bluuebunny/crossref_metadata_2025 huggingface dataset:
Postural and Neurological Deficits in Broiler Chicks after Cervical Vaccination with Live Vaccine
Authors: Gustafson C. R. , Cooper G. L. , Charlton B. R. , Bickford A. A. , Nordhausen R.
Published: May 2011

A disease characterized by paresis and paralysis was seen in 7–9-day-old broiler chicks after vaccination in the neck area at day-of-age with a live virus vaccine containing viruses of Marek's disease, fowl pox, and infectious bursal disease. Affected birds presented with variable signs of ataxia, lateral recumbency, leg paralysis, and twisting or S-shaped flexure of the neck. Gross lesions noted at necropsy included swelling and edema of the subcutaneous tissues and muscles of the neck at the injection site area. A heavy mononuclear inflammatory cell infiltration was seen in the subcutaneous tissues, connective tissues, and muscles of the neck at the injection site. In some cases, the inflammatory process extended along fascial planes to involve the epidural spaces surrounding the spinal cord. Fatty changes with possible demyelination of nerve fibers were noted in some sections of the spinal cord adjacent to the inflammatory lesions. Clusters of poxviruses were found within some inflammatory lesions on transmission electron photomicrographs.
Porcine Circovirus 2–Associated Disease in Eurasian Wild Boar
Authors: Ellis John , Spinato Maria , Yong Choon , West Keith , McNeilly Francis , Meehan Brian , Kennedy Seamus , Clark Edward , Krakowka Steven , Allan Gordon
Published: May 2011

Porcine circovirus 2 (PCV2) was first identified in high-health herds of domestic swine and was associated with a debilitating disease called postweaning multisystemic wasting syndrome (PMWS). Most subsequent studies have indicated that PCV2 infects only swine but there is little information on porcids other than improved breeds of domestic swine. Multisystemic disease was reported in a group of Eurasian wild boars raised under free-range conditions. Affected young pigs had pneumonia and enteritis and were cachectic. Porcine circovirus 2 was identified in affected tissue by immunohistochemistry and in situ hybridization, and a PCV2-like virus was isolated from pooled organs. The open reading frame (ORF2) of the isolated PCV2 had a 98.7% homology with the ORF2 of a reference PCV2 isolate. These diagnostic data indicate that PCV2 can infect and cause disease in Sus scrofa subspecies other than domestic swine.
Evaluation of the Brucella Abortus Species–Specific Polymerase Chain Reaction Assay, an Improved Version of the Brucella AMOS Polymerase Chain Reaction Assay for Cattle
Authors: Bricker Betsy J. , Ewalt Darla R. , Olsen Steven C. , Jensen Allen E.
Published: May 2011

In a blind test, 344 samples representing 80 bacterial isolates were analyzed by the Brucella abortus species–specific polymerase chain reaction (BaSS PCR) assay for the identification and discrimination of B. abortus field strains (wild-type biovars 1, 2, and 4) from 1) B. abortus vaccine strains, 2) other Brucella species, and 3) non- Brucella bacteria. Identical samples were tested in 2 laboratories. Half the samples were fully viable, and half were bacteria that had been killed by methanol fixation. The results in 1 laboratory correctly identified 100% of the samples, resulting in a predictive value of 100% for all categories and 100% sensitivity and specificity under the prescribed conditions. The second laboratory misidentified 31 samples, resulting in a range of 66.7–100% sensitivity, 93.2–99.7% specificity, and 77.3–98.2% predictive values depending on the category. There was no significant difference in viable versus fixed bacteria for either laboratory. Subsequent review of the protocol indicated that contamination was the likely cause of 26 of the 31 erroneous identifications. The results show that the BaSS PCR assay has the potential to be a very reliable screening tool for B. abortus identification. However, the data also provide a cautionary reminder of the importance of preventing contamination in diagnostic PCR.
Comparison of Phenotypic Traits and Genetic Relatedness of Salmonella Enterica Subspecies Arizonae Isolates from a Colony of Ridgenose Rattlesnakes with Osteomyelitis
Authors: Bemis David A. , Owston Michael A. , Lickey Adrienne L. A. , Kania Stephen A. , Ebner Paul , Rohrbach Barton W. , Ramsay Edward C.
Published: May 2011

Reptiles are well-known sources of human Salmonella infections; however, little is known about the ability of Salmonella to cause disease in reptiles. Thirty-seven isolates of Salmonella enterica subspecies arizonae ( S. arizonae) were obtained from retrospective and prospective studies of a closed colony of ridgenose rattlesnakes ( Crotalus willardi) with osteomyelitis. All isolates ( N = 7) from bone lesions were of a single serotype, 56:z4,z23, and this serotype was found on only 1 occasion among 8 other serotypes isolated from 21 cloacal and intestinal samples. The remainder ( N = 7) of serotype 56:z4,z23 isolates were from other extraintestinal sites, including liver, ovary, blood, and testis. S. arizonae isolates were susceptible to most antimicrobials, and plasmid profiles did not correlate with serotype or antimicrobial resistance. Isolates of the 56:z4,z23 serotype ( N = 14) formed a tight cluster with 95% similarity by XbaI macrorestriction analysis. Individual isolates of serotypes, 56:z4,z23, 38:(k)-z35, and 48:i-z invaded HeLa cells but an isolate of serotype 50:r-z did not. The same individual isolates of serotype 56:z4,z23 and 48:i-z also invaded viper heart cells. The Salmonella InvA gene was detected by polymerase chain reaction (PCR) in all S. arizonae serotypes tested, including 5 serotype 56:z4,z23 isolates and individual isolates of serotypes 48:i-z and 50:r-z. A source or possible explanation for increased virulence of S. arizonae serotype 56:z4,z23 in this unique host has not been found.
Clinical Coccidiosis in a Boar Stud
Authors: Yaeger Michael J. , Holtcamp Andrew , Jarvinen Julie A.
Published: May 2011

