Distinct neutrophil C5a receptor inflammatory events in cows initiated by chemoattractant C5a and lipopolysaccharide around parturition and in mid lactation.

In neutrophils, toll-like receptor and complement component 5a (C5a) signaling are critical pathways regulating innate immunity. In cows, not much is known about the second C5a receptor, complement component 5a receptor 2 (C5AR2). It is an interesting player in sepsis treatment because it is considered to have an anti-inflammatory effect during normal inflammation. Periparturient cows are prone to severe infections, and the objectives of this study were to investigate the expression and functionality of C5AR2 during peripartum. We investigated the effect of 2 major inflammatory stimuli, C5a and lipopolysaccharide (LPS), on the expression of a selected number of genes (C5AR1, C5AR2, TLR4, ITGAM, COX2, and CXCL8) and functions linked to these receptors. Overall, TLR4, ITGAM, and C5AR2, all of which are involved in early inflammation, showed a lower expression in periparturient cows. However, an overall lower expression seems not to be the only explanation for the increased risk of sepsis in periparturient cows. Normally, in response to inflammation and as seen in the mid-lactation group, the expression of these genes increases after stimulation with LPS. However, in periparturient cows, stimulation with LPS led to a decrease in expression of these receptors, indicating a different response of neutrophils in response to LPS during this period. A decrease in ITGAM (coding for CD11b) expression complicates correct neutrophil localization and phagocytosis. Its downregulation upon stimulation might be detrimental for adequate eradication of the pathogen and might increase the risk of an imbalanced inflammation; C5AR2 seems to play a central role in this altered response. In addition, myeloperoxidase (MPO) activity in periparturient cows is lower in response to C5a stimulation. It has been suggested that MPO plays an important role in neutrophil shutdown and, thereby, timely resolution of inflammation. A decreased MPO activity might thus prolong the inflammatory reaction of the neutrophils. This finding was supported by the increased viability of the neutrophils obtained from periparturient cows. Even after stimulation, we found a lower caspase-3 activity in this group, indicating that they might be activated for a longer time compared with the neutrophils from mid-lactation cows. Accordingly, these alterations might contribute to a temporal mismatch in inflammatory responses, as often seen in severe periparturient infections.

Germ-free sea bass Dicentrarchus labrax larval model: a valuable tool in the study of host-microbe interactions.

A thorough understanding of host-microbe interactions is crucial for more efficient disease management in the marine larviculture industry. As demonstrated in terrestrial animal research, gnotobiotic systems (involving animals cultured in germ-free conditions or inoculated with known microorganisms) are excellent tools to extend our understanding of the mechanisms involved in host-microbe interactions and allow the evaluation of new treatments for diseases. In this study, we introduce a germ-free European sea bass Dicentrarchus labrax larval model, independent of the continuous addition of antimicrobial agents. This model has an experimental set-up that allows addition of live feed to the larvae without compromising the germ-free status. This model will facilitate and render aquaculture research more effective in terms of mitigation fish larval diseases.

Clearance of Escherichia coli O157:H7 infection in calves by rectal administration of bovine lactoferrin.

Enterohemorrhagic Escherichia coli (EHEC) strains, of which E. coli O157:H7 is the best-studied serotype, are an important group of foodborne pathogens causing severe illness in humans worldwide. The main reservoirs for EHEC are ruminants, mostly cattle, which harbor the bacteria in their intestinal tracts without showing clinical symptoms. In this study, we used bovine lactoferrin, a natural occurring bactericidal and immunomodulating protein, as an antibacterial agent against EHEC infection in cattle. Nine 3-month-old Holstein-Friesian calves were experimentally infected with EHEC (strain NCTC12900). Three animals received a daily rectal spray treatment with bovine lactoferrin, three animals received an oral treatment, and three animals served as a control group. Blood samples were collected weekly and fecal samples twice weekly to monitor antibody responses and fecal excretion, respectively. Animals in the rectal group ceased shedding within 26 days of the experimental treatment and remained negative. This beneficial effect of bovine lactoferrin was not observed in the oral group, where animals were still shedding at the time of euthanasia (day 61). All groups developed serum responses, but no clear differences could be observed between the groups. However, the results indicate that the use of bovine lactoferrin as a rectal treatment can be a useful strategy to preclude further transmission of EHEC infections from cattle to humans.

