Continuity in ovine Johne's disease vaccination practices despite a decline in clinical disease.

The Gudair® vaccine has been commercially available in Australia for almost two decades for the control of ovine Johne's disease, but concerns have been raised about potential discontinuation of vaccination by producers after a decline in the incidence of clinical disease. An online questionnaire was distributed to Australian sheep producers to identify the proportion of respondents discontinuing the Gudair vaccine and reasons for discontinuation. Results revealed that 88% of sheep producers surveyed have continued to vaccinate their sheep with Gudair, with continuation greater for predominantly Merino sheep flocks. Reasons for discontinuing vaccination stemmed from management, economic or health concerns. These results suggest that Gudair is still widely used by Australian sheep producers and concerns about large-scale discontinuation are unfounded. These findings have implications for ovine Johne's disease control programs in Australia.

A survey of ewe mortality on the Central Tablelands of New South Wales in 2010, a year of above-average rainfall.

BACKGROUND: Sheep producers reported elevated ewe mortality following a wetter-than-average season on the tablelands of New South Wales in 2010. We conducted a survey of local producers to determine the extent and cause of ewe losses, as well as associated risk factors. METHODS: A questionnaire was designed and posted to sheep producers running at least 1000 sheep. Data from questionnaires were analysed using linear mixed models to identify risk factors associated with ewe mortality. Data on the likely causes of deaths, particularly those associated with foot abscess, were analysed by ordinal generalised linear mixed models to determine the most prevalent causes of ewe death. RESULTS: The mean annual ewe mortality was 5.4% (range 0.1%-28.6%). Higher mortality was associated with older ewes and lambing in paddocks with longer pastures. The two most important causes of ewe death reported by farmers were 'died during lambing' and 'foot abscess'. Mortality due to foot abscess was more commonly reported in adult ewes than maiden ewes (odds ratio and 95% confidence interval: 5.61; 3.52, 8.94), when lambing on pasture longer than 5 cm than ≤5 cm (2.96; 1.54, 5.70) and when lambing in spring than in autumn (4.87; 1.36, 17.41). CONCLUSIONS: The level of ewe mortality observed was at the upper end of that reported previously in Australia. Risk factor analysis suggested that, in wet years, losses due to foot abscess could be reduced by lambing older heavier ewes in autumn on pasture swards less than 5 cm high.

Post-weaning growth of beef heifers drenched with long- or short-acting anthelmintics.

OBJECTIVE: To determine whether (i) the drenching practices of beef producers in the Central Tablelands of NSW utilising short-acting anthelmintics limit the post-weaning growth of heifers and (ii) a single weaning treatment of a long-acting anthelmintic improves post-weaning growth. METHODS: Commencing at weaning, 20/100 heifers in each of five herds were injected four times at 90-day intervals with long-acting moxidectin to suppress gastrointestinal nematodes. In year 1 the balance was drenched using the owner's usual commercial practice with short-acting anthelmintics (5 herds) or left undrenched (1 herd). In year 2 the balance was drenched with long-acting moxidectin within 3 months of weaning (4 herds) or with short-acting drenches only (2 herds). At each visit live weight (LW) was recorded and a subsample tested for worm egg count (WEC) and plasma pepsinogen. RESULTS: In year 1 by 3 and 6 months' post weaning, suppressed heifers were significantly heavier than the undrenched and short-acting groups, and by 12 months were 40 and 28 kg heavier, respectively. In year 2, the LW of the suppressed and long-acting heifers was not different, but by 3 and 6 months both groups were significantly heavier than the short-acting group and by 12 months were 22 kg heavier. CONCLUSION: The post-weaning growth of beef heifers treated with short-acting drenches as adopted by Tablelands beef producers was slower than heifers drenched suppressively or with a single long-acting drench at weaning.

Effectiveness of Gudair™ vaccine for the control of ovine Johne's disease in flocks vaccinating for at least 5 years.

