bluuebunny/crossref_metadata_2025 huggingface dataset:
Identification and Differentiation of European and African/Australian Strains of Mycoplasma Mycoides Subspecies Mycoides Small-Colony type Using Polymerase Chain Reaction Analysis
Authors: Miles Katie , Churchward Colin P. , McAuliffe Laura , Ayling Roger D. , Nicholas Robin A. J.
Published: May 2011

Mycoplasma mycoides subspecies mycoides small-colony type ( M. m. m. SC) is the cause of the economically important contagious bovine pleuropneumonia. Isolates from Africa and Australia have previously been documented to have a fragment of approximately 8.84 kb, which is absent in European strains. A set of polymerase chain reaction (PCR) primers over this region was designed to identify M. m. m. SC isolates and separate European strains from those of Africa/Australia. Specificity of the PCR assay was achieved through the positioning of an oligonucleotide within the insertion sequence IS1296, upstream of this deletion, which then was paired with a reverse primer, upstream of the deletion, within the 8.84 kb-deleted region or downstream of the deletion, generating fragments of 1.1 kb (all M. m. m. SC strains), 1.4 kb (African/Australian strains only) and 1.3 kb (European strains only), respectively. Identification and differentiation was specific for DNA from M. m. m. SC with no amplification of DNA from other cluster members or closely related species. The PCR products did not require differentiation by use of a restriction endonuclease, and have potential for use in detection of this organism in clinical samples.
Development of a Polymerase Chain Reaction Assay for the Detection of Antibiotic Resistance Genes in Community DNA
Authors: Patterson Sheila K. , Singer Randall S.
Published: May 2011

Many methods are used to detect antibiotic resistance genes in samples. The objective of the study reported here was to compare polymerase chain reaction (PCR) analysis of community DNA with fecal culturing for detecting antibiotic resistance genes in cattle samples. In the laboratory-based portion of this study, known concentrations of an Escherichia coli strain with 3 antibiotic resistance genes ( cmy-2, flo, and cat) were added to feces from dairy cattle. These genes were used to assess the effect of various primer pairs, chromosomally versus plasmid-encoded genes, and gene copy number on the sensitivity of PCR amplification. Gene-specific PCR amplification was performed on the community DNA extracted from the feces. Feces were cultured for the inoculated strain. In the field-based portion of the study, 80 cattle fecal samples of unknown gene status were compared by use of similar methods. Culture and PCR amplification from community DNA extractions produced variable results, and this variability was most noticeable at dilutions that approached the detection limit of the assay. Typically, PCR amplification had a higher sensitivity than did culture for detecting the gene of interest. However, the sensitivity of culture was improved by plating on selective media containing antibiotics. The community DNA approach enables assessment of bacterial communities in complex samples such as feces, a task that can be prohibitive by budget or time constraints associated with culture methods. Through a strategic combination of culture and community DNA approaches, the relationship between specific selection pressures and the persistence and dissemination of specific resistance genes can be elucidated.
Evaluation of the Gamma Interferon ELISA in Sheep Subclinically Infected with Mycobacterium Avium Subspecies Paratuberculosis Using a Whole-Cell Sonicate or a Johnin Purified-Protein Derivative
Authors: Robbe-Austerman Suelee , Stabel Judith R. , Palmer Mitchell V.
Published: May 2011

The aim of the study reported here was to estimate the sensitivity and specificity of the gamma interferon (IFN-γ) ELISA for paratuberculosis in sheep using receiver-operating characteristic analysis. Bacteriologic culture of tissues was used to define the reference positive population ( n = 33). Two reference negative populations were used: culture-negative sheep from infected flocks ( n = 77), and sheep from noninfected flocks ( n = 358). We also evaluated the accuracy of 2 Mycobacterium avium subspecies paratuberculosis (MAP) antigen preparations, a whole-cell sonicate (MpS) and a johnin purified-protein derivative (PPDj). The source of the reference negative sheep used in the analysis affected overall accuracy of the IFN-γ ELISA. The area under the curve was 0.683 (95% confidence interval 0.574–0.787), using culture-negative sheep from infected flocks, was 0.831 (0.764–0.889), using sheep from noninfected flocks for the MpS, and was 0.809 (0.726–0.881) and 0.897 (0.862–0.925) for the PPDj, respectively. Using the MpS, the cut point that classified the most sheep correctly was an optical density reading of 0.20, for sensitivity of 40.7% (19.4–57.6) and specificity of 88.7% (77.0–95.7) or 97.6% (93.04–99.5), depending on the reference negative population used. Using the PPDj, the cut point that classified the most sheep correctly was 0.25 for sensitivity of 66.7% (47.2–82.7) and specificity of 93.5% (85.5–97.9) or 98.3% (96.4–99.4), respectively. The PPDj was more accurate at identifying MAP-infected sheep than was the MpS ( P = 0.034).
Interstitial Pneumonia in Neonatal Canine Pups with Evidence of Canine Distemper Virus Infection
Authors: Pandher Karamjeet , Podell Brendan , Gould Daniel H. , Johnson Bill J. , Thompson Sheri
Published: May 2011