Assessment of welfare of Brazilian and Belgian broiler flocks using the Welfare Quality protocol.

The Welfare Quality consortium has proposed a science-based protocol for assessing broiler chicken welfare on farms. Innovative features make the protocols particularly suited for comparative studies, such as the focus on animal-based welfare measures and an integration procedure for calculating an overall welfare status. These protocols reflect the scientific status up to 2009 but are meant to be updated on the basis of inter alia implementation studies. Because only few such studies have been done, we applied the Welfare Quality protocol to compare the welfare of broiler flocks in Belgium (representing a typical European Union (EU) country which implies stringent animal welfare legislation) versus Brazil (the major broiler meat exporter to the EU and with minimal animal welfare legislation). Two trained observers performed broiler Welfare Quality assessments on a total of 22 farms in Belgium and south Brazil. All of the farms produced for the EU market. Although the overall welfare was categorized as 'acceptable' on all farms, many country differences were observed at the level of the welfare principles, criteria, and measures. Brazilian farms obtained higher scores for 3 of the 4 welfare principles: 'good feeding' (P = 0.007), 'good housing' (P < 0.001), and 'good health' (P = 0.005). Four of the 10 welfare criteria scores were, or tended to be, higher on Brazilian than Belgian farms: 'absence of prolonged thirst' (P < 0.001), 'ease of movement' (P < 0.001), 'absence of injuries' (P = 0.002), and 'positive emotional state' (P = 0.055). The only criteria with a higher score for the Belgian farms than their Brazilian counterparts were 'absence of prolonged hunger' (P = 0.048) and 'good human-animal relationship' (P = 0.002). Application of the Welfare Quality protocol has raised several concerns about the validity, reliability, and discriminatory potential of the protocol. The results also call for more research into the effect of animal welfare legislation as broiler welfare on the south Brazilian farms appeared to be superior to that on the Belgian farms. Animal-based welfare assessments on a larger sample of farms are needed to evaluate to what extent these findings may be generalized.

Impact of a trace element supplementation programme on health and performance of cross-breed (Bos indicus x Bos taurus) dairy cattle under tropical farming conditions: a double-blinded randomized field trial.

Small-scale urban dairy farms (n = 16) in and around Jimma, Ethiopia with cross-bred (Bos indicus × Bos taurus) cows were enrolled in a double-blinded intervention study to investigate the effect of a trace element supplementation programme on trace element status and milk concentrations as well as performance [body condition score (BCS), milk yield, leptin], milk composition, antioxidant status (ferric-reducing ability of plasma (FRAP), thiobarbituric acid-reactive substances (TBARS)], blood biochemistry, serum proteins and immune response (antibody titre upon rabies vaccination). The farms were allocated to a (1) placebo or (2) Cu, Zn, Se, Co and I supplementation treatment for 150 d. On days 0 and 120, four lactating cows per farm were sampled for milk and plasma, and on day 150 for serum, following primo-vaccination. Cu deficiency was present in 17% and marginal Se deficiency in 30% of initially sampled cows, while no Zn shortage was detected. Over 120 days, trace element supplementation caused a bigger increase in plasma Se and Cu concentrations, but also a larger decrease of plasma Fe concentrations. A larger increase in milk Se concentrations was observed in the supplemented group, whereas none of the other elements were affected. BCS decreased more over time in the supplemented group. None of the other parameters of performance and antioxidant status nor milk composition or blood biochemistry was affected by treatment. Antibody response to rabies vaccination did not differ between groups, whereas α1-globulins tended to be lower and ß-globulins tended to be higher in the supplemented group. In conclusion, despite improved Cu and Se status and Se concentrations in milk, cows on tropical urban dairy farms did not seem to benefit from trace element supplementation, with respect to the parameters investigated.

