bluuebunny/crossref_metadata_2025 huggingface dataset:
Optimized Processing of Fine-Needle Lymph Node Biopsies for Automated Immunostaining
Authors: Aulbach Adam D. , Swenson Cheryl L. , Kiupel Matti
Published: May 2011

A straightforward, reliable technique for postcollection processing and evaluation of cytologic specimens for antigen detection using an automated immunostainer was developed. Visual assessment of cell suspension turbidity was used in parallel with light microscopic examination of concentrated cytospin preparations to verify the diagnostic utility of samples for immunocytochemical staining. Fine-needle lymph node biopsies from 81 dogs with lymphadenomegally and a cytologic or histologic diagnosis of lymphoma were introduced into ethylenediamine tetra-acetic acid tubes containing standardized storage media. Cell suspension turbidity was assessed to estimate cell concentration and resultant volume required for cytospin preparations with optimal cellularity. Preliminary cytospin preparations (using estimated volumes based upon turbidity) were stained using modified Wright stain and examined microscopically for intact neoplastic cell concentration. Once an optimal volume for cytospin preparations was established, additional concentrated slides were prepared for immunophenotyping, using an automated immunostainer and antibodies specific for cluster of differentiation (CD)79a and CD3e. All cell suspension samples with adequate gross turbidity had ample intact neoplastic cell concentration for immunocytochemical staining. Based on CD79a and CD3e expression, 51 (63%) B cell, 19 (23%) T cell, 3 mixed T and B cells (4%), and 3 non-T- and non-B-cell lymphomas (4%), as well as 5 (6%) nondiagnostic samples were identified. Three out of 5 of the nondiagnostic samples were submitted early in the investigation prior to the establishment of gross specimen turbidity guidelines. Immunocytochemical staining results were in complete agreement with all 6 available immunohistochemical correlates. The ability to visually assess sample adequacy prior to sample submission may encourage more widespread use of immunocytochemical techniques.
Identification of Atypical Scrapie in Canadian Sheep
Authors: Mitchell Gordon B. , O'Rourke Katherine I. , Harrington Noel P. , Soutyrine Andrei , Simmons Marion M. , Dudas Sandor , Zhuang Dongyue , Laude Hubert , Balachandran Aru
Published: May 2011

Scrapie, a transmissible spongiform encephalopathy of sheep and goats, exists in most small ruminant-producing countries of the world. A novel form of this disease was recently recognized and is known by various names, including Nor98, Nor98-like, and atypical scrapie. Differing from classic scrapie in epidemiology, histopathology, and biochemical characteristics, atypical scrapie cases have been identified throughout Europe and in the United States. Enhanced scrapie surveillance efforts recently identified 3 cases of atypical scrapie in Canada.
Molecular Detection of Autosomal-Dominant Feline Polycystic Kidney Disease by Multiplex Amplification Refractory Mutation System Polymerase Chain Reaction
Authors: Lee Ya-Jane , Chen Hsin-Yu , Wong Min-Liang , Hsu Wei-Li
Published: May 2011

Feline autosomal-dominant polycystic kidney disease (ADPKD), with its characteristic growth of fluid-filled cysts of different sizes, is the most prevalent inherited genetic disease of cats. The point mutation (C→A transversion) in exon 29 of the PKD1 gene is known to contribute to ADPKD development and can thus serve as a target for the molecular genetic diagnosis of ADPKD. To this end, a simple amplification refractory mutation system (ARMS) polymerase chain reaction (PCR) was designed with 3 primers: 2 forward primers specifically targeting either the mutant or normal allele, and 1 universal reverse primer for amplification of both alleles. The new method was tested on the DNA from 35 feline blood samples, which included 15 mutant cats and 20 wild type cats. As verified by direct DNA sequencing, both sensitivity and specificity of this tri-primer ARMS PCR were 100%. As the multiplex ARMS PCR test can be performed in a single PCR reaction without other post-PCR procedures, it is a simple and accurate method for molecular studies of feline ADPKD.
Amplification of the Second Internal Transcribed Spacer Ribosomal DNA of Individual Trichostrongylid Nematode Larvae by Nested Polymerase Chain Reaction
Authors: Sim Kathrin A. , Hoar Bryanne , Kutz Susan J. , Chilton Neil B.
Published: May 2011

The second internal transcribed spacer (ITS2) of ribosomal DNA (rDNA) is used as a genetic marker to identify trichostrongylid nematodes. However, it is often difficult to amplify by polymerase chain reaction (PCR) the ITS2 rDNA of a single trichostrongylid nematode larva or egg. A nested PCR (nPCR) assay was, therefore, developed to amplify the ITS2 from individual trichostrongylid nematode larvae. The results show that the ITS2 rDNA of a significantly greater proportion of individual larvae was amplified using nPCR compared with a standard PCR. There was also no need to column-purify the genomic DNA before nPCR, which is more time and cost effective for studies involving large sample sizes. The amplicons produced from the secondary phase of the nPCR were subjected to single-strand conformation polymorphism analyses and DNA sequencing to confirm the species identity of the larvae used in the current study as Ostertagia gruehneri. The nPCR assay was also used to amplify the ITS2 from individual trichostrongylid eggs. The ability to amplify the ITS2 rDNA from large numbers of individual nematode eggs and larvae has important implications for diagnostic testing and for conducting epidemiological studies on these parasites of veterinary importance.
An Outbreak of Lethal Toxoplasmosis in Pigs in the Gansu Province of China
Authors: Li Xuerui , Wang Yanhua , Yu Fenggang , Li Tongxiang , Zhang Delin
Published: May 2011