This report describes an outbreak of coccidiosis in a boar stud. A live, untreated, adult boar with a history of diarrhea was submitted to the Iowa State University Veterinary Diagnostic Laboratory, Ames, IA. For a 3-month period, approximately 40% of the boars in this stud had developed gray to brown diarrhea that lasted 1–3 days. Affected boars did not lose condition, and antibiotic therapy did not appear to affect the clinical course of the disease. At necropsy, the distal ileum was palpably thickened and covered by a thick, yellow-green, fibrinous exudate. Microscopic changes in the ileum consisted of an erosive enteritis associated with the presence of numerous coccidia within mid to superficial villus enterocytes. The mucosa was covered by a fibrinous exudate admixed with numerous nonsporulated coccidian oocysts. A light growth of Salmonella enterica serovar Derby was isolated from the small intestine of this animal, but laboratory tests were negative for Lawsonia and Brachyspira spp. Individual or paired fecal samples were obtained from 6 additional boars experiencing similar clinical signs. Numerous Eimeria spinosa oocysts were identified in these samples. Neither Salmonella nor Brachyspira spp. were cultured from submitted fecal samples. Necropsy of a live boar and examination of feces from 6 additional animals confirmed that the mild, sporadic, transient diarrhea in this boar stud was due to coccidiosis.
Identification of Trichomonadid Protozoa from the Bovine Preputial Cavity by Polymerase Chain Reaction and Restriction Fragment Length Polymorphism Typing
Authors: Hayes Dawn C. , Anderson Rebecca R. , Walker Richard L.
Published: May 2011

Accurate identification of the bovine pathogen Tritrichomonas foetus is sometimes complicated by the presence of other trichomonadid protozoa in clinical samples. A highly specific and reproducible approach for differentiating 3 common types of bovine trichomonadid protozoa found in the bovine preputial cavity, T. foetus, Pentatrichomonas hominis, and a Tetratrichomonas species, was developed using polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) analysis. Universal trichomonadid protozoa primers, TFR1 and TFR2, were used to amplify the 5.8S rRNA gene and internal transcribed spacer regions (ITSRs), and the products were digested with the restriction enzyme HpyCH4IV. Restriction fragment length polymorphism analysis was performed on 55 trichomonad isolates from bovine preputial washing and scraping samples. The RFLP results correlated 100% with 5.8S rRNA gene and ITSR sequence results and PCR results with primers specific for T. foetus. The results of this study demonstrate that PCR and RFLP analysis can be used in lieu of DNA sequencing to identify the specific trichomonadid protozoa isolated from the bovine preputial cavity.
Severe Persistent Orf in Young Goats
Authors: de la Concha-Bermejillo A. , Guo J. , Zhang Z. , Waldron D.
Published: May 2011

Orf (contagious ecthyma) is a viral disease of small and wild ruminants, humans, and less frequently other species. In sheep and goats, the disease is characterized by the formation of vesiculo-proliferative lesions in the skin of lips and nostril. Here, a form of generalized orf in 16 goat kids from 2 different locations in west Texas is described. The disease was characterized by multifocal, severe, proliferative dermatitis that persisted from about 2 months of age until the goat kids were euthanized 3 months later. All affected goats were Boer or Boer crosses under 1 year of age. The mean immunoglobulin concentration in sera of affected goats was elevated compared with healthy control goats. Severe to moderate lymphadenomegaly of the nodes draining the areas of the skin affected with orf lesions was present in all 16 goat kids. Suppurative arthritis, chronic fibrinous pneumonia, and premature thymic involution were found in 3, 5, and 7 of the goat kids, respectively. The skin lesions of 3 goat kids were infested with larvae of the opportunistic black garbage fly ( Ophira sp.). The orf virus was identified in skin lesions by isolation in Marbin—Darby ovine kidney cells, electron microscopy, and amplification of viral DNA by polymerase chain reaction. The orf virus was not detected in peripheral blood or lymph node mononuclear cells of any of the goats. Cross-neutralization experiments showed that an ovine orf virus antiserum raised in sheep was more effective in neutralizing a sheep orf virus isolate than a caprine orf virus isolate. The clinical and epidemiological characteristics of these orf cases may be the result of susceptibility factors within some individuals of the Boer breed of goats.
Ascaridoid Nematodes and Associated Lesions in Stomachs of Subadult Male Northern Fur Seals (Callorhinus Ursinus) on St. Paul Island, Alaska: (1987—1999)
Authors: Spraker T. R. , Lyons E. T. , Tolliver S. C. , Bair H. D.
Published: May 2011

Stomachs from subadult male northern fur seals (SAMs) ( Callorhinus ursinus) (n = 5,950) from St. Paul Island, Alaska, were examined from 1987 to 1999 for lesions and parasites. On gross examination of these stomachs, parasite nodules were evident in 92% of the stomachs and active and healing ulcers were found in 14% and 10%, respectively. Prevalence of nematodes from 1987 to 1997 was 88% for 5,700 SAMs for which numbers of parasites were estimated but not identified to the genus level. All nematodes recovered from 250 SAMs examined in 1998 ( n = 124; 99% infected) and in 1999 ( n = 126; 91% infected) were identified and counted. Prevalences in 1998 and 1999 were 5% and 0% for Anisakis spp., 52% and 18% for Contracaecum spp., and 96% and 89% for Pseudoterranova spp., respectively.
Preparation and Characterization of Polyclonal and Monoclonal Antibodies against Lawsonia Intracellularis
Authors: Guedes Roberto M. C. , Gebhart Connie J.
Published: May 2011