Long term outcome after lichtenstein hernia repair using general, locoregional or local anaesthesia.

BACKGROUND: Chronic pain or discomfort after hernia surgery is nowadays a more challenging concern than recurrence. This study aimed to evaluate the long-term impact of local anaesthetic repair (LA) on pain, discomfort, paraesthesia and functional outcome after Lichtenstein hernia repair as compared to locoregional (LRA) and general anaesthesia (GA). METHODS: patients with primary or recurrent inguinal hernia underwent Lichtenstein repair with a polypropylene mesh. All patients with a follow-up of at least three years were sent a detailed questionnaire and offered an outpatient visit. Kaplan-Meier estimates and Cox proportional hazard regressions were used to analyse the relationship between time to event variables and explanatory variables including anaesthesia type. RESULTS: Between 1994 and 2006, in two cohorts, 330 patients answered the questionnaire: 100 under GA, 35 under LRA, and 195 under LA. This represented a response rate of 95, 94, and 98% respectively. Compared to GA and LRA, LA resulted in less long term pain, discomfort and paraesthesia. Moreover, resumption of social and professional activities was faster after LA. Recurrence rates were 1, 0, and 0.5% respectively. CONCLUSIONS: After Lichtenstein inguinal hernia repair, LA results in beneficial effects beyond the immediate postoperative period.

Effect of artificial insemination protocol and dose of frozen/thawed stallion semen on pregnancy results in mares.

Deep intra-uterine insemination is commonly accepted as a routine procedure for artificial insemination in horses. The motives and principles of deep insemination are well described, but the equipment used may differ. In this trial, the efficiency of two different insemination pipettes for deep intra-uterine insemination in the mare was compared with insemination into the uterine body using commercially available frozen-thawed semen of two stallions of proven fertility. These inseminations were performed using two different doses. The semi-flexible Minitube pipette was compared with a newly designed insemination device with a more flexible telescopic insemination catheter (Ghent device). The semi-flexible Minitube pipette performed better than the newly designed insemination device with respect to pregnancy outcome (p = 0.008). The superiority of deep horn insemination over uterine body insemination was reflected by the better pregnancy rates obtained after deep insemination using the same low doses (30.6% better pregnancy rates) (p = 0.0123).

Performance of an animal-based test of thirst in commercial broiler chicken farms.

Animal-based measures of thirst are currently absent from animal welfare monitoring schemes due to the lack of a well-validated indicator applicable for on-farm use. In the present study, an on-farm test based on voluntary water consumption from an unfamiliar open drinker was validated in a (semi-)commercial setting. To investigate the effect of thirst on water consumption, we subjected 4 flocks of 1,500 broilers to either 0 or 12 h of water deprivation and subsequently measured the amount of water that small subgroups consumed after the deprivation period (first experiment). Broilers that were water deprived before the test drank more than control broilers (P < 0.001). In a second experiment, a similar test was performed using 20 commercial broiler flocks in Belgium and Brazil. After a pretreatment water consumption test, the birds were subjected to 0 or 6 h of water deprivation, and a posttreatment water consumption test was conducted. Only in Brazil, deprived birds drank significantly more than controls in the posttreatment water consumption test (P < 0.001). A tendency for a difference was found in Belgium (P = 0.083). Pre- and posttreatment water consumption was higher in Brazil than in Belgium (P < 0.001). Stocking density and temperature influenced, respectively, the pretreatment and the control's posttreatment water consumption in Brazil, but not in Belgium. These results indicate that the water consumption test is sufficiently sensitive to discriminate between control and 12 h deprived flocks, and in Brazil even between control and 6 h deprived birds. The location of the test within the house did not affect the amount of water consumed in either experiment, suggesting that this variable does not have to be standardized. However, the amount of water consumed by broilers able to drink freely for a long period depended on indoor climatic variables (in Brazil only) and possibly genotype. This suggests that these variables need to be considered when interpreting the test outcome in terms of the thirst level experienced by the broilers.

