bluuebunny/crossref_metadata_2025 huggingface dataset:
Caprine Herpesvirus-2–Associated Malignant Catarrhal Fever in White-Tailed Deer (Odocoileus Virginianus)
Authors: Li Hong , Wunschmann Arno , Keller Janice , Hall D. Greg , Crawford Timothy B.
Published: May 2011

A subacute disease presenting primarily as alopecia and weight loss occurred in 2 white-tailed deer ( Odocoileus virginianus) on farms in Minnesota and in Texas. A presumptive diagnosis of malignant catarrhal fever (MCF) was made on the basis of histological lesions. Antibody against an epitope conserved among the MCF group viruses was detected in the serum of both deer. DNA samples from the deer were subjected to a variety of PCR amplifications. Alignment of the amplified sequences from the diseased animals revealed that they were 100% identical to each other and to the same DNA fragment from the newly recognized member of the MCF virus group endemic in domestic goats ( Capra hircus), provisionally named caprine herpesvirus 2 (CpHV-2). A seroprevalence survey from one of the deer farms showed a high rate of subclincal infection in the deer population. This study provides further confirmation that CpHV-2 is a pathogen, at least for deer, and emphasizes the risk of loss from MCF when mixing cervids with goats.
Identification and Characterization of Fowlpox Virus Strains Using Monoclonal Antibodies
Authors: Singh Pratik , Kim Tae-Joong , Tripathy Deoki N.
Published: May 2011

The use of 2 monoclonal antibodies (MAbs), P1D9 and P2D4, which recognize different fowlpox virus (FPV) antigens, for the identification and characterization of FPV strains was evaluated. Initially, the MAbs were used in conjunction with a dot blot assay that enabled FPV to be differentiated from the avian herpesvirus, infectious laryngotracheitis virus. Confirmation of the specificity of these MAbs was provided by the demonstration that only FPV antigens were recognized by a combination of both antibodies when used for immunoblotting proteins contained in various avipoxviruses. Later, an antigenic characterization of 11 FPV field isolates, 6 FPV vaccine strains, and 3 pigeonpox virus vaccines was performed by Western blotting with the individual MAbs. Whereas MAb P2D4 consistently recognized a protein with an apparent molecular weight of 60 kD, there was variability in the size of the antigen that was immunoreactive with the other MAb. For example, MAb P1D9 recognized an antigen of apparent molecular weight of 46 kD in all vaccine strains except 2 of FPV origin. In these exceptions, either only a 39-kD or both a 42- and 46-kD protein were immunoreactive. As for the field isolates, a 39-kD antigen was recognized in 8 of them, whereas a 42-kD antigen was detected in the remaining 3. Therefore, the more extensive immunoblotting technique may facilitate FPV strain differentiation, whereas routine diagnosis of fowlpox could be accomplished by using the MAb-based dot blot assay.
Encephalitozoon Cuniculi Placentitis and Abortion in a Quarterhorse Mare
Authors: Patterson-Kane J. C. , Caplazi P. , Rurangirwa F. , Tramontin R. R. , Wolfsdorf K.
Published: May 2011

Encephalitozoon cuniculi is a microsporidial parasite, which has rarely been reported to cause placentitis in animals. A late-term aborted fetus and placenta from a Quarterhorse were presented to the Livestock Disease Diagnostic Center, University of Kentucky, for diagnostic examination. There was a necrotizing placentitis, with distension of many chorionic epithelial cells by intracytoplasmic vacuoles containing 1–2-μm-diameter, elongated, gram-positive organisms. The organisms were identified as E. cuniculi by electron microscopy and by polymerase chain reaction using primers to microsporidial ribosomal DNA. Joints of the fetus were swollen, with gross and microscopic lesions of synovitis; however, E. cuniculi DNA was not detected.
Papular Dermatitis Induced in Guinea Pigs by the Biting Midge Culicoides Sonorensis (Diptera: Ceratopogonidae)
Authors: O'Toole D. , de León A. A. Pérez , Hearne C. , McHolland L. , Yun L. , Tabachnick W.
Published: May 2011

Histological, ultrastructural, and virological examinations were performed on abdominal skin from guinea pigs after a blood meal by colony-bred biting midges, Culicoides sonorensis. Small, superficial, cutaneous, crateriform ulcers with necrosis of superficial dermis developed at feeding sites and healed within 24–48 hours. Animals developed nonpruritic erythematous papules 5 days after feeding that persisted until the study ended at 12 days after feeding. Papules corresponded histologically to foci of epidermal hyperplasia and superficial interstitial dermatitis with intraepidermal micropustules and scattered intraepidermal polykaryons. The principal ultrastructural changes were spongiosis in germinal epithelium and neutrophilic-histiocytic exocytosis. No viral agents or broken mouthparts were identified in lesions. The dermatitis may represent a host reaction to persisting insect salivary secretion and should be considered as an additional consequence of blood feeding in future studies involving biting midges.
Malignant Ovarian Teratoma in a Red-Eared Slider (Trachemys Scripta Elegans)
Authors: Newman Shelley J. , Brown Cyndi J. , Patnaik Amiya K.
Published: May 2011

