Encephalitis and neuronal necrosis in a 3-month-old suckled beef calf.

A 3-month-old suckled beef calf from the west coast of Scotland showed neurologic clinical signs for 1 week and was euthanized after failing to respond to treatment. Blood and tissue samples, including the brain, were submitted for diagnosis. Histologic examination of the brain showed neuronal chromatolysis and necrosis in the hind brain and loss of Purkinje cells in the cerebellum, accompanied by mild nonsuppurative encephalitis in the hind brain with a striking lack of inflammation in the cerebellar layers. Other microscopic lesions present were mild nonsuppurative meningitis with perivascular cuffs, diffuse hypergliosis, and occasional foci of neuronophagia. Polymerase chain reaction amplification of viral nucleic acids and specific immunohistochemical labeling allowed the identification of louping ill virus, and serology showed high titers of immunoglobulin M, indicating a recent infection.

Diagnostic exercise: generalized alopecia and mural folliculitis in a goat.

An 18-month-old cross-bred goat was presented with generalized erythema and thinning of the hair coat, as well as localized moderate scaling. Histopathological evaluation of skin biopsies showed hyperplasia and marked disruption of the infundibular epithelium owing to a predominant infiltrate of macrophages with multinucleated histiocytic giant cells and some lymphocytes, plasma cells, and eosinophils. Examination of peripheral blood and skin by polymerase chain reaction gave positive results for ovine herpesvirus type 2 consistent with a diagnosis of malignant catarrhal fever.

Typology of obsessive-compulsive symptoms in children and adolescents with anorexia nervosa.

BACKGROUND: Previous studies have shown that symptoms of obsessive compulsive disorder are common in both adults and children and adolescents with anorexia nervosa. Until now, no study has explored the specific obsessive compulsive symptoms shown in children and adolescents with AN. METHOD: In this study we report types of symptoms displayed by young people with AN and explore similarities and differences with adults with AN and with people with OCD. RESULTS: Common obsessions concerned contamination, aggressive and somatic concerns, and common compulsions were related to ordering/arranging, and checking. CONCLUSION: It is important for clinicians to be aware of the obsessions and compulsions they are most likely to encounter when working with these patients. If missed or ignored, such symptoms may interact with eating disorder symptoms and impede treatment.

First cases of squirrelpox in red squirrels (Sciurus vulgaris) in Scotland.

Squirrelpox, caused by a poxvirus, is a major threat to the remaining UK red squirrel population. The spread of antibody-positive grey squirrels has been monitored in the UK for the past decade. In 2005 grey squirrels that had been exposed to the virus appeared in the south of Scotland for the first time, followed approximately two years later by the appearance of squirrelpox disease in the local red squirrels. Four squirrels were examined. They all had gross external lesions and histological lesions typical of squirrelpox disease, but no significant internal lesions. The diagnosis was confirmed by PCR, electron microscopy and serology.

Proximal femoral osteotomy in children with cerebral palsy: the perspective of the trainee.

BACKGROUND: There are a range of implants for fixation of proximal femoral osteotomies (PFOs) in children. We investigated the training experiences and preferences of orthopaedic residents and fellows who were learning PFO, using a fixed angled blade plate (ABP) or a locking, cannulated blade plate (LCBP). We also studied short-term technical and radiographic outcomes. METHODS: This was a prospective, parallel-group, cohort study of 90 consecutive children and adolescents with cerebral palsy who underwent bilateral PFOs with ABP or LCBP. Surgical trainees completed a questionnaire to document the ease or difficulty of each operative step. RESULTS: There were 48 boys and 42 girls, with a mean age of eight years and a mean follow-up of 25 months. Trainees preferred the LCBP system for: insertion of the guidewire, the seating chisel and the blade plate, as well as overall technical ease of use (p < 0.001). Radiographic outcomes were similar with no between-group differences for migration percentage (p = 0.996) or neck shaft angle (p = 0.849), but there was a higher prevalence of technical errors in the ABP group. CONCLUSIONS: Trainee surgeons expressed a preference for LCBPs when learning PFO in children with cerebral palsy. Radiographic outcomes were similar in both groups, with close attending surgeon supervision.

Development of a real time RT-PCR to detect and type ovine pestiviruses.