In October 2004, a swine farm in Jinchang, Gansu Province, China, experienced an outbreak of toxoplasmosis. Most of the affected pigs had a rectal temperature greater than 40°C and gradually lost their appetite. Morbidity reached 57%, and mortality was approximately 2%. Analysis of blood samples from affected pigs using hemagglutination inhibition (HI) assay, immunoglobulin G—enzyme-linked immunosorbent assay (IgG-ELISA), and IgM-ELISA tests showed high titers of anti— Toxoplasma gondii antibody. Tachyzoites of T. gondii were found in body fluids of mice inoculated intraperitoneally with ground samples from the heart, liver, spleen, and brain of 2 sick pigs. In addition, the inoculation of 5 pigs with T. gondii tachyzoites caused death in 2 of the pigs. The origin of this outbreak was concluded to be food-borne T. gondii infection.
Uterine Endometrial Polyp with Severe Hemorrhage and Cystic Endometrial Hyperplasia—Pyometra Complex in a Dog
Authors: Gumber Sanjeev , Springer Nora , Wakamatsu Nobuko
Published: May 2011

The current report describes an unusual presentation of uterine endometrial polyp with severe hemorrhage and cystic endometrial hyperplasia—pyometra complex in a 9.5-year-old female Doberman Pinscher. The dog presented with a 2-day history of bloody discharge from the vulva and an enlarged abdomen. The postmortem examination revealed a markedly distended right uterine horn with a large pedunculated mass (17 cm × 9 cm × 4 cm) and blood. Based on the histological findings, the diagnosis of uterine endometrial polyp was made.
Urinary Bladder Agenesis in an Alpaca (Vicugna Pacos) Cria
Authors: Melidone Raffaele , Parry Nicola M. A. , Gruntman Alisha
Published: May 2011

A 10-day-old female alpaca ( Vicugna pacos) cria with a history of urinary straining and dribbling was presented for evaluation. The animal had markedly elevated blood fibrinogen (800 mg/dl), mildly elevated phosphorus (9.3 mg/dl), and minimally elevated blood urea nitrogen (38 mg/dl) concentrations. The total protein (5.0 g/dl) concentration was mildly decreased. These findings were suggestive of mild renal disease. An abdominal ultrasound revealed bilateral hydronephrosis and hydroureter, and no urinary bladder was identified. Gross postmortem examination revealed urinary bladder agenesis and bilateral hydronephrosis and hydroureter, with both ureters opening into a sinus in the caudal vagina. Histologic examination of the kidneys showed necrosuppurative pyelonephritis with pelvic dilation, and both ureters had mild lymphoplasmacytic and histiocytic inflammation.
Congenital Cervical Bronchogenic Cyst in a Calf
Authors: Lee Jae Y. , Yoon In H. , Cho Seong W. , Jeong Seong M. , Park Chang S. , Kim Myung C.
Published: May 2011

The present report describes an extrathoracic bronchogenic cyst in a 30-day-old female calf. Histologically, the cyst wall was lined by a layer of ciliated pseudostratified columnar epithelium with peripheral arrangement of cartilage, glands, and smooth muscle fascicles. The mass was successfully removed by simple surgical excision.
Avian Bornavirus is Present in Many Tissues of Psittacine Birds with Histopathologic Evidence of Proventricular Dilatation Disease
Authors: Raghav Raj , Taylor Michael , DeLay Josepha , Ojkic Davor , Pearl David L. , Kistler Amy L. , DeRisi Joseph L. , Ganem Donald , Smith Dale A.
Published: May 2011

Proventricular dilatation disease (PDD) is a neurologic disease of psittacine birds suspected to be caused by a recently identified Avian bornavirus (ABV). In the current report, data supporting the causal association of ABV with PDD are presented. Immunohistochemistry (IHC) with rabbit polyclonal antiserum raised against ABV nucleocapsid protein was used to identify cell and organ distribution of viral antigen. The ABV antigen was most consistently detected in brain, spinal cord, adrenal gland, pancreas, and kidney. Histopathologic evaluation was correlated with ABV-specific polymerase chain reaction (PCR) and immunohistochemical tests in multiple tissues from 16 psittacine birds with and without PDD. Using histopathologic diagnosis as the gold standard, the sensitivity and specificity of IHC for ABV antigens were found to be 100% and 100%, respectively. Many more tissues were positive for ABV RNA by reverse transcription PCR than were positive for pathologic changes or viral antigens by IHC, indicating the presence of subclinical or asymptomatic infection at many sites.
Blood Steroid Concentrations in Domestic Mongolian Horses
Authors: Haffner John C. , Fecteau Kellie A. , Eiler Hugo , Tserendorj Tsek , Hoffman Rhonda M. , Oliver Jack W.
Published: May 2011

Traditionally, analysis of blood cortisol alone has been used to evaluate adrenal function. Currently, multisteroid analyses are considered more informative than analysis of a single hormone to assess adrenal function. The objective of the present research was to create a database for steroid reference values for domestic Mongolian horses. Seven adrenal steroid levels were determined in the blood of 18 colts, 34 stallions, 25 geldings, 17 fillies, and 29 mares. Results were as follows (lowest and highest group median, in nanograms per milliliter): progesterone: <0.030 (fillies), 4.30 (mares), and 0.070 (all horses); 17-OH-progesterone: 0.070 (colts), 0.520 (mares), and 0.110 (all horses); androstenedione: 0.101 (colts), 0.256 (stallions), and 0.181 (all horses); testosterone: <0.040 (mares, stallions, and fillies), 0.040 (geldings and colts), and <0.40 (all horses); estradiol: 0.066 (stallions), 0.093 (fillies), and 0.085 (all horses); cortisol: 23.040 (colts), 70.210 (geldings), and 50.770 (all horses); and aldosterone: 0.018 (colts), 0.297 (geldings), and 0.191 (all horses). Overall medians indicate that cortisol (98.70%) is the predominant steroid, followed by aldosterone (0.37%), androstenedione (0.35%), 17-OH-progesterone (0.21%), estradiol (0.17%), progesterone (0.14%), and testosterone (0.06%). This information provides adrenal and gonadal steroid reference concentrations to assist in physiological characterization and diagnosis of endocrine disorders in domestic Mongolian horses.
Evaluation of Three Automated Human Immunoturbidimetric Assays for the Detection of C-Reactive Protein in Dogs
Authors: Klenner Stefanie , Bauer Natali , Moritz Andreas
Published: May 2011