OBJECTIVE: Examine the prevalence of shedding of Mycobacterium avium subsp. paratuberculosis (Mptb) at least 5 years after starting vaccination with Gudair™ in flocks of varying initial prevalence of ovine Johne's disease (OJD) and identify risk factors for variation in vaccine efficacy. METHODS: Pooled faecal culture (PFC) was conducted for 41 flocks from southern NSW and Victoria to determine estimates of current OJD prevalence. The data were compared to estimates of prevalence at or prior to commencement of vaccination at least 5 years earlier, based on available serological or PFC tests when vaccination commenced. A cross-sectional study was conducted to identify risk factors for differing prevalence levels in 36 of the flocks. RESULTS: Historical data enabled classification of 37 flocks as high (13; 35.1%), medium (10; 27.0%) or low (14; 37.8%) estimated initial OJD prevalence. Results of PFC in 2008-09 identified that 81.1% (30/37) of flocks had detectable shedders, with 48.6% (18/37) flocks still classified as medium or high OJD prevalence, including 50% (7/14) of flocks initially classified as low prevalence. Shedding was not detected in 18.9% (7/37) flocks. Flocks with OJD prevalence exceeding 1% at 5 years or more following the commencement of vaccination were associated with reports of sheep straying and introduction of new sheep. CONCLUSION: Despite significant declines in estimated OJD prevalence following vaccination for ≥5 years, 81.1% of flocks were shedding Mptb and considered at risk of spreading the disease or suffering recrudescence of losses if vaccination were to cease. Flock managers are advised to persist with vaccination.

Effect of unvaccinated Merino wether lambs on shedding of Mycobacterium avium subspecies paratuberculosis in flocks vaccinating for ovine Johne's disease.

OBJECTIVE: To determine the prevalence of Mycobacterium avium subspecies paratuberculosis (Mptb) shedding by unvaccinated wethers compared with vaccinated ewes. METHODS: Six self-replacing, ovine Johne's disease (OJD)-infected Merino flocks that had been vaccinating lambs with Gudair(TM) for at least 5 years, but had not vaccinated their last two drops of wethers were studied. Faecal samples were collected in pools of 50 from 350 vaccinated ewes (1-2 years old) and from 350 unvaccinated wethers of the same age and were cultured to determine the presence of viable Mptb. RESULTS: The estimated prevalence of shedding sheep among the unvaccinated wethers was 6-fold greater than among the vaccinated female siblings (0.21% vs 1.27%), with faecal pools collected from unvaccinated wethers being 19-fold more likely to be positive than those from the vaccinated ewes. CONCLUSION: Leaving wethers unvaccinated is likely to considerably increase pasture contamination with Mptb and potentially jeopardise disease control.

Association of microsatellite polymorphisms with immune responses to a killed Mycobacterium avium subsp. paratuberculosis vaccine in Merino sheep.

AIM: To study the association of polymorphisms at five microsatellite loci with immune responses to a killed Mycobacterium avium subsp. paratuberculosis (Map) vaccine. METHODS: Merino sheep (504 vaccinates and 430 unvaccinated controls) from a long-term Johne's vaccine trial undertaken on three different properties in the Central Tablelands of New South Wales, Australia, were genotyped for five microsatellite markers located in three immunologically significant chromosome regions. The marker loci included three from the major histocompatibility complex (MHC), namely DYMS1, OLADRB and SMHCC1; and one each from the solute carrier family 11 member 1 (SLC11A1), OVINRA1, and the interferon-γ (IFN-γ), o(IFN)-γ, gene regions. Associations between immune responses and genetic polymorphisms at the marker loci were examined by analysing both allelic and genotypic effects. RESULTS: The o(IFN)-γ locus had only two alleles, whereas the other four loci exhibited extensive polymorphism, with the number of alleles ranging from 10 (OVINRA1) to 21 (DYMS1), resulting in 30-92 genotypes per locus. Heterozygosities varied between 37% (o(IFN)-γ) and 87% (SMHCC1), while information on polymorphic contents ranged from 0.31 (o(IFN)-γ) to 0.87 (DYMS1). Each of the three properties exhibited unique allelic and genotypic frequencies. Analysis of immune response data revealed strong antibody and IFN-γ responses as early as 2 months post-vaccination. Immune responses in control animals on all three properties remained consistently low, except for slightly elevated IFN-γ responses at a few time-points on two properties, concomitant with exposure to natural infection. Genotype-phenotype association analyses revealed a number of marker genotypes/alleles to be significantly associated with antibody and IFN-γ responses. However, the effects of only five genotypes (one each at DYMS1, OLADRB, SMHCC1, OVINRA1 and o(IFN)-γ) and three alleles (one each at o(IFN)-γ, DYMS1and OLADRB) on IFN-γ responses were consistent across the three properties. CONCLUSION: Considering the significance of IFN-γ responses in protection against Map, it is possible that the genotypes/alleles identified might have a role in protective immune responses to natural Map infections, and further studies are warranted to confirm this.