Proliferative enteropathy is an intestinal infectious disease caused by the obligate intracellular bacterium Lawsonia intracellularis. Immunohistochemistry staining has superior sensitivity over hematoxylin and eosin and silver staining for detecting L. intracellularis in histological sections. A L. intracellularis—specific monoclonal antibody (MAb) produced in the UK (IG4 MAb) has been described in the literature. However, no monoclonal or polyclonal antibodies are commercially available. Therefore, the objective of this study was to produce and characterize new polyclonal and monoclonal antibodies against L. intracellularis that are suitable for diagnostic use. The new monoclonal (2001 MAb) and polyclonal antibodies (1999 PAb) were compared with the IG4 MAb using Western blot analysis of outer membrane proteins (OMPs) of 6 L. intracellularis isolates, Bilophila wadsworthia and Brachyspira hyodysenteriae and using immunohistochemistry of known positive and negative histologic samples and pure cultures of L. intracellularis, B. wadsworthia, B. hyodysenteriae, Salmonella choleraesuis, S. typhimurium, and Escherichia coli K88. Immunogold staining using 2001 MAb was performed to show the specificity of the antibody against an L. intracellularis surface protein. Western blot analysis showed that the 2001 MAb targeted an OMP of 77 kD, which made it different from the IG4 MAb that targeted an 18-kD OMP. The immunogold stain demonstrated the specificity of the 2001 MAb to a surface protein of L. intracellularis. The polyclonal antibody (1999 PAb) targeted 5 OMPs (77, 69, 54, 42, and 36 kD). Both the 2001 MAb and 1999 PAb stained known positive, but not negative, histologic samples. Both the 2001 MAb and 1999 PAb reacted with a pure culture of L. intracellularis but not with any other common enteric pathogens. These two new antibodies will be useful for immunodiagnosis of L. intracellularis.
Neuronal Ceroid-Lipofuscinosis in a Labrador Retriever
Authors: Rossmeisl John H. , Duncan Robert , Fox Jonathan , Herring Erin S. , Inzana Karen D.
Published: May 2011

An 8-year-old Labrador Retriever with an 11-month history of progressive partial seizures and necropsy examination findings characteristic of the lamellar form of canine neuronal ceroid-lipofuscinosis (NCL) is presented. The clinical, light microscopic, and ultrastructural features of this case most closely resemble human adult-onset NCL (Kufs disease). This is the first report of NCL occurring in the Labrador Retriever breed.
Rhodococcus Equi and Genetic Susceptibility: Assessing Transferrin Genotypes from Paraffin-Embedded Tissues
Authors: Mousel Michelle R. , Harrison Lenn , Donahue J. Michael , Bailey Ernest
Published: May 2011

Rhodococcus equi is a bacterial pathogen, ubiquitous in the soil, that infects many foals and is lethal to some. Transferrin is an iron-binding protein that has bacteriostatic properties in the blood. Transferrin is also highly polymorphic in most species, with 15 variants identified for horses using biochemical methods, and may be responsible for variation in susceptibility to bacterial pathogens. The objectives of this study were 1) to compare biochemical typing with DNA typing of transferrin, 2) to determine if transferrin DNA from archival paraffin-embedded tissue samples can be recovered and typed with molecular methods, and 3) to determine if there was an association between foal death caused by R. equi and transferrin type. Comparing biochemical methods and DNA sequencing for 41 horses demonstrated correspondence between the typing methods. The allele frequency of archival paraffin-embedded tissue samples from 34 Thoroughbred foals that succumbed to R. equi showed an excess of the F allele and a deficiency of the D allele ( P < 0.05). Year of collection ( P > 0.8), age of foal ( P > 0.3), and sex of foal ( P > 0.6) were not statistically associated with transferrin type. The archival material was successfully transferrin typed using DNA sequencing, and there may be an association between foal death caused by R. equi and transferrin type.
Cerebral Cysticercosis by Taenia Crassiceps in a Domestic Cat
Authors: Wünschmann Arno , Garlie Virginia , Averbeck Gary , Kurtz Harold , Hoberg Eric P.
Published: May 2011

Cerebral cysticercosis by Taenia crassiceps was diagnosed in an adult female domestic shorthair cat. The animal was euthanized 6 weeks after the initial presentation with signs of vomiting, lethargy, and ataxia. The disease took an intermittent relapsing course with the neurological signs progressing eventually to recumbancy and coma. At necropsy, numerous cysticerci were found in the dilated left lateral ventricle and the adjacent brain parenchyma. The cysticerci were identified as metacestodes of T. crassiceps larvae based on size and morphology of the cysts; shape, number, and size of the rostellar hooks; and mode of proliferation, including endogenous and exogenous budding. Cerebral cysticercosis by T. crassiceps is rare in atypical intermediate hosts and has not been described in cats.
Conceptual Foundations for Infectious Disease Surveillance
Authors: Thurmond Mark C.
Published: May 2011