Four dead canine pups (5–12 days old) from 3 litters in Douglas County of north central Colorado were submitted to the Colorado State University Diagnostic Laboratory for necropsy. Pups were originally presented to the referring clinics for respiratory tract illness, with or without diarrhea. At necropsy, the lungs from all pups had similar lesions, including random foci of hemorrhage and failure to collapse on opening of the thoracic cavity. The lungs were histologically characterized by subacute interstitial pneumonia, with alveolar septa expanded by a histiocyte-rich infiltrate with a few lymphocytes and neutrophils. The alveolar spaces were filled with moderate amounts of proteinaceous fluid, foamy macrophages, and a few neutrophils. Lungs from 3 of the 4 pups were test positive for canine distemper virus (CDV) by use of reverse transcriptase-polymerase chain reaction (RT-PCR) analysis. Immunohistochemically stained lungs, including those from the pup that were CDV negative, by use of RT-PCR analysis, were test positive for CDV antigen in bronchial and bronchiolar epithelial cells and in a few alveolar macrophages. Central nervous system lesions were not observed in any of the 4 pups. These cases represent an unusual presentation of canine distemper in neonatal pups marked by respiratory tract lesions without central nervous system involvement. Canine distemper should be considered in the differential diagnosis of neonatal canine respiratory tract illness.
Detection and Characterization of Naturally Acquired West Nile Virus Infection in a Female Wild Turkey
Authors: Zhang Z. , Wilson F. , Read R. , Pace L. , Zhang S.
Published: May 2011

An adult female wild turkey exhibiting disorientation and failure to flee when approached was submitted to the Mississippi Veterinary Research and Diagnostic Laboratory. Gross pathologic examination revealed evidence of dehydration and the presence of modest numbers of adult nematodes in the small intestine. Histologic examination revealed extensive multifocal perivascular lymphocytic infiltration in brain, marked heterophilic hyperplasia in bone marrow, and multifocal interstitial lymphocytic infiltration in heart, pancreas, ventriculus, and skeletal muscles. West Nile virus (WNV) was isolated from the brain, lung, and kidney tissues using cultured Vero cells. Higher copies of viral RNA were detected from brain, lung, and kidney than from heart, liver, or spleen by quantitative real-time reverse transcription-polymerase chain reaction (RRT-PCR) analysis. Immunohistochemical (IHC) analysis detected WNV antigen in various tissues including neurons, kidney, respiratory tract epithelium, heart, and bone marrow. On the basis of the data from this investigation, it is concluded that WNV caused encephalitis along with many other pathologic changes in the affected wild turkey.
Novel Iridovirus in a Nautilus (Nautilus Spp.)
Authors: Gregory Christopher R. , Latimer Kenneth S. , Pennick Kate E. , Benson Keith , Moore Tiffany
Published: May 2011

Intracytoplasmic inclusion bodies suggestive of iridovirus infection were observed in formalin-fixed, paraffin-embedded tissues from a nautilus ( Nautilus spp.) that died without premonitory signs. Transmission electron microscopy revealed enveloped, hexagonal, viral particles that measured approximately 176 nm in diameter. Virions contained a dense central core and morphology typical of iridoviruses. Extracted DNA was amplified using primers homologous to conserved iridovirus sequences. The amplicons were cloned, sequenced, and determined to be approximately 60% similar to reported amphibian iridovirus sequences. A polymerase chain reaction-generated digoxigenin probe was used to detect viral nucleic acid in tissue sections by DNA in situ hybridization and high-affinity cytochemistry. The detected nucleic acid corresponded to the inclusion bodies observed microscopically. This represents a novel iridovirus of mollusks.
Streptococcus Pneumoniae-Associated Cellulitis in a Two-Month-Old Domestic Shorthair Kitten
Authors: Zhang Shuping , Wilson Floyd , Pace Lanny
Published: May 2011

An approximately 2-month-old, reproductively intact female Domestic Shorthair kitten was presented to the Mississippi Veterinary Research and Diagnostic Laboratory with a history of possible trauma to the left shoulder region while playing with children, and was found dead the following day. Marked swelling, with subcutaneous edema and hemorrhages, was observed in the left forelimb. Severe pleocellular, but largely suppurative cellulitis, fasciitis, and interstitial myositis with edema were observed microscopically in sections from the affected limb. Massive numbers of gram-positive diplococci also were observed. Other pathologic changes included moderate interstitial pneumonia, mild cholangitis, lymph node hemorrhage, gastrointestinal nematodiasis, mild enteritis, and mild interstitial nephritis. Bacteriologic culture identified Streptococcus pneumoniae as the causative agent, which was confirmed by polymerase chain reaction amplification of the pneumolysin gene from chromosomal DNA of the isolate.
Use of Age and Milk Production Data to Improve the Ability of Enzyme-Linked Immunosorbent Assay Test Results to Predict Mycobacterium Avium ssp. Paratuberculosis Fecal Culture Status
Authors: Berghaus Roy D. , Farver Thomas B. , Anderson Randall J. , Adaska John M. , Gardner Ian A.
Published: May 2011

Cows from 2 California dairies were tested for paratuberculosis at the end of lactation by using fecal culture and a commercially available serum enzyme-linked immunosorbent assay (ELISA) test kit. Individual cow characteristics and production variables were evaluated along with ELISA testing results as predictors of fecal culture status. In multivariable logistic regression analysis, age and a herd-standardized version of 305-day mature equivalent (305 ME) milk production were significant predictors of fecal culture status after adjusting for herd, quarter of the study year, and ELISA sample-to-positive (S/P) ratio. The area under a nonparametric receiver operating characteristic curve was significantly greater for a multivariable model that included age and the level of milk production when compared with a model without these covariates. In conclusion, consideration of cow-level covariates was useful as an aid in predicting Mycobacterium avium ssp. paratuberculosis (MAP) fecal culture status. For a given ELISA S/P ratio, older cows and those with lower 305 ME milk production relative to other cows in the herd were significantly more likely to be shedding MAP in their feces at the end of lactation.
Evaluation of Nephelometry for Albumin Measurement in Serum and Cerebrospinal Fluid: Experiences with an Indwelling Subarachnoidal Catheter System for Repetitive Cerebrospinal Fluid Collection in Horses
Authors: Goehring L. S. , Kessels B. G. F. , van Maanen C. , Voorbij H. A. M. , van Oldruitenborgh-Oosterbaan M. M. Sloet
Published: May 2011