C-reactive protein (CRP) is a major, acute-phase protein in dogs; however, there is a need for automated assays to ensure in-time patient monitoring. Three automated immunoturbidimetric assays (Randox, Thermo, and Wako) developed for human beings were evaluated for their ability to detect canine CRP, including method validation, evaluation of diagnostic use, and establishment of exploratory reference intervals. Sera from 36 healthy dogs and 82 diseased dogs were included for method comparison with the enzyme-linked immunosorbent assay (ELISA; Tridelta) serving as the reference method. A nonparametric estimate of the 1-sided 95% reference interval was established ( n = 36). Precision study revealed good intraassay coefficients of variation (CVs) of 1–10%, 0–9%, and 2–13% for the Randox, Thermo, and Wako assays, respectively. Interassay CVs were 18%, 24%, and 19% respectively. Because of a low linear range, the Thermo test was considered unsuitable for use with canine specimens. No significant differences were present between the results obtained with the Randox and Wako assays with CRP concentrations less than 15 mg/l; however, median CRP results differed significantly between the Thermo test and the ELISA ( P = 0.03). Bland–Altman analysis detected a proportional bias of 0.28, −0.59, and 0.61 mg/l for the Randox, Thermo, and Wako assays, respectively. For all tests, median CRP values were significantly different between healthy dogs and dogs with neoplasia. The upper limit of the reference intervals were 8.2 and 9.9 mg/l for the Randox and Wako assays, respectively. In contrast to the Thermo test, the Randox and Wako assays were suitable for detection of abnormally high canine CRP concentrations; however, improvement of assay precision and evaluation of accuracy are warranted before their clinical use with canine specimens.
Detection of Perinuclear Antineutrophil Cytoplasmic Antibodies and Antinuclear Antibodies in the Diagnosis of Canine Inflammatory Bowel Diseas
Authors: Mancho Carolina , Sainz Ángel , García-Sancho Mercedes , Villaescusa Alejandra , Tesouro Miguel A. , Rodríguez-Franco Fernando
Published: May 2011

In recent years, serologic markers for diagnosis and classification of inflammatory bowel disease (IBD) have been used in human medicine. Perinuclear, antineutrophil, cytoplasmic antibodies (p-ANCA) are the most important of these markers. Because of their similar pattern of fluorescence, antinuclear antibodies (ANA) could cause misleading interpretations. The aim of the present study was to evaluate the use of an indirect fluorescent antibody test to detect p-ANCA in dogs with IBD, to compare the presence of p-ANCA in dogs with IBD with the presence of the same antibodies in other dogs, and to analyze the presence of ANAs in the p-ANCA-positive samples. Using a 1:10 dilution as a cutoff point, a sensitivity of 0.34 and a specificity of 0.86 was obtained when dogs with IBD were compared with the other groups as a whole, and specificity increased to 0.94 when dogs with IBD were compared with animals with other chronic gastrointestinal disorders. The lowest specificity value, 0.76, was obtained when the group of dogs with IBD was compared with that of dogs with different inflammatory and infectious disorders. Globally, 78 dogs were positive for p-ANCA when the cutoff was 1:10. Only 1 dog from these 78 animals was also seropositive to ANA. The results suggest that 1) detection of p-ANCA might be included in the IBD diagnostic protocol as another test to differentiate between this disease and other digestive diseases with similar clinical signs, and 2) most p-ANCA- positive dogs are not ANA positive.
Use of Real-Time Quantitative Polymerase Chain Reaction to Monitor Antibiotic Therapy in a Dog with Naturally Acquired Mycoplasma haemocanis Infection
Authors: Hulme-Moir K. Lisa , Barker Emily N. , Stonelake Anna , Helps Chris R. , Tasker Séverine
Published: May 2011

Mycoplasma haemocanis is a hemotropic bacterium that can be associated with acute hemolytic disease in immunocompromised or splenectomized dogs. The present case report describes for the first time the use of real-time quantitative polymerase chain reaction (qPCR) to monitor M. haemocanis infection in a splenectomized dog. The report also describes the application of real-time qPCR for the analysis of deoxyribonucleic acid extracted from stained blood films. The analysis of blood films from the time of initial presentation allowed a retrospective confirmation of M. haemocanis infection. The M. haemocanis copy numbers remained high throughout antibiotic treatment of this dog. A decline in copy numbers was only recorded after 11 months of therapy, when improvements in clinical and hematological indices were also noted. Clearance of infection was not achieved, and the dog remained positive for M. haemocanis at 3.5 months postcessation of antibiotic therapy. Cytological examination of blood films for the presence of organisms was insensitive for the detection of parasitemia.
Development of a Polymerase Chain Reaction Assay for Quantification of Lawsonia Intracellularis
Authors: Wattanaphansak Suphot , Gebhart Connie J. , Anderson Janet M. , Singer Randall S.
Published: May 2011