The purpose of this report is to offer concepts for consideration in developing infectious disease surveillance systems, defined here as active, formal, and systematic processes intentionally directed to rapidly seek out and identify infectious disease agents or disease. Performance of surveillance systems can be judged by their accuracy (sensitivity and specificity), precision (repeatability), timeliness, multiple utility, and value. Surveillance system operation and function necessary to achieve high performance are defined in part by characteristics of the specific infectious disease, including disease transition state dynamics, that define probabilities of being in the latent, infectious, or clinical phase of disease. Two key components of surveillance are the sampling scheme, which is intended to maximize the probability of capturing an infected animal or specimen as soon as possible after the herd has been exposed, and the diagnostic assays, which should maximize the probability of detecting the agent, or evidence of the agent, if it is present in the specimen, while minimizing the likelihood of a false-positive result. Proportional risk sampling, targeted sampling, and repeated sampling are strategies that can improve overall surveillance system accuracy and particularly the temporal sensitivity related to early detection. Hierarchical sampling schemes and multiplexed assays can maximize efficiency and improve utility by serving multiple surveillance systems and purposes. Development of the surveillance systems needed to address emerging and foreign animal diseases will necessarily require design and architecture that are highly probability-driven to maximize surveillance sensitivity and specificity and to minimize cost.
Molecular Detection of Anaplasma Platys in Dogs Using Polymerase Chain Reaction and Reverse Line Blot Hybridization
Authors: Sparagano Olivier A. E. , de Vos Alexander P. , Paoletti Barbara , Cammà Cesare , de Santis Paola , Otranto Domenico , Giangaspero Annunziata
Published: May 2011

Several polymerase chain reactions (PCRs) and a reverse line blot hybridization (RLB) method were used to identify Anaplasma platys in dogs held in a kennel in Italy. Whereas PCR techniques confirmed the presence of A. platys, the RLB method not only correlated the results obtained by PCR but also ruled out the presence of other species such as Ehrlichia canis or E. chaffeensis. There was no correlation between infection status and age or breed of the dogs. Polymerase chain reaction performed on the Rhipicephalus sanguineus ticks collected from those dogs showed that they were also infected with A. platys. Sequences obtained from some samples and compared with those within the GenBank also confirmed the presence of A. platys.
Short-Term Repeatability of Measurements of Antimicrobial Susceptibility of Escherichia Coli Isolated from Feces of Feedlot Cattle
Authors: Wagner Bruce , Morley Paul S. , Dargatz David A. , Wittum Thomas E. , Keefe Thomas J. , Salman Mo D.
Published: May 2011

Short-term stability of measurements of antimicrobial susceptibility of Escherichia coli isolated from feces of feedlot cattle is important in developing monitoring and surveillance programs. Frequent evaluations (i.e., daily) are resource intensive and in some situations may be impractical for long-term sampling protocols. Consequently, a point-in-time measurement will need to be used to represent conditions in the perisampling period. In this study, 30 fecal samples were collected from each of 6 cattle pens on a commercial cattle feedlot on 2 occasions separated by 48 hours. Escherichia coli was isolated from single and pooled samples. The isolates were tested for antimicrobial susceptibility against a panel of 17 antimicrobials. Resistance to 5 antimicrobials (ampicillin, nalidixic acid, streptomycin, sulfamethoxazole, and tetracycline) was detected in single and pooled samples from both sampling periods (days 1 and 3). The prevalence of isolates resistant to these 5 antimicrobials was 2% or higher in all treatment combinations except for pools obtained from day 3 samples. Lower levels of resistance to 6 more antimicrobials were detected inconsistently across the single and pooled samples. Logistic models constructed for the antimicrobials to which the E. coli isolates were most commonly resistant demonstrated that there were no significant differences between periods ( P > 0.10) and between single and pooled samples ( P > 0.20). The distribution of the number of antimicrobials to which isolates were resistant was consistent for the single samples across periods, but there appeared to be a lower prevalence of any resistance in day 1 pooled samples. A larger number of resistant phenotypes were detected in the single samples than in the pooled samples, and resistant phenotypes with prevalence of less than 2% were detected inconsistently across periods and single and pooled samples. Resistance to individual antimicrobials was consistent by all measures when the prevalence was at least 2%. Inconsistent results were obtained for antimicrobials to which resistance rarely occurred. The apparent inconsistencies do not appear to be related to external factors but rather to sampling intensity. Short-term stability is a plausible assumption under sampling strategies that are designed to detect specific levels of prevalence. However, when resistance levels fall below these levels, there will likely be fluctuations in the presence or absence of rare resistant phenotypes and in their prevalence and central tendency measures.
Reproduction of Postweaning Multisystemic Wasting Syndrome in Pigs Experimentally Inoculated with a Swedish Porcine Circovirus 2 Isolate
Authors: Allan Gordon , McNeilly Francis , Meehan Brian , McNair Irene , Ellis John , Krakowka Steve , Fossum Caroline , Wattrang Eva , Wallgren Per , Adair Brian
Published: May 2011

In recent years, porcine circovirus type 2 (PCV2)—associated postweaning multisystemic wasting syndrome (PMWS) has been reported worldwide. However, to date, PMWS has not been reported in Sweden despite the demonstration of serum antibodies to a PCV2-like virus in Swedish pigs. This communication reports the experimental reproduction of clinical PMWS after inoculation of colostrum-deprived (CD) pigs, derived from a Northern Ireland herd, with an isolate of PCV2 virus recovered from a clinically normal Swedish pig that was necropsied in 1993. The clinical disease and histological lesions observed in CD pigs inoculated with this virus were indistinguishable from those observed in previous studies on CD pigs inoculated with a PCV2 virus isolate recovered from pigs with PMWS. These results highlight the disease potential of PCV2 isolated from regions apparently free of PMWS and suggest that the status of the host and its environment is an important factor in the development of clinical PMWS.
Results of a New Serologic Test Suggest an Association of Waddlia Chondrophila with Bovine Abortion
Authors: Dilbeck-Robertson Pam , McAllister Milton M. , Bradway Dan , Evermann James F.
Published: May 2011

