bluuebunny/crossref_metadata_2025 huggingface dataset:
Detection of Lawsonia Intracellularis in Swine using Polymerase Chain Reaction Methodology
Authors: Jordan Dianna M. , Knittel Jeffrey P. , Roof Michael B. , Schwartz Kent , Larson David , Hoffman Lorraine J.
Published: May 2011

The polymerase chain reaction (PCR) was evaluated for its usefulness as a diagnostic tool to detect Lawsonia (ileal symbiont) intracellularis. Porcine ilea were collected from swine cases submitted to the Iowa State University Veterinary Diagnostic Laboratory between December 1, 1994, and June 30, 1995. Sampling was random, with no regard to health status. There were 621 ileum scrapings evaluated using the PCR technique. Thirty-five of the samples were positive, either by PCR or conventional diagnostic methods such as histology and Warthin-Starry silver stain. These 35 samples were further evaluated by Warthin-Starry silver stain and indirect immunofluorescent antibody test (IFAT) to confirm the presence of L. intracellularis in the tissue sections. Of the 26 samples positive by PCR, 22 were positive by IFAT. Sixteen of the 22 were also positive when stained with Warthin-Starry and evaluated microscopically for typical bacteria. Nine of the original samples were negative by all 3 techniques. PCR appears more sensitive and specific for L. intracellularis detection than Warthin-Starry stain and IFAT. This study provides evidence that PCR may be useful as a reference standard for the detection of L. intracellularis. PCR may be an appropriate monitoring tool for swine herds because it is a rapid procedure that could be applied to batch testing. Although the test is currently too laborious and expensive for routine diagnostic use, there may be situations in which it is justified because of the advantages of greater sensitivity and specificity.
The Prevalence of Cryptosporidium and Giardia Spp. in Fecal Samples from Free-Ranging White-Tailed Deer (Odocoileus Virginianus) in the Southeastern United States
Authors: Rickard Lora G. , Siefker Carla , Boyle Carolyn R. , Gentz Edward J.
Published: May 2011

The prevalence of Cryptosporidium sp. and Giardia sp. in fecal samples collected from free-ranging white-tailed deer ( Odocoileus virginianus) was determined for animals in Virginia and Mississippi. The 34 Virginia samples came from young of the year (≤6 months of age) presented to the Wildlife Center of Virginia, and the 360 Mississippi samples came from animals (≥6 months of age) collected from 42 different sites across the state. Cryptosporidial oocysts were detected in 8.8% and 5.0% and Giardia sp. cysts were detected in 2.9% and 1.1% of the samples from Virginia and Mississippi, respectively. Analysis of data by collection site was not possible because of the small number of samples from some sites. Therefore, collection sites in Mississippi were grouped by the stream basin in which they occurred. Binary logistic regression analysis indicated the probability of infection with Cryptosporidium sp. decreased with increasing age of animal in all positive stream basins except 1, in which the pattern was reversed. This is the first report of either Cryptosporidium sp. or Giardia sp. in free-ranging white-tailed deer. The role of white-tailed deer in the epidemiology of these parasites is unknown. However, this study demonstrates that white-tailed deer shed cysts and oocysts of both parasites in the environment and must be considered potential sources of contamination.
Toxicosis in Cattle from Concurrent Feeding of Monensin and Dried Distiller's Grains Contaminated with Macrolide Antibiotics
Authors: Basaraba R. J. , Oehme F. W. , Vorhies M. W. , Stokka G. L.
Published: May 2011

Consumption of monensin-containing feed contaminated with macrolide antibiotic residues resulted in the death of cattle from multiple feedlots in south-central Kansas. Cattle were fed milo dried distiller's grains (DDG) with solubles from a common source in conjunction with the ionophore antibiotic, monensin. Deaths occurred as early as 72–96 hours after feeding and were preceded by either no premonitory signs or 1 or more of the following: anorexia, depression, dyspnea, locomotor deficits, and recumbency. Significant gross lesions were pulmonary and mesenteric edema, hepatomegaly, and generalized myocardial and skeletal muscle pallor that was confirmed histologically as acute myodegeneration and necrosis. Other significant histologic lesions included centrolobular hepatocellular necrosis, congestion, and pulmonary interstitial and alveolar edema with fibrin exudation. Animals that survived beyond 6 weeks had poor weight gain and coalescing foci of myocardial fibrosis with residual myocardial degeneration. Analysis of trace mineral supplements for monensin were within the manufacturer's label range. The DDG samples from affected feedlots had 50–1,500 ppm of erythromycin, clarithromycin, and related macrolide antibiotic analogues, which originated in the alcohol residue. In a preliminary feeding trial, cattle fed this contaminated DDG in combination with monensin had clinical signs and died with gross and histologic findings comparable to those of the field cases. Even though rations supplemented with the contaminated DDG contained approved levels of monensin, the clinical and postmortem findings were consistent with those expected for monensin toxicosis. The presence of macrolide antibiotic residues in the contaminated feed appeared to affect the biotransformation of otherwise nontoxic levels of monensin, leading to clinical ionophore toxicosis.
Diagnosis of Malignant Catarrhal Fever by PCR Using Formalin-Fixed, Paraffin-Embedded Tissues
Authors: Crawford Timothy B. , Li Hong , O'Toole Donal
Published: May 2011