The measurement of albumin concentrations in cerebrospinal fluid (CSF) and serum for albumin quotient (AQ) calculations in normal horses was performed by 2 methods: 1) total protein measurement, followed by electrophoresis of the samples to obtain an albumin percentage; and 2) albumin immunoprecipitation quantitated by nephelometry. The results of both methods correlated well, and nephelometry was chosen to determine the albumin concentrations in CSF samples obtained from an indwelling subarachnoidal catheter for daily sampling. Because the use of an indwelling catheter to collect repetitive CSF samples is a novel technique, routine cytological CSF analysis was performed along with daily clinical evaluation to ascertain the well-being of the horses. The catheters were placed in 2 horses for periods of 14 and 17 days. One horse exhibited pleocytosis on cytological evaluation of CSF on 2 occasions for a 1–2-day duration; however, the AQ showed a significant increase on only 1 occasion. The other horse had a normal cell count in CSF but showed 2 sudden changes in the AQ value; however, these values remained within the 95% confidence interval for AQ in horses. Albumin quotient values of the second horse were consistently below the lower range of the confidence interval. Results from this study indicate that nephelometry can be used for albumin determination in serum and CSF samples from horses. Furthermore, an indwelling subarachnoidal catheter system can provide serial CSF samples in horses, thus obviating the need for repetitive centesis for serial CSF sampling.
Inability of Kaolin Treatment to Remove Nonspecific Inhibitors from Equine Serum for the Hemagglutination Inhibition Test Against Equine H7N7 Influenza Virus
Authors: Boliar Saikat , Stanislawek Wlodek , Chambers Thomas M.
Published: May 2011

The hemagglutination inhibition test is used by many diagnostic and surveillance laboratories for detection of antibodies to influenza viruses. It is well known that the hemagglutination inhibition test is affected by nonspecific inhibitors present in equine serum. Several serum treatments are in use to remove these inhibitors, including treatment with kaolin. Discrepant results were observed in the authors' laboratories when using kaolin treatment before testing equine sera for antibodies against equine influenza virus (EIV) subtype-1 (H7N7). It is demonstrated here that kaolin treatment leads to false positive results when testing for antibodies against EIV subtype-1, as compared to other standard serum treatments (trypsin-periodate, receptor-destroying enzyme). Against EIV subtype-2 (H3N8), however, false positive results were not evident. Trypsin-periodate and receptor-destroying enzyme (RDE) treatments appear to be superior to kaolin for removal of nonspecific inhibitors from equine serum and should be used for serological diagnosis and surveillance of equine influenza virus.
Application of a Dot Enzyme-Linked Immunosorbent Assay for Evaluation of the Immune Status to Canine Parvovirus and Distemper Virus in Adult Dogs before Revaccination
Authors: Waner Trevor , Mazar Shlomit , Keren-Kornblatt Ephraim
Published: May 2011

A growing body of literature has been published indicating that the current practice of annual vaccination of dogs may not be beneficial and in some cases may even be harmful. A number of publications have proposed assessing the immune status of dogs before annual revaccination. In this study the usefulness of a commercially available dot-ELISA kit was evaluated to determine the duration IgG antibody titers to canine parvovirus (CPV) and canine distemper virus (CDV) in 158 dogs vaccinated at least one year ago. Overall, the percentage of dogs with protective antibody titers to both CPV and CDV was 84%. The percentage of dogs with borderline antibody titers was 11% for CPV and 10% for CDV. Four percent of the dogs had no detectable antibody to CPV and 6% had no antibody to CDV. The results reported here are in good agreement with other studies measuring IgG antibody levels. It is concluded that the kit offers veterinarians the opportunity of determining antibody titers and revaccinating only those pets whose antibody titers to specific diseases have waned.
Detection of Early Stages of Myxobolus Cerebralis in Fin Clips from Rainbow Trout (Onchorynchus Mykiss)
Authors: Skirpstunas Ramona T. , Hergert John M. , Baldwin Thomas J.
Published: May 2011

A nested polymerase chain reaction (PCR) assay was used to detect early stages of Myxobolus cerebralis in caudal and adipose fin samples from rainbow trout (RT). To determine sensitivity, groups of 10 RT were exposed to 2,000 M. cerebralis triactinomyxons/fish for 1 hour at 15°C and subsequently moved to clean recirculating water. Fish were held for 2 and 6 hours and 1, 2, 3, 5, 7, 10, 30, and 60 days before sampling by nonlethal fin biopsy. Nested PCR performed on fin clips showed that M. cerebralis DNA was detected in caudal fin tissue in 100% of fish up to 5 days postexposure. At days 7 and 10 postexposure, 80% of fish were positive, and at 60 days postexposure, 60% of fish were positive using this technique. Conversely, testing on adipose fin clips proved less sensitive, as positive fish dropped from 80% at day 7 to below 20% at day 10 postinfection. Since detection of M. cerebralis infection using caudal fin samples coupled with nested PCR is an effective method for detection of early parasite stages, use of this technique provides for accurate, nonlethal testing.
Falcon Adenovirus Infection in Breeding Taita Falcons (Falco Fasciinucha)
Authors: Dean Jason , Latimer Kenneth S. , Oaks J. Lindsay , Schrenzel Mark , Redig Patrick T. , Wünschmann Arno
Published: May 2011