The objective of the present study was to develop a quantitative polymerase chain reaction (qPCR) assay using SYBR Green for quantification of Lawsonia intracellularis in cell culture and pig fecal samples. Specific primers were designed and tested using the aspartate ammonia-lyase ( aspA) gene as a target. Serial 10-fold dilutions of cell culture samples and several sets of spiked feces were used for qPCR optimization. The lower limit of the linear range of the assay in cell culture was 5.1 × 102 L. intracellularis/ml. A concentration of between 2.55 × 104 and 2.55 × 103 L. intracellularis/g was the lower limit of the linear range when testing community DNA from spiked fecal samples. From both cell culture and fecal samples, L. intracellularis could be detected but not accurately quantified at levels approximately 1 log below the linear range. No cross-reactivity of qPCR was found when the assay was tested using the DNA extracted from 16 species of enteric bacteria commonly found in pig feces or closely related to L. intracellularis. The new qPCR assay might prove to be a sensitive, specific, precise, and accurate method for the detection and quantification of L. intracellularis in field samples.
Molecular Identification of Bovine Papillomaviruses Associated with Cutaneous Warts in Southern Brazil
Authors: Silva Mariana Sá e , Weiss Marcelo , Brum Mário Celso Sperotto , dos Anjos Bruno Leite , Torres Fabricio Dias , Weiblen Rudi , Flores Eduardo Furtado
Published: May 2011

Bovine papillomaviruses (BPVs) are widespread pathogens mainly associated with benign, self-limiting, cutaneous lesions (warts). At least 8 viral types, defined by serology or nucleotide sequences of the L1 gene, have been identified to date. Different serotypes are associated with the specific type and morphology of the lesion and with particular geographical regions. This article describes the molecular identification of papillomaviruses from Brazilian cattle ( n = 48) and horses ( n = 1) through partial amplification and sequencing of the L1 gene. Bovine papillomavirus–1 (BPV-1) was identified in warts from 29 cattle (59%), BPV-6 from 9 cattle (18%), and BPV-2 in 8 lesions (16%). Warts of 2 cattle harbored L1 sequences of a new BPV type (BAA5), otherwise identified almost exclusively in healthy skin. The newly proposed BPV type “BR-UEL-4” was identified in a sarcoid tumor of a horse. Thus, the present report provides information on the main types of BPV involved in bovine papillomatosis in Brazil and reveals a new viral type associated with equine sarcoid, which to date has been attributed exclusively to BPV-1 and BPV-2.
Characterization of the ptfA Gene of Avian Pasteurella Multocida Strains by Allele-Specific Polymerase Chain Reaction
Authors: Sellyei Boglárka , Bányai Krisztián , Magyar Tibor
Published: May 2011

Pasteurella multocida is the causative agent of fowl cholera in domesticated and wild birds. The disease outcome is affected by various host- and pathogen-specific determinants. Several putative virulence factors have been proposed to play a key role in this interaction, including the ptfA gene, the products of which assemble to form type 4 fimbriae on the bacterial surface. One way to understand more precisely how ptfA contributes to pathogenesis is to gather molecular features of this gene in circulating avian P. multocida strains. Therefore, molecular characterization of the ptfA gene of P. multocida strains isolated from domestic poultry was performed using the combination of nucleotide sequence analysis and a newly developed allele-specific polymerase chain reaction assay. Two major ptfA alleles were identified among 31 strains, representing various serogroups and somatic serotypes. It was noteworthy that allele specificity and case severity of a subset of strains correlated with the available gross pathology data. Therefore, the acquisition of comprehensive clinical and epidemiological data together with molecular characteristics of individual strains will help to design and implement adequate preventive and intervention strategies.
Polyomavirus Infection in a Free-Ranging California Sea Lion (Zalophus Californianus) with Intestinal T-Cell Lymphoma
Authors: Colegrove Kathleen M. , Wellehan James F. X. , Rivera Rebecca , Moore Peter F. , Gulland Frances M. D. , Lowenstine Linda J. , Nordhausen Robert W. , Nollens Hendrik H.
Published: May 2011

An adult female California sea lion ( Zalophus californianus) that stranded in central California was found to have a small glossal polypoid mass on gross necropsy. Histologically, the mass was consistent with a fibropapilloma, and intranuclear inclusions were found within endothelial cells lining small arterioles within the mass. Electron microscopy revealed 40-nm virions within endothelial intranuclear inclusions. Rolling circle amplification was used to obtain a partial viral genomic sequence. Sequence analysis identified the virus as a novel polyomavirus, tentatively named California sea lion polyomavirus 1. In addition, the sea lion had a severely thickened small intestine and swollen pale kidneys on gross examination. Severe renal amyloidosis with chronic interstitial nephritis was diagnosed histologically as well as T-cell intestinal lymphoma, which was confirmed via immunophenotyping and molecular clonality. The relationship, if any, between polyomavirus infection and the other disease processes in this sea lion is not known, but it is considered unlikely that the polyomavirus induced the lymphoma.
Granular Cell Tumor in a Lumbar Spinal Nerve of a Dog
Authors: Rao Deepa , Rylander Helena , Drees Randi , Schwarz Tobias , Steinberg Howard
Published: May 2011

A 2-year-old Great Dane dog with a 2.5-week history of progressive paraparesis was presented to the Veterinary Medical Teaching Hospital at the University of Wisconsin-Madison. Neurologic examination revealed nonambulatory paraparesis with reduced to absent withdrawal hind-limb reflexes and lumbar pain. Magnetic resonance imaging and gross pathology confirmed a larger regional lumbar mass and a second smaller extradural mass within the spinal canal. The left lumbar mass was associated with extensive hemorrhage; dissection showed a dark-red, soft, well-circumscribed mass measuring 2 cm × 1.5 cm × 0.5 cm within the left fourth lumbar spinal nerve. Histopathological evaluation with immunohistochemistry revealed sheets of round to polygonal cells with diffuse granular cytoplasm demonstrating diastase-resistant periodic acid–Schiff reactivity and positive immunoexpression of S100 and neuron-specific enolase. The smaller extradural mass within the spinal canal exhibited similar morphology. Based on gross, histological, and immunohistochemical evidence, the masses were diagnosed as granular cell tumor.
Duodenal Gland Cysts and Pseudodiverticula in Sheep
Authors: Mariola Penadés , Guerrero Irene , Alberto Benito-Peña , Corpa Juan M.
Published: May 2011