Evaluation of the use of a heparin dose-response test in dogs to determine the optimal heparin dose during intravascular procedures and assessment of the in vitro heparin response in healthy dogs.

BACKGROUND: No guidelines for administering and monitoring anticoagulants intraprocedurally are currently available in dogs, despite the prevalence of procedures necessitating systemic anticoagulation with heparin. OBJECTIVES: To evaluate an activated clotting time (ACT)-based heparin dose-response (HDR) test to predict the individual required heparin dose in dogs during intravascular procedures, and to investigate both the in vitro heparin - ACT and in vitro heparin - factor anti-Xa activity (anti-Xa) relationships in dogs. METHODS: Blood was collected from eight healthy beagles undergoing a cardiac procedure and utilised to establish baseline ACT and for in vitro evaluation. Subsequently, 100 IU/kg heparin was administered intravenously (IV) and ACT was remeasured (HDR test). The required heparin dose for an ACT target response ≥300 s was calculated for each individual and ACT was remeasured after administration of this dose. For in vitro testing, a serial heparin blood dilution (0-0.5-1-2-4 international unit (IU)/mL) was prepared and ACT and anti-Xa were determined using whole blood and frozen plasma, respectively. RESULTS: The HDR test overestimated the required heparin dose in 3/7 dogs. In vitro, ACT and anti-Xa increased significantly with increasing blood heparin concentration. Heparin - ACT was nonlinear in 4/8 dogs at heparin concentrations >2 IU/mL, whereas heparin - anti-Xa remained linear throughout the tested range. CONCLUSIONS: The HDR test poorly estimated the required heparin dose in dogs. This is most likely attributed to a nonlinear heparin - ACT relationship, as observed in vitro. Anti-Xa is a promising alternative for ACT; however, unavailability as a point-of-care test and lack of in vivo target values restrict its current use.

Automated assessment of footpad dermatitis in broiler chickens at the slaughter-line: evaluation and correspondence with human expert scores.

Footpad dermatitis is increasingly used as an indicator of decreased broiler welfare, and automation of dermatitis monitoring potentially reduces the effort needed to monitor commercial flocks. In this study we evaluated a prototype system for the automatic assessment of footpad dermatitis in broiler chickens by comparing the automatic assessment with a human expert assessment. The expert aimed at selecting 2 times (different period) 20 broilers per footpad dermatitis category (5 categories in total), from 2 different flocks of 38-d-old broilers on an experimental farm. Two days later these broilers were transported to the slaughterhouse, where footpad dermatitis was assessed by the automatic system. Subsequently the footpads were reassessed by the same expert that had selected the birds. Automatic scores were only weakly correlated with scores given by the expert on-farm (r = 0.54) and at the slaughterhouse (r = 0.59). Manual evaluation of the photographs on which the automatic system based its scores revealed several errors. For 41.1% of the birds, the automatic system assessed only one of the footpads, whereas for 15.2% neither footpad was assessed. For 49.4% of the birds, scores were based on partially incorrectly identified areas. When data from such incomplete and obviously incorrect assessments were discarded, stronger correlations between automatic and expert scores were found (r = 0.68 and r = 0.74 for expert scores given on-farm and at-slaughter, respectively). Footpads that were missed by the automatic system were more likely to receive a high expert score at slaughter (P = 0.02). However, average flock scores did not differ greatly between automatic and expert scores. The prototype system for automatic dermatitis assessment needs to be improved on several points if it is to replace expert assessment of footpad dermatitis.

Dehydration indicators for broiler chickens at slaughter.