A 7-year-old intact female red-eared slider ( Trachemys scripta elegans) presented with anorexia of 2–3-weeks duration. Coelomic radiographs demonstrated 3 shelled eggs in the right cavity. Hormone therapy failed to induce oviposition. Exploratory laparotomy revealed 3 calcified eggs in the right oviduct and a large mass obliterating the left ovary. The large, multilobulate mass had scattered, firm, white nodules and weighed 450 g. Histopathology revealed a malignant teratoma composed of immature embryonal and extraembryonal elements derived from all 3 germ layers with multifocal epithelial anaplasia. This is the first report of a teratoma in chelonian gonadal tissue. Teratomas and reptilian gonadal neoplasia are reviewed.
Comparison of Two DNA Extractions and Nested PCR, Real-Time PCR, a New Commercial PCR Assay, and Bacterial Culture for Detection of Mycobacterium Avium Subsp. Paratuberculosis in Bovine Feces
Authors: Christopher-Hennings Jane , Dammen Matthew A. , Weeks Shelleen R. , Epperson William B. , Singh Shri N. , Steinlicht Gina L. , Fang Ying , Skaare Jessica L. , Larsen Jill L. , Payeur Janet B. , Nelson Eric A.
Published: May 2011

In this study, 5 combinations of 2 DNA extractions and 3 polymerase chain reaction (PCR) techniques were compared with culture for the detection of Mycobacterium paratuberculosis directly from bovine feces. These combinations included a new commercial extraction technique combined with a commercial PCR/Southern blot technique, nested PCR (nPCR), or real-time PCR, and a university-developed extraction combined with nPCR or real-time PCR. Four of the 5 combinations had statistically similar sensitivities between 93% and 100% and specificity between 95% and 100%, when compared with culture results from 63 bovine fecal samples. These results indicated that using a commercial extraction with a commercial PCR/Southern blot, nPCR, or real-time PCR, or a university-developed extraction with real-time PCR would result in similar sensitivities to culture for the identification of M. paratuberculosis from bovine feces and are valid alternatives to culture.
Experimental Reproduction of Postweaning Multisystemic Wasting Syndrome in Cesarean-Derived, Colostrum-Deprived Piglets Inoculated with Porcine Circovirus Type 2 (PCV2): Investigation of Quantitative PCV2 Distribution and Antibody Responses
Authors: Okuda Y. , Ono M. , Yazawa S. , Shibata I.
Published: May 2011

Sixteen cesarean-derived, colostrum-deprived piglets were inoculated intranasally with porcine circovirus type 2 (PCV2), originally isolated from a pig affected with postweaning multisystemic wasting syndrome (PMWS). At 1 day postinoculation (PI), 3 of the 5 piglets in the uninoculated control group were moved to the room of inoculated piglets for contact exposure. Porcine circovirus type 2 was detected by polymerase chain reaction (PCR) in swabs from inoculated piglets from 1 day PI and from contact piglets from 2 days after cohabitation. Porcine circovirus type 2 was also detected in all serum samples but not in control piglets 7 days PI. Until the end of study, PCV2 was detected in swabs and serum samples by PCR but not in the control piglets. One inoculated piglet died suddenly without clinical signs 19 days PI. Beginning at 14 days PI, 5 piglets, including 1 contact piglet, had clinical signs of depression, anorexia, and icterus, and 1 inoculated piglet died 21 days PI. Most of the piglets exhibiting the above clinical signs became moribund and were necropsied 21 and 28 days PI. In the piglets that showed clinical signs, gross lesions, including icterus of liver and hemorrhage in stomach, and typical histopathological lesions of PMWS, such as lymphoid depletion and basophilic intracytoplasmic inclusion bodies in lymph nodes and other tissues, were observed. Porcine circovirus type 2 was detected by PCR in all tissue samples except in those of the control piglets. Porcine circovirus type 2 was recovered from several tissue samples of the piglets necropsied until 35 days PI. In particular, PCV2 was recovered in high titer from most of the tissue samples of the piglets exhibiting clinical signs. Serum antibody against PCV2 was mostly detected in inoculated piglets and in contact piglets 14 and 21 days PI by an indirect fluorescence antibody test but was not detected in the piglets exhibiting clinical signs until 28 days PI. These results indicate that PCV2 was able to induce clinical PMWS in the absence of other swine pathogens and that there were significant differences in both the quantitative PCV2 distribution in tissues and the antibody response between the piglets that were infected and developed PMWS and those that were infected but remained healthy.
Rate of Apoptosis in Feline Mammary Tumors is not Predictive of Postsurgical Survival
Authors: Sarli Giuseppe , Preziosi Rosario , Benazzi Cinzia , Bazzo Roberta , Mandrioli Luciana , Marcato Paolo Stefano
Published: May 2011

Invasion, cell proliferation and apoptosis are important biological features of neoplasia, bearing prognostic importance. Histological stage, mitotic index, and apoptotic index have been assessed in 33 feline malignant mammary tumors. Histological stage ( P < 0.01) and mitotic index ( P < 0.001) had a significant association with prognosis in univariate analysis. Apoptotic index did not correlate with survival ( P = 0.44), and histological stage ( P = 0.48) did not correlate with mitotic index ( P = 0.39). In feline malignant mammary tumors the apoptotic index seems unable to predict survival and lacks any correlation with proliferation assessed as mitotic index. A possible explanation for the lack of correlation between apoptotic index and survival may be due to the rapid acquisition of pathways of apoptosis resistance in feline mammary tumors or to rapid hormone receptors loss.
Neurological Disorder in Dairy Cattle Associated with Consumption of Beer Residues Contaminated with Aspergillus Clavatus
Authors: Loretti Alexandre Paulino , Colodel Edson Moleta , Driemeier David , Corrěa Andre Mendes , Bangel Jorge José , Ferreiro Laerte
Published: May 2011

