Identification and genotyping of feline infectious peritonitis-associated single nucleotide polymorphisms in the feline interferon-γ gene

Feline infectious peritonitis (FIP) is an immune-mediated, highly lethal disease caused by feline coronavirus (FCoV) infection. Currently, no protective vaccine or effective treatment for the disease is available. Studies have found that some cats survive the challenge of virulent FCoV isolates. Since cellular immunity is thought to be critical in preventing FIP and because diseased cats often show a significant decrease in interferon-γ (IFN-γ) production, we investigated whether single nucleotide polymorphisms (SNP) in the feline IFN-γ gene (fIFNG) are associated with the outcome of infection. A total of 82 asymptomatic and 63 FIP cats were analyzed, and 16 SNP were identified in intron 1 of fIFNG. Among these SNP, the fFING + 428 T allele was shown to be a FIP-resistant allele (p = 0.03), and the heterozygous genotypes 01C/T and +408C/T were found to be FIP-susceptible factors (p = 0.004). Furthermore, an fIFNG + 428 resistant allele also showed a clear correlation with the plasma level of IFN-γ in FIP cats. For the identification of these three FIP-related SNP, genotyping methods were established using amplification refractory mutation system PCR (ARMS-PCR) and restriction fragment length polymorphisms (RFLP), and the different genotypes could easily be identified without sequencing. The identification of additional FIP-related SNP will allow the selection of resistant cats and decrease the morbidity of the cat population to FIP.     

Description of a regional anaesthesia technique for the dorsal cranium in the dog: a cadaveric study

Objective

To identify landmarks and to describe a technique for nerve blockade of the dorsal cranium in dogs.

Establishment and Application of a Nested PCR Assay for Detection of Fowl Adenovirus Group Ⅰ

According to the sequence of hexon gene of fowl adenovirus groupⅠ(FAVⅠ) strain published in GenBank,two pairs of primers were designed and synthesized.The outer primers amplified a fragment of 475 bp in length, and the inner primers amplification fragment was 237 bp in length. A nested PCR assay for rapid detection of FAVⅠ was established.A specific 237 bp fragment was amplified from DNA templates of FAVⅠstrain,but no bands were amplified with templates extracted respectively from avian influenza virus (AIV) subtype H9,Newcastle disease virus (NDV), infectious bursal disease virus (IBDV),duck plague virus (DPV), reticuloendotheliosis (REV), avian reovirus (ARV), Marek's disease virus (MDV). Sensitivity of the 1st and 2nd amplifications by the nested PCR assay were 100 pg and 1 fg,respectively.The sensitivite of the 2nd amplifications increased by 10⁵ times.The results showed that the nested PCR was specific,sensitive,rapid,accurate,and could be used as a routine assay for the detection of FAVⅠ.This method had good reproducibility, specificity and sensitivity, and might detect low content FAVⅠ accurately and rapidly. This method could be used as a method for the diagnosis and detection of clinical cases,and molecular epidemiological investigation of FAVⅠ.     

Active surveillance for scrapie in the Netherlands: effect of a breeding programme on the prevalence of scrapie in sheep (2002-2010)?

The susceptibility of sheep to scrapie is modulated by the prion protein (PrP) genotype of the animal. An ambitious voluntary scrapie control programme was started in the Netherlands in 1998, based on selection of rams with theARR/ARR genotype for breeding. This programme was followed by an obligatory programme in 2004; the programme has been voluntary since 2007. We monitored the prevalence of PrP genotype frequencies and the prevalence of scrapie in the Dutch sheep population between 2002 and June 2010. Results showed that selection for scrapie-resistant sheep resulted in an increase in the ARR allele frequency in the Dutch national flock from 37.5% in 2005 to 61.4% in 2009. Moreover, surveillance data showed that there was a significant decrease in the prevalence of scrapie a few years after the start of the obligatory breeding programme, from more than 0.2% in 2004 to 0.015% in 2009. This decrease is a consequence of the increased number of scrapie-resistant sheep in the Dutch sheep population. To date, the results and the models based on the data show that the selective breeding programme should be continued for several years in order to successfully eradicate scrapie. It will be important to monitor the PrP frequency and scrapie prevalence in the Dutch sheep population in the coming years.     