Waddlia chondrophila is a little-known intracellular organism belonging to the order Chlamydiales that has twice been isolated from aborted bovine fetuses. To initiate an investigation of the possibility that W. chondrophila may be an abortifacient pathogen, a serologic test was developed and used to screen bovine sera that were submitted to the Washington Animal Disease Diagnostic Laboratory (Pullman, WA). A highly significant statistical association ( P < 0.00001) was observed when comparing antibody titers in cows that had aborted with those in other classes of cattle. Although this result is consistent with the possibility that infection with W. chondrophila may be a cause of bovine abortion, it is also possible that seroprevalence simply increases with age or that exposure rates differ between cows and other classes of cattle. Future serologic studies should specifically compare antibody titers between aborting cows and carefully matched nonaborting cohorts.
Molecular and Antigenic Characterization of Bordetella Bronchiseptica Isolated from a Wild Southern Sea Otter (Enhydra Lutris Nereis) with Severe Suppurative Bronchopneumonia
Authors: Staveley Clare M. , Register Karen B. , Miller Melissa A. , Brockmeier Susan L. , Jessup David A. , Jang Spencer
Published: May 2011

Bordetella bronchiseptica was isolated in pure culture from the lung, abdomen, and intestine of a wild free-ranging southern sea otter ( Enhydra lutris nereis) with severe, suppurative bronchopneumonia. Immunohistochemistry, using antiserum raised to B. bronchiseptica, revealed strong positive staining of bacteria attached to bronchial ciliated epithelia as well as scattered positive staining in affected alveoli. Western blot analysis demonstrated that virulence factors, filamentous hemagglutinin, pertactin, and adenylate cyclase toxin are produced by the sea otter B. bronchiseptica isolate. Ribotype analysis using Pvu II restriction digests indicated that this isolate is most similar to strains commonly obtained in domestic dogs and cats.
In Vitro Evaluation of the Susceptibility of Edwardsiella Ictaluri, Etiological Agent of Enteric Septicemia in Channel Catfish, Ictalurus Punctatus (Rafinesque), to Florfenicol
Authors: McGinnis Anissa , Gaunt P. , Santucci T. , Simmons R. , Endris R.
Published: May 2011

In vitro studies were conducted to assess the sensitivity of Edwardsiella ictaluri, the etiological agent of enteric septicemia of catfish (ESC), to the antibacterial drug florfenicol (FFC). Twelve different E. ictaluri isolates from cases submitted between 1994 and 1997 to the Thad Cochran National Warmwater Aqua-culture Center fish diagnostic laboratory (Stoneville, MS) were used for testing. These isolates originated from channel catfish ( Ictalurus punctatus) infected with E. ictaluri through natural outbreaks of ESC in the commercial catfish ponds in Mississippi. Seven hundred sixty-seven additional cultures of E. ictaluri were obtained from channel catfish infected experimentally with E. ictaluri. In some of these experimental infections, FFC was used for treatment. These cultures of E. ictaluri were identified by morphological and biochemical tests. Kirby-Bauer zones of inhibition (in mm) for FFC against E. ictaluri were determined using standard methods. The minimum inhibitory concentration (MIC) of FFC was determined for the natural outbreak E. ictaluri isolates and arbitrarily selected experimental cultures. The zones of inhibition for FFC tested with E. ictaluri ranged from 31 to 51 mm. The MIC for FFC tested with E. ictaluri was consistently 0.25 μg/ml. Edwardsiella ictaluri tested in these studies were highly sensitive to FFC in vitro.
A Reverse Transcriptase—Polymerase Chain Reaction Assay for the Diagnosis of Turkey Coronavirus Infection
Authors: Velayudhan Binu T. , Shin Hyun-Jin , Lopes Vanessa C. , Hooper Tom , Halvorson David A. , Nagaraja Kakambi V.
Published: May 2011

This study reports on the development of a reverse transcriptase—polymerase chain reaction (RT-PCR) for the specific detection of turkey coronavirus (TCoV). Of the several sets of primers tested, 1 set of primers derived from the P gene and 2 sets derived from the N gene of TCoV could amplify the TCoV genome in the infected samples. The RT-PCR was sensitive and specific for TCoV and did not amplify other avian RNA and DNA viruses tested except the infectious bronchitis virus (IBV). To overcome the problem of IBV amplification, a set of separate primers was designed from the spike protein gene of IBV. The RT-PCR under the same conditions as above could effectively differentiate between TCoV and IBV. The closely related bovine coronavirus and transmissible gastroenteritis virus of pigs were differentiated from TCoV using the same RT-PCR with slight modifications. The results of RT-PCR correlated well with the results of the immunofluorescent test for the same samples tested at the Purdue University Animal Disease Laboratory, West Lafayette, Indiana. The nucleotide sequence and projected amino acid sequence comparison of the P gene of different isolates of TCoV from 5 different states in the United States revealed a close association among the different isolates of TCoV.
Seroprevalence of Antibodies to Sarcocystis Neurona in Equids Residing in Oklahoma
Authors: Bentz Bradford G. , Ealey Katie A. , Morrow Jennifer , Claypool P. L. , Saliki Jeremiah T.
Published: May 2011

A sampling of equids from the state of Oklahoma produced an estimate of seroprevalence of antibody to Sarcocystis neurona to be about 89.2%. This figure represents the highest currently reported regional seroprevalence of antibody to this organism. Regional differences in seroprevalence were found in the western quadrants of the state relative to the eastern quadrants of the state, with a significantly higher seroprevalence in the eastern regions. Thoroughbreds were found to exhibit a statistically significant lower seroprevalence as a breed group when compared with other breeds sampled.
Application and Evaluation of RT-PCR—ELISA for the Nucleoprotein and RT-PCR for Detection of Low-Pathogenic H5 and H7 Subtypes of Avian Influenza Virus
Authors: Dybkær Karen , Munch Mette , Handberg Kurt J. , Jørgensen Poul H.
Published: May 2011