A neurological syndrome in dairy cattle associated with consumption of moldy beer residues is described. The disease occurred on 1 farm in late June 2001, during winter. Six heifers and 1 cow out of 45 cattle were affected during a 3-week period. The affected animals died spontaneously or were euthanized approximately 2–14 days after the onset of clinical signs. The clinical signs were characterized by flaccid paralysis and gait abnormalities. Clinical signs were more pronounced after exercise and included stiff and unsteady gait, knuckling at the fetlocks of the hind limbs, frequent falling, inability to rise, muscular tremors, especially of the head and the hindquarters, and drooling. Main necropsy findings included degenerative and necrotic changes of the larger medial muscle groups of the hindquarters, i.e., adductor, pectineus, quadriceps femoris, rectus femuris, sartorius, semimembranosus, semitendinosus, and vastus medialis, and of the fore-quarters, including pectoralis descendens, pectoralis ascendens, and transversus pectoralis. The main histologic findings consisted of degenerative and necrotic neuronal changes (chromatolysis) of varying severity and extent affecting selected nuclei of the brainstem and neurons of the ventral horns of the spinal cord. Similar microscopic lesions were observed in the neurons of the spinal cord of 1 experimental sheep force-fed for 35 days with 1 kg/day of the same batch of foodstuff that was originally fed to the cattle. Coarse white or gray lumps, interpreted as mycelia, were observed in the beer by-product. Aspergillus clavatus was the dominant fungus isolated. Deaths ceased after the consumption of beer residue was discontinued. Recovery from illness was observed in 1 animal. The diagnosis was based on epidemiological data, clinical signs, necropsy findings, histological lesions, dosing trial, and mycology. A similar condition caused by consumption of barley byproducts, sprouted wheat, corn sprouts, and beetroot screenings contaminated with A. clavatus has been reported in cattle and sheep worldwide.
Application of Immunoperoxidase-Based Techniques to Detect Herpesvirus Infection in Tortoises
Authors: Origgi F. C. , Klein P. A. , Tucker S. J. , Jacobson E. R.
Published: May 2011

Indirect (IIP) and direct (DIP) immunoperoxidase assays were developed for the serological and histological diagnoses of herpesvirus infection in tortoises, respectively. A mouse monoclonal antibody (MAb HL1546), specific for the heavy chain of tortoise IgY, was used as the secondary antibody in the IIP assay. Rabbit polyclonal antisera raised against 2 sucrose gradient–purified tortoise herpesvirus isolates (HV4295/7R/95 and HV1976) were used as primary antibodies for the detection of herpesvirus antigen either in infected cell cultures or in formalin-fixed, paraffin-embedded tissues. The IIP and DIP assays could detect either the presence of anti-herpesvirus antibody in the plasma of exposed tortoises or the presence of herpesvirus antigen in infected tissues, respectively. Although the IIP test complements the enzyme-linked immunosorbent assay and the serum neutralization test already available for measuring herpesvirus-specific antibody in tortoises, the DIP test is useful for the histological diagnosis of herpesvirus infection in tortoises.
Absence of Evidence of Porcine Circovirus Infection in Piglets with Congenital Tremors
Authors: Kennedy Seamus , Segalés Joaquim , Rovira Albert , Scholes Sandra , Domingo Mariano , Moffett Deborah , Meehan Brian , O'Neill Ronan , McNeilly Francis , Allan Gordon
Published: May 2011

Porcine circovirus types 1 (PCV1) and 2 (PCV2) have been associated with congenital tremors (CTs) in piglets in the United States. In this study, central nervous system and nonneural tissues of 40 CT piglets from Spain, the United Kingdom, Ireland, and Sweden were investigated for the presence of PCV1 and PCV2 using in situ hybridization and immunohistochemical labeling on paraffin sections. The polymerase chain reaction for PCV2 was also carried out on sera from the Spanish CT cases. No evidence of circovirus nucleic acid or antigen was found in any CT piglet. Although these results do not support the hypothesis that PCV1 or PCV2 are linked to porcine CT, they cannot disprove it.
Immunohistochemical Detection and Distribution of Prion Protein in a Goat with Natural Scrapie
Authors: Valdez Reginald A. , Rock Matthew J. , Anderson Anne K. , O'Rourke Katherine I.
Published: May 2011

Formalin-fixed, paraffin-embedded tissue sections from a 3-year-old female Angora goat suffering from clinical scrapie were immunostained after hydrated autoclaving using a monoclonal antibody (mAb, F99/97.6.1; IgG1) specific for a conserved epitope on the prion protein. Widespread and prominent deposition of the scrapie isoform of the prion protein (PrPSc) was observed in the brain, brainstem, spinal cord, retina, postganglionic neurons associated with parasympathetic ganglia of myenteric and submucosal plexuses, Peyer's patches, peripheral lymph nodes, and pharyngeal and palatine tonsils. The goat was homozygous for PrP alleles encoding 5 octapeptide repeat sequences in the N-terminal region of the prion protein and isoleucine at codon 142, a genotype associated with high susceptibility and short incubation times in goats. The results of this study indicate that mAb F99/97.6.1 is useful for detection of PrPSc deposition, and this is a specific and reliable immunohistochemical adjunct to histopathology for diagnosis of natural caprine scrapie, although precise determination of the diagnostic sensitivity and specificity of the assay as a diagnostic test for scrapie in goats will require examination of a sufficiently large sample size. As with ovine scrapie, prion protein is widely distributed in the central and peripheral nervous systems, gastrointestinal tract, and lymphoid tissues in natural caprine scrapie.
Plasma Cell Myeloma Producing IgG, IgM, and IgA Immunoglobulins in a Cow
Authors: Kameyama Mamoru , Ishikawa Yoshiharu , Shibahara Tomoyuki , Kadota Koichi
Published: May 2011