A previously described polymerase chain reaction (PCR) assay (amplification of a 238-bp fragment of ovine herpesvirus 2 [OHV-2] genomic DNA) for diagnosis of sheep-associated malignant catarrhal fever (MCF) was adapted for use on formalin-fixed, paraffin-embedded tissues. Variables affecting its use were examined. Archived tissues from cattle, white-tailed deer ( Odocoileus virginianus), and bison ( Bison bison) diagnosed with MCF by clinical signs or histologic lesions were obtained from 2 veterinary diagnostic laboratories. Tissues from healthy animals and from animals diagnosed with other common bovine viral diseases were examined as controls. A total of 86 blocks from 37 suspect MCF cases were examined. Forty-one blocks from healthy animals and animals with unrelated viral diseases were examined as controls. The assay was specific for sheep-associated MCF and did not yield false-positive signals from healthy animals or from cases of infectious bovine rhinotracheitis, bovine virus diarrhea, mucosal disease, or parainfluenza-3 virus infection. A wide variety of tissues were suitable substrates, including spleen, lymph node, intestine, brain, lung, and kidney. Extracted DNA provided a more suitable target than did unextracted tissue lysate. The highest levels of viral DNA were present in lymphoid organs and intestine, but the data indicate that in acute clinical cases, most organs contain sufficient viral DNA to serve as a suitable diagnostic specimen. Fixation of 0.5-cm3 blocks of tissue in 10% neutral buffered formalin was deleterious to the target DNA, and PCR signals progressively diminished after fixation for >45 days. Detection of genomic DNA of OHV-2 by PCR was successful for archived tissues that were 15 years old.
Comparison of PCR, Virus Isolation, and Indirect Fluorescent Antibody Staining in the Detection of Naturally Occurring Feline Herpesvirus Infections
Authors: Burgesser Kent M. , Hotaling Stephanie , Schiebel Anita , Ashbaugh Scott E. , Roberts Steven M. , Collins James K.
Published: May 2011

Cats with clinical signs suggestive of ocular infection with feline herpesvirus type 1 (FHV 1) and cats without such signs were assayed by 3 methods to detect FHV. Comparison of polymerase chain reaction (PCR), virus isolation, and indirect fluorescent antibody staining techniques for the detection of FHV demonstrated higher sensitivity of PCR in detecting this common infectious agent of cats. Compared with PCR, sensitivity and specificity for virus isolation was 49% and 100%, respectively, and those of indirect immunofluorescence were 29% and 96%, respectively. FHV was detected in 13.7% of client-owned cats with conjunctivitis and in 31% of shelter cats with no ocular signs. The use of FHV PCR as a diagnostic test for FHV-associated disease is limited because of the occurrence of healthy carriers.
Encephalitis Induced by Bovine Herpesvirus 5 and Protection by Prior Vaccination or Infection with Bovine Herpesvirus 1
Authors: Cascio Karin E. , Belknap Ellen B. , Schultheiss Patricia C. , Ames Abbe D. , Collins James K.
Published: May 2011

Calves were intranasally challenged with bovine herpesvirus 5 (BHV5) and followed for the development of viral infection, clinical encephalitis, histologic lesions in the brain, and viral sequences in the trigeminal ganglia. Calves that were previously vaccinated with bovine herepesvirus 1 (BHV1, n = 4) or previously infected with BHV1 ( n = 5) or that had not been exposed to either virus ( n = 4) were compared. No calf developed signs of encephalitis, although all calves developed an infection as indicated by nasal secretion of BHV5 and seroconversion to the virus. Histologic lesions of encephalitis consisting of multifocal gliosis and perivascular cuffs of lymphocytes were observed in calves not previously exposed to BHV1. BHV5 sequences were amplified from the trigeminal ganglia of calves previously vaccinated and from calves not previously exposed to BHV1; calves sequentially challenged with BHV1 and later BHV5 had exclusively BHV1 sequences in their trigeminal ganglia. Administration of dexamethasone 28 days after BHV5 challenge did not influence clinical disease or histologic lesions in either previously unexposed calves ( n = 2) or previously immunized calves ( n = 2), although it did cause recrudescence of BHV5, as detected by nasal virus secretion.
Genomic Fingerprinting and Development of a Dendrogram for Brucella spp. Isolated from Seals, Porpoises, and Dolphins
Authors: Jensen Allen E. , Cheville Norman F. , Thoen Charles O. , MacMillan Alastair P. , Miller W. George
Published: May 2011

Genomic DNA from reference strains and biovars of the genus Brucella was analyzed using pulsed-field gel electrophoresis (PFGE). Fingerprints were compared to estimate genetic relatedness among the strains and to obtain information on evolutionary relationships. Electrophoresis of DNA digested with the restriction endonuclease XbaI produced fragment profiles for the reference type strains that distinguished these strains to the level of species. Included in this study were strains isolated from marine mammals. The PFGE profiles from these strains were compared with those obtained from the reference strains and biovars. Isolates from dolphins had similar profiles that were distinct from profiles of Brucella isolates from seals and porpoises. Distance matrix analyses were used to produce a dendrogram. Biovars of B. abortus were clustered together in the dendrogram; similar clusters were shown for biovars of B. melitensis and for biovars of B. suis. Brucella ovis, B. canis, and B. neotomae differed from each other and from B. abortus, B. melitensis, and B. suis. The relationship between B. abortus strain RB51 and other Brucella biovars was compared because this strain has replaced B. abortus strain 19 for use as a live vaccine in cattle and possibly in bison and elk. These results support the current taxonomy of Brucella species and the designation of an additional genomic group(s) of Brucella. The PFGE analysis in conjunction with distance matrix analysis was a useful tool for calculating genetic relatedness among the Brucella species.
Evaluation of the Baculovirus-Expressed S Glycoprotein of Transmissible Gastroenteritis Virus (TGEV) as Antigen in a Competition ELISA to Differentiate Porcine Respiratory Coronavirus from TGEV Antibodies in Pigs
Authors: Sestak Karol , Zhou Zhongfang , Shoup David I. , Saif Linda J.
Published: May 2011