Freedom of (prolonged) thirst is considered to be of paramount importance for animal welfare. This emotion normally results from dehydration, which can be measured using physiological indicators. Because no reliable physiological indicator for thirst was available for broilers, we aimed to identify such a measure in this study. This indicator would ideally be integrated into quality control systems in commercial slaughter plants. In the first experiment, water deprivation was manipulated systematically by withdrawing water for different durations (total water withdrawal for 0 (control), 24, 36, or 48 h, or a 10-d period with restricted access to water for 2 times 10 min per day). A significant decrease in drained blood content and BW occurred from 36 h of total water deprivation onward (both P = 0.03), whereas long-term restricted access tended to decrease drained blood content (P = 0.05). No effect of water deprivation or restriction on skin turgor was found. In the second experiment, water was withdrawn for 0 (control), 6, 12, 24, or 48 h. Plasma chloride concentration was increased after 6 h of water withdrawal, but did not rise further with longer withdrawal. If assessed at slaughter, chloride will thus mainly reflect the catching-to-slaughter interval. In contrast, plasma creatinine and hematocrit levels showed a numerical decrease after 6 h of water withdrawal, but rose again after prolonged withdrawal. Plasma creatinine values were significantly higher in 24-h-deprived birds than in 6-h-deprived birds (P < 0.01), allowing for discernment between water withdrawal during catching and transport from dehydration that had occurred on the farm. Blood sodium concentrations and plasma osmolality showed a steady increment between 0 and 24 h of water deprivation (P < 0.001 and P < 0.001 for both), and may thus be used to assess the combined effects of water deprivation on farm and during the catching-to-slaughter interval. These findings may form the basis of an on-farm or at-slaughter test that could be included in integrated animal welfare assessment schemes.

Differential gene expression of the toll-like receptor-4 cascade and neutrophil function in early- and mid-lactating dairy cows.

During early lactation, neutrophils display several reduced immune functions. Particularly, a delayed recruitment of neutrophils into the infected udder seems to be one of the underlying events involved in the severity of postpartum Escherichia coli intramammary infections. The purpose of this study was to analyze the effect of in vitro chemotaxis and diapedesis on the expression of toll-like receptor-4 (TLR4)-related genes in bovine blood neutrophils isolated from 10 early-lactating (EL) and 10 mid-lactating (ML) cows. Functional characterization of the neutrophil population was performed by measuring phagocytosis and production of reactive oxygen species (chemiluminescence). Messenger RNA was extracted from neutrophils, and the expression of TLR4 and associated genes in EL and ML cows was analyzed by reverse-transcription quantitative PCR. To study the effect of chemotaxis and diapedesis on the expression of genes of the TLR4 cascade, neutrophils were stimulated to (trans)migrate in response to C5a using in vitro models. Our salient findings were that both neutrophil migration in vitro and lactation stage induced significant changes in the expression of several genes of the TLR4 signaling cascade. Before migration, expression of TRAF6, ATF3, RELA, IL8, and C5aR were lower in EL than in ML cows. Diapedesis and chemotaxis induced an increase in expression of TLR4, ATF3, and IL8 in both EL and ML cows. Diapedesis resulted in a downregulation of Syk, a TLR4-associated gene, in ML cows. This study shows that the perturbations in neutrophil functions during EL are accompanied by modulation of TLR4 pathway genes. These data can contribute to the understanding of the mechanisms explaining the relationship between stage of lactation and risk of severe E. coli mastitis.

Anaphylatoxin C5a-induced toll-like receptor 4 signaling in bovine neutrophils.