A real time one-step RT-PCR was designed to detect and type border disease virus (BDV), bovine viral diarrhea virus (BVDV) type 1 and BVDV type 2 in ovine samples. The real time RT-PCR was shown to behave in a linear manner and had limits of detection of 100-1000 copies of viral RNA as judged by in vitro transcribed RNA. The real time RT-PCR was validated on 50 clinical samples from UK flocks and was more sensitive than a virus isolation and a classical nested RT-PCR (nRT-PCR). The results of real time RT-PCR virus typing agreed completely with sequencing. The majority of ovine isolates were BDV; a small proportion were BVDV type 1. BVDV type 2 was not detected in any sample. This test appears reliable and can be used for the typing of ovine pestiviruses in the UK.

Detection of Louping ill virus in clinical specimens from mammals and birds using TaqMan RT-PCR.

The identification of Louping ill virus (LIV) in clinical specimens has been routinely achieved by virus isolation using susceptible pig kidney cells and subsequent serological analysis. While this method is sensitive and detects infectious virus, it is relatively labour intensive and time-consuming. In view of the veterinary and potential medical importance of LIV, a rapid and precise detection method for routine use that employs the TaqMan reverse transcription polymerase chain reaction (RT-PCR) has been developed to detect LIV RNA extracted from field samples. The TaqMan assay was evaluated against virus isolation using 22 cell culture grown LIV isolates, which had previously been partially characterised by sequencing, and material from 63 suspect field cases. Histopathological and/or serological reports were available for 39 of the suspect cases, providing additional diagnostic information to evaluate the results obtained from the TaqMan RT-PCR assay. The TaqMan assay was as sensitive as the cell culture infectious virus assay currently used and had the advantage that it was able to detect LIV in clinical specimens from which infectious virus could not be isolated possibly due to the presence of high levels of LIV antibody.

Ionizing radiation modulates vascular endothelial growth factor (VEGF) expression through multiple mitogen activated protein kinase dependent pathways.

We investigated the role of radiation-induced mitogen activated protein kinase (MAPK) pathway activity in the regulation of proliferation, cell survival and vascular endothelial growth factor (VEGF) production in primary astrocytes and in T9 and RT2 glioblastoma cells derived from Fisher 344 rats. In these cells, ionizing radiation (2 Gy) caused activation of the MAPK pathway which was blocked by specific inhibitor drugs. Blunting of radiation-induced MAPK activity weakly enhanced radiation-induced apoptosis 24 h after exposure in RT2 cells. Furthermore, blunting of MAPK activation weakly enhanced the ability of radiation to reduce RT2 cell growth in clonogenic growth assays. These findings argue that inhibition of MAPK signaling reduces proliferation and enhances cell killing by ionizing radiation in transformed astrocytes. Proliferation and survival of cancer cells has been linked in vivo to enhanced expression of angiogenic growth factors. Recently we demonstrated that the gene product of a novel rodent radiation-responsive gene, progression elevated gene 3 (PEG-3), could enhance vascular endothelial growth factor (VEGF) promoter activity in rodent fibroblasts, leading to increased VEGF protein levels and tumorigenic behavior in vivo. Thus PEG-3 and VEGF expression could be expected to directly correlate with the oncogenic potential of transformed cells. RT2 cells expressed more PEG-3 and VEGF protein than T9 cells, and were more tumorigenic in vivo than T9 cells. Radiation activated the PEG-3 promoter via MAPK signaling and ectopic over-expression of PEG-3 enhanced both basal MAPK activity and basal VEGF promoter activity. Basal MAPK activity partially correlated with basal VEGF promoter activity and VEGF protein levels in primary astrocytes, T9 and RT2 cells. Radiation increased the activity of the VEGF promoter and VEGF protein levels in primary astrocytes, T9 and RT2 cells which were dependent upon MAPK function. Furthermore, inhibition of AP-1 transcription factor signaling by dominant negative c-Jun (TAM67) also significantly reduced basal, and to a lesser extent radiation-induced, VEGF promoter function in RT2 cells. Collectively, our data demonstrate that radiation-induced MAPK signaling can both protect cells from radiation-induced cell death as well as enhance protein levels of pro-angiogenic factors such as VEGF. Enhanced VEGF expression in RT2 cells may be mediated via MAPK and JNK pathway signaling which converges upon the AP-1 transcription factor complex.