Four female and 3 male Taita falcons ( Falco fasciinucha) out of a breeding colony of 14 Taita falcons (7 pairs) died during the breeding season after showing lethargy and anorexia for 1 to 2 days. All animals were submitted for necropsy. Gross lesions in the female falcons were characterized by anemia secondary to marked hemorrhage into the ovary and oviduct, serofibrinous effusion into the cardioabdominal cavity and serosal petechiae. In addition, marked necrotizing splenitis and pulmonary hemorrhage were present. Histologically, the female falcons had mild necrotizing hepatitis with numerous intranuclear inclusion bodies and necrotizing splenitis with rare inclusion bodies. There were no gross lesions in the male falcons, and the histological lesions were characterized by urate deposition and rare intranuclear inclusion bodies in the renal tubular epithelial cells. Adenoviral particles were found by electron microscopy in the cloacal contents of the female Taita falcons but not in the male falcons. DNA in situ hybridization revealed widespread aviadenoviral nucleic acid within the nuclei of hepatocytes, renal tubular epithelial cells, and adrenal cells in the female falcons but no aviadenoviral nucleic acid in 1 male falcon and only a low quantity of adenoviral nucleic acid in the liver and kidney of another male Taita falcon. PCR amplified aviadenoviral DNA in the liver and intestine of all Taita falcons. The amplicons were sequenced, and the virus was identified as falcon adenovirus. The deaths of the female and male birds were attributed to the aviadenovirus infection.
Use of Sentinel Chickens to Evaluate the Effectiveness of Cleaning and Disinfection Procedures in Noncommercial Poultry Operations Infected with Exotic Newcastle Disease Virus
Authors: McCluskey Brian J. , Burgess Brandy , Glover James , Kinde Hailu , Hietala Sharon
Published: May 2011

The use of sentinel chickens in establishing the negative status of commercial poultry flocks depopulated due to exotic Newcastle disease (END) is considered to be an economically beneficial process. However, the costs and benefits of using sentinel chickens in noncommercial operations are in question. The objective of this study was to use sentinel chickens to evaluate whether adequate cleaning and disinfection coupled with an appropriate time period without susceptible poultry species on the premises would eliminate END virus from a noncommercial poultry operation and preclude the need for placement of sentinels in previously infected operations before declaring them free of virus. Noncommercial poultry operations were selected from the 2002 to 2003 END outbreak database. Operations included in the study had one or more isolations of END virus (ENDV) from cloacal or oropharyngeal swabs of birds on the premises. A total of 546 birds were placed on 53 premises. All sentinel birds sampled after placements were negative by virus detection methods and serologic tests. Results of this study indicate that time and the application of appropriate cleaning and disinfection procedures will adequately mitigate the risk of viable virus persisting in noncommercial poultry operations. In the future, this information may eliminate the need for sentinel bird placement to ensure virus free status of premises before repopulation, thereby decreasing the costs of END eradication.
Cerebellar Degeneration in Cattle Grazing Solanum Bonariense (“Naranjillo”) in Western Uruguay
Authors: Verdes José Manuel , Moraña Antonio , Gutiérrez Fernando , Battes Daniel , Fidalgo Luis Eusebio , Guerrero Florentina
Published: May 2011

Cattle in western Uruguay that were eating Solanum bonariense developed periodic episodes of ataxia, hypermetria, hyperesthesia, head and thoracic limb extension, opisthotonus, nystagmus, and falling to the side or backward. Similar clinical signs were experimentally reproduced in cattle by administration of S. bonariense via rumen cannula at a dose of 1,024 g/kg body mass. No significant gross lesions were observed in field cases or experimentally induced cases. Spontaneous and induced histologic lesions were similar and included vacuolation, degeneration, and loss of Purkinje cells. Axonal spheroids, microcavitations, and other changes of wallerian-type degeneration in cerebellar white matter were also observed. Ultrastructural changes included increased number of electron-dense residual storage bodies in membrane-bound vesicles in affected Purkinje cells, and similar vesicles and mitochondria in axonal spheroids. No histologic lesions were detected in the other examined tissues. The Purkinje-cell swelling and vacuolation with subsequent cerebellar degeneration are suggestive of Purkinje-cell specific toxin that produces abnormal lysosome function and cell specific axonal transport. This is the first report of S. bonariense toxicity.
Onchocercosis of an Intervertebral Joint Capsule Causing Cervical Vertebral Stenotic Myelopathy in a Horse
Authors: Hestvik Gete , Ekman Stina , Lindberg Ronny
Published: May 2011

A novel case where onchocercosis was identified as a cause of cervical myelopathy in the horse is described. A 15-year-old Connemara mare was euthanized due to progressive locomotion disturbance. Postmortem examination revealed soft-tissue swelling in the intervertebral joint capsule of C6-7 with narrowing of the vertebral canal. On light microscopy, axonopathy was pronounced in the corresponding segment of the spinal cord. Fibrous tissue and eosinophilic granulomas were found in the joint capsule, together with parasites identified histologically as Onchocerca sp.
Maxillary Osteosarcoma in a Prairie Dog (Cynomys Ludovicianus)
Authors: Mouser Pamela , Cole Alex , Lin Tsang Long
Published: May 2011