Six cases of acquired duodenal diverticulitis (pseudodiverticula) were found in a flock of sheep over a short period of time. All the animals had duodenal lesions characterized by the presence of multiple saccular dilations filled with feed material. The mucosal surface was elevated by multiple small nodules that histologically corresponded to cystic dilatations of the duodenal glands, which likely caused the displacement, atrophy, and disappearance of the muscular layer, leading to pseudodiverticula. The gross appearance, microscopic findings, and epidemiological characteristics suggest that this is a different pathological process to that described for diverticula in animals to date.
Aelurostrongylus Abstrusus Larvae in the Colon of Two Cats
Authors: Ellis Angela E. , Brown Cathy A. , Yabsley Michael J.
Published: May 2011

A 12-year-old male domestic shorthair cat with a history of depression, aggression, and fever was presented for necropsy, and formalin-fixed tissues from a second male domestic shorthair cat of unknown age were submitted for histopathologic examination to the Athens Veterinary Diagnostic Laboratory. Both cats had granulomatous pneumonia associated with large numbers of parasite eggs and larvae, consistent with Aelurostrongylus abstrusus. Both cats also had larvae within glands in the colon that were identified morphologically as A. abstrusus first-stage larvae.
Soft-Tissue Mineralization of Bullfrog Larvae (Rana Catesbeiana) at a Wastewater Treatment Facility
Authors: Keel M. Kevin , Ruiz Alina M. , Fisk Aaron T. , Rumbeiha Wilson K. , Davis Andrew K. , Maerz John C.
Published: May 2011

Bullfrog tadpoles (Rana catesbeiana) from a wastewater treatment facility were identified with severe lesions consisting of large, up to 1-cm in diameter, mineralized nodules protruding from the tail or gular region. Sectioning of formalin-fixed specimens revealed more extensive mineralization involving the vertebrae or muscles of the head and tail. Nodules examined microscopically were not associated with parasitic or infectious agents. Large nodules consisted of mineralized aggregates surrounded by a margin of granulomatous inflammation. Individual connective-tissue fibers and muscle cells were mineralized at some foci. The nodules consisted entirely of calcium phosphate, and the lesions appeared to be novel. Total serumcalcium concentrations of tadpoles and calcium concentrations in water samples did not differ significantly with increasing distance from the discharge site. Affected tadpoles had elevated cholecalciferol (25-OH-vitamin D3) levels. Effluent from this wastewater treatment facility is divided into 3 streams, each passing through a separate series of wetlands allowing for replicated evaluation of tadpoles with increasing distance from the proximate inputs of treated wastewater. The prevalence of lesions was correlated with proximity of cells to the initial wastewater discharge site, and 28.5% of bullfrog larvae in the first cells had lesions. None were affected in the fifth cells. Southern leopard frog larvae (Rana sphenocephala), the only other species affected, had a much lower prevalence of lesions (<1%) than bullfrog tadpoles and were only affected in the first cells. To date, the primary cause of elevated cholecalciferol is undetermined, but it appears to be remediated by passage of water through the wetlands.
Detection of PrPSc in Formalin-Fixed, Paraffin-Embedded Tissue by Western Blot Differentiates Classical Scrapie, Nor98 Scrapie, and Bovine Spongiform Encephalopathy
Authors: Loiacono Christina M. , Beckwith Nadine , Kunkle Robert A. , Orcutt Dennis , Hall S. Mark
Published: May 2011

Transmissible, spongiform encephalopathies including bovine spongiform encephalopathy (BSE) and scrapie are fatal neurodegenerative disorders associated with the presence of an infectious abnormal isoform of normal mammalian proteins called prions. Identification of the prion protein associated with scrapie (PrPSc) in the central nervous system is typically based upon immunoassays including immunohistochemistry (IHC) using formalin-fixed tissues or Western blot (WB) assays using fresh and/or frozen, non–formalin-fixed tissues. Each assay can discriminate between BSE, classical scrapie, and a previously reported strain of scrapie recently identified in the United States named Nor98 scrapie. Different tissue samples are required from the same animal to run these 2 different immunoassays. This may result in inconsistent test results for the same animal. Sampling problems such as collecting insufficient volumes of fresh tissue or less than optimal anatomic location of brainstem for IHC can affect the ability of the test procedures to offer definitive and discriminatory results. Recently, a WB method using formalin-fixed, paraffin-embedded (FFPE) tissue to identify PrPSc was developed that successfully identified PrPSc in sheep affected by classical scrapie. In the current study, the use of this technique to produce discriminatory results identifying classical BSE in bovine tissue and both classical and Nor98 scrapie in ovine tissue using paraffin-embedded brain samples is described. Protein-banding patterns from WB using FFPE tissue were similar to protein-banding patterns produced by WB assays utilizing fresh tissues from the same animals, and results correlated well with the IHC PrPSc-positive staining present in the cerebellum and obex regions of brain samples from these animals.
Comparison of Two Methods for Blood Lead Analysis in Cattle: Graphite-Furnace Atomic Absorption Spectrometry and Leadcare® II System
Authors: Bischoff Karyn , Gaskill Cynthia , Erb Hollis N. , Ebel Joseph G. , Hillebrandt Joseph
Published: May 2011