A diagnosis of plasma cell myeloma was made in a 6-year-old Holstein cow that showed continuous nosebleed, progressing emaciation, and hyperglobulinemia. Necropsy revealed enlargement of the caudal mesenteric lymph nodes. Lymph nodes and spleen were infiltrated by neoplastic plasma cells. Renal lymph nodes, bone marrow, and kidneys also were affected. Amyloid-like material that was negative for Congo red was present in most of the affected lymph nodes. Congo red–positive amyloid material was noted in spleen, kidneys, liver, and adrenal glands. Neoplastic plasma cells exhibited immunohistochemical reactions for λ-light chain and 3 classes of heavy chain but not for κ-light chain. Nonamyloid deposits were labeled with antisera against λ-light chain and heavy chains. Plasmacytoid cells, which stained positively for IgG, IgM, or IgA, were intermingled in all lesions. Production of 3 types of immunoglobulins may be caused by heavy-chain class switching, constantly occurring in certain proportions.
Pulmonary Carcinosarcoma in a Cat
Authors: Ghisleni Gabriele , Grieco Valeria , Mazzotti Marcello , Caniatti Mario , Roccabianca Paola , Scanziani Eugenio
Published: May 2011

A Domestic Shorthaired cat was presented with coughing and severe respiratory distress. Thoracic radiographs revealed a lobar mass and numerous additional cavitated intrapulmonary masses. The cat was euthanized and submitted for necropsy. Histological examination of the large mass revealed 2 distinct neoplastic components consisting of bronchial adenocarcinoma admixed with neoplastic areas composed of highly atypical undifferentiated spindle cells (sarcomatous component). Simultaneous expression of vimentin and cytokeratin by a subpopulation of neoplastic epithelial cells and by rare neoplastic spindle cells was identified. On the basis of histology and immunohistochemical results, a diagnosis of primary pulmonary carcinosarcoma with intrapulmonary epithelial metastases was made. Pulmonary carcinosarcoma is a well-known pathological entity in humans. It is a rare tumor in animals and has not been previously reported in cat.
Indirect Method for Prediction of Hemagglutination Inhibition Antibody Titers to Newcastle Disease Virus in Chickens by Titration of Antibodies in Egg Yolk
Authors: Yeo SangGeon , Nagy Eva , Krell Peter J.
Published: May 2011

Attempts were made to establish methods for indirect prediction of hemagglutination inhibition (HI) antibody titers to Newcastle disease virus (NDV) in sera of laying hens and day-old chicks by determining if these are correlated to HI titers in egg yolks. For this purpose, geometric means of HI antibody titers in sera from 60 hens, yolks from 60 matched eggs, and sera from 180 day-old chicks of an identical vaccination program were measured and plotted. There was a significant correlation between HI antibody titers in yolks (X) and hens (Y), with a linear regression of Y = 23.24 + 0.47X and a correlation coefficient of r = 0.65. The linear regression between HI antibody titers in yolks (X) and chicks (Y) was Y = 6.33 + 0.36X ( r = 0.58). Immunity to NDV in hens and their offspring can be maintained effectively, and the proper time for the vaccination or booster can be determined by reference to HI titers predicted from the linear regression in the present study. The approach of testing egg yolk for HI titers provides a feasible alternative to determining HI titers from blood samples and eliminates stress in birds during blood sampling.
Evaluation of a Digestion Assay and Determination of Sample Size and Tissue for the Reliable Detection of Trichinella Larvae in Walrus Meat
Authors: Leclair Daniel , Forbes Lorry B. , Suppa Sandy , Gajadhar Alvin A.
Published: May 2011

A digestion assay was validated for the detection of Trichinella larvae in walrus ( Odobenus rosmarus) meat, and appropriate samples for testing were determined using tissues from infected walruses harvested for food. Examination of muscles from 3 walruses showed that the tongue consistently contained approximately 2–6 times more larvae than the pectoral and intercostal muscles. Comparison of numbers of larvae in the root, body, and apex of the tongue from 3 walruses failed to identify a predilection site within the tongue, but the apex was considered an optimal tissue because of the high larval density within the tongue and the ease of collection. All 31 spiked samples weighing 50 g each and containing between 0.1 and 0.4 larvae per gram (lpg) were correctly identified as infected, indicating that the sensitivity of this procedure is adequate for diagnostic use. A sample size of 10 g consistently detected larvae in 2 walrus tongues containing ≥0.3 lpg ( n = 40), and until additional data are available, sample sizes from individual walrus tongues should be a minimum of 10 g. This study provides the preliminary data that were used for the development of a food safety analytical protocol for the detection of Trichinella in walrus meat in arctic communities.
Temporal Patterns of Diagnostic Results in Serial Samples from Cattle with Advanced Paratuberculosis Infections
Authors: Barrington George M. , Gay John M. , Eriks Inge S. , Davis William C. , Evermann James F. , Emerson Carlene , O'Rourke Jennifer L. , Hamilton Mary J. , Bradway Daniel S.
Published: May 2011