To date, few tumors have been identified in prairie dogs, with odontoma being the most common. Osteosarcoma has been documented in a wide range of species, including a number of rodents. In this case, a locally invasive maxillary osteosarcoma was diagnosed in a prairie dog. Gross examination revealed a pale, tan, lobulated, sessile maxillary mass extending ventrally into the oral cavity from the hard palate and the gingiva surrounding the upper right cheek teeth. The mass invaded the right nasal cavity and retrobulbar space causing exophthalmia. Microscopically, the mass consisted of densely packed spindle-shaped cells with occasional multinucleated giant cells. Brightly eosinophilic osteoid was multifocally scattered in the tumor mass. To the authors' knowledge, this is the first documented report of maxillary osteosarcoma in a prairie dog.
Disseminated Mycobacteriosis Due to Mycobacterium Avium in Captive Bengal Tiger (Panthera Tigris)
Authors: Cho Ho-Seong , Kim Yong-Hwan , Park Nam-Yong
Published: May 2011

A 2-year-old captive female Bengal Tiger ( Panthera tigris) died after prolonged anorexia in the Gwangju Uchi Park Zoo, Gwangju, Republic of Korea. Necropsy revealed multiple nodules of varying sizes in the lung, liver, kidney, and spleen. Histopathologic examination revealed a typical granuloma composed of caseous necrotic areas surrounded by lymphocytes with a few giant cells and foamy macrophages. Periodic acid-Schiff stain and Gomori methenamine silver stain did not reveal any fungal bodies. The Ziehl-Neelsen acid-fast stain revealed few acid-fast organisms in the lung, liver, kidney, and spleen. A polymerase chain reaction assay of the lung, liver, kidney, and spleen yielded a positive result for Mycobacterium avium subsp. avium. This is an unusual case of disseminated infection of a wild mammal with avian mycobacteriosis, and is believed to be most likely associated with the feeding of tigers with culled chickens infected with M. avium.
A Specific Method for Measurement of Equine Active Myeloperoxidase in Biological Samples and in in Vitro Tests
Authors: Franck Thierry , Kohnen S. , Deby-Dupont G. , Grulke S. , Deby C. , Serteyn D.
Published: May 2011

An original method called SIEFED (specific immunological extraction followed by enzymatic detection) was developed for the specific detection of the activity of equine myeloperoxidase (MPO). The method consists of the extraction of MPO from aqueous solutions by immobilized anti-MPO antibodies followed by washing (to eliminate proteins and interfering molecules) and measurement of MPO activity using a detection system containing a fluorogenic substrate, hydrogen peroxide, and nitrite as reaction enhancer. The SIEFED technique was applied to study active MPO in horse biological fluids and the effects of 2 polyphenolic molecules, curcumin and resveratrol, on MPO activity. The detection limit of the SIEFED was 0.23 mU/ml. The SIEFED exhibited good precision with intra-assay and interassay coefficients of variation below 10% and 20%, respectively, for MPO activities ranging from 0.25 to 6.4 mU/ml. The activity of MPO was generally higher than 1 mU/ml in the fluids collected from horses with inflammatory diseases. Curcumin and resveratrol exerted a dose-dependent inhibition on MPO activity and, as they were removed before the enzymatic detection of MPO, the results suggest a direct drug-enzyme interaction or an enzyme structure modification by the drug. The SIEFED is a new tool that would be useful for specific detection of active MPO in complex media and for selection of MPO activity modulators.
Control Panels of Meat Juice Samples for a Salmonella Enzyme-Linked Immunosorbent Assay
Authors: Bak Hanne , Sørensen Vibeke
Published: May 2011

In the Danish pig production system, an indirect enzyme-linked immunosorbent assay (ELISA) for detection of antibodies in meat juice is used for Salmonella surveillance. Quality control (QC) of this ELISA was previously based on repeated testing of control serum samples. The purpose of the study reported here was to collect, characterize, and implement a panel of meat juice pools for supplemental internal QC. Muscle samples for extraction of meat juice were collected from slaughter pigs of 5 herds infected with Salmonella spp. and from 4 herds without Salmonella infection. A QC panel with 39 pools of meat juice, yielding ELISA optical density (OD) values covering the full range of expected OD values, was prepared and tested repeatedly to determine mean and SD OD values. Each pool was tested twice on each microtitration plate, and the results were used to determine limits for validity of future tests. This QC panel was included as an internal QC to be tested every month. Besides the QC panel, 2 panels containing 100 samples of meat juice with OD above the positive cut-off value and 100 samples with OD below that value were prepared for quarterly control of the diagnostic sensitivity (DSe) and the diagnostic specificity (DSp) of the ELISA. The inclusion of these panels in the QC system will provide information about drifts in DSe and DSp of the test. The procedures described here can be applied to other tests where meat juice samples are used for testing.
Evaluation of Telomerase in Canine Mammary Tissues by Immunohistochemical Analysis and a Polymerase Chain Reaction-Based Enzyme-Linked Immunosorbent Assay
Authors: Panarese Serena , Brunetti Barbara , Sarli Giuseppe
Published: May 2011

The enzyme telomerase is considered a potential marker for neoplastic tissue and is used as a diagnostic and prognostic tool in clinical medicine and therapeutics. For this reason, the possible role of telomerase activation in the process of malignant transformation is currently the subject of intense research efforts. The focus of the study reported here was to detect telomerase in 37 canine mammary samples, by comparing two methods: immunohistochemical (IHC) analysis for detecting the catalytic subunit of the enzyme, telomerase reverse transcriptase (TERT), and the telomeric repeat amplification protocol–enzyme-linked immunosorbent assay (TRAP-ELISA), a polymerase chain reaction (PCR)-based technique that uses a colorimetric detection method. Using the TRAP-ELISA, samples were considered positive when they yielded a difference of at least 0.2 absorbance units between the readings at 450 nm versus 690 nm wavelength. On the basis of this criterion, 18 negative and 19 positive cases were obtained. Specific immunohistochemical staining was observed mainly in the nucleoli, to a lesser extent in the nuclei, and rarely in the cytoplasm of epithelial cells. A sample was considered positive when at least 10% of the epithelial cells had specific staining. The Pearson correlation between the TRAP-ELISA and IHC results was significant only when IHC nucleolar ( r = 0.53, P < 0.01) or nuclear ( r = 0.36, P < 0.05) staining or their combination ( r = 0.58, P < 0.01) was considered. Thus, IHC staining of nucleoli and nuclei can be considered as an alternative method to the TRAP-ELISA. The detection of telomerase in normal mammary gland and fibrocystic mastopathy using both methods does not support the idea that telomerase may be used as a specific marker of mammary neoplasia in dogs.
Nested Polymerase Chain Reaction Detection and Duration of Porcine Circovirus Type 2 in Semen with Sperm Morphological Analysis from Naturally Infected Boars
Authors: McIntosh Kathleen A. , Harding John C. S. , Parker Sarah , Ellis John A. , Appleyard Greg D.
Published: May 2011