Salmonella Brandenburg — emergence of a variant strain on a sheep farm in the South Island of New Zealand

AIM: To compare the rectal and I/V administration of tramadol in dogs, to assess both its pharmacokinetic properties and absolute bioavailability.

METHODS: After rectal administration via suppositories and I/V injection of tramadol (4 mg/kg), the concentration of tramadol and its main metabolites, O-desmethyl-tramadol (M1), N-desmethyl-tramadol (M2) and N,O-didesmethyl-tramadol (M5), were determined in plasma, using high-performance liquid chromatography (HPLC). A balanced cross-over study was used, involving six male Beagle dogs.

RESULTS: Plasma concentrations after rectal and I/V administration were fitted on the basis of mono- and bi-compartmental models, respectively. Following rectal administration tramadol was detected from 5 minutes up to 10 hours, in lesser amounts than M5 and M2, while M1 was detected in negligible amounts. Following I/V administration tramadol was detected up to 10 hours, M2 and M5 were detected at similar concentrations, and M1 was present at low concentrations. The area under the curve (AUC) of the three metabolites did not differ significantly after either route of administration of tramadol. The absolute bioavailability of tramadol via rectal administration was 10 (SD 4)%.

CONCLUSIONS: After rectal administration of tramadol suppositories, absorption of the active ingredient was rapid, but its metabolism quickly transformed the parent drug to high levels of M2 and M5.

CLINICAL RELEVANCE: In the dog, rectal pharmaceutical formulation of tramadol would have a different pharmacokinetic behaviour than in humans.     

Technical note: A novel method for routine genotyping of the G allele of β-casein (CSN2) and T allele of κ-casein (CSN3) in a sheep population using LightCycler

The goal of this work was to develop a protocol for rapid genotyping of A and G variants at the CSN2 locus and genotyping of T and C variants at the CSN3 locus in sheep breeds (Sumava and Valachian) by means of PCR and LightCycler analysis. The LightCycler technique combines rapid and efficient in vitro amplification of DNA in glass capillaries with melting curve analysis based on fluorescence resonance energy transfer for the sensitive detection of point mutation. The A variant had a greater frequency (Sumava, 0.778; Valachian, 0.835) than did variant G (Sumava, 0.222; Valachian, 0.165) in both sheep breeds. The CSN3 locus was found to be monomorphic, with no polymorphism identified in either population.     

Breeding for resistance to footrot – the use of hoof lesion scoring to quantify footrot in sheep

So that genetic studies can be undertaken on footrot in sheep, it is necessary that a reliable and repeatable method to categorise the phenotype is available. This paper summarises the methods used and results obtained from 1600 hoof lesion scores of 100 mixed-age ewes independently scored twice by two trained operators. Using a 5-pont scale describing the severity of foot lesions, residual correlations were used to assess agreement between scorers and scoring occasions. Data were analysed using both zero-1 and continuous data methods. The average prevalence of any score >0 was 15%, and of scores >1 was 12%. The residual correlation between scorers for SUM_FR was 0.87 and between scoring occasions it was also 0.87, indicating high repeatability or agreement both within and between scorers. No significant differences were detected between scorers or between scoring occasions for any of the traits analysed, or different analytical methods used.     

Evaluation of farmers’ diagnostic performance for detection of diarrhoea in nursery pigs using digital pictures of faecal pools

Background

Overconsumption of antibiotics in the pig industry is of concern in relation to antimicrobial resistance. False positive disease diagnosis may result in the treatment of healthy animals. In Denmark, diarrhoea is the most common cause of antibiotic treatment in pigs. Farm personnel are not professional clinicians, which could result in inappropriate antibiotic treatments of diarrhoea.The primary objectives of this pilot study using digital pictures of faecal pools was to evaluate farmers’ diagnostic performance in the assessment of faecal consistency in nursery pigs and to investigate the effect of different co-variables, including practical experience. A secondary objective was to compare the diagnostic performance of farmers with that of veterinarians.At a pig congress, observers (farm personnel and veterinarians) working professionally with pigs participated in a faecal consistency test consisting of 16 pictures of faecal pools (eight diarrhoeic and eight non-diarrhoeic). The faecal pools had previously been collected and subjected to faecal dry matter determination. The true status of the faecal pools was determined by the faecal dry matter content (diarrhoea: faecal dry matter ≤ 18%). The true status was used to evaluate the farmers’ and veterinarians’ diagnostic performance.

