrassment, probably due to tracheal collapse. Mildly-affected individuals had a short, blocky stature and some survived to breeding age.

PATHOLOGICAL FINDINGS: Gross and microscopic lesions of variable severity were present in the tracheal, articular, epiphyseal and physeal cartilages. In severe cases, articular cartilage in major joints was eroded from weight-bearing surfaces. The trachea was flaccid, abnormally kinked, and had thickened cartilaginous rings and a narrow lumen. Affected sheep that survived to breeding age eventually developed severe degenerative joint disease. Histologically, chondrocytes were disorganised, surrounded by concentric rings of abnormal fibrillar material, and the matrix often contained focal to coalescing areas of chondrolysis.

DIAGNOSIS: Inherited chondrodysplasia of Texel sheep.

CLINICAL RELEVANCE AND CONCLUSIONS: This chondrodysplasia differs from those previously described in sheep and is considered to be a newly-recognised, recessively-inherited genetic disease of the Texel breed. A defect in the synthesis of glycosaminoglycans in cartilage matrix is suspected. This disease of sheep may provide a suitable model for studying various forms of therapy for human chondrodysplasias.     

The effect of progesterone supplementation post mating and shearing of ewes in early pregnancy on the reproductive performance of ewes and birthweight of lambs

AIM: To determine whether short-term progesterone supplementation post mating or shearing of ewes in early pregnancy affected either the proportion of ewes that lambed or that had multiple lambs, or the birthweight of lambs.

METHODS: Romney ewes (n=457) were synchronised in oestrus using controlled internal drug-releasing (CIDR) devices containing progesterone, and mated to Romney rams over a 5-day period. The mid-point of mating (Day 0) occurred 2 days after the withdrawal of CIDR devices. Ewes mated (n=397) were randomly allocated to one of four treatment groups: shearing at Day 5, shearing at Day 30, no shearing, and no shearing plus progesterone supplementation using a CIDR device inserted on Day 3 for 6 days. During the period from Day 5 to Day 27, six harnessed Suffolk rams were placed with the ewes and matings recorded. At Day 48, all ewes that did not return to the Suffolk rams were scanned for pregnancy using ultrasound. At Day 140, single- and multiple-bearing ewes were set-stocked at 15.1 and 12.2 ewes/ha, respectively, and equivalent numbers of ewes from each treatment group were placed in each paddock. Blood samples from 10 unshorn and 10 progesterone-supplemented ewes were collected on Days 3, 6 and 9, and analysed for plasma progesterone concentrations. Lambs were identified to their dam and weighed within 12 h of birth, and again at 27 and 93 days after the mid-point of lambing. The ewes were weighed at regular intervals throughout the trial.

RESULTS: Plasma progesterone concentrations of supplemented ewes were higher than those of unsupplemented ewes (3.28 vs 1.75 ng/ml) on Day 6 (p=0.02) but not on Day 9 (4.58 vs 4.63 ng/ml). Treatment of ewes had no effect on either the proportion of ewes which lambed to the synchronised mating period or that had multiple lambs. Lambs born to ewes shorn at Day 30 tended (p=0.09) to be heavier at birth (by 0.28 kg) than those born to unshorn ewes but this effect was not evident when data were corrected for length of gestation. Neither shearing at Day 5 nor progesterone supplementation had any effect on the birthweight of lambs, and the treatment of ewes had no effect on the survival rate of lambs to weaning.

CONCLUSIONS: Progesterone supplementation for 6 days beginning 3 days post mating did not increase either the proportion of ewes that lambed or that had multiple lambs, or the birthweight of lambs. Shearing 5 days after mating had no significant effect on the reproductive performance of ewes and need not be avoided, but is unlikely to result in an increase in lamb birthweight. Shearing ewes at Day 30 may result in an increase in the birthweight of lambs but, ideally, ewes should be further advanced in pregnancy before shearing is undertaken.     