It is well known that signaling in neutrophils through both the complement component 5a (C5a) and C5a receptor (C5aR) and the toll-like receptor 4 (TLR4) pathways plays an essential role in innate defense. Neutrophil dysfunction, as seen during sepsis in severe mastitis during the periparturient period, is correlated with elevated concentrations of anaphylatoxin C5a. The aim of the current study was to elucidate the effect of C5a on TLR4 signaling in bovine neutrophils. Neutrophils were incubated with a high (but physiological) dose of purified C5a, and mRNA was extracted from neutrophils at different time points postincubation (PI). The incubation with C5a resulted in a biphasic C5aR expression profile, a phenomenon that might be explained by internalization (at 10 min PI) with subsequent reconstitution (starting at 40 min PI) of this receptor. The expression of TLR4, as well as its coreceptor, CD14, showed a similar biphasic change as observed with C5aR. In addition, changes in the mRNA expression levels of several genes belonging to the TLR4 pathway, such as TICAM-1, IKKα, and MAP3K7 were noted. The maximal expression of TLR4, CD14, and C5aR mRNA at 80 min PI was accompanied by a peak in IL8 mRNA, indicating that C5a is able to induce IL-8 production in neutrophils in vitro without the need of a costimulatory factor such as lipopolysaccharide. Moreover, a relatively constant expression of RELA was accompanied by increased expression of ATF3, an endogenous inhibitor of nuclear factor-κB mediated transcription, implying that C5a regulates TLR4 signaling and IL-8 synthesis independently. A significant time-dependent correlation was found between C5aR and TLR4, with the majority of the selected TLR4-dependent genes showing a significant correlation with C5aR at 80 min PI, when C5aR and TLR4 mRNA expression reached its maximum, suggesting crosstalk between both receptors. Taken together, this study showed that C5a is able to (1) alter the expression of genes belonging to the TLR4 pathway and (2) induce IL8 gene expression in bovine neutrophils. In addition, indications for cross-talk between complement activation and TLR4 signaling were found in the present study.

Association between navicular bone fragmentation and shape in Belgian Warmblood horses.

OBJECTIVES: To investigate whether in navicular bones (NB) from warmbloods, distal border fragmentation is associated with the shape of the proximal articular border or other radiological findings. METHODS: Radiographs of the front feet of 325 normal, subadult horses presented for admission as breeding stallions were reviewed. RESULTS: The proximal articular border of the total of 650 NB was classified as straight (n = 278), convex (n = 184), undulating (n = 147) or concave (n = 41). Distal border fragments were present in 57 NB (8.8%). They were significantly more prevalent in NB with a concave (9/41; 22%) or undulating (19/147; 13%) proximal articular border compared to NB with a straight (17/278; 6%) or convex shape (12/184; 7%). No other significant associations were found. CLINICAL SIGNIFICANCE: Hypothetically, since a shape-fragment association was found and the distribution of biomechanical forces exerted on the navicular bone are assumed to be shape dependent, distal border fragments may be a result of unfavourable loading of the navicular region.

The lipopolysaccharide model for the experimental induction of transient lameness and synovitis in Standardbred horses.

An established lipopolysaccharide (LPS) model previously described in Warmbloods, was inconsistent in Standardbred horses, where lameness was not detected despite the presence of synovitis. The present study aimed to determine the dose of LPS from E. coli O55:B5 required to induce mild to moderate lameness following middle carpal joint injection in Standardbred horses and to quantitate the induced lameness over time, with and without anti-inflammatory pre-treatment. In a baseline trial, eight healthy, clinically sound Standardbred horses were used in a rule-based dose-escalation design trial, starting at a dose of 10 endotoxin units (EU). Lameness at trot was evaluated visually and quantitatively (using an inertial-sensor system and pressure plate analysis). Synovial fluid aspirates were analysed for total nucleated cell counts, total protein and prostaglandin E2 (PGE2). Following 2 months wash-out, the effective LPS-dose determined in the baseline trial was used to evaluate the effect of anti-inflammatory treatment. A mixed model for repeated measures with horse as random effect was used for analysis. After injection of 10 EU LPS, the desired degree of lameness was observed in the baseline trial, with maximal lameness at post-injection hour (PIH) 4, followed by a rapid decline and return to baseline by PIH 48. No lameness was observed following pre-treatment with meloxicam. In synovial fluid, PGE2 was significantly higher at PIH 8 and PIH 24 in the baseline trial compared with following meloxicam pre-treatment. In conclusion, injection of the middle carpal joint with 10 EU LPS consistently induces a transient lameness and synovitis in Standardbred horses.