Results

A total of 119 farmers and 18 veterinarians were included in the statistical analysis. For the farmers, the mean proportion of faecal pools assessed as diarrhoeic was 0.48, the mean proportion of correctly classified faecal pools was 0.84, the mean diagnostic sensitivity was 0.83 and the mean diagnostic specificity was 0.86. Farmers with less than four years of practical experience detected clinical diarrhoea more accurately than farmers with more than four years of practical experience (p < 0.05). No significantly differences between farmers and veterinarians was observed (p > 0.20).

Conclusions

The results, using digital pictures of faecal pools, suggest that farmers and veterinarians have similar diagnostic performance in relation to diarrhoea. False positive classification of non-diarrhoeic pigs appears to be a larger problem than false negative classification of diarrhoeic pigs under Danish conditions. If these results can be confirmed under practical conditions, training in, and validation of, clinical diagnoses may be an important factor in reducing antibiotic consumption in the pig industry.     

Control of canine leptospirosis in Europe: time for a change?

Changes in the formulation of the Leptospira components of dog vaccines are being considered in Europe, following changes in North America. This article discusses the options for change and recommends the continued inclusion of serovars Icterohaemorrhagiae and Canicola plus the inclusion of serovars Bratislava and Grippotyphosa (for mainland Europe only). If other serovars, such as Pomona, are to be considered in the future, then there is a need for additional clinical, cultural and serological studies across Europe to support their inclusion.     

The effects on cardio-respiratory and acid-base variables of the anaesthetic alfaxalone in a 2-hydroxypropyl-β-cyclodextrin (HPCD) formulation in sheep

The objective of this study was to determine the pharmacodynamic effects in sheep of the anaesthetic alfaxalone in a 2-hydroxypropyl-β-cyclodextrin formulation. Seven Ripollesa sheep, weighing 43.0±6.6 kg, were used in the study. Twenty-four hours after instrumentation, the sheep were anesthetised with alfaxalone (2 mg/kg bodyweight IV) in cyclodextrin. Heart rate, arterial blood pressure, respiratory rate and arterial blood gases were recorded. Alfaxalone administration resulted in minimal cardio-respiratory depression. Time to standing from anaesthesia was 22.0±10.6 min. Apnoea was not observed in any of the sheep. Significant differences from baseline were not observed in respiratory rate or arterial blood pressure. Heart rate increased significantly (P<0.05) immediately after administration, returning to control values at 20 min. The calculated haemoglobin saturation (SO2) decreased significantly during the first 15 min after alfaxalone administration. The arterial pH decreased significantly during the first 30 min of the study, although no significant differences from basal values were observed in the arterial partial pressure of carbon dioxide (PaCO2). The results showed that alfaxalone in 2-hydroxypropyl-β-cyclodextrin administered as an IV bolus at 2 mg/kg produced minimal adverse effects and an uneventful recovery from anaesthesia in sheep.     

Development of a Real-time PCR Assay for Detection of Chicken IL-1β, IL-18 and Tumor Necrosis Factor α Genes

In order to develop a SYBR GreenⅠ Real-time PCR assay for detection of chicken IL-1β,IL-18 and tumor necrosis factor-α (TNF-α) genes, four specific primer pairs were designed according to the chicken's IL-1β,IL-18, TNF-α and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) gene sequences in GenBank. The four fragments were amplified by RT-PCR from chicken embryo fibroblasts, cloned and sequenced. The recombinant plasmids containing the target gene were constructed and used as the Real-time PCR standard templates. Real-time PCR assays based on SYBR GreenⅠfor detection of chicken IL-1β,IL-18, TNF-α and GAPDH were established. The results showed that each gene's melting curve also had a single peak,each gene's amplification efficiency was 101.2%, 95.6%, 100.1% and 98.2%, R² was 0.9996,0.9998,0.9957 and 0.9989. Moreover,the assays were highly sensitive,the detection limit of 100 copies in 35 Ct and each gene's coefficient of variation less than 1.4 percent for intra-assay. This reliable Real-time PCR assay might be used for decting chicken's IL-1β,IL-18 and TNF-α mRNA expressing and provided the basis for quantitative analysis of cytokine expression in host cell after virus infect which cause immunosuppressive diseases.     