As part of investigating diagnostic strategies for Mycobacterium avium subsp. paratuberculosis ( Map), serial results from polymerase chain reaction (PCR) on extraintestinal tissues (blood, milk, and liver) were compared with those from more conventional detection methods including serum enzyme-linked immunosorbent assay (ELISA), fecal culture, and fecal PCR. Three cows previously identified as being subclinically infected with Map were selected for the study. Blood, milk, and feces were collected daily and liver biopsies were obtained weekly for a 30-day period. Unexpectedly, a substantial daily variation in serum ELISA sample to positive (S/P) ratios was observed in all 3 cows. In contrast, fecal culture results were consistently positive. However, whereas fecal culture colony counts were consistently high for 2 cows throughout the study, colony counts from the third cow varied from day to day. Diagnostic sensitivity of PCR for fecal, blood, milk, and liver samples in these advanced subclinically infected cows was 87%, 40%, 96%, and 93%, respectively.
A Canine Distemper Outbreak in Alaska: Diagnosis and Strain Characterization Using Sequence Analysis
Authors: Maes Roger K. , Wise Annabel G. , Fitzgerald Scott D. , Ramudo Albert , Kline Joseph , Vilnis Aivars , Benson Cherie
Published: May 2011

Vaccination with modified-live vaccines has been very effective in reducing the incidence of canine distemper, a disease that can be devastating in unvaccinated populations. A diagnostic submission to the Animal Health Diagnostic Laboratory at Michigan State University, East Lansing, Michigan, involved a case in which several hundred dogs in an Alaskan town died in a suspected canine distemper outbreak. Cytoplasmic and intranuclear eosinophilic inclusion bodies, consistent with canine distemper virus (CDV) infection, were found in urinary bladder, spleen, lung, and salivary gland. Direct fluorescent antibody test gave results that could be considered positive for canine distemper. Because of the condition of the tissues received, the histopathology and fluorescent antibody–staining results were suggestive but not conclusive of CDV. In this study, immunohistochemistry, reverse transcriptase–polymerase chain reaction (RT-PCR), and DNA sequencing were used to confirm the presence of canine distemper virus in these tissues and to perform molecular characterization of the virus. Immunohistochemistry showed the presence of the virus in spleen, lung, and salivary gland. Viral RNA was detected by RT-PCR in brain, spleen, liver, lung, and kidney, both with nucleoprotein and phosphoprotein (P)-gene–specific primers. Sequence alignment and phylogenetic analysis of a 540-bp P-gene fragment of the Alaskan strain with corresponding sequences of 2 vaccine and 7 wild-type CDV strains showed that the virus responsible for the outbreak was closely related to a virulent strain of distemper virus from Siberia.
A Novel Multiplex Polymerase Chain Reaction Approach for Detection of Four Human Infective Cryptosporidium Isolates: Cryptosporidium Parvum, types H and C, Cryptosporidium Canis, and Cryptosporidium Felis in Fecal and Soil Samples
Authors: Lindergard Gabriella , Nydam Daryl V. , Wade Susan E. , Schaaf Stephanie L. , Mohammed Hussni O.
Published: May 2011

A nested multiplex polymerase chain reaction (PCR) approach was adopted for the simultaneous detection of 4 human infective genotypes of the protozoan parasite Cryptosporidium. Specific PCR primers were designed for the heat shock protein 70 gene of 2 genotypes of Cryptosporidium parvum (human and bovine types), Cryptosporidium canis, and Cryptosporidium felis. These 4 genotypes have all been found in human fecal samples. The primers amplified DNA fragments of specific sizes, each representing a unique genotype. The limit of detection of the method was found to vary between 10 and 100 oocysts per 1 ml fecal material. There appeared to be no cross-reactivity with other organisms commonly present in feces and soil, and the approach has a high specificity. The rapid identification of various human infective Cryptosporidium isolates is a part of the authors' long-term aim of determining the routes of infection with oocysts and thereby increase their epidemiological understanding of Cryptosporidium infection in humans and animals.
The Use of a Mimic to Detect Polymerase Chain Reaction– Inhibitory Factors in Feces Examined for the Presence of Lawsonia Intracellularis
Authors: Jacobson Magdalena , Englund Stina , Ballagi-Pordány András
Published: May 2011

Lawsonia intracellularis is an intracellular organism that causes proliferative enteritis in pigs. This bacterium is difficult to culture, and antemortem demonstration of the microbe is therefore often performed on fecal samples by polymerase chain reaction (PCR). Polymerase chain reaction is sensitive and specific, but inhibitory factors in feces might cause false-negative results. This article describes the construction and use of an internal standard, a mimic. The mimic is amplified by the same primers as those used for L. intracellularis DNA and thus could indicate false-negative results in clinical samples. The amplicon was clearly visible when as few as 10 mimic molecules were added per amplification reaction and when no inhibitors were present. When fecal samples were spiked with the mimic, the detection limit was 102 molecules per PCR. Sixty clinical samples, 20 from wild boars, 20 from growing pigs with diarrhea, and 20 from pigs without diarrhea, were prepared by a boiling procedure and subjected to PCR together with 103 mimic molecules. Nine samples were positive, of which 7 originated from pigs with diarrhea and 2 from pigs without diarrhea. In 14 samples from wild boars, in 8 samples from pigs without diarrhea, and in 3 samples from pigs with diarrhea, neither the mimic nor the target DNA was visible. This indicated the presence of inhibitors in these samples. It is concluded that the mimic can be used as an internal control in the diagnosis of L. intracellularis to indicate inhibition of PCR.
Comparison of Targeting F and G Protein Genes to Detect Bovine and Ovine Respiratory Syncytial Viruses
Authors: Eleraky Nasser Z. , Kania Stephen A. , Evermann James F. , Potgieter Leon N. D.
Published: May 2011