Three 1-tube Reverse Transcriptase Polymerase Chain Reactions (RT-PCR) directed against the genes encoding the nucleoprotein (NP) and the H5 and H7 hemagglutinin (HA) gene, respectively, were used for detection of avian influenza virus (AIV) in various specimens. A total of 1,040 samples originating from chickens experimentally infected with 2 different low pathogenic avian influenza viruses, from domestic ducks and from wild aquatic birds were examined. The outcome of 1) the universal AIV RT-PCR including a PCR-enzyme-linked immunosorbent assay (ELISA) procedure directed against NP (NP RT-PCR-ELISA) and 2) the subtype specific RT-PCR for H5 and H7 were compared to the results obtained by inoculation of the same specimens into the allantoic cavity of embryonated specific pathogen free (SPF) hen's eggs. Using inoculation in SPF fowl eggs as standard the sensitivity of the NP RT-PCR-ELISA and the RT-PCR for H5 or H7 was 91% and 94%, and the corresponding specificity 98% and 96%. In comparison with inoculation into eggs an additional of 9 samples were positive by NP RT-PCR-ELISA and 13 samples were positive by RT-PCR for one of the HA subtypes. Hence, the 3 RT-PCR procedures described are fast, sensitive and specific for detecting AIV and subtyping H5 and H7 and they are obvious alternatives when testing large numbers of samples.
Escherichia coli O157:H7 in the Gallbladders of Experimentally Infected Calves
Authors: Stoffregen William C. , Pohlenz Joachim F. L. , Dean-Nystrom Evelyn A.
Published: May 2011

Fifteen weaned calves (age 89–141 days) were treated with dexamethasone (0.25 mg/kg, IV) for 3 days before, the day of, and the day after inoculation with 10 colony-forming units of either Escherichia coli O157:H7 (strain 86–24, which produces Shiga toxin 2 and intimin; n = 13) or nonpathogenic E. coli (strain 123, which does not produce Shiga toxin or intimin; n = 2). All calves were necropsied 4 days after inoculation. Histologic lesions of attaching and effacing bacteria were observed in the large intestine (12/13) and in the gallbladder mucosa (5/13) of calves inoculated with E. coli 86–24. Cholecystitis was present in 12 of 13 calves that received E. coli 86–24. Inoculum bacteria were recovered from the distal colons or feces (13/13) and gallbladders (3/4) of calves inoculated with 86–24.
Localized Cutaneous Infection with Francisella Tularensis Resembling Ulceroglandular Tularemia in a Cat
Authors: Valentine Beth A. , DeBey Brad M. , Sonn Robert J. , Stauffer Larry R. , Pielstick Leon G.
Published: May 2011

A chronically draining subcutaneous mass was removed from the ventral cervical region of a 6-year-old spayed female Domestic Shorthair cat. The histopathologic diagnosis was severe locally extensive pyogranulomatous and necrotizing cellulitis. Bacterial culture yielded Francisella tularensis subsp. tularensis as the causative agent. Immunohistochemical evaluation of sections for F. tularensis was negative. One year later, the cat was euthanized because of progressive lethargy found to be due to hypertrophic cardiomyopathy with pulmonary thromboembolism. No evidence of cutaneous or systemic infection by F. tularensis was found at necropsy. This case appears to be a localized form of tularemia resembling the ulceroglandular form of tularemia in humans and suggests that bacterial culture may be more sensitive than immunohistochemistry in detecting organisms in cases of localized F. tularensis infection.
Detection of c-kit Mutations in Canine Mast Cell Tumors using Fluorescent Polyacrylamide Gel Electrophoresis
Authors: Cameron L. R. Jones , Grahn Robert A. , Chien May B. , Lyons Leslie A. , London Cheryl A.
Published: May 2011

Mutations consisting of internal tandem duplications (ITDs) in exons 11 and 12 of the proto-oncogene c- kit are found in 30–50% of malignant canine mast cell tumors (MCTs). Traditionally, identification of such mutations in tumor specimens has been undertaken using standard polymerase chain reaction (PCR) and agarose gel electrophoresis. This procedure is limited to the detection of insertions and deletions larger than 9 base pairs in size. The purpose of this study was to compare the efficiency and accuracy of standard agarose gel electrophoresis with fluorescent polyacrylamide gel electrophoresis (PAGE) for the detection of ITDs in canine MCTs. The results of this study demonstrate that PAGE of labeled PCR products accurately predicts the size of the ITD in each tumor. In addition, other small insertions and deletions were not identified, suggesting that if they occur in canine MCTs, they do so infrequently. Because fluorescent and polyacrylamide formats are automated and have better resolution than agarose gels, fluorescent PAGE provides a more accurate, economical, and higher throughput method for the detection of c- kit mutations in canine MCTs.
Comparison of Reverse Transcriptase–Polymerase Chain Reaction, Virus Isolation, and Immunoperoxidase Assays for Detecting Pigs Infected with Low, Moderate, and High Virulent Strains of Classical Swine Fever Virus
Authors: Handel Kathy , Kehler Helen , Hills Kevin , Pasick John
Published: May 2011