Re: Regional variations in the nematode worm populations of breeding ewes in New Zealand

Abstract

Extract

The paper by Hervé et al (2003) Hervé, M, McAnulty, RW, Logan, CM and Sykes, AR. 2003. Regional variations in the nematode worm populations of breeding ewes in New Zealand. New Zealand Veterinary Journal, 51: 159–164. [Taylor &; Francis Online], [Web of Science ®] , [Google Scholar] which appeared in the August 2003 issue of the New Zealand Veterinary Journal reported on regional differences in the relative proportions of nematode parasite species cultured from faecal samples collected from ewes during lactation.     

Farm management practices associated with macrocyclic lactone resistance on sheep farms in New Zealand

AIM: To identify farm practices associated with the presence of resistance to a macrocyclic lactone (ML) anthelmintic on sheep farms in New Zealand.

METHODS: A cross-sectional study was conducted to test for associations between the presence of resistance to an ML anthelmintic (ivermectin) and management practices on sheep farms in New Zealand. Selection of farms was both random (n=80) and purposive (n=32; being farms with a history of suspected ML resistance). Resistance was inferred from faecal nematode egg count (FEC) reduction (FECR) tests (FECRTs) when there was <95% reduction in FEC 7–10 days after treatment with a half dose of ivermectin (0.1 mg/kg). A logistic regression model was built to identify farm-level factors that were associated with the presence or absence of ML resistance.

RESULTS: Of the 112 flock managers that were approached for interview, 103 (92%) returned useable questionnaires. The odds of ML resistance were increased: on farms that had used long-acting ML products in ewes as a pre-lambing treatment for ≥3 of the previous 5 years (odds ratio (OR) = 7.2; 95% confidence interval (CI) = 1.7–30.3); on farms where <70% of the total stock units mid-winter were from sheep (OR=6.5; 95% CI=1.6–25.6); on farms which over the year purchased ≥10% of the number of sheep present mid-winter (OR=7.1; 95% CI=1.5–34.7); and on farms where the average wool diameter of the main flock was <37 (OR=4.1; 95% CI=1.1–14.7) microns. The model provided a good fit to the data (pseudo R²=0.64; Hosmer-Lemeshow statistic = 0.38).

CONCLUSIONS: Explanatory factors identified as associated with the presence of ML (ivermectin) resistance on farms included the use of long-acting anthelmintic formulations in ewes pre-lambing, sources of refugia of unselected parasites on the farm, breed of sheep and their requirements for anthelmintic treatments, and the importing of resistant parasites with purchased stock. The study provides support for controls that aim to provide refugia of susceptible worms and that minimise the risk of introduction of resistance through effective quarantine-drenching.     

Prevalence of anthelmintic resistance on 62 beef cattle farms in the North Island of New Zealand

AIM: To establish the prevalence of anthelmintic resistance in parasitic nematodes on a random sample of beef cattle herds in the North Island of New Zealand.

METHODS: A cross-sectional prevalence study was conducted using a standardised faecal nematode egg count (FEC) reduction (FECR) test (FECRT) for ivermectin, levamisole and albendazole on 60 calves on each of 62 farms in the North Island chosen at random from farms that conformed with the selection criteria. Resistance to an anthelmintic was inferred when there was <95% reduction in FEC 7-10 days after treatment. Larval cultures were performed for all control groups and for treated groups for which resistance was evident.

RESULTS: Of the farms that completed the FECRT, 4/61 (7%) showed ≥95% reduction in FEC for all anthelmintics tested. Resistance to ivermectin was evident on 56/61 (92%) farms, to albendazole on 47/62 (76%) farms, and to both ivermectin and albendazole on 45/61 (74%) farms. Resistance to levamisole was evident on only 4/62 (6%) farms. The parasites most prevalent in resistant populations cultured were Cooperia spp. On 45/61 (74%) farms where Cooperia spp were present in suffi cient numbers, resistance to both ivermectin and albendazole was evident. No cases of levamisole-resistant Cooperia spp were detected. Resistance of Ostertagia spp to ivermectin was evident on 4/45 (9%) farms, to albendazole on 15/46 (35%) farms, and to levamisole on 4/46 (9%) farms.

CONCLUSION: Anthelmintic resistance in parasitic nematodes of cattle is common in the North Island of New Zealand. Beef farmers need to be aware of the risks posed by anthelmintic resistance, and routine FECR testing is recommended to ensure optimal productivity and to guide decision-making when purchasing anthelmintics to be used on-farm.