Replication kinetics of neurovirulent versus non-neurovirulent equine herpesvirus type 1 strains in equine nasal mucosal explants.

Equine herpesvirus type 1 (EHV-1) is the causative agent of equine herpes myeloencephalopathy, of which outbreaks are reported with increasing frequency throughout North America and Europe. This has resulted in its classification as a potentially emerging disease by the US Department of Agriculture. Recently, it was found that a single nucleotide polymorphism (SNP) in the viral DNA polymerase gene (ORF30) at aa 752 (N-->D) is associated with the neurovirulent potential of EHV-1. In the present study, equine respiratory mucosal explants were inoculated with several Belgian isolates typed in their ORF30 as D(752) or N(752), to evaluate a possible difference in replication in the upper respiratory tract. In addition, to evaluate whether any observed differences could be attributed to the SNP associated with neurovirulence, the experiments were repeated with parental Ab4 (reference neurovirulent strain), parental NY03 (reference non-neurovirulent strain) and their N/D revertant recombinant viruses. The salient findings were that EHV-1 spreads plaquewise in the epithelium, but plaques never cross the basement membrane (BM). However, single EHV-1-infected cells could be observed below the BM at 36 h post-inoculation (p.i.) for all N(752) isolates and at 24 h p.i. for all D(752) isolates, and were identified as monocytic cells and T lymphocytes. Interestingly, the number of infected cells was two to five times higher for D(752) isolates compared with N(752) isolates at every time point analysed. Finally, this study showed that equine respiratory explants are a valuable and reproducible model to study EHV-1 neurovirulence in vitro, thereby reducing the need for horses as experimental animals.

Identification of polymorphisms in the ovine Shadow of prion protein (SPRN) gene and assessment of their effect on promoter activity and susceptibility for classical scrapie.

Shadow of prion protein (SPRN) is an interesting candidate gene thought to be involved in prion pathogenesis. In humans, an association has already been discovered between mutations in SPRN and the incidence of variant and sporadic Creutzfeldt-Jakob disease. However, in sheep, the effect of mutations in SPRN is largely unknown. Therefore, we analysed the presence of mutations in the entire ovine SPRN gene, their association with scrapie susceptibility and their effect on SPRN promoter activity. In total, 26 mutations were found: seven in the promoter region, four in intron 1, seven in the coding sequence and eight in the 3' untranslated region. The mutations detected in the coding sequence and the promoter region were subsequently analysed in more detail. In the coding sequence, a polymorphism causing a deletion of two alanines was found to be associated with susceptibility for classical scrapie in sheep. Furthermore, a functional analysis of deletion constructs of the ovine SPRN promoter revealed that the region 464 to 230 bp upstream of exon 1 (containing a putative AP-2 and putative Sp1 binding sites) is of functional importance for SPRN transcription. Six mutations in the SPRN promoter were also found to alter the promoter activity in vitro. However, no association between any of these promoter mutations and susceptibility for classical scrapie was found.

Urinary markers in healthy young and aged dogs and dogs with chronic kidney disease.

BACKGROUND: Blood urea nitrogen and serum creatinine concentrations only detect a decrease of > 75% of renal functional mass. Therefore, there is a need for markers that allow early detection and localization of renal damage. HYPOTHESIS: Urinary albumin (uALB), C-reactive protein (uCRP), retinol binding protein (uRBP), and N-acetyl-beta-D-glucosaminidase (uNAG) concentrations are increased in dogs with chronic kidney disease (CKD) compared with healthy controls and in healthy older dogs compared with young dogs. ANIMALS: Ten dogs with CKD, 10 healthy young dogs (age 1-3 years), and 10 healthy older dogs (age > 7 years) without clinically relevant abnormalities on physical examination, hematology, biochemistry, and urinalysis. METHODS: Urinary markers were determined using an ELISA (uALB, uCRP, and uRBP) or a colorimetric test (uNAG). Results were related to urinary creatinine (c). The fixed effects model or the Wilcoxon rank sum test were used to compare the different groups of dogs. RESULTS: uALB/c, uRBP/c, and uNAG/c were significantly higher in CKD dogs than in healthy dogs. No significant difference was found for uCRP, which was not detectable in the healthy dogs and only in 3 of the CKD dogs. Between the healthy young and older dogs, no significant difference was detected for any of the markers. CONCLUSION: The urinary markers uALB/c, uRBP/c, and uNAG/c were significantly increased in dogs with CKD compared with healthy controls. Additional studies are needed to evaluate the ability of these markers to detect renal disease before the onset of azotemia.