Pigs were experimentally inoculated with Glentorf, Lelystad/97, and Alfort/187: representative low, moderate, and high virulent strains of classical swine fever virus (CSFV). Animals were tested for viremia using virus isolation and reverse transcriptase–polymerase chain reaction (RT-PCR) assays run under routine diagnostic conditions. The virus was detected in the peripheral blood by virus isolation and RT-PCR assays of all Glentorf- and Lelystad/97-infected pigs beginning at 3 days postinoculation (dpi) and in all Alfort/187-infected pigs beginning at 2 dpi. Viremia, as determined by virus isolation, remained detectable in Lelystad/97- and Alfort/187-infected pigs until the last animal within each cohort was euthanized on days 12 and 7 postinoculation, respectively. In contrast, the virus could be isolated from the blood of all Glentorf-infected pigs between 3 and 7 dpi but not from 10 to 21 dpi when the experiment was concluded. Viremia, as determined by RT-PCR, became apparent in Alfort/187-infected pigs at 2 dpi and in Glentorf- and Lelystad/97-infected pigs at 3 dpi. All pigs, regardless of the CSFV strain used, remained RT-PCR positive until they were euthanized. Tonsils were harvested from all the pigs and frozen sections tested for the presence of the CSFV antigen using polyclonal pestivirus and monoclonal CSFV horseradish peroxidase (HRPO) conjugates. Immunostaining reactions were positive for all the Alfort/187- and Lelystad/97-infected pigs. By contrast, tonsils from the Glentorf-infected pigs gave negative to equivocal results. These data suggest that an RT-PCR assay performed on blood may be the best test when dealing with pigs infected with low virulent strains of CSFV.
Erysipelothrix Septicemia in a Little Blue Penguin (Eudyptula Minor)
Authors: Boerner Leslie , Nevis Kathleen R. , Hinckley Lynn S. , Weber E. Scott , Frasca Salvatore
Published: May 2011

On June 25, 2002, aquarium veterinarians treated a 5-year-old, male little blue penguin ( Eudyptula minor) that was acutely recumbent and dull, with inappetence of 24-hour duration. The penguin died within 10 minutes of presentation despite emergency resuscitation efforts. Gross pathologic findings consisted of pulmonary congestion and intestinal hemorrhage. Histopathologic findings included necrosis of tips of intestinal villi, increased numbers of mononuclear cells in pulmonary interstitium and hepatic sinusoids, and gram-positive bacteria in systemic microvasculature. Transmission electron microscopic examination revealed short gram-positive bacilli located in lumina of glomerular capillaries and in cytoplasm of mononuclear phagocytic cells in the lung and liver. Erysipelothrix rhusiopathiae was recovered from the lung, liver, and intestine by bacteriologic culture. Amplicons from polymerase chain reaction (PCR) tests using Erysipelothrix genus–specific primers and total genomic DNA extracted from formalin-fixed, paraffin-embedded tissue sections of lung and intestine demonstrated 99% nucleotide sequence identity with 16S small-subunit ribosomal DNA of E. rhusiopathiae and E. tonsillarum. The source of infection was speculated to be fish in the diet; however, repeated attempts to detect Erysipelothrix spp. from the mucous layer of food fish using bacteriologic culture and PCR were unsuccessful. This is the first report of erysipelas in a captive aquatic bird. Details of the isolation of E. rhusiopathiae and the application of molecular testing to identify Erysipelothrix DNA in formalin-fixed, paraffin-embedded tissue sections are given.
Evaluation of Five Diagnostic Methods for the Detection and Quantification of Myxobolus Cerebralis
Authors: Kelley Garry O. , Zagmutt-Vergara Francisco J. , Leutenegger Christian M. , Myklebust Karin A. , Adkison Mark A. , McDowell Terry S. , Marty Gary D. , Kahler Alex L. , Bush Arla L. , Gardner Ian A. , Hedrick Ronald P.
Published: May 2011

Diagnostic methods were used to identify and quantify Myxobolus cerebralis, a myxozoan parasite of salmonid fish. In this study, 7-week-old, pathogen-free rainbow trout ( Oncorhynchus mykiss) were experimentally infected with M. cerebralis and at 7 months postinfection were evaluated with 5 diagnostic assays: 1) pepsin–trypsin digest (PTD) to detect and enumerate spores found in cranial cartilage, 2) 2 different histopathology grading scales that provide a numerical score for severity of microscopic lesions in the head, 3) a conventional single-round polymerase chain reaction (PCR), 4) a nested PCR assay, and 5) a newly developed quantitative real-time TaqMan PCR. There were no significant differences ( P > 0.05) among the 5 diagnostic assays in distinguishing between experimentally infected and uninfected control fish. The 2 histopathology grading scales were highly correlated ( P < 0.001) for assessment of microscopic lesion severity. Quantification of parasite levels in cranial tissues using PTD and real-time TaqMan PCR was significantly correlated r = 0.540 ( P < 0.001). Lastly, 104 copies of the 18S rDNA gene are present in the M. cerebralis genome, a feature that makes this gene an excellent target for PCR-based diagnostic assays. Also, 2 copies of the insulin growth factor–I gene are found in the rainbow trout genome, whose detection can serve both as an internal quality control for amplifiable DNA and as a basis to quantify pathogen genome equivalents present in quantitative PCR assays.
Age-Stratified Validation of an Indirect Salmonella Dublin Serum Enzyme-Linked Immunosorbent Assay for Individual Diagnosis in Cattle
Authors: Nielsen Liza Rosenbaum , Ersbøll Annette Kjár
Published: May 2011