The radiological assessment of pelvic obliquity in cerebral palsy and the impact on hip development.

Pelvic obliquity is a common finding in adolescents with cerebral palsy, however, there is little agreement on its measurement or relationship with hip development at different gross motor function classification system (GMFCS) levels. The purpose of this investigation was to study these issues in a large, population-based cohort of adolescents with cerebral palsy at transition into adult services. The cohort were a subset of a three year birth cohort (n = 98, 65M: 33F, with a mean age of 18.8 years (14.8 to 23.63) at their last radiological review) with the common features of a migration percentage greater than 30% and a history of adductor release surgery. Different radiological methods of measuring pelvic obliquity were investigated in 40 patients and the angle between the acetabular tear drops (ITDL) and the horizontal reference frame of the radiograph was found to be reliable, with good face validity. This was selected for further study in all 98 patients. The median pelvic obliquity was 4° (interquartile range 2° to 8°). There was a strong correlation between hip morphology and the presence of pelvic obliquity (effect of ITDL on Sharpe's angle in the higher hip; rho 7.20 (5% confidence interval 5.59 to 8.81, p < 0.001). This was particularly true in non-ambulant adolescents (GMFCS IV and V) with severe pelvic obliquity, but was also easily detectable and clinically relevant in ambulant adolescents with mild pelvic obliquity. The identification of pelvic obliquity and its management deserves closer scrutiny in children and adolescents with cerebral palsy.

Alterations in calcium-mediated signal transduction after traumatic injury of cortical neurons.

Calcium influx and elevation of intracellular free calcium ([Ca2+]i), with subsequent activation of degradative enzymes, is hypothesized to cause cell injury and death after traumatic brain injury. We examined the effects of mild-to-severe stretch-induced traumatic injury on [Ca2+]i dynamics in cortical neurons cultured on silastic membranes. [Ca2+]i was rapidly elevated after injury, however, the increase was transient with neuronal [Ca2+]i returning to basal levels by 3 h after injury, except in the most severely injured cells. Despite a return of [Ca2+]i to basal levels, there were persistent alterations in calcium-mediated signal transduction through 24 h after injury. [Ca2+]i elevation in response to glutamate or NMDA was enhanced after injury. We also found novel alterations in intracellular calcium store-mediated signaling. Neuronal calcium stores failed to respond to a stimulus 15 min after injury and exhibited potentiated responses to stimuli at 3 and 24 h post-injury. Thus, changes in calcium-mediated cellular signaling may contribute to the pathology that is observed after traumatic brain injury.

A new model for rapid stretch-induced injury of cells in culture: characterization of the model using astrocytes.

The purpose of this study was to develop a simple, reproducible model for examining the morphologic, physiologic, and biochemical consequences of stretch-induced injury on tissue-cultured cells of brain origin. Rat cortical astrocytes from 1- to 2-day-old rats were cultured to confluency in commercially available 25-mm-diameter tissue culture wells with a 2-mm-thick flexible silastic bottom. A cell injury controller was used to produce a closed system and exert a rapid positive pressure of known amplitude (psi) and duration (msec). The deformation of the membrane, and thus the stretch of the cells growing on the membrane, was proportional to the amplitude and duration of the air pressure pulse. Extent of cell injury was qualitatively assessed by light and electron microscopy and quantitatively assessed by nuclear uptake of the fluorescent dye propidium iodide, which is excluded from cells with intact membranes. Lactate dehydrogenase (LDH) enzyme release was measured spectrophotometrically. Cell injury was found to be proportional to the extent of the silastic membrane deformation. Increasing cell stretch caused mitochondrial swelling and vacuolization as well as disruption of glial filaments. Stretching also caused increased dye uptake, with maximum dye uptake occurring with a 50 msec pressure pulse duration, whereas deformations produced over longer periods of time (seconds) caused little dye uptake. With increasing postinjury survival fewer cells took up dye, implying cell repair. LDH release was also proportional to the amplitude of cell stretch, with maximum release occurring within 2 h of injury. In summary we have developed a simple, reproducible model to produce graded, strain-related injuries in cultured cells. Our continuing experiments suggest that this model can be used to study the biochemistry and physiology of injury as well as serve as a tool to examine the efficacy of therapeutic agents.