A nested polymerase chain reaction (nPCR) protocol was applied to porcine semen to demonstrate the porcine circovirus type 2 (PCV2) shedding patterns and duration in naturally infected boars. Sperm morphology analysis was performed on a subset of samples to determine if the presence of PCV2 DNA in semen was associated with reduced semen quality. Semen was collected serially from 43 boars representing 6 breeds, aged 33.9 to 149.3 weeks. Of the 903 semen samples collected, 30 samples (3.3%) were positive for PCV2 DNA by nPCR from 13 boars. Boars shedding PCV2 DNA in semen ranged between 35.9 and 71.0 weeks of age, and shedding occurred during a period of up to 27.3 weeks. A semen nPCR test was 2.6 times more likely to be positive when collected from pigs that were ≤52 weeks of age, and 3.0 times more likely to be positive when collected from pigs that were ≤26 weeks from time of entry into the stud main unit (generalized estimating equations: P = 0.02; 95% confidence interval [CI] of the odds ratio 1.2 to 5.5, and P = 0.01; 95% CI of the odds ratio 1.3 to 6.9, respectively). These results demonstrate a sporadic and long-term shedding pattern of PCV2 DNA in semen from naturally infected boars. PCV2 DNA in semen does not appear to have detrimental effects on sperm morphology; however, boar age and, possibly, breed may contribute to the persistence of PCV2-shedding in semen.
A Multiplex Polymerase Chain Reaction for Discriminating Erysipelothrix Rhusiopathiae from Erysipelothrix Tonsillarum
Authors: Yamazaki Yoshinao
Published: May 2011

Erysipelothrix rhusiopathiae is the causative agent of swine erysipelas, and it causes great economic losses in Japan and worldwide. In meat inspection, it is very important to distinguish E. rhusiopathiae from other bacteria showing similar clinical signs of disease or similar bacterial characteristics. To distinguish E. rhusiopathiae from Erysipelothrix tonsillarum, 2 polymerase chain reaction (PCR) systems were combined. The primer sets ERY-1F and ERY-2R were designed to amplify 2,210 base pairs (bp) of nucleotide sequence specific for E. rhusiopathiae chromosomal DNA, and the primer sets MO101 and ERS-1R were designed to amplify 719 bp of nucleotide sequence including a highly conserved region of genus Erysipelothrix 16S rRNA. Two fragments were amplified when E. rhusiopathiae was used as the PCR template using the primer sets, whereas a single fragment was amplified when E. tonsillarum was used as the template.
Detection of Antibodies to West Nile Virus in Equine Sera Using Microsphere Immunoassay
Authors: Balasuriya Udeni B. R. , Shi Pei-Yong , Wong Susan J. , Demarest Valerie L. , Gardner Ian A. , Hullinger Pamela J. , Ferraro Gregory L. , Boone Joshua D. , Cino Casey L. De , Glaser Amy L. , Renshaw Randall W. , Ledizet Michel , Koski Raymond A. , MacLachlan N. James
Published: May 2011

One hundred and ninety-one sera from horses that recently were exposed to West Nile virus (WNV) by either vaccination or natural infection or that were not vaccinated and remained free of infection were used to evaluate fluorescent microsphere immunoassays (MIAs) incorporating recombinant WNV envelope protein (rE) and recombinant nonstructural proteins (rNS1, rNS3, and rNS5) for detection of equine antibodies to WNV. The rE MIA had a diagnostic sensitivity and specificity, respectively, of 99.3% and 97.4% for detection of WNV antibodies in the serum of horses that were recently vaccinated or naturally infected with WNV, as compared to the plaque reduction neutralization test (PRNT). The positive rE MIA results were assumed to be WNV-specific because of the close agreement between this assay and the PRNT and the fact that unvaccinated control horses included in this study were confirmed to be free of exposure to the related St Louis encephalitis virus. The NS protein–based MIA were all less sensitive than either the rE MIA or PRNT (sensitivity 0–48.0), although the rNS1 MIA distinguished horses vaccinated with the recombinant WNV vaccine from those that were immunized with the inactivated WNV vaccine ( P < 0.0001) or naturally infected with WNV ( P < 0.0001). The rE MIA would appear to provide a rapid, convenient, inexpensive, and accurate test for the screening of equine sera for the presence of antibodies to WNV.
Assessment of a Bovine Viral Diarrhea Virus Antigen Capture ELISA and a Microtiter Virus Isolation ELISA Using Pooled Ear Notch and Serum Samples
Authors: Cleveland Susan M. , Salman M. D. , Van Campen Hana
Published: May 2011