Clinical and electrophysiological characterization of myokymia and neuromyotonia in Jack Russell Terriers.

BACKGROUND: Generalized myokymia and neuromyotonia (M/NM) in Jack Russell Terriers (JRTs) is related to peripheral nerve hyperexcitability syndrome in humans, a symptom complex resulting from diverse etiologies. OBJECTIVE: Clinical and electrodiagnostic evaluation is used to narrow the list of possible etiological diagnoses in JRTs with M/NM. ANIMALS: Nine healthy JRTs and 8 affected JRTs. METHODS: A prospective study was conducted comparing clinical and electrophysiological characteristics in 8 JRTs affected by M/NM with 9 healthy JRT controls. RESULTS: All affected dogs except 1 had clinical signs typical of hereditary ataxia (HA). In 6 dogs, neuromyotonic discharges were recorded during electromyogram. Motor nerve conduction studies showed an axonal neuropathy in only 1 affected dog. Compared with controls, brainstem auditory-evoked potentials (BAEP) showed prolonged latencies (P<.05) accompanied by the disappearance of wave components in 3 dogs. Onset latencies of tibial sensory-evoked potentials (SEP) recorded at the lumbar intervertebral level were delayed in the affected group (P<.001). The BAEP and SEP results of the only neuromyotonic dog without ataxia were normal. CONCLUSIONS AND CLINICAL IMPORTANCE: The BAEP and spinal SEP abnormalities observed in JRTs with M/NM were associated with the presence of HA. Therefore, these electrophysiological findings presumably arise from the neurodegenerative changes characterizing HA and do not directly elucidate the pathogenesis of M/NM. An underlying neuronal ion channel dysfunction is thought to be the cause of M/NM in JRTs.

Elbow lameness in dogs of six years and older: arthroscopic and imaging findings of medial coronoid disease in 51 dogs.

OBJECTIVES: To report on the frequency and distribution of lesions of the medial coronoid process in dogs of six years of age and older, and to describe the arthroscopic findings in the affected joints. To compare those lesions seen in 'old' dogs with those seen in 'young' dogs between the ages of five and 18 months. METHODS: A retrospective study of dogs six-years-old and older admitted for elbow lameness and subsequent elbow arthroscopy. The dogs were divided into groups according to the lesions of the medial coronoid process diagnosed during arthroscopic examination and computed tomography (CT). The radiographic degree of osteoarthritis (OA) and arthroscopically diagnosed lesions on the medial humeral condyle are described. RESULTS: In 51 'old' dogs, five types of lesions could be identified on arthroscopy and CT: chondromalacia-like lesions (2%), fissures (27.5%), non-displaced fragments (12%), displaced fragments (27.5%), and erosions within the medial compartment without fragmentation (31%). A significantly different distribution of lesions was seen in 'young' dogs: fissures (23%), non-displaced fragments (45%), displaced fragments (29%), and erosions within the medial compartment without fragmentation (3%). No difference in radiographic degree of OA was seen between the two groups. CLINICAL SIGNIFICANCE: This study demonstrates the relatively high incidence of medial coronoid disease in dogs older than six years of age, and it highlights one particular problem in 'old' dogs: the complete erosion of the medial compartment.