Astrocytes generate isoprostanes in response to trauma or oxygen radicals.

Previous studies have shown that oxygen radical scavengers prevent the reduced cerebral blood flow that occurs following experimental traumatic brain injury. The exact chemical species responsible for the posttraumatic reduction in flow is unknown. We tested whether isoprostanes, which are formed by non-cyclooxygenase-dependent free radical attack of arachidonic acid and are vasoconstrictors of the cerebral circulation, are increased in astrocytes following stretch-induced trauma or injury with a free radical generating system. Isoprostane (8-epi-prostaglandin F2alpha) was analyzed in cells and in media by immunoassay. Confluent rat cortical astrocytes in culture were injured by a hydroxyl radical generating system consisting of hydrogen peroxide and ferrous sulfate or by rapid stretch of astrocytes grown on a deformable silastic membrane. Some cells were treated with the iron chelator deferoxamine for 1 h before injury. The hydroxyl generating system caused free and cell-bound isoprostanes to increase to more than 400% of control. After trauma, free and membrane bound isoprostanes increased to 321 +/- 34% and 229 +/- 23% of control, respectively, and posttraumatic increases were prevented by deferoxamine. Since astrocytes are in close proximity to cerebral vessels, posttraumatic free radical formation may increase the formation of isoprostanes, which in turn produce vasoconstriction and decrease cerebral blood flow.

Influence of phenylmethylsulfonyl fluoride on anandamide brain levels and pharmacological effects.

The endogenous cannabinoid anandamide produces cannabimimetic effects similar to those produced by delta9-tetrahydrocannabinol (delta9-THC), but has a much shorter duration of action due to its rapid metabolism to arachidonic acid and polar metabolites via action of fatty acid amide hydrolase (FAAH). Our earlier observations that anandamide's effects persisted after brain levels of anandamide itself had substantially dropped prompted us to examine the influence of the irreversible amidase inhibitor, phenylmethyl sulfonyl fluoride (PMSF), on the brain levels and pharmacological effects of anandamide. As shown previously, pretreatment with PMSF resulted in a leftward shift of the anandamide dose effect curves for antinociception and hypothermia in male mice. Brain and plasma levels of anandamide, arachidonic acid and polar metabolites peaked at 1 min after i.v. injection with 3H-anandamide and remained high at 5 min post-injection, with levels falling sharply thereafter. Pretreatment with PMSF (30 mg/kg, i.p.) prior to an injection of 1 or 10 mg/kg 3H-anandamide resulted 5 min later in enhanced brain levels of anandamide compared to those obtained with 3H-anandamide plus vehicle injection. Levels of arachidonic acid and polar metabolites in brain were not significantly increased. The clear correspondence between brain levels of anandamide following pretreatment with PMSF and pharmacological activity suggests that this parent compound is responsible for the antinociception and hypothermia that occurred 5 min after injection. These results further suggest that metabolite contribution to anandamide's effects, if any, would occur primarily at later times.

Herpesviruses of carnivores.

This review focuses on felid herpesvirus 1 (FHV-1), the most studied of the carnivore herpesviruses. Canid herpesvirus (CHV-1) and phocid (seal) herpesvirus 1 (PhHV-1) are also included where information is available. FHV-1 is a member of the Varicellovirus genus of the Alphaherpesvirinae, which appears to be closely related phylogenetically to both CHV-1 and PhHV-1. FHV-1 infects both domestic and some wild Felidae, such as cheetahs, and is predominantly a respiratory pathogen of cats. As in other herpesviruses, infection with FHV-1 is characterised by a latent carrier state, during which intermittent shedding of infectious virus may occur. Typical of an alphaherpesvirus, the primary site of FHV-1 latency is neurological tissue (trigeminal ganglion), though recent studies using the polymerase chain reaction have suggested that some latency may occur in non-neurological sites. Latently infected carriers are epidemiologically important as sources of infection for susceptible animals. Though conventional modified live and inactivated vaccines have been available for a number of years, they do not protect against infection nor the development of latency. Recently, work has focused on molecular characterisation of FHV-1, detecting genes such as glycoproteins or regulatory genes. Such work will enable better understanding of the interaction of FHV-1 with the natural host. Deletion mutants of some of these genes may also have potential as vaccine strains.