In this study, 2 reverse transcriptase–polymerase chain reaction (RT-PCR) assays were developed and compared for simultaneous detection of bovine and ovine respiratory syncytial viruses (RSVs). One assay was based on a set of primers, which amplified a 426-bp fragment of either bovine or ovine RSV F gene (RT-PCR F). The F products could be distinguished by EcoRI or BstYI restriction endonuclease cleavage. In the other assay, a set of primers amplified a 542-bp fragment of either ovine or bovine RSV G gene (RT-PCR G). EcoO1091 and RsaI restriction enzymes were used to differentiate between the ovine and bovine PCR-G products. Sequencing of the PCR products confirmed the fidelity of both assays. The 2 assays were evaluated using 18 bovine RSV isolates, 1 ovine RSV, 1 bighorn sheep RSV isolate, 1 caprine RSV isolate, 2 human RSV isolates, and several other viruses associated with bovine respiratory tract disease. RT-PCR G may be more sensitive in detecting viral RNA. Because the target sequence of the F gene is more conserved than that of the G gene, RT-PCR F followed by the appropriate restriction enzyme cleavage may be superior to RT-PCR G to discriminate between the 2 ruminant RSV subgroups. This assay should prove useful for determining the relative contribution of ovine and bovine RSV to the pathogenesis of bovine respiratory tract disease.
Lightning Injury in an Outdoor Swine Herd
Authors: Van Alstine William G. , Widmer William R.
Published: May 2011

Three pigs, weighing 63 kg–70 kg each, from a group of 8 pigs in an outdoor pen that was struck by lightning were necropsied. All 3 pigs presented with hind limb paralysis. The only lesions identified were multiple fractures of the last (seventh) lumbar vertebral body and first sacral vertebral segment, with dorsal displacement of the sacrum and transection of the distal spinal cord and spinal nerves. Hemorrhages extended from the fracture sites into muscles immediately surrounding the lumbosacral junction and retroperitoneally into the pelvic cavity. These hemorrhages were not clearly visible until the pelvic region was dissected. Lesions commonly found in human lightning-strike victims were not present in these pigs. Because vertebral fractures may be the only lesions and may be grossly subtle in heavily muscled pigs, careful pelvic and vertebral dissection is recommended in cases of suspected lightning strike and electrocution.
The Immunologic Response of Dogs to Bartonella Vinsonii Subspecies Berkhoffii Antigens: as Assessed by Western Immunoblot Analysis
Authors: Breitschwerdt Edward B. , Suksawat Jiraporn , Chomel Bruno , Hegarty Barbara C.
Published: May 2011

Bartonella vinsonii subspecies berkhoffii is a recently recognized zoonotic pathogen that causes endocarditis, granulomatous rhinitis, and granulomatous lymphadenitis in dogs. Isolation of B. vinsonii ( berkhoffii) from blood or tissue samples is frequently unsuccessful; therefore, diagnosis is primarily dependent on serologic or molecular testing modalities. Because previous canine serologic studies have used an indirect immunofluorescence assay (IFA), without Western immunoblot (WI) confirmation, the overall objective of this study was to examine the diagnostic use of WI for confirmation of B. vinsonii ( berkhoffii) infection in dogs. To confirm that agar-grown and cell culture–grown organisms yielded similar patterns of WI antigenic protein recognition, the 2 preparations were compared using IFA-reactive sera obtained from dogs experimentally infected with B. vinsonii ( berkhoffii). Temporal changes in the pattern of antigenic protein recognition were characterized using sera obtained from dogs at various time points after experimental B. vinsonii ( berkhoffii) infection. The specificity of B. vinsonii ( berkhoffii) WI was examined by testing canine sera that were reactive to B. henselae, B. clarridgeiae, Ehrlichia canis, Rickettsia rickettsii, Babesia canis, Anaplasma phagocytophilum (previously E. equi), or Brucella canis antigens. Clinical accessions including serum samples obtained from B. vinsonii ( berkhoffii) culture–positive dogs and B. vinsonii ( berkhoffii) culture–negative dogs that were IFA seroreactive to B. vinsonii ( berkhoffii) antigens were examined by WI. The results of this study indicate that WI using agar-grown or cell culture–grown B. vinsonii ( berkhoffii) antigens produce identical patterns of antigenic protein recognition. After experimental infection, there is a progressive increase in the number of antigenic proteins that are recognized by WI, with the 33-kD antigen representing the first and the most persistent antigen recognized by B. vinsonii ( berkhoffii)–infected dogs. Regarding specificity, sera from dogs that were reactive to various heterologous antigens did not recognize B. vinsonii ( berkhoffii) antigens by IFA or WI, and sera from dogs experimentally infected with B. henselae did not recognize B. vinsonii ( berkhoffii) antigens by WI. Regarding clinical accessions, there was good agreement between B. vinsonii ( berkhoffii) IFA test results and WI analysis. Western immunoblot analysis can be used to detect or confirm exposure to B. vinsonii ( berkhoffii) in dogs.
Thermoregulatory Response of Dairy Cows Fed Ergotized Barley during Summer Heat Stress
Authors: Al-Tamimi Hosam J. , Rottinghaus George E. , Spiers Donald E. , Spain Jim , Chatman Darryl , Eichen Peggy A. , Carson Thomas L.
Published: May 2011