To reduce the cost of whole herd screening for bovine viral diarrhea virus persistently infected animals, the sensitivity and specificity of an antigen-capture enzyme-linked immunosorbent assay (AC-ELISA) and a microtiter virus isolation ELISA using saline from ear notch samples or pooled serum was determined. Pooled saline from ear notch samples, assayed by AC-ELISA, gave a sensitivity and specificity of 98% and 94%, respectively, for pools containing 2 samples and 72% and 100%, respectively, for pools of 5. The sensitivity of pooled ear notch or serum samples for bovine viral diarrhea virus detection by microtiter virus isolation (sensitivity < 5%) or serum samples for detection by AC-ELISA (sensitivity < 15%) is too low to be used for whole herd screening. Pooling saline from ear notch samples from 2 animals tested by AC-ELISA, however, could provide a less expensive, reliable method for whole herd screening for bovine viral diarrhea virus.
First Report of Bluetongue Virus Serotype 1 Isolated from a White-Tailed Deer in the United States
Authors: Johnson Donna J. , Ostlund Eileen N. , Stallknecht David E. , Goekjian Virginia H. , Jenkins-Moore Melinda , Harris Steven C.
Published: May 2011

In November 2004, tissues were collected from a hunter-killed white-tailed deer in St. Mary Parish, Louisiana. Bluetongue virus (BTV) was isolated from the tissues; however, the isolate could not be identified as any of the US domestic serotypes. Subsequent testing by virus neutralization using serotype-specific antiserum tentatively identified the isolate as BTV serotype 1 (BTV-1), which had not previously been found in the United States. Primers were designed based on the sequence of an outer capsid protein gene of a South African BTV-1 strain. Reverse transcription–polymerase chain reaction testing with the BTV-1 primers and product sequencing confirmed the Louisiana isolate as BTV-1. This is the first report of BTV-1 in the United States.
Panniculitis in a Horse with Peripancreatitis and Pancreatic Fibrosis
Authors: Waitt Laura H. , Cebra Christopher K. , Tornquist Susan J. , Löhr Christiane V.
Published: May 2011

A 22-year-old pinto mixed breed mare was admitted for evaluation of severe colic signs and gastric reflux. Multiple nonpainful, variably sized hard masses were palpated in the subcutis over the thorax, abdomen, and hindquarters of the horse. The mare was diagnosed with sterile peritonitis and had systemically high γ-glutamyltransferase, amylase, and lipase. Three days into treatment she became febrile with signs of persistent and mild abdominal discomfort; euthanasia was elected. Necropsy revealed peripancreatitis, pancreatic fibrosis, abdominal steatitis and panniculitis. Panniculitis associated with peripancreatitis has been described in humans and dogs but not in horses. Pathogenic models for this entity are discussed.
Complete Diphallia, Imperforate Ani (Type 2 Atresia Ani), and an Accessory Scrotum in a 5-Day-Old Calf
Authors: Loynachan Alan T. , Jackson Carney B. , Harrison Lenn R.
Published: May 2011

A 5-day-old Angus calf was submitted to the necropsy service at the University of Kentucky Livestock Disease Diagnostic Center. At birth, the calf was noted to have 2 scrota. Five days post-calving the calf developed severe tenesmus and an umbilical infection. Clinical examination revealed atresia ani. At necropsy, complete diphallus, imperforate ani (type 2 atresia ani), and remnants of 2 scrota were identified. This report describes the gross pathologic appearance of a rare case of complete diphallia, imperforate ani (type 2 atresia ani), and an accessory scrotum in a 5-day-old calf.
Congenital Diaphragmatic Eventration in a Stillborn Foal
Authors: Jackson Carney , Collyer Petra Borchers , Loynachan Alan
Published: May 2011

A stillborn full term foal was presented at necropsy. The dilated duodenum and the dorsal and ventral left colon and cecum extended into the thoracic cavity and were encased by a thin transparent membrane originating from the diaphragm. The congenital condition was diagnosed as a developmental anomaly with diaphragmatic eventration. To the authors' knowledge, this is the first report of diaphragmatic eventration in a Thoroughbred stillborn foal.
Rib Osteoblastic Osteosarcoma in an African Hedgehog (Atelerix Albiventris)
Authors: Benoit-Biancamano Marie-Odile , d'Anjou Marc-André , Girard Christiane , Langlois Isabelle
Published: May 2011

A 3-year-old African hedgehog ( Atelerix albiventris) was presented to the Exotic Animal Clinic of the University of Montreal for evaluation of a mass growing on the right thoracic wall. The diagnostic workup, which included helical computed tomography, confirmed the presence of a large mass, originating from the right 7th rib, infiltrating the thoracic wall and cavity. The animal was euthanized due to the poor prognosis. At necropsy, a well-demarcated mass penetrated the thoracic wall and incorporated the 6th to 8th ribs. Cut sections of the tumor were white, glistening, firm, and gritty. Microscopically, it was composed of polyhedral to elongated cells with interspersed trabeculae of osteoid and large areas of coagulative necrosis. On the basis of histopathologic findings, a diagnosis of osteoblastic osteosarcoma was made. To the authors' knowledge, this is the first report of an osteoblastic osteosarcoma on the thoracic wall of an African hedgehog, as well as the first report of the use of helical computed tomography in that species.
Histologic Features and Clinical Outcomes of Melanomas of Lip, Haired Skin, and Nail Bed Locations of Dogs
Authors: Schultheiss Patricia C.
Published: May 2011