The spike (S) glycoprotein of the Miller strain of transmissible gastroenteritis virus (TGEV) was recently cloned and expressed in baculovirus. The recombinant S protein was used as the coating antigen in a competition (blocking) enzyme-linked immunosorbent assay (ELISA) in combination with monoclonal antibodies to the S protein epitope A (conserved on TGEV and porcine respiratory coronavirus [PRCV]) or epitope D (present on TGEV only) to differentiate PRCV- from TGEV-induced antibodies. One set (set A) of 125 serum samples were collected at different times after inoculation of caesarean-derived, colostrum-deprived ( n = 52) and conventional young pigs ( n = 73) with 1 of the 2 porcine coronaviruses or uninoculated negative controls (TGEV/PRCV/negative = 75/30/20). A second set (set B) of 63 serum samples originated from adult sows inoculated with PRCV and the recombinant TGEV S protein or with mock-protein control and then exposed to virulent TGEV after challenge of their litters. Sera from set A were used to assess the accuracy indicators (sensitivity, specificity, accuracy) of the fixed-cell blocking ELISA, which uses swine testicular cells infected with the M6 strain of TGEV as the antigen source (ELISA 1) and the newly developed ELISA based on the recombinant S protein as antigen (ELISA 2). The sera from set B (adults) were tested for comparison. The plaque reduction virus neutralization test was used as a confirmatory test for the presence of antibodies to TGEV/PRCV in the test sera. The accuracy indicators for both ELISAs suggest that differential diagnosis can be of practical use at least 3 weeks after inoculation by testing the dual (acute/convalescent) samples from each individual in conjunction with another confirmatory (virus neutralization) antibody assay to provide valid and complete differentiation information. Moreover, whereas ELISA 1 had 10–20% false positive results to epitope D for PRCV-infected pigs (set A samples), no false-positive results to epitope D occurred using ELISA 2, indicating its greater specificity. The progression of seroresponses to the TGEV S protein epitopes A or D, as measured by the 2 ELISAs, was similar for both sets (A and B) of samples. Differentiation between TGEV and PRCV antibodies (based on seroresponses to epitope D) was consistently measured after the third week of inoculation.
Experimental Infection of Calves with Bovine Viral Diarrhea Virus Genotype II (NY-93)
Authors: Odeón Anselmo C. , Kelling Clayton L. , Marshall Donald J. , Estela E. Sonia , Dubovi Edward J. , Donis Ruben O.
Published: May 2011

To ascertain the virulence of bovine viral diarrhea virus (BVDV) genotype II, isolate NY-93 was inoculated intranasally into 3 calves, 2 of which were treated with a synthetic glucocorticoid prior to and after virus inoculation. Anorexia, fever (up to 42 C), dyspnea, and hemorrhagic diarrhea developed 6 days after intranasal inoculation with BVDV NY-93. The condition of all calves deteriorated further until the end of the study on day 14 postinoculation. The most significant postmortem macroscopic changes in all calves were limited to the gastrointestinal tract and consisted of moderate to severe congestion of the mucosa with multifocal hemorrhages. Microscopic lesions found in the gastrointestinal tract were similar to those observed in mucosal disease, including degeneration and necrosis of crypt epithelium and necrosis of lymphoid tissue throughout the ileum, colon, and rectum. The basal stratum of the epithelium of tongue, esophagus, and rumen had scattered individual necrotic cells. Spleen and lymph nodes had lymphocytolysis and severe lymphoid depletion. Severe acute fibrinous bronchopneumonia was present in dexamethasone-treated calves. Abundant viral antigen was detected by immunohistochemistry in the squamous epithelium of tongue, esophagus, and forestomachs. BVDV antigen was prominent in cells of the media of small arteries and endothelial cells. The presence of infectious virus in tissues correlated with an absence of circulating neutralizing antibodies. These findings highlight the potential of BVDV genotype II to cause severe disease in normal and stressed cattle.
Dot Immunobinding Assay for Detection of Bovine Herpesvirus 4 Antibodies in Rabbits
Authors: Essmail Mohamed , Baker Dale , Collins James , Vandewoude Sue , Salman Mohamed , Hegazy A. A.
Published: May 2011

A dot immunobinding assay (DIA) was used for the detection of antibody against bovine herpesvirus 4 (BHV-4) in experimentally infected specific-pathogen-free male and female rabbits. A semipurified virus preparation was used as the antigen, and protein A/G—horseradish—peroxidase conjugate and diaminobenzidine tetrahydrochloride were used as the detection system. Results of the DIA procedure on serum samples of experimentally infected male and female rabbits were compared with those of a complement-dependent virus neutralization (VN) test. None of the tested sera (0/60 samples) from either male or female rabbits were positive by the complement-dependent VN test. Results of the DIA procedure for the same tested sera were positive in 35 of 60 samples (58%) from BHV-4 infected rabbits, indicating higher sensitivity of DIA procedure as compared with the complement-dependent VN test.
Application of a Nested Polymerase Chain Reaction Assay to Detect Mycoplasma Hyopneumoniae from Nasal Swabs
Authors: Calsamiglia Maria , Pijoan Carlos , Trigo Alicia
Published: May 2011