Detection of Subclinical Mastitis in Dromedary Camels (Camelus dromedarius) using Somatic Cell Counts and the N-Acetyl-β-D-glucosaminidase Test

Somatic cell counts, N-acetyl--D-glucosaminidase (NAGase) activity and the infection status of the udder were determined in quarter milk samples (n = 86) from 22 multiparous, clinically healthy camels, traditionally managed by Bedouin nomads in the Negev desert, Israel. Seventy (81.4%) of the 86 samples examined contained bacteria, of which 35 (40.7%) gave mixed isolations of two or more bacteria, suggesting the existence of subclinical mastitis in the camel herds studied. Sixteen samples (18.6%) yielded no growth of bacteria. Staphylococcus aureus, Micrococcus spp., Bacillus spp., Streptococcus dysgalactiae and Escherichia coli were the main organisms isolated. The somatic cell count (SCC) ranged from 1.01×10⁵ to 11.78×10⁶ cells/ml. NAGase values were between 41.4 and 372 NAGase units. Quarter milk samples that contained bacteria had significantly (p<0.01) higher mean values for SCC but the mean NAGase levels were not significantly different for the bacteriologically negative and positive samples. There was a low correlation coefficient (r ² = 0.097) between the SCC and NAGase in the quarter milk samples from which bacteria were not isolated (n = 16) and a low negative correlation (r ² = –0.038) with the samples that contained bacteria (n = 70). The type of bacteria had a significant effect (p<0.01) on the SCC but not on the NAGase activity. Quarter samples from which Staphylococcus aureus (coagulase positive) was isolated showed the highest mean SCC and this organism is therefore suspected to be the underlying cause of the subclinical mastitis. The SCC gave a better indication of the presence of pathogenic microorganisms in milk samples than did NAGase.     

Bordetella bronchiseptica experimental model development in pigs and efficacy evaluation of a single intramuscular injection of gamithromycin (Zactran® for Swine) against Bordetella bronchiseptica-associated respiratory disease in experimentally infected piglets

In the Bordetella bronchiseptica infection model development study, twenty-eight piglets were inoculated with B. bronchiseptica strain of either canine (10⁹ CFU/ml) or swine (10⁸ and 10⁹ CFU/ml) origin; swine origin strain at 10⁹ CFU/ml was chosen for the efficacy assessment study due to higher incidence and severity of gross and histopathological lesions compared with other strains. To assess efficacy of gamithromycin against B. bronchiseptica, forty piglets were experimentally inoculated on Day 0 and clinical signs were scored as per severity. Animals were then treated either with gamithromycin or saline on Day 3. The Global Clinical Scores in gamithromycin-treated group were consistently lower than the saline-treated control group from Day 4 onwards and were 0 and 40 in the gamithromycin-treated and saline-treated control groups, respectively, on Day 6. Severity and frequency of gross and histopathological observations were significantly lower in gamithromycin-treated animals compared with saline-treated controls. The efficacy of Zactran® for Swine at the label dose for the treatment of B. bronchiseptica–associated respiratory disease was demonstrated based on the faster reduction in clinical signs as early as 1 day post-gamithromycin treatment and based on the significant difference in the severity of macroscopic and microscopic lung lesions 10 days post-gamithromycin treatment.     

Evaluation of ‘best practice’ (SCOPS) guidelines for nematode control on commercial sheep farms in England and Wales