Lesions attributed to vaccination of sheep with Gudair for the control of ovine paratuberculosis: post farm economic impacts at slaughter.

OBJECTIVE: To determine i) the prevalence of lesions at the site of vaccination with Gudair, and ii) the incidence of discounting of slaughtered sheep carcasses due to the presence of these lesions. DESIGN: A survey of the prevalence of injection lesions and actual discounts applied to 20 consignments of vaccinates slaughtered in abattoirs in New South Wales. PROCEDURE: Consignments of sheep previously vaccinated against ovine Johne's disease (OJD) were assessed on the slaughter chain for the prevalence of vaccination site lesions and any costs associated with carcass trimming were estimated. In addition a telephone survey was conducted to determine the experiences and risk attitudes of 8 abattoirs in south east New South Wales likely to have previously processed vaccinates. RESULTS: The prevalence of lesions observed was 18% for adult (mutton) and 65% for lamb carcasses. The value of the trim removed was insignificant, the labour cost of its removal was nil and no carcass was downgraded to a lower value grade. CONCLUSION: Under the market conditions existing at the time the study was conducted, in sheep vaccinated at the recommended site high on the neck, it is unlikely that OJD vaccination site lesions will be a significant cost to producers or the processing industry and will represent only a very small proportion of the total cost of OJD control by vaccination.

Lesions in sheep following administration of a vaccine of a Freund's complete adjuvant nature used in the control of ovine paratuberculosis.

CASE HISTORIES: Occurrences of adverse reactions in seven sheep flocks in Australia following vaccination against paratuberculosis where veterinary attention was requested are reviewed. All cases occurred within the 3-year period following commencement of use of a vaccine of a Freund's complete adjuvant nature, at a time when approximately six million doses of vaccine had been administered. CLINICAL FINDINGS: In the first case, 26/58 (45%) Merino sheep vaccinated as adults had palpable tissue reactions at or near the site of vaccination; enlarged prescapular lymph nodes were palpated in 17 (29%), and nine (16%) sheep had both palpable lesions at the site of vaccination and enlarged prescapular lymph nodes. The reactions included caseous nodules up to 5.5 cm in diameter. In the other cases, fistulating or granulomatous wounds were occasionally found at the recommended site of injection behind the ear, and myiasis was rare. Occurrences of inappropriate choice of injection site were recorded, including injection into the axilla of two Merino rams, and lesions in the tissues of the maxilla and nose of almost 50% of 350 Border Leicester lambs. Four outbreaks of progressive paralysis due to injection into cervical musculature were reported, described as "OJD staggers" by producers. DIAGNOSIS: Granulomatous cellulitis and lymphadenitis associated with oil droplets typical of "oil granulomata". Injection of vaccine into the dorsal cervical area resulted in progressive paralysis due to myonecrosis and suspected granulomatous leptomeningitis. CLINICAL RELEVANCE AND CONCLUSIONS: Although lesions at and near the site of injection are common, adverse reactions to vaccination were rare and included mortality from cervical spinal injection, production losses from injection in the maxilla or axilla or if myiasis resulted, and potential marketing losses if animals or carcasses are discounted as a result of the lesions. Risk factors for adverse reactions included inadequate restraint of sheep, breed of sheep, experience of the operator, poor injection technique, and inappropriate placement of vaccine. Increasing attention to the proper restraint of animals, restricting vaccination to the recommended site behind the ear, careful placement of the vaccine into subcutaneous tissue to avoid drainage of vaccine material into tissues such as the spinal cord, and post-vaccination supervision to address welfare concerns should adverse reactions occur are recommended.

Efficacy of a killed vaccine for the control of paratuberculosis in Australian sheep flocks.