The porcine respiratory disease complex (PRDC) is an increasingly important cause of decreased swine productivity and is characterized by slow growth, decreased feed efficiency, anorexia, cough, and dyspnea. Mycoplasma hyopneumoniae is among the most prevalent and important infectious agents associated with PRDC. Understanding of mycoplasmal pneumonia has been hindered by inadequate diagnostic methods. Many of the currently available tests are relatively insensitive or nonspecific when used in a diagnostic laboratory setting or are too costly or difficult for routine diagnostic use. Several polymerase chain reaction (PCR) assays have been described, but they are not sensitive enough to detect the microorganisms in live pigs, from either nasal or tracheal swabs. A nested PCR using 2 species-specific sets of primers from the 16S ribosomal DNA gave positive results with as little as 80 microorganisms and did not cross-react with other mycoplasma species or with other microorganisms commonly found in the respiratory tract of pigs. This assay was better suited for detection of M. hyopneumoniae from nasal swabs than was conventional PCR. Nasal swab samples were taken at different time periods following experimental challenge of 10 susceptible pigs. Only 2 of the 55 swabs examined gave a positive result with conventional PCR, whereas 30 of the 55 swabs gave a positive result using the nested PCR. Twenty of 40 (50%) nasal swabs from pigs experiencing a respiratory disease outbreak where M. hyopneumoniae had been diagnosed also gave a positive result with the nested PCR. To confirm that the amplified product was specific, 4 nested PCR products were purified, sequences were determined and aligned, and they were confirmed to be from M. hyopneumoniae.
Isolation of Cilia-Associated Respiratory (CAR) Bacillus from Pigs and Calves and Experimental Infection of Gnotobiotic Pigs and Rodents
Authors: Nietfeld Jerome C. , Fickbohm Barry L. , Rogers Douglas G. , Franklin Craig L. , Riley Lela K.
Published: May 2011

Filamentous, gram-negative bacteria morphologically similar to cilia-associated respiratory (CAR) bacillus of rodents and rabbits were isolated from the tracheas of 5 pigs and 4 calves. All pigs but none of the calves had histologic lesions of chronic tracheitis. In silver-stained histologic sections, CAR bacilli were adhered to the tracheal epithelium of each pig but were not found in the calves. Like CAR bacillus of rats, the bacteria displayed gliding motility and grew only in cell culture or cell culture medium supplemented with fetal serum. Initially, all isolates were contaminated by Mycoplasma spp. This contamination was eliminated from 4 pig isolates by limiting dilutions, and mycoplasma-free isolates were used to intranasally inoculate gnotobiotic pigs and CAR bacillus-free mice and rats and to immunize guinea pigs. The gnotobiotic pigs remained healthy, and when they were necropsied 4 and 7 weeks after infection no macroscopic or microscopic lesions were found in the respiratory tract. However, CAR bacillus was isolated at both times from the nasal cavities and tracheas of inoculated pigs, and the ciliated tracheal epithelium of infected pigs necropsied 7 weeks after infection was colonized by low numbers of CAR bacillus-like bacteria. The rats and mice remained healthy through week 12 postinoculation, and evidence of short- or long-term colonization was not detected by histologic examination or culture. When used as primary antibody for immunohistochemical staining, sera from guinea pigs immunized with pig CAR bacillus specifically stained CAR bacilli colonizing the respiratory epithelium of naturally infected pigs, whereas sera collected prior to immunization failed to react with the bacteria. These results indicate that CAR bacilli are unlikely to be primary pathogens of pigs or cattle and that rodents do not act as reservoirs.
A Lysosomal Storage Disease Induced by Ipomoea Carnea in Goats in Mozambique
Authors: de Balogh Katalin K. I. M. , Dimande Alberto P. , van der Lugt Jaco J. , Molyneux Russell J. , Naudé Theuns W. , Welman Wilhelmina G.
Published: May 2011

A novel plant-induced lysosomal storage disease was observed in goats from a village in Mozambique. Affected animals were ataxic, with head tremors and nystagmus. Because of a lack of suitable feed, the animals consumed an exotic hedge plant growing in the village that was identified as Ipomoea carnea (shrubby morning glory, Convolvulaceae). The toxicosis was reproduced by feeding I. carnea plant material to goats. In acute cases, histologic changes in the brain and spinal cord comprised widespread cytoplasmic vacuolation of neurons and glial cells in association with axonal spheroid formation. Ultrastructurally, cytoplasmic storage vacuoles in neurons were membrane bound and consistent with lysosomes. Cytoplasmic vacuolation was also found in neurons in the submucosal and mesenteric plexuses in the small intestine, in renal tubular epithelial cells, and in macrophage-phagocytic cells in the spleen and lymph nodes in acute cases. Residual alterations in the brain in chronic cases revealed predominantly cerebellar lesions characterized by loss of Purkinje neurons and gliosis of the Purkinje cell layer. Analysis of I. carnea plant material by gas chromatography—mass spectrometry established the presence of the mannosidase inhibitor swainsonine and 2 glycosidase inhibitors, calystegine B2 and calystegine C1, consistent with a plant-induced α-mannosidosis in the goats. The described storage disorder is analogous to the lysosomal storage diseases induced by ingestion of locoweeds ( Astragalus and Oxytropis) and poison peas ( Swainsona).
Neuronal Vacuolation in Raccoons from Oregon
Authors: Hamir Amir N. , Fischer Kay A.
Published: May 2011

During a 2-year period (1995–1997), vacuoles were detected in neurons of 21/50 (42% prevalence) raccoons ( Procyon lotor) in Oregon. Age or sex predisposition was not apparent. Twenty of these raccoons were from within a radius of 40 km of Corvallis in western Oregon. Microscopically, the vacuoles were variable in size, were in the perikarya, and were consistently present in pontine nuclei. Brain tissues were negative for rabies virus antigen by fluorescent antibody test and for the protease-resistant protein prion by immunohistochemistry. Electron microscopic examination of the brain stem of selected animals revealed accumulation of electron-dense material within neuronal perikarya. Light and electron microscopic examination indicated that the accumulated intracellular material had a high lipid content. These lesions suggest a form of neuronal storage condition. Further research is required to identify the composition of the intracellular lipid material, to elucidate the mechanism of neuronal vacuolation in raccoons, and to understand the basis for the apparent geographic restriction of this lesion.
Development of a Blocking ELISA for Screening Antibodies to Porcine Rubulavirus, La Piedad Michoacan Virus
Authors: Nordengrahn A. , Svenda M. , Moreno-Lopez J. , Bergvall A. , Hernandez P. , McNeilly F. , Allan G. , Merza M.
Published: May 2011