Outcomes, signalments, and the relationship of histologic features with the outcome of melanomas located in lip, nail bed, and haired skin of dogs were reviewed. These melanomas were diagnosed as benign or malignant based on histologic features. Melanomas of the lip arose from mucous membrane in most cases. 32 dogs with lip melanomas that had histologic features of malignancy, 22 died because of the tumor within 1 year and 10 were tumor free for at least 1 year following removal. Of 10 dogs with melanomas with benign histologic features on the mucous membrane of the lip, 9 were tumor free for at least 1 year. Of 4 dogs with benign appearing tumors of the haired skin of the lip, 3 were tumor free for at least 1 year. Melanomas with histologic features of malignancy occurred in many locations of haired skin, and 11 of 24 dogs were tumor free for at least 1 year. All nail bed melanomas had histologic features of malignancy and all invaded the third phalanx, but 6 of 14 dogs were tumor free for at least 1 year after amputation of the digit. Among these dogs, the 1-year survival rates for tumors classified as malignant by histologic features were 69% for lip, 46% for haired skin, and 43% for nail bed. However, the clinical outcome of an individual malignant tumor could not be predicted accurately by any specific histologic features.
Test Strategies in Bovine Viral Diarrhea Virus Control and Eradication Campaigns in Europe
Authors: Houe H. , Lindberg A. , Moennig V.
Published: May 2011

Several European countries have initiated national and regional control-and-eradication campaigns for bovine viral diarrhea virus (BVDV). Most of these campaigns do not involve the use of vaccines; in Germany, vaccination is used only in states in which it is considered necessary because of high BVDV prevalence. In European countries without organized BVDV control programs, vaccination is commonly used to control BVDV. Diagnostic test strategies are fundamental to all control-and-eradication campaigns; therefore, the purpose of this review is to describe how the available diagnostic tests are combined into test strategies in the various phases of control-and-eradication campaigns in Europe. Laboratory techniques are available for BVDV diagnosis at the individual animal level and at the herd level. These are strategically used to achieve 3 main objectives: 1) initial tests to classify herd status, 2) follow-up tests to identify individual BVDV-infected animals in infected herds, and 3) continued monitoring to confirm BVDV-free status. For each objective or phase, the validity of the diagnostic tests depends on the mode of BVDV introduction and duration of infection in test-positive herds, and on how long noninfected herds have been clear of BVDV. Therefore, the various herd-level diagnostic tools—such as antibody detection in bulk milk or in blood samples from young stock animals, or BVDV detection in bulk milk—need to be combined appropriately to obtain effective strategies at low cost. If the individual diagnostic tests are used with due consideration of the objectives of a specific phase of a BVDV control program, they are effective tools for controlling and eradicating BVDV in regions not using vaccination and where vaccination is a part of the control or eradication program.
Comparison of Milk and Serum Enzyme–Linked Immunosorbent Assays for Diagnosis of Mycobacterium Avium Subspecies Paratuberculosis Infection in Dairy Cattle
Authors: Lombard Jason E. , Byrem Todd M. , Wagner Bruce A. , McCluskey Brian J.
Published: May 2011

Milk and serum samples from 35 dairy herds in 17 states were evaluated for cow- and herd-level Mycobacterium avium subspecies paratuberculosis (MAP) antibody test agreement. Evaluation of 6,349 samples suggested moderate agreement between milk and serum enzyme-linked immunosorbent assay (ELISA) results, with a kappa value of 0.50. Cow-level sensitivity (Se) for 18 dairy operations with 1,921 animals was evaluated relative to fecal culture results. At the cow level, the milk ELISA relative Se was not significantly different from that of the serum ELISA (21.2 and 23.5%, respectively). Logistic regression models revealed a positive association between lactation number and milk ELISA status. Non-Holstein cows were more likely to test milk ELISA positive than Holstein cows. Cows in the first 2 weeks of lactation and after week 45 of lactation were more likely to test milk ELISA positive than cows between 3 and 12 weeks of lactation. Milk production > 80% of herd average was negatively associated with testing milk ELISA positive. Animals in the West and Midwest regions were less likely than animals in the Southeast region to test ELISA positive by either test. Estimates for herd-level sensitivity for the milk and serum ELISA, relative to fecal culture results, ranged from 56 to 83%. At the cow and herd levels, milk ELISA performed equivalent to serum ELISA using fecal culture as a reference for MAP infection and has the advantage of decreased labor costs on farms that use Dairy Herd Improvement Association testing.
A New Fluorogenic Real-Time RT-PCR Assay for Detection of Lineage 1 and Lineage 2 West Nile Viruses
Authors: Jiménez-Clavero Miguel Angel , Agüero Montserrat , Rojo Gema , Gómez-Tejedor Concepción
Published: May 2011

West Nile virus represents an emerging threat for animal and human health worldwide. This virus exhibits a marked genetic variation, with at least 2 distinct evolutionary lineages. Lineage 1 has been recognized in Africa, Asia, Europe, Oceania, and more recently in the Americas, whereas lineage 2 is restricted to Africa. Perhaps for this reason, the available real-time RT-PCR methods for detecting West Nile virus genome have mainly focused on lineage 1. However, both viruses may potentially be spread beyond their endemic areas by migratory birds. This report describes a new real-time reverse transcription-PCR (RT-PCR) method based on a 59-Taq nuclease-39 minor groove binder DNA probe (TaqMan MGBE) that allows the detection of a wide range of West Nile virus isolates, including both lineages 1 and 2. This method was able to detect West Nile viruses from different origins (North and Central Africa, Middle East, Europe, and North America), whereas other flaviviruses (Usutu, Dengue, Yellow fever) analyzed in parallel remained negative. The sensitivity achieved by this assay was 1022-1023 pfu/tube. This method, which can be performed in 96-well format, could be suitable for the large-scale surveillance of West Nile virus in areas where both lineages can potentially spread.