Claviceps purpurea infects the seed heads of cereal grains and grasses and produces ergopeptine alkaloids that cause hyperthermia and agalactia in cattle during periods of heat stress. A field experiment was undertaken to examine the effects of ergopeptine alkaloids found in barley on thermal status of dairy cattle during periods of heat stress. Production end points were also measured to identify the effect of the change in thermal status. Contaminated barley screenings containing known levels of ergopeptine alkaloids were fed to lactating Holstein cattle (10 μg total ergopeptine alkaloids/kg BW/day) for 10 days during summer heat stress. Air temperature increased 14.4 C during the first 8 days of treatment and then declined the same during the last 2 days. Extreme daily values for rectal temperature and respiration rate, using averages of all animals, showed maximum increases of 2.3 C and 56.8 breaths/minute, respectively, during this period. Group afternoon milk production decreased 2 kg/day during the heat stress period, with no measurable change in feed intake. A greater level of hyperthermia occurred in cattle consuming the diet with ergopeptine alkaloids, with only marginal symptoms of ergot toxicosis reflected in feed intake and milk production. Therefore, the ergopeptine alkaloid dose used in this study represents a level for minimal induction of the ergot toxicity response.
Detection of Porcine Circovirus Type 2 in Feces of Pigs with or without Enteric Disease by Polymerase Chain Reaction
Authors: Yang Jeong S. , Song Dae S. , Kim So Y. , Lyoo Kwang S. , Park Bong K.
Published: May 2011

To establish the sensitive polymerase chain reaction(PCR) method and detect porcine circovirus type 2 (PCV2) from intestines and feces of commercial swine herds with or without enteric disease, intestinal samples from 68 pigs and 29 fecal samples from commercial swine farms were collected. A primer set, forward primer 5′-GAAGAATGGAAGAAGCGG-3′ and reverse primer 5′-CTCACAGCAGTAGACAGGT-3′, could detect the virus at a concentration as low as 2 infectious virions per milliliter under controlled conditions using PK-15 cell-adapted PCV2. The genomic nucleotide sequences of open reading frame 1 (ORF1) PCR products from fecal samples were found to have complete homology with other PCV2s deposited in the GenBank database. Transmissible gastroenteritis virus (TGEV) and porcine epidemic diarrhea virus (PEDV) as the other enteric pathogens were also investigated by performing duplex reverse transcription-PCR (RT-PCR). Among 63 pigs with clinical enteric disease, 18 PCV2s (14 from intestines and 4 from feces), 7 TGEVs from intestines, and 18 PEDVs (14 from intestines and 4 from feces) were detected by PCR and the duplex RT-PCR. In 34 pigs (14 from intestines and 20 from feces) without clinical enteric disease, only PCV2 was detected in 19 pigs (3 from intestines and 16 from feces). Both PEDV and PCV2 were found in 6 pigs with clinical enteric disease. Among 15 PCV2 samples that were PCR-positive, 4 were culture-positive at passage level 3 in PK-15 cells. These results reveal that PCV2 is shed through the feces of pigs without clinical enteric disease, which suggests the potentiality of the fecal–oral transmission of PCV2 in feces.
Prevalence of a Gene Encoding Adhesin Involved in Diffuse Adherence among Escherichia Coli Isolates in Pigs with Postweaning Diarrhea or Edema Disease
Authors: Ha Seung-Kwon , Choi Changsun , Chae Chanhee
Published: May 2011

A total of 604 Escherichia coli strains isolated from weaned pigs with diarrhea or edema disease on 653 swine farms were screened for the presence of the adhesin involved in diffuse adherence (AIDA) gene by polymerase chain reaction (PCR). Escherichia coli isolates that carried AIDA genes were also tested by PCR for the detection of 5 fimbriae (F4, F5, F6, F18, and F41), 3 heat-stable (STa, STb, and EAST1) and 1 heat-labile (LT) enterotoxin, and Shiga toxin 2e (Stx2e) genes. Forty-five (7.5%) of the 604 E. coli isolates carried the gene for AIDA. Of these 45 isolates, 5 (11.1%) carried EAST1 genes only, 1 (2.2%) carried genes for at least one of the fimbrial adhesins, 12 (26.7%) carried genes for at least one of the toxins, and 27 (60%) carried genes for at least one of the fimbrial adhesins and toxins. Fifty-one percent of strains that carried AIDA genes carried Stx2e genes, and 40% of strains that carried AIDA genes carried F18ab. The isolation rate of enterotoxigenic E. coli strain carrying genes for AIDA was 87%, and the isolation rate of Shiga toxin-producing E. coli strain carrying genes for AIDA was 49%. AIDA may represent an important virulence determinant in pigs with postweaning diarrhea or edema disease.
Identification of Canine Transmissible Venereal Tumor Cells Using in Situ Polymerase Chain Reaction and the Stable Sequence of the Long Interspersed Nuclear Element
Authors: Liao Kuang-Wen , Lin Zei-Yi , Pao Hai-Nie , Kam Sook-Yee , Wang Fun-In , Chu Rea-Min
Published: May 2011