A blocking enzyme-linked immunosorbent assay (ELISA) was developed to detect antibodies to porcine rubulavirus (La Piedad Michoacan Virus [LPMV]) in serum samples from pigs. The test, based on a monoclonal antibody against the LPMV hemagglutinin-neuraminidase glycoprotein, had a sensitivity of 99% and a specificity of 97%. The results of this test were in agreement with those obtained by an indirect ELISA and hemagglutination inhibition, indirect immunofluorescence, and virus neutralization tests. The blocking ELISA is considered the most suitable test for routine screening for antibodies against LPMV.
Evaluation of an Immunofluorescent Antibody Test to Detect Bovine Herpesvirus 1-Specific IgM
Authors: Graham D. A. , Foster J. C. , German A. , McLaren I. E. , Adair B. M. , Merza M.
Published: May 2011

An indirect immunofluorescent antibody test (IIFAT) was developed to detect bovine herpesvirus 1 (BHV-1)-specific IgM. All sera were treated with protein-G agarose prior to testing to eliminate the possibility of false-positive results due to IgM-isotype rheumatoid factor (IgM-RF). Specific IgM was first detected 8 days after experimental infection of 3 calves free of maternally derived antibody, with peak responses occurring 2–7 days later. Seroconversion was detected in all 3 calves using a single-dilution enzyme-linked immunosorbent assay. Following reinfection at 30 days postinfection, a low-level IgM response was detected in only 1 calf. Seroconversion was detected in 2 calves. There was no evidence of activation of IgM-RF by infection or reinfection with BHV-1. When 87 acute and convalescent serum pairs collected from 21 outbreaks of respiratory disease were tested, specific IgM was detected in 58 animals (66.6%) from 19 (90.5%) outbreaks. Seroconversion was detected in 44 of these animals (50.6%) from 17 outbreaks (81.0%). The correlations between these 2 assays on a calf and outbreak basis were 79.3% and 90.5%, respectively. Specific IgM was detected in 17/20 sera (85.0%) collected from an additional outbreak. No virus was detected by virus isolation or immunofluorescent staining in nasal mucus samples collected at the same time. Detection of specific IgM by IIFAT is a useful technique for the serodiagnosis of BHV-1 infection.
Comparison of Visual Microscopic and Computer-Automated Fluorescence Detection of Rabies Virus Neutralizing Antibodies
Authors: Péharpré Daniel , Cliquet Florence , Sagné Eveline , Renders Caroline , Costy Françoise , Aubert Miche
Published: May 2011

The rapid fluorescent focus inhibition test (RFFIT) and the fluorescent antibody virus neutralization test (FAVNT) are both diagnostic tests for determining levels of rabies neutralizing antibodies. An automated method for determining fluorescence has been implemented to reduce the work time required for fluorescent visual microscopic observations. The automated method offers several advantages over conventional visual observation, such as the ability to rapidly test many samples. The antibody titers obtained with automated techniques were similar to those obtained with both the RFFIT ( n = 165, r = 0.93, P < 0.001) and the FAVNT ( n = 52, r = 0.99, P < 0.001).
An Enzyme-Linked Immunosorbent Assay Using Nuclear Antigen for Detection of Feline Herpesvirus 1 Antibody
Authors: Satoh Y. , Iizuka K. , Fukuyama M. , Kishikawa S. , Nishino Y. , Ikeda T. , Kiuchi A. , Hara M. , Tabuchi K.
Published: May 2011

To detect antibody against feline herpesvirus 1 (FHV-1) in the sera of cats, the sensitivity and specificity of an enzyme-linked immunosorbent assay (ELISA) using nuclear antigen was investigated. The standardized optical density readings (ODs) of the ELISA obtained by the 1-step serum dilution (1:80) method were compared with the serum neutralization test (SNT) results, with a correlation of 0.993, and with the hemagglutination inhibition (HI) test results, with a correlation of 0.851. The ODs for the ELISA titers were obtained using the serial serum dilution method and were compared with the SNT results, with a correlation of 0.933, and with the HI test results, with a correlation of 0.987. In the experimental infection of 4 specific-pathogen-free cats, the results of different serologic tests (SNT and HI) and the ELISA using the serial serum dilution method revealed rapid production of antibodies after inoculation, whereas the ELISA using the one-step serum dilution method indicated that titers increased more slowly. These results indicate that with the present ELISA using nuclear antigen, there are fewer demands on time and labor, making the method convenient for monitoring FHV-1 infection.
Selenium Elimination in Pigs after an Outbreak of Selenium Toxicosis
Authors: Davidson-York Dorothy , Galey Francis D. , Blanchard Patricia , Gardner Ian A.
Published: May 2011