Parasitic diseases are a major constraint to optimum livestock production and are the major cause of economic loss in UK sheep flocks, with farmers remaining dependant on anthelmintics for control. In the UK, research and evidence based, “best practice” guidelines for sustainable control of parasites in sheep (SCOPS) were first produced in 2004 and have been regularly updated since. This study was designed to evaluate the effect of these best practice guidelines for worm control on lamb production and infection levels, compared with more traditional management. Sixteen farms were selected based on a 2 cube factorial design with 3 factors known to affect worm epidemiology: control regimen; farm type; and climatic region. A formalised plan for worm control using 7 potential resistance-delaying practices was prepared for each of the 8 best practice (SCOPS) farms, in conjunction with the farms veterinarians. The 8 farms in the traditional management group (CONTROL farms) were selected based on ongoing evidence of them using worm control strategies deemed to be “higher-risk”. A cohort of 40–50 study lambs at each farm was monitored from birth to finishing, allowing evaluation of lamb productivity, worm infection levels and for comparison of numbers of anthelmintic treatments. Birth and mid-season weights were used to calculate daily live-weight gain. Birth and finish dates were used to calculate time to finish and finish weights were also compared. Faecal egg counts, larval culture and species differentiation were undertaken throughout the year to assess the impact of the control strategies on worm burdens. There was no significant difference in results for any of the 3 production responses when comparing predicted means accounting for the differences in birth weight. In fact SCOPS farms had, on average, a higher daily weight gain and finish weight than CONTROL farms when comparing observed means. Statistical analysis of infection levels clearly showed no significant effect according to farm type (p = 0.71) or treatment (p = 0.81). In contrast the effect of region (p = 0.08), although not significant, had a much larger effect size (standardised mean difference) with lower parasite burdens based on faecal egg counts on Northern farms compared to Southern farms. For both ewes and lambs, significantly fewer treatments were carried out on the SCOPS farms. The data collected from this study suggests that farms implementing SCOPS principles use less anthelmintic than other farms, without loss of animal performance or increased worm burden.     

Development of a sedation protocol using orally administered tiletamine–zolazepam–acepromazine in free-roaming dogs

Objective

To investigate the sedative effects in dogs of tiletamine–zolazepam–acepromazine (TZA) or ketamine–flunitrazepam (KF) administered orally and to evaluate the effectiveness of encapsulated TZA for capturing free-roaming dogs.

Estimating sample size for a small‐quadrat method of botanical survey for application in mixed Bushveld

Abstract

Species density and grass density were measured using a small‐quadrat method in eight plant communities in the Nylsvley Nature Reserve, South Africa. The presence of new species in each additional quadrat was used to calculate sample size. A sample size of 20 quadrats of 0.25 m² per site was found to be sufficient to measure species richness and density, and grass density in most plant communities in Mixed Bushveld. A sample size of approximately 30 quadrats was found to be necessary to survey plant communities with a relatively high species richness.     

Hybrid technique for ventricular septal defect closure in a dog using an Amplatzer® Duct Occluder II

A left-to-right shunting muscular ventricular septal defect (VSD) was diagnosed in a 4-month-old, female, 1.8 kg Bichon Frise – poodle mix dog. Echocardiographic evidence of cardiac remodeling, calculated pulmonary blood flow (Qp) to systemic blood flow (Qs) ratio of 2.8, and radiographic evidence of pulmonary edema supported the diagnosis of a hemodynamically important VSD. Using a combination of surgery and interventional catheter-based techniques to approach the VSD through the right ventricle, the VSD was occluded with an Amplatzer^® Duct Occluder (ADO) II device. The ADO II is a low profile, flexible device originally developed for patent ductus arteriosus closure in humans that has been used to close muscular and perimembranous VSD in children. This report describes the hybrid procedure and imaging that was essential for successful occlusion of the VSD in this dog.     

Selection for food conversion in broilers: Body composition of birds selected for increased body‐weight gain,food consumption and food conversion ratio

1. Body composition was determined in 9‐week‐old chickens sampled from four lines selected either for increased body‐weight gain (line W), for increased food consumption (line F), for decreased food conversion ratio (line E), or at random (line C), after three, four and five generations of selection.

2. After five generations of selection the proportions of carcass water (678 g/kg) and protein (187 g/kg) were highest in line E and lowest (636, 180 g/kg respectively) in line F. The proportion of fat was lowest (83 g/kg) in line E and highest (134 g/kg) in line F.

3. Lines W and C were similar and intermediate to lines E and F for all the carcass constituents.

4. There was more fat in females (115 vs 98 g/kg) and less protein (177 vs 190 g/kg) and water (652 vs 665 g/kg) than in males.

5. Line differences in carcass composition were not appreciably altered if birds were killed at equal weight rather than equal age.