Enzyme-linked immunosorbent assays (ELISAs) are routinely used for cattle herd diagnosis of Salmonella Dublin infection in many countries. It is also possible to use such tests for individual diagnosis. Passively transferred immunoglobulins may cause false-positive test results in young calves. Also, false-positive test results may be seen in recovered animals several months after infection. False-negative results are seen in acutely infected animals, especially immature animals that are unable to produce a humoral antibody response to infection. To be able to interpret the individual animal test results, it is necessary to take age into account when validating the ELISA. In the present study an age-stratified validation of an indirect Salmonella Dublin serum ELISA as predictor of bacterial excretion was performed using receiver-operating characteristic (ROC) analysis. Three age groups were formed according to the results of an exploratory analysis of the age effect on area under curve (AUC) of ROC curves. The AUC for the youngest age group (0–99 days) was 0.816 (SE = 0.033), which was significantly ( z = 4.23, P < 0.0001) smaller than the 0.977 (SE = 0.019) estimated for the next age group (100–300 days). The oldest age group (>300 days) had an AUC of 0.905 (SE = 0.023), which was significantly different from the AUC of both the other age groups ( z = 2.21, P = 0.027 when compared with the youngest age groups and z = 2.41, P = 0.016 when compared with the age group of 100–300 days). The results showed that the indirect Salmonella Dublin serum ELISA is most valid for detection of infection in individual cattle from the age of 100 days. Purpose-related test sensitivities and specificities were evaluated at different cutoff values.
Increased Concentration of GM1-Ganglioside in Cerebrospinal Fluid in Dogs with GM1- and GM2-Gangliosidoses and its Clinical Application for Diagnosis
Authors: Satoh Hiroyuki , Yamato Osamu , Asano Tomoya , Yamasaki Masahiro , Maede Yoshimitsu
Published: May 2011

GM1- and GM2-gangliosidoses are lethal lysosomal diseases that are caused by a defect of acid hydrolases, resulting in the intralysosomal accumulation of the specific physiological substrates, GM1- and GM2-gangliosides, respectively. In the present study a method for the diagnosis of canine GM1-gangliosidosis was established using canine cerebrospinal fluid (CSF). The concentration of GM1-ganglioside in CSF was determined by thin-layer chromatography–enzyme immunostaining using biotin-conjugated cholera toxin B, which specifically binds with GM1-ganglioside. The concentration of CSF GM1-ganglioside was increased in Shiba dogs with GM1-gangliosidosis, and the increased level was approximately proportional to the age of the dogs. The concentration was high in the affected dog even at 5 months of age, when Shiba dogs with GM1-gangliosidosis first manifest neurologic signs. In addition, the concentration of CSF GM1-ganglioside in a dog with the GM2-gangliosidosis 0 variant (Sandhoff disease) was also 7 times the normal level. From these results it was concluded that this laboratory technique enables a definitive and early diagnosis of canine GM1-gangliosidosis even if tissues and organs cannot be obtained. However, because GM1-ganglioside can also be elevated in cases of GM2-gangliosidosis, it is necessary to assay for specific enzyme deficiencies to definitively separate GM1- from GM2-gangliosidosis.
Malignant Neoplasia in Four Alpacas
Authors: Sartin Eva A. , Crowe D. Ralph , Whitley Elizabeth M. , Treat Robert E. , Purdy Stephen R. , Belknap Ellen B.
Published: May 2011

Malignant neoplasia in 4 alpacas was characterized by acute onset of clinical signs and rapidly deteriorating condition. Postmortem examination revealed metastatic or multicentric neoplasia in the abdominal organs of alpacas 1, 3, and 4 and an extensive thoracic mass in alpaca 2. Immunohistochemical stains supported a diagnosis of B-cell lymphosarcoma in alpacas 1–3 and a neuroendocrine neoplasm in alpaca 4.
Electron Microscopic Study of Canine Babesia Gibsoni Infection
Authors: Radi Zaher A. , Styer Eloise L. , Frazier Ken S.
Published: May 2011

Canine babesiosis is a tick-borne parasitic disease caused by the intraerythrocytic parasites, Babesia canis and Babesia gibsoni. A lethargic, weak, American Staffordshire Terrier (pit bull) dog, which had regenerative, normocytic, normochromic anemia, was shown by polymerase chain reaction analysis to be infected with B. gibsoni. Transmission electron microscopy of ethylenediamine tetraacetic acid–treated blood disclosed many well-preserved, intraerythrocytic babesia trophozoites. Four morphologic forms of babesia trophozoites are described (small spheres, small rods, irregular forms lacking pseudoinclusions, and large spheres having pseudoinclusions) and are compared with intraerythrocytic forms of B. canis and B. gibsoni described in other light and electron microscopic studies of in vivo and in vitro Babesia infections. This is the first detailed transmission electron microscopic study of canine B. gibsoni–infected red blood cells in North America.
Adaptation and Limitations of Established Hemagglutination Inhibition Assays for the Detection of Porcine Anti—Swine Influenza Virus H1N2 Antibodies
Authors: Long Bradley C. , Goldberg Tony L. , Swenson Sabrina L. , Erickson Gene , Scherba Gail
Published: May 2011

Hemagglutination inhibition (HI) has been a reliable method for determining porcine antibody levels to the well-characterized swine influenza virus (SIV) H1N1 and H3N2 subtypes. However, the recent emergence of the novel H1N2 serotype of SIV and the persistence of 2 other serotypes (H1N1 and H3N2) in the United States swine population represents a significant challenge to diagnostics. Both standardized and modified HI protocols were used in a blinded study to examine a collection of 50 control sera representing a total of 12 swine that were experimentally inoculated with one of the 3 SIV subtypes. Using these control sera data, a statistical basis for analysis was established in an attempt to classify 30 field sera with known case histories or seroprevalance into SIV serotype categories. By this approach 57% of the field sera could be classified into specific categories. The remaining samples that could not be classified reliably were most likely composed of heterogeneous anti-SIV antibody populations. These results indicate that although serological differentiation might be possible in a controlled environment, applications of these methods to field samples are currently problematic. Approaches other than HI will be required to fulfill the current need for SIV diagnostics and surveillance when specific serotype identification is required.