A field trial was undertaken from 1999 until 2004 to determine the efficacy of a killed M. a. paratuberculosis vaccine, Gudair, for the control of ovine Johne's disease (OJD) in merino sheep run under Australian pastoral conditions. On each of three farms experiencing significant OJD losses (5-15% per annum), 200 merino lambs (age 1-4 months) were vaccinated, and 200 lambs were left unvaccinated. Animal assessments and sample collections were conducted twice yearly until 4 or 5 years of age. The impact of vaccination on mortality rate, faecal shedding of M. a. paratuberculosis (by pooled and individual faecal culture), liveweight, wool productivity, vaccine injection site lesions and cellular (BOVIGAM) and humoral (PARACHEK) immunity was examined. Vaccination stimulated cell-mediated and humoral immune responses, reduced mortalities due to OJD by 90% and delayed faecal shedding for the first year post-vaccination. Thereafter, the prevalence of shedders among vaccinates was reduced by 90%. The numbers of M. a. paratuberculosis excreted by the vaccinated groups were also reduced by at least 90% at most sampling times. However, high levels of excretion by vaccinates occurred on some occasions, and although only 7 of 600 vaccinates died from OJD, all 7 had multibacillary lesions. Thus there remains a risk that some vaccinated sheep will transfer the disease. Small reductions in liveweight were found in vaccinated lambs in the first year, but there was little effect on wool production. Vaccine injection site lesions were detected in almost 50% of sheep after 2 months, and these persisted for at least 4 years in 20-25% of vaccinates. Data from this trial enabled the registration of Gudair in Australia in 2002 and underpins the pivotal role of vaccination in the current management of OJD.

Injury caused by self-inoculation with a vaccine of a Freund's complete adjuvant nature (Gudair) used for control of ovine paratuberculosis.

OBJECTIVE: To document the occurrence and consequences of accidental self-inoculation of vaccinators (producers, farm employees, contractors) with the recently registered Gudair vaccine for the control of ovine paratuberculosis in Australia. DESIGN AND PROCEDURE: A survey of the first 50 primary producers permitted to use the vaccine in sheep and a description of six cases of accidental self-inoculation for which medical attention was sought, and which occurred after the vaccine became widely available. RESULTS: The survey recorded that, of 37 respondents vaccinating 155,523 sheep, there were 21 incidents of exposure to the vaccine, an overall rate of one incident per 7406 vaccinations. In five of these incidents there was only superficial skin contact with vaccine; in 16 there was needle penetration without vaccine injection. There were no reports of self-inoculation with vaccine. Six cases of self-inoculation with Gudair vaccine that required medical intervention are described. Of these five were in males and one in a female; four involved injection of vaccine into the leg and single cases involved a foot or hand. Most cases required surgical removal of the injected vaccine to allow wound repair; three required extensive surgery and open drainage. Even with surgery recovery took as long as 9 months. Possible risk factors for self-inoculation and the resulting outcome are discussed. CONCLUSIONS: Gudair ovine paratuberculosis vaccine can cause prolonged granulomatous inflammation if inadvertently injected into human tissue. After.self-inoculation, early surgical debridement of the damaged tissue and drainage to remove the vaccine material are advised to avoid progression to extensive necrosis.

The viability of transferred sheep embryos after long-term cryopreservation.

The survival to term of 414 sheep embryos, thawed and transferred after conventional cryopreservation and storage for 13 years, was evaluated. A concurrent experiment involving the transfer of vitrified embryos to 91 ewes and artificial insemination of 51 ewes with frozen-thawed semen from sires of the long-term cryopreserved embryos provided forms of control treatments. The donor ewes had a mean ovulation rate of 10.9, and 7.1 embryos per ewe were cryopreserved. Each recipient ewe received two embryos and pregnancy was assessed at Day 18, Day 54 and term. The pregnancy rate was lower in the long-term embryo group than the artificial insemination group at Day 18 (P<0.01) and Day 54 (P<0.05), although the difference at term (31% v. 49%) was not significant, with the vitrified embryo group being similar to the long-term group. Embryo survival to birth was 21%, with the majority of loss (80%) occurring by Day 18. The later stage of development and higher grade of transferred embryos and the older age of donor ewes resulted in a significantly higher (P<0.01) pregnancy rate at Day 54 and term, and percentage of lambs born and weaned. Other effects of donor ewes (genotype, superovulation treatment, number of ovulations and embryos cryopreserved) were not significant. Implications for the design of genetic evaluation and germplasm conservation programmes using embryo cryopreservation technology are discussed.

Fertility and its relationship to motility characteristics of spermatozoa in ewes after cervical, transcervical, and intrauterine insemination with frozen-thawed ram semen.