The current study compared the LeadCare® II test kit system with graphite-furnace atomic absorption spectrometry for blood lead (Pb) analysis in 56 cattle accidentally exposed to Pb in the field. Blood Pb concentrations were determined by LeadCare II within 4 hr of collection and after 72 hr of refrigeration. Blood Pb concentrations were determined by atomic absorption spectrometry, and samples that were coagulated ( n = 12) were homogenized before analysis. There was strong rank correlation ( R 2 = 0.96) between atomic absorption and LeadCare II (within 4 hr of collection), and a conversion formula was determined for values within the observed range (3–91 mcg/dl, although few had values >40 mcg/dl). Median and mean blood pb concentrations for atomic absorption were 7.7 and 15.9 mcg/dl, respectively; for LeadCare II, medians were 5.2 mcg/dl at 4 hr and 4.9 mcg/dl at 72 hr, and means were 12.4 and 11.7, respectively. LeadCare II results at 4 hr strongly correlated with 72 hr results ( R 2 = 0.96), but results at 72 hr were lower ( P < 0.01). There was no significant difference between coagulated and uncoagulated samples run by atomic absorption. Although there have been several articles that compared LeadCare with other analytical techniques, all were for the original system, not LeadCare II. The present study indicated that LeadCare II results correlated well with atomic absorption over a wide range of blood Pb concentrations and that refrigerating samples for up to 72 hr before LeadCare II analysis was acceptable for clinical purposes.
Border Disease Virus Shedding and Detection in Naturally Infected Pyrenean Chamois (Rupicapra Pyrenaica)
Authors: Cabezón Oscar , Rosell Rosa , Velarde Roser , Mentaberre Gregorio , Casas-Díaz Encarna , Lavín Santiago , Marco Ignasi
Published: May 2011

Pyrenean chamois ( Rupicapra pyrenaica) populations of the central and eastern Pyrenees have been affected by severe outbreaks associated with Border disease virus (BDV) since 2001. Eight Pyrenean chamois (7 males and 1 female) from 1 to 8 years of age with clinical signs consistent with BDV infection were studied. At necropsy, whole blood, tissue samples (skin, brain, prescapular lymph node, thyroid gland, lung, liver, spleen, kidney, small intestine, bone marrow, and testicle), urine, and nasal, oral, and rectal swabs were obtained. The fetus from a pregnant female was also studied. Reverse transcription polymerase chain reaction (RT-PCR) was used to detect the virus in all samples, and virus isolation was performed. Sera and tissue samples were positive to RT-PCR, and the virus was isolated from all chamois. The nasal, oral, and rectal swabs and urine samples were RT-PCR positive in 100%, 85.71%, 71.43%, and 100% of chamois, respectively, confirming the excretion of the virus via these 4 routes. In addition, sera were tested for BDV antibodies using enzyme-linked immunosorbent assay and seroneutralization techniques, with negative results. Sequence analysis of the 5′ untranslated region in 7 of the chamois confirmed that the virus is grouped into the BDV-4 genotype, the same BDV previously described in Pyrenean chamois. To the authors' knowledge, this is the first study of naturally infected Pyrenean chamois, providing evidence that infected animals shed BDV through nasal, oral, fecal, and urinary excretion routes.
Further Diversity of the Staphylococcus Intermedius Group and Heterogeneity in the MboI Restriction Site Used for Staphylococcus Pseudintermedius Species Identification
Authors: Slettemeås Jannice Schau , Mikalsen Jarle , Sunde Marianne
Published: May 2011

Species identification of 200 beta-hemolysin–producing canine staphylococcal isolates was performed using a recently described polymerase chain reaction–restriction fragment length polymorphism (PCR-RFLP) method (based on MboI restriction of a pta gene fragment), supplemented with biochemical testing and sequencing of housekeeping genes. The PCR-RFLP method misclassified a small fraction (approximately 1%) of the Staphylococcus pseudintermedius population as a result of heterogeneity in the MboI restriction site. A potentially novel species within the Staphylococcus intermedius group (SIG) was found, having closest similarity to S. intermedius based on sequence comparison to the genes sodA, pta, hsp60, tuf, and full-length 16S ribosomal DNA, thus demonstrating further species diversity within the SIG.
Atherosclerosis Associated with Pericardial Effusion in a Central Bearded Dragon (Pogona Vitticeps)
Authors: Schilliger Lionel , Lemberger Karin , Chai Norin , Bourgeois Aude , Charpentier Maud
Published: May 2011

Atherosclerosis is a common disease in pet birds, particularly in psittacines, and is frequently found when performing postmortem examinations on adult and old dogs, in which it is mainly associated with endocrine diseases, such as hypothyroidism and diabetes mellitus. However, atherosclerosis is poorly documented in reptiles and consequently poorly understood. In the current case report, atherosclerosis and pericardial effusion were diagnosed in a 2-year-old male central bearded dragon ( Pogona vitticeps) based on ultrasound visualization, necropsy, and histologic examination.
Phalaris Arundinacea (Reed Canarygrass) Grass Staggers in Beef Cattle
Authors: Binder Ellen M. , Blodgett Dennis J. , Currin John F. , Caudell David , Cherney Jerry H. , LeRoith Tanya
Published: May 2011