In May 1996, 150 grower pigs in 5 California counties were exposed to selenium-contaminated feed distributed by a single feed company. Feed samples from 20 herds had a mean selenium concentration of 121.7 ppm dry weight (range, 22.1–531 ppm). In San Luis Obispo County, 52 pigs in 24 herds were exposed to the feed, and 8 pigs died with signs of paralysis. Bilateral symmetrical poliomyelomalacia involving the ventral horns of the cervical and lumbar intumescence was evident on histologic examination of spinal cord from affected pigs. Of 44 surviving exposed pigs, 33 (75%) exhibited signs of selenosis, including anorexia, alopecia, and hoof lesions. Thirty-nine of 44 pigs (88.6%) had elevated (>1 ppm) blood selenium concentrations. Surviving exposed pigs were changed to a standard commercial ration containing approximately 0.5 ppm (dry weight) selenium. Blood selenium concentrations were determined weekly for 46 days following removal of the contaminated feed and were compared with values of 20 control pigs fed a standard commercial ration. Mean (±SD) blood selenium concentrations of exposed pigs were 3.2 ± 2.6 ppm at the initial sampling and 0.4 ± 0.1 ppm after 46 days. Mean blood selenium concentrations of ≤ 0.3 ppm for control pigs at all samplings were significantly lower ( P < 0.001) than concentrations for exposed pigs. Muscle and liver samples of 22 of the 44 exposed pigs were collected at slaughter approximately 72 days after withdrawal of the selenium-contaminated feed. Muscle samples had a mean selenium concentration of 0.36 ppm (wet weight). Liver samples had a mean selenium concentration of 1.26 ppm (wet weight). One liver sample had a selenium value in the toxic range for pigs (3.3 ppm wet weight; reference range, 0.4–1.2 ppm). A 1-compartment pharmacokinetic model of selenium elimination in exposed pigs was generated, and the geometric mean blood selenium elimination half-life was estimated to be 12 days. The 60-day withdrawal time recommended by the Food Animal Residue Avoidance Database was considered sufficient to allow safe human consumption of tissues from exposed pigs.
Immunohistochemical Detection of Canine Distemper Virus in Haired Skin, Nasal Mucosa, and Footpad Epithelium: A Method for Antemortem Diagnosis of Infection
Authors: Haines Deborah M. , Martin Karen M. , Chelack Brian J. , Sargent Ronald A. , Outerbridge Catherine A. , Clark Edward G.
Published: May 2011

A reliable antemortem diagnostic method is needed for determining infection with canine distemper virus (CDV). The utility of immunohistochemical detection of CDV antigen was examined was examined for samples of nasal and footpad epithelium and haired skin in dogs with and without detectable CDV antigen in the lung and/or brain. Tissues from 57 dogs at risk of CDV infection were tested. Viral antigen was found in the lung and/or brain of 28 dogs. Among these dogs, viral antigen was demonstrated in the epithelial cells of the nasal mucosa in 24 of 27 dogs, in the footpad epithelium in 24 of 26 dogs, and in the haired skin of the dorsal neck in 26 of 27 dogs. Among the 29 dogs without CDV antigen in either the lung or brain, 1 dog had positive staining for viral antigen in the skin and nasal mucosa. Biopsies of haired skin of the dorsal neck, which is relatively simple to sample, can be used for antemortem immunohistochemical testing for acute and subacute infection with CDV.
Bovine Viral Diarrhea Virus Isolated from Fetal Calf Serum Enhances Pathogenicity of Attenuated Transmissible Gastroenteritis Virus in Neonatal Pigs
Authors: Woods R. D. , Kunkle R. A. , Ridpath J. F. , Bolin S. R.
Published: May 2011

A bovine viral diarrhea virus (BVDV-C) was isolated from swine tissue culture cells used to attenuate the transmissible gastroenteritis virus (TGEV) after 68 passes. Piglets given a pure culture of BVDV-C developed clinical signs similar to those of a mild TGEV infection and recovered by 10 days postexposure. Villous blunting and fusion was observed in the small intestine, and a lymphocyte depletion was observed in Peyer's patches in the ileum. Piglets given a combination of BVDV-C and attenuated TGEV developed clinical signs similar to those of a virulent TGEV infection and were euthanized. The combined infection induced a generalized lymphocyte depletion throughout the lymphatic system and villous atrophy in the intestinal tract. Piglets exposed to a another type I strain of BVDV (NY-1) either alone or in combination with the attenuated TGEV had mild clinical signs similar to those of a TGEV infection. Moderate villous atrophy in the ileum and a lymphocyte depletion in the mesenteric lymph node were observed in these piglets postmortem. The data indicate a potential problem for diagnostic laboratories in relation to a diagnosis of virulent TGEV infections and in the field for young piglets exposed to a BVDV-contaminated TGEV vaccine.
Application of a Quantitative Algorithm to Restriction Endonuclease Analysis of Aujeszky's Disease (Pseudorabies) Virus from a Geographically Localized Outbreak
Authors: Scherba Gail , Wiemers John F. , Siegel Arthur M. , Jin Ling , Austin Connie C. , Bowman Laurel , Redeford Bonnie , Johnston Nancy A. , Weigel Ronald M.
Published: May 2011

A retrospective epidemiologic study was conducted to evaluate the application of an objective quantitative algorithm for estimating genetic similarity from restriction endonuclease analysis data. The analysis was performed to assist the determination of chronologic trends in an Aujeszky's disease viral epidemic in a geographic region. DNA from each viral isolate obtained during the epidemic was digested with 4 restriction endonucleases and molar ratio labeled to generate separate fragment patterns that were simultaneously compared using the algorithm. The resultant estimates of genetic similarity were then used in conjunction with time of virus isolation and specific geographic location of the outbreaks to identify the probable sources of infection and the patterns of spread among swine herds. This type of quantitative analysis enabled a more precise and objective approach than previously has been applied to the interpretation of restriction endonuclease data, thereby demonstrating the benefit of this methodology for the investigation of infectious disease outbreaks.
Detection and Identification of Mycoplasma from Bovine Mastitis Infections Using a Nested Polymerase Chain Reaction
Authors: Baird Shane C. , Carman Jane , Dinsmore R. Page , Walker Richard L. , Collins James K.
Published: May 2011