The fertility of ewes after artificial insemination and the relationship between fertility and motility characteristics assessed by a computerized motility analysis system were examined with ram semen frozen in diluents reported to improve postthaw motility. The percentages of motile and progressive spermatozoa were better when frozen in proline- or glycine betaine-containing or HEPES-based, rather than Tris-based, diluents (P < 0.01). The fertility of spermatozoa frozen in diluents containing proline or glycine betaine was slightly reduced, whereas when both compatible solutes were present, the reduction was more pronounced, in comparison with semen frozen in Tris- or HEPES-based diluents (9.5 versus 71.1 and 66.6%; P < 0.01). Fertility of frozen-thawed spermatozoa was higher after laparoscopic insemination than after cervical or transcervical insemination (P < 0.01). Similarly, higher fertility was obtained after cervical insemination with fresh than with frozen-thawed semen (32.4 versus 11.3%; P < 0.01). Furthermore, loss of embryos was lower after laparoscopic insemination of ewes with semen frozen in a Tris diluent than with semen frozen in proline diluents, in glycine betaine diluents, or in proline-plus-glycine betaine diluents (0.0 versus 26.0, 38.5, and 60.0%; P < 0.001). A wide variation in the postthaw percentage of motile (31.6-59.7%) and progressive (22.6-43.1%) spermatozoa and in the fertility of spermatozoa from individual rams was also observed after laparoscopic (29.2-59.7%) or cervical insemination (8.7-30.5%). Postthaw motility results from immediately after thawing and fertility results from experiments where intrauterine insemination was performed with semen frozen in proline- or glycine betaine-containing or HEPES- or Tris-based diluents were pooled and subjected to a pairwise correlation procedure. The correlation analysis showed relationships between some of the motility characteristics (P < 0.01), but there were no relationships between the motility characteristics and fertility.

Sources of variation in the reproductive performance of ewes inseminated with frozen-thawed ram semen by laparoscopy.

Fertility data from 8 artificial insemination programs, involving more than 5000 ewes and 110 rams in 3 flocks, were analyzed to determine variation due to individual AI program and ram in the reproductive performance of ewes inseminated with frozen-thawed semen by laparoscopy. The semen had been previously frozen by commercial AI centers in either pellets or straws. Both AI program and individual ram affected the proportion of ewes pregnant and the number of fetuses per ewe inseminated, but not the number of fetuses per pregnant ewe. Semen samples from 97 of the rams used were analyzed on a Hamilton Thorn HTM 2000 image analyzer for sperm concentration, percentage of motile and progressively motile spermatozoa, mean progressive velocity, and mean linear index. The correlations between these traits and reproductive performance obtained after insemination were calculated. There was large variation in the quantity and quality of the frozen semen, but only the number of total and motile spermatozoa inseminated per ewe was correlated with fertility (0.25 and 0.26, respectively). Regression analysis showed that none of the traits measured were useful for predicting fertility.

Effect of anti-oestradiol-17B antibodies on the reproductive response of ewes superovulated with PMSG.

A field experiment was conducted to examine the effect of anti-oestradiol-17B antibody titre on the oestrous and ovulatory responses of ewes to low (600 i.u.) or high (1200 i.u.) doses of pregnant mare's serum gonadotrophin (PMSG). Merino ewes were treated with intravaginal sponges and were subsequently used as vehicle-treated controls or were immunized to produce reciprocal anti-oestradiol-17B antibody titres less than 1000 or greater than 1000. Ewes were then treated with PMSG and the incidence of oestrus and ovulation, ovulation rate, and yield of embryos recorded. Treatment of immune ewes with 1200 i.u. PMSG resulted in both a higher proportion of ewes ovulating and a higher ovulation rate than in immune ewes treated with 600 i.u. (86% v. 67% and 13.4 v. 6.0 respectively). As anti-oestradiol-17B titres increased there was a reduction in the proportion of ewes exhibiting oestrus. The proportion of ewes ovulating decreased as antibody increased in ewes treated with 600 i.u. PMSG but not in those treated with 1200 i.u., suggesting an increased positive feedback of oestradiol with high PMSG doses. Fertilization rates were highest at the lower PMSG dose (68% v. 42%) and increased with increasing titre. Overall, there was no increase in ovulation rate or in yield of embryos over control values from either low (less than 1000) or high (greater than 1000) antibody titres.