Four adult mixed-breed beef cows from a cow–calf operation in West Virginia were referred to the Virginia-Maryland Regional College of Veterinary Medicine in March 2009 with weakness, ataxia, hind limb paresis progressing to lateral recumbency, and death within 2–3 days. Histologically, there was accumulation of light brown, granular pigment in neurons of the ventral gray horns of the spinal cord (more severe in thoracic and lumbar sections), brain stem, and pons, resulting in distortion and bulging of the cell body and displacement of the Nissl substance, suggestive of Phalaris sp. grass toxicosis. The most severely affected cow had accumulation of dark green–brown pigment in renal tubular epithelial cells. Reed canarygrass ( Phalaris arundinacea) was identified in pastures, and the concentration of tryptamine alkaloids in new leaf blades was approximately 0.2% on a wet weight basis. These alkaloids are serotonergic receptor agonists, resulting in neurologic “staggers” in ruminants. Delayed onset times of up to 4–5 months have been reported in sheep after removal from Phalaris sp. pastures. Distribution of pigment in serotonergic tracts of the midbrain, brain stem, and spinal cord with Phalaris sp. toxicoses is distinct and differs from lipofuscin. Electron microscopy confirmed that the pigment was not lipofuscin. From these findings, a diagnosis of delayed P. arundinacea toxicosis was made. Over a 2-month period, 18 cows died with similar clinical signs.
Mast Cell Tumors in a Llama (Lama Glama)
Authors: Lin Tzu-Yin , Hamberg Alexander , Pentecost Rebecca , Wellman Maxey , Stromberg Paul
Published: May 2011

A 9-year-old female llama ( Lama glama) that served as a blood donor at The Ohio State University Veterinary Teaching Hospital developed multiple small, raised, firm, non-haired cutaneous masses on the right hip, left cheek, and right and left shoulders. Cytological evaluation of fine-needle aspirates from the cutaneous mass from the left shoulder and right hip comprised many well-differentiated, highly granulated mast cells with moderate numbers of eosinophils. Occasional mast cells exhibited erythrophagocytosis and contained a small amount of hemosiderin or several variably sized vacuoles. A cytologic diagnosis of mast cell tumor with evidence of prior hemorrhage was made, and the masses were surgically removed. Microscopically, each mass consisted of sheets of neoplastic round cells that formed nonencapsulated nodules in the dermis and infiltrated into the adjacent dermal collagen. Eosinophils were scattered among the mast cells at the periphery of the nodules. Neoplastic mast cells, but not eosinophils, exhibited positive membrane KIT expression and cytoplasmic vimentin staining. A final diagnosis of mast cell tumor was made based on cytology, histology, and immunohistochemistry.
Effect of Sampling Method and Storage Conditions on Albumin, Retinol-Binding Protein, And N-Acetyl-β-D-Glucosaminidase Concentrations in Canine Urine Samples
Authors: Smets Pascale M. Y. , Meyer Evelyne , Maddens Bert , Duchateau Luc , Daminet Sylvie
Published: May 2011

Urinary markers for renal dysfunction are gaining interest, but effects of sampling method, storage conditions, and urinary tract inflammation or infection on these markers are unclear. Therefore, the objectives of the current study were to determine the difference in urinary albumin (uALB), urinary retinol-binding protein (uRBP), and urinary N-acetyl-β-D-glucosaminidase (uNAG) concentrations in cystocentesis and voided samples and to investigate concentration changes after storage at −20°C and at −80°C. Effects of a protease inhibitor were also assessed in samples stored at −80°C for 12 months. In a pilot experiment, influence of in vitro hematuria, pyuria, and bacteriuria on the urinary markers was evaluated. A mixed model was used to calculate mean differences and 95% confidence intervals. Urinary ALB, uNAG, and uRBP concentrations were similar in voided and cystocentesis samples. After storage for 4 months at −20°C, uALB concentration was not affected, and uRBP concentration showed a mild and clinically irrelevant decrease, whereas uNAG activity was significantly lower compared with fresh samples. After storage for 12 months at −80°C, uALB and uRBP concentrations did not differ from fresh samples, but uNAG activity was severely decreased. Protease inhibitor addition did not preserve uNAG activity. Experimental hematuria, pyuria, and bacteriuria did not seem to affect urinary markers, although further research is needed.
Bi-Antigenic Immunoassay Models Based on the Recombinant PvpA Proteins for Mycoplasma Gallisepticum Diagnosis in Chickens
Authors: Büyüktanır Özlem , Genç Oktay , Yurdusev Nevzat
Published: May 2011

The present study aimed to produce the relatively conserved central fragment of the Mycoplasma gallisepticum PvpA cytadhesin as recombinant antigen and to determine its species-specific diagnostic potential in comparison with the full-length recombinant rPvpA336 protein. For this purpose, a recombinant protein (rPvpA134) consisting of 134 amino acids with apparent molecular mass of 27 kD was produced and highly purified. The rPvpA134 protein was composed of the amino acid residues at positions 133–265 with respect to the wild-type PvpA. Two bi-antigenic diagnostic models based on Western blot and enzymatic rapid immunofiltration assay (ERIFA) were developed to compare simultaneously the diagnostic potential of the recombinant antigens rPvpA134 and rPvpA336. Although 40% of the confirmed rPvpA336-positive chicken sera were detected as reactive with rPvpA134, this protein would be a useful secondary diagnostic antigen with which to confirm species-specific antibody response for monitoring M. gallisepticum infections. It can be concluded from the present study that 2 bi-antigenic models were successfully adapted to the specific diagnosis of chicken M. gallisepticum. Furthermore, by virtue of its simplicity and rapidity, the ERIFA model has multi-antigenic application potential, making it an alternative field test that is widely applicable in the veterinary diagnostic field.
Histologic and Bacteriologic Findings in Valvular Endocarditis of Slaughter-Age Pigs
Authors: Jensen Henrik E. , Gyllensten Johanna , Hofman Carolina , Leifsson Páll S. , Agerholm Jørgen S. , Boye Mette , Aalbæk Bent
Published: May 2011