A polymerase chain reaction (PCR) test was compared with culture for the detection and diagnosis of bovine Mycoplasma intramammary infection. The PCR test was applied to 24-hour Mycoplasma enrichment cultures of milk from cows with suspected mastitis and from bulk tank milk. In comparison to culture, the sensitivity and specificity of the PCR method were 96.2% and 99.1% for individual cow milk and 100% and 99.8% for the bulk tank milk, respectively. However, in discrepant cases where PCR was positive and culture was negative, the PCR test was correct; subsequent PCR tests and culturing of the individual cow's milk yielded positive results. The PCR test simultaneously detected and differentiated among 11 bovine Mycoplasma species.
Mycobacterium Paratuberculosis Detection in Bovine Feces is Improved by Coupling Agar Culture Enrichment to an IS900-Specific Polymerase Chain Reaction Assay
Authors: Secott T. E. , Ohme A. M. , Barton K. S. , Wu C. C. , Rommel F. A.
Published: May 2011

The feasibility of coupling an agar culture enrichment step with gene amplification (ACE-PCR) as a means to improve turnaround time and detect Mycobacterium paratuberculosis (Mpt) in the presence of contaminants was investigated. Fecal samples from 463 Pennsylvania dairy cows were cultured in duplicate sets. One replicate from each set was processed and interpreted according to standard culture (SC) protocol, whereas cultures from the second replicate were harvested at 6 weeks postinoculation; DNA extracts from the harvested material were evaluated by a polymerase chain reaction (PCR) test for the Mpt-specific IS 900 gene. One hundred seventy-six of 463 culture sets were positive by either method. One hundred sixty-five of these (94%) were ACE-PCR positive, and 151 (86%) were positive by SC. Eleven SC-positive samples were ACE-PCR negative, and 9 ACE-PCR-positive samples were negative by SC; these discrepancies could be a consequence of a low organism burden (≤5 organisms/g) or slow growth rate of Mpt in cultures of these samples. One hundred thirty-nine of 463 culture sets (30%) were reported as inconclusive because of culture contamination according to SC protocol; 16 of these (11.5%) were ACE-PCR positive. Seventy-four ACE-PCR-positive sets (42% of all positives) were negative or inconclusive by SC at 6 weeks postinoculation. Agar culture enrichment prior to IS 900 PCR testing significantly improves Mpt culture turnaround time and sensitivity.
Dose Response of Sheep Poisoned with Locoweed (Oxytropis Sericea)
Authors: Stegelmeier Bryan L. , James Lynn F. , Panter Kip E. , Gardner Dale R. , Pfister James A. , Ralphs Michael H. , Molyneux Russell J.
Published: May 2011

Locoweed poisoning occurs when livestock consume swainsonine-containing Astragalus and Oxytropis species over several weeks. Although the clinical and histologic changes of poisoning have been described, the dose or duration of swainsonine ingestion that results in significant or irreversible damage is not known. The purpose of this research was to document the swainsonine doses that produce clinical intoxication and histologic lesions. Twenty-one mixed-breed wethers were dosed by gavage with ground Oxytropis sericea to obtain swainsonine doses of 0.0, 0.05, 0.1, 0.2, 0.4, 0.8, and 1.0 mg/kg/day for 30 days. Sheep receiving ≥0.2 mg/kg gained less weight than controls. After 16 days, animals receiving ≥0.4 mg/kg were depressed, reluctant to move, and did not eat their feed rations. All treatment groups had serum biochemical changes, including depressed α-mannosidase, increased aspartate aminotransferase and alkaline phosphatase, as well as sporadic changes in lactate dehydrogenase, sodium, chloride, magnesium, albumin, and osmolarity. Typical locoweed-induced cellular vacuolation was seen in the following tissues and swainsonine doses: exocrine pancreas at ≥0.05 mg/kg; proximal convoluted renal and thyroid follicular epithelium at ≥0.1 mg/kg; Purkinje's cells, Kupffer's cells, splenic and lymph node macrophages, and transitional epithelium of the urinary bladder at ≥0.2 mg/kg; neurons of the basal ganglia, mesencephalon, and metencephalon at ≥0.4 mg/kg; and cerebellar neurons and glia at ≥0.8 mg/kg. Histologic lesions were generally found when tissue swainsonine concentrations were ≅150 ng/g. Both the clinical and histologic lesions, especially cerebellar lesions are suggestive of neurologic dysfunction even at low daily swainsonine doses of 0.2 mg/kg, suggesting that prolonged locoweed exposure, even at low doses, results in significant production losses as well as histologic and functional damage.
Isolation of a Bovine Adenovirus Serotype 10 from a Calf in the United States
Authors: Lehmkuhl H. D. , Cutlip R. C. , DeBey B. M.
Published: May 2011

Virus isolated from the lung, liver, kidney, and small intestine of a 3-month-old Holstein heifer with a clinical history of pneumonia and lesions in multiple organs was identified as an adenovirus on the basis of morphological and physicochemical characteristics. The adenovirus was determined to be a serotype 10 bovine adenovirus and represents the first reported isolation of this serotype in the United States. Inoculation of calves with this isolate resulted in mild to moderate clinical response consisting of fever, inappetence, increased respiratory rate, cough, and listlessness. Gross lesions were minimal in the respiratory tract and consisted of fibrin in the airways and small areas of consolidation in the cranial lobes of the lung. Mucofibrinous foci were present on the mucosa of the upper small intestine.
Experimental Model of Type II Bovine Viral Diarrhea Virus-Induced Thrombocytopenia in Neonatal Calves
Authors: Walz Paul H. , Bell Thomas G. , Steficek Barbara A. , Kaiser Lana , Maes Roger K. , Baker John C.
Published: May 2011