Molecular phylogenetic analysis of felid herpesvirus 1.

The position of felid herpesvirus 1 within the alphaherpesvirus subfamily was investigated using molecular phylogenetic techniques applied to multiple sequence alignments of recently reported FHV-1 gene homologs (glycoprotein B, ribonucleotide reductase and DNA polymerase). FHV-1 was most closely related to other carnivore alphaherpesviruses, (phocid herpesvirus 1 and canid herpesvirus 1) and to the equid herpesviruses 1 and 4.

A field trial to assess the effect of vaccination against feline herpesvirus, feline calicivirus and feline panleucopenia virus in 6-week-old kittens.

A trivalent (feline panleucopenia, feline herpesvirus, feline calicivirus), modified live, commercially available cat vaccine was used at either 6, 9 and 12 weeks of age (early schedule) or 9 and 12 weeks of age (conventional schedule), and the serological response to vaccination was assessed. The level of maternally derived antibody present at 6 weeks of age was also established. The use of early vaccination at 6 weeks of age induced an antibody response to each virus by 9 weeks of age in a significant proportion of kittens compared with unvaccinated littermates. There was no difference between the conventionally and early-vaccinated groups in terms of antibody response to any antigen by 12 and 15 weeks of age.

Feline leucocyte antigen class II polymorphism and susceptibility to feline infectious peritonitis.

There are four outcomes to feline coronavirus (FCoV) infection: the development of feline infectious peritonitis (FIP, which is immune-mediated), subclinical infection, development of healthy lifelong carriers and a small minority of cats who resist infection (Addie and Jarrett, Veterinary Record 148 (2001) 649). Examination of the FCoV genome has shown that the same strain of virus can produce different clinical manifestations, suggesting that host genetic factors may also play a role in the outcome of infection. FIP is most prevalent amongst pedigree cats, although how much of this is due to them living in large groups (leading to higher virus challenge and stress which predisposes to FIP) and how much is due to genetic susceptibility is not known. If host genetics could be shown to play a role in disease, it may allow the detection of cats with a susceptibility to FIP and the development of increased population resistance through selective breeding. The feline leucocyte antigen (FLA) complex contains many genes that are central to the control of the immune response. In this preliminary study, we used clonal sequence analysis or reference strand conformational analysis (RSCA) to analyse the class II FLA-DRB of 25 cats for which the outcome of FCoV exposure was known. Individual cats were shown to have between two and six FLA-DRB alleles. There was no statistically significant association between the number of alleles and the outcome of FCoV infection. No particular allele appeared to be associated with either the development of FIP, resistance to FCoV, or the carrier status. However, the analysis was complicated by apparent breed variation in FLA-DRB and the small number of individuals in this study.

Hereditary cerebellar degeneration in three full sibling kittens.

Two domestic shorthair littermate kittens had signs of cerebellar dysfunction, first observed between seven and eight weeks of age; a third littermate was unaffected. The signs were progressive and the more severely affected kitten was euthanased after six days. A postmortem examination revealed no gross lesions but the kitten had cerebellar cortical degeneration with extensive loss of Purkinje cells. The second kitten was euthanased at 10 months of age with similar, though more pronounced, changes. One of the two kittens in the next litter of the same parents had similar clinical signs and histopathological findings. The lesions in the cerebellum are interpreted as probably due to genetically determined abiotrophy. In addition, the two older kittens had medullary neuronal changes interpreted as probable neuraxonal dystrophy, and focal vacuolation of the neuropil in the medulla and cervical spinal cord.

Spongiform encephalopathy in a captive puma (Felis concolor).

A captive adult puma developed ataxia, a hypermetric gait and whole body tremor. The signs progressed over a period of six weeks. Histopathological examination following euthanasia demonstrated spongiform encephalopathy, gliosis and mild non-suppurative meningoencephalitis. Immunostaining with a polyclonal antiserum revealed prion protein (PrP) associated with these changes in sections of cervical spinal cord and medulla. This is the first confirmed case of a scrapie-like spongiform encephalopathy described in a non-domestic cat in the United Kingdom.