Endocarditis lesions from 117 slaughter pigs were examined pathologically and etiologically in addition to 90 control hearts with cardiac valves. Lesions were located on the valves; however, the lesions had extended to the walls in 21 cases (18%). Lesions predominated on the mitral valve (59%). A total of 28 cases, from which no growth was obtained or a contamination flora was grown, were screened by fluorescence in situ hybridization (FISH) for bacteria (general bacterial probe) and probes specific for Streptococcus suis and Erysipelothrix rhusiopathiae, respectively. Using FISH, an additional 10 cases of endocarditis due to S. suis and E. rhusiopathiae were disclosed. Within lesions, streptococci predominated (53%) followed by E. rhusiopathiae (30%). Distinct features of both the lesions and the shape and localization of bacterial colonies were related to streptococci and E. rhusiopathiae. The propensity for streptococci to be localized on more than 1 valve in single hearts may be because S. suis–infected pigs tend to have been infected for a longer period compared with E. rhusiopathiae. Mineralization of endocarditis lesions was significantly associated with infection by streptococci, and was seen in 71% of the cases, whereas it was present in only 28% of lesions caused by E. rhusiopathiae. In addition, areas with mineralization were significantly correlated to the presence of a granulomatous reaction. Granulomatous endocarditis is likely a result of a foreign body reaction due to dystrophic mineralization. Local proliferation of valvular endothelial cells, found in 9 hearts in the current study, may increase the risk of developing thrombosing endocarditis in pigs.
Diagnostic Application of H3N8-Specific Equine Influenza Real-Time Reverse Transcription Polymerase Chain Reaction Assays for the Detection of Canine Influenza Virus in Clinical Specimens
Authors: Lu Zhengchun , Dubovi Edward J. , Zylich Nancy C. , Crawford P. Cynda , Sells Stephen , Go Yun Young , Loynachan Alan T. , Timoney Peter J. , Chambers Thomas M. , Balasuriya Udeni B. R.
Published: May 2011

The objective of the current study was to determine the capability of 3 recently described one-step TaqMan real-time reverse transcription polymerase chain reaction (real-time RT-PCR) assays targeting the nucleoprotein (NP), matrix (M), and hemagglutinin (HA) genes of H3N8 Equine influenza virus (EIV NP, EIV M, and EIV HA3 assays, respectively) to detect Canine influenza virus (CIV). The assays were initially evaluated with nucleic acid extracted from tissue culture fluid (TCF) containing the A/canine/FL/43/04 strain of Influenza A virus associated with the 2004 canine influenza outbreak in Florida. The EIV NP, EIV M, and EIV HA3 assays could detect CIV nucleic acid at threshold cycle (Ct) values of 16.31, 23.71, and 15.28, respectively. Three assays using TCF or allantoic fluid (AF) samples containing CIV ( n = 13) and archived canine nasal swab samples ( n = 20) originally submitted for laboratory diagnosis of CIV were further evaluated. All TCF and AF samples, together with 10 nasal swab samples that previously tested positive for virus by attempted isolation in embryonated hens' eggs or Madin–Darby canine kidney cells, were positive in all 3 real-time RT-PCR assays. None of the 3 assays detected the H1N1 Swine influenza virus strain in current circulation. These findings demonstrate that previously described real-time RT-PCR assays targeting NP, M, and H3 HA gene segments of H3N8 EIV are also valuable for the diagnosis of CIV infection in dogs. The assays could expedite the detection and identification of CIV.
Consistent Detection of Felis Domesticus Papillomavirus 2 DNA Sequences Within Feline Viral Plaques
Authors: Munday John S. , Peters-Kennedy Jeanine
Published: May 2011

Viral plaques are well recognized skin lesions of cats. They are thought to be caused by papillomavirus infection; however, the causative papillomavirus is uncertain. In the current study, polymerase chain reaction using 2 consensus primer sets and 1 primer set specific for Felis domesticus papillomavirus 2 (FdPV-2) was used to amplify DNA from a series of 14 feline viral plaques. The FdPV-2 sequences were detected in all 14 viral plaques by the specific primers but in only 1 of 14 feline cutaneous trichoblastomas. Papillomavirus DNA was amplified from 8 plaques using the consensus primers. Sequences from FdPV-2 were amplified using the consensus primers from 4 plaques. In addition, 3 plaques contained papillomavirus DNA sequences from Felis domesticus papillomavirus sequence MY1, and a previously unreported papillomavirus DNA sequence was amplified from 1 plaque. As FdPV-2 was consistently present within the plaques, this suggests that this papillomavirus is the likely etiologic agent. Feline viral plaques can undergo neoplastic transformation to Bowenoid in situ carcinomas (BISCs). As FdPV-2 DNA is frequently present within BISCs, this suggests that FdPV-2 induces viral plaque formation and then remains detectible after neoplastic transformation.
Retrospective Study of Necropsy-Associated Coagulase-Positive Staphylococci in Horses
Authors: Haenni Marisa , Targant Hayette , Forest Karine , Sévin Corinne , Tapprest Jackie , Laugier Claire , Madec Jean-Yves
Published: May 2011

Coagulase-positive staphylococci (CoPS) are potential causative agents of equine infections, but they are rarely responsible for the death of the animal. In the current study, staphylococci implicated in the death or euthanasia of horses were retrospectively studied in 3,457 necropsies performed over a decade (1995-2006). Morbidity associated with CoPS was 1.7%, representing 60 isolates of CoPS, which were identified as Staphylococcus aureus (59) and Staphylococcus pseudintermedius (1). Coagulase-positive staphylococci (alone or in association with another bacterial species) were associated with the death or euthanasia of 90% of the cases (54/60). Proportions of antibiotic resistance to penicillin G and tetracycline reached 62.7% and 23.7%, respectively. Virulence genes were detected in 91.7% of the strains, with a majority of seh or sei enterotoxin genes. Finally, 3 methicillin-resistant Staphylococcus aureus (MRSA) isolates belonging to the t064 spa-type were identified. One strain was isolated in 2003 and might thus be one of the first cases of equine MRSA in France.