Thrombocytopenia has been associated with type II bovine viral diarrhea virus (BVDV) infection in immunocompetent cattle, but the mechanism is unknown. The purpose of the present study was to develop and characterize a model of type II BVDV-induced thrombocytopenia. Colostrum-deprived Holstein calves were obtained immediately after birth, given a BVDV-negative and BVDV antibody-negative plasma transfusion, housed in an isolation facility, and randomly assigned to either control ( n = 4) or infected ( n = 5) groups. Infected calves were inoculated by intranasal instillation on day 3 of age with 107 TCID50 of the prototype type II isolate, BVDV 890, whereas control calves were sham inoculated. Blood counts and virus isolations from serum, white blood cells, and platelets were performed daily until day 12 after infection, at which time all experimental calves were euthanatized, and pathologic, virologic, and immunohistochemical examinations were performed. On physical examination, the control calves remained normal, but the infected calves developed pyrexia and diarrhea characteristic of type II BVDV infection. The platelet count decreased in all infected calves, and a statistically significant difference in the platelet count between control and infected calves was observed on days 7–12 after infection. In addition, the mean platelet volume and white blood cell counts also decreased. Examination of the bone marrow from the infected calves revealed immunohistochemical staining for BVDV antigen in megakaryocytes and evidence of concurrent megakaryocyte necrosis and hyperplasia.
Evaluation of Excretory-Secretory Products and Somatic Worm Antigens for the Serodiagnosis of Experimental Parelaphostrongylus Tenuis Infection in White-Tailed Deer
Authors: Ogunremi Oladele , Lankester Murray , Loran Steven , Gajadhar Alvin
Published: May 2011

Three different antigen preparations of Parelaphostrongylus tenuis were assessed for their effectiveness in an indirect enzyme-linked immunosorbent assay (ELISA) to diagnose experimental infection of white-tailed deer (WTD). The antigen preparations were the excretory-secretory products of third-stage larvae (ES-L3), somatic antigens of third-stage larvae (sL3), and somatic antigens of the adult stage (sA) of P. tenuis. The relative sensitivities of the antigen preparations in indirect ELISA were ES-L3 > sL3 > sA. Immunoglobulin G (IgG) antibodies to ES-L3 and sL3 could be detected 14 days postinfection and were consistently present in all infected animals from the first month to the end of the experiment at 5 months. In contrast, IgG antibodies to sA could not be detected at any time in 2 infected WTD. ES-L3 and sL3 proved reliable in the early detection of anti- P. tenuis antibodies and in the serological monitoring of experimentally infected animals. Significant cross-reactivity between all P. tenuis antigen preparations and sera from animals infected with parasites other than P. tenuis may preclude their use for field diagnosis. Nevertheless, isolation of unique P. tenuis antigen(s) should lead to the development of a specific serological test for infected white-tailed deer.
Normal and Toxic Zinc Concentrations in Serum/Plasma and Liver of Psittacines with Respect to Genus Differences
Authors: Puschner Birgit , St. Judy Leger , Galey Francis D.
Published: May 2011

Determination of zinc concentration in serum/plasma and tissue of caged and aviary birds is commonly requested by practitioners because of an increased awareness of zinc toxicity. However, interpretation of zinc levels is often based on normal zinc concentrations established for poultry. Also, it is likely that intergenus differences exist in normal zinc concentrations of pet birds. In an attempt to determine normal and toxic concentration ranges, zinc concentrations in liver ( n = 276) and serum/plasma ( n = 260) collected from psittacines between 1990 and 1998 were analyzed. Zinc concentrations were determined by inductively coupled argon plasma emission spectroscopy analysis. The results were categorized by genus and, when available, by history. Birds that were diagnosed with zinc toxicosis (on the basis of history, clinical examination, pathology, and laboratory findings) were exempt and not included in establishing normal ranges. The results indicate that important differences occur with respect to genera. For example, cockatoos and Eclectus parrots have higher normal zinc concentrations in serum or plasma than other psittacines. In addition, analysis of all the submitted cases suggests that potentially toxic zinc concentrations in livers of psittacines can be well below the range considered toxic in chickens (>200 ppm).
Evidence Retention Policies in U.S. Law Enforcement Agencies: Implications for Unsolved Cases and Postconviction DNA Testing
Authors: Strom Kevin J. , Hickman Matthew J. , Ropero-Miller Jeri D.
Published: May 2011

The use of forensic evidence in the criminal justice system has grown appreciably in the United States. Yet policies that dictate how state and local agencies maintain and store forensic evidence have not kept pace. This study examined the prevalence of evidence retention policies, as well as storage locations and tracking systems, in a nationally representative sample of state and local law enforcement agencies. Less than half of U.S. police departments have a policy for preserving biological evidence from convicted offenders. Among agencies having a policy, the responsibility for retaining evidence was most commonly placed with the investigating law enforcement agency. Implications of these findings and policy directions are discussed.
Advanced Scientific Methods and Procedures in the Forensic Investigation of Clandestine Graves
Authors: Larson Daniel O. , Vass Arpad A. , Wise Marc
Published: May 2011

Our goal is to discuss the new technologies and procedures that we have developed for the discovery and recovery of buried victims. We argue that forensic investigations of clandestine graves must be grounded in the most advanced scientific methods and evidence-collection techniques available. A structured program that includes an interdisciplinary team of forensic scientists and law enforcement experts is proposed to facilitate all aspects of the investigative and legal process. Such issues are of great relevance because most legal jurisdictions have a number of cases each year and present operating procedures are not standardized. There is a clear need for national dialog to improve our investigative efforts and insure best practices in forensic science across legal jurisdictions and law enforcement agencies.