Canine transmissible venereal tumor (CTVT) is a unique tumor that can be transplanted across the major histocompatibility complex (MHC) barrier by viable tumor cells. In dogs, CTVT grows progressively for a few months and then usually regresses spontaneously. A long interspersed nuclear element (LINE) insertion is found specifically and constantly in the 5′ end of the CTVT cell c-myc gene, outside the first exon. The rearranged LINE—c-myc gene sequence has been used with polymerase chain reaction (PCR) to diagnose CTVT. However, in CTVT cells, the total length of the inserted LINE gene is not constant. In this experiment, variation in the inserted LINE gene was studied to determine which parts of the LINE sequence can be used as primers to identify CTVT cells with in situ PCR (IS PCR). The LINE gene was inserted between the TATA boxes in the promoter region of c-myc. In CTVT cells, deletions of different lengths are frequent in this gene. However, the 550-bp segment at the 5′ end of the LINE—c-myc gene was stable. Thus, primers were designed to cover the stable 0.55-kb segment from the 5′ end outside the first exon of the c-myc gene to the 5′ end of LINE gene stable segment. With these primers and IS PCR, individual CTVT cells in formalin-fixed tissue sections and CTVT cultures were identified. Cells from other canine tumors were negative for this gene. In addition, the CTVT-specific, 0.55-kb segment was not found in any spindle-shaped cells from progressive or regressive phase CTVT. The IS PCR technique also did not detect any positive spindle-shaped cells in CTVT cell cultures. Thus, fibroblastic terminal differentiation is less likely to be a mechanism for spontaneous regression of CTVT cells.
Multiple Cutaneous Leiomyomas in the Perineum of a Horse
Authors: Bailey Keith L. , Kinsel Michael J. , Connell Kathy A.
Published: May 2011

Multiple cutaneous masses developed in the perineum of a 14-year-old Saddlebred stallion over a period of approximately 5 years. Clinically, the masses ranged in size from 3- to 9-mm diameter and were not ulcerated, painful, or pruritic. Three of the masses were surgically excised and submitted for microscopic evaluation. The masses were dome shaped to nodular, located in the superficial dermis, and composed of haphazardly arranged bundles of plump spindle-shaped cells. The tumor cells immunoreacted with monoclonal antibodies directed against desmin, muscle-specific actin, and smooth muscle actin, confirming a smooth muscle origin. Multiple cutaneous leiomyomas have not been reported previously in horses.
16S Ribosomal RNA Sequence—Based Identification of Veterinary Clinical Bacteria
Authors: Cai Hugh , Archambault Marie , Prescott John F.
Published: May 2011

This study evaluated 16S rRNA gene sequence analysis methods as tools for identification of 22 phenotypically difficult to identify veterinary clinical bacterial isolates in a veterinary diagnostic laboratory. The study compared 16S rRNA gene sequencing and conventional phenotypic identification methods. Using 16S rRNA full-gene sequencing, 95% (21/22) of the isolates were identified to the genus level and 86% (19/22) to the species level. The conventional or commercially available manual identification phenotypic characterization methods presumptively identified 91% (20/22) of the isolates to the genus level and 1 isolate to the species level. However, only 55% (12/22) or 4.5% (1/22) of the phenotypic identifications were correct at the genus or species level when they were compared with the 16S rRNA full-gene sequencing. This study also compared 16S rRNA full-gene and partial-gene sequencing. The results demonstrated that the best 16S rRNA gene—sequencing approach is full-gene sequencing because it gives the most precise species identification. Sequencing of the variable regions 1, 2, and 3 of the 16S rRNA gene could be used for tentative identification because the ability of this sequencing to identify bacteria to the genus level is similar to that of the 16S rRNA full-gene sequencing. This method identified only 14% (3/22) isolates differently to the species level compared with the 16S rRNA full gene sequence. Sequencing of the variable regions 7, 8, and 9 is not recommended because it gives more ambiguous identifications. The cost of a 16S RNA full-gene—sequencing analysis was Can $160 and Can $60 for a partial 16S rRNA gene sequence, i.e., sequencing of variable regions 1, 2, and 3 or variable regions 7, 8 and 9.
Valvular Endocarditis Associated with Helcococcus Ovis Infection in a Bovine
Authors: Post Karen W. , Rushton Steven D. , Billington Stephen J.
Published: May 2011

A 12-month-old Angus bull calf with a history of fever and lethargy of several weeks' duration was necropsied. Macroscopic findings included general dehydration, congestion, and edema within the craniodorsal lobes of the lung, multifocal ecchymotic hemorrhages on the dorsal epaxial and gluteal muscles, bloody ingesta within the gastrointestinal tract, and a 4- × 4- × 5-cm irregular plaque located on the right atrioventricular heart valve. Microscopically, there were focally extensive pulmonary alveolar infiltrates of histiocytes and neutrophils, large numbers of necrotic hypereosinophilic hepatocytes located within the centrilobular and midzonal regions of the liver, and, within the plaque from the right atrioventricular valve, a large mass formed by abundant laminated fibrin that contained numerous small multifocal aggregates of Gram-positive cocci. This report describes the first apparent isolation of Helcococcus ovis from cattle.
The Seroprevalence of Johne's Disease in Georgia Beef and Dairy Cull Cattle
Authors: Pence Mel , Baldwin Charles , Black C. Carter
Published: May 2011

Beef and dairy cattle serum samples, collected during 2000 at sale barns throughout Georgia, were obtained from the Georgia State Brucellosis Laboratory and were used to conduct a retrospective epidemiological study. Statistical samplings of 5,307 sera, from over 200,000 sera, were tested for antibodies to Mycobacterium avium ssp. paratuberculosis, (Johne's disease) using a commercial enzyme-linked immunosorbent assay test kit. An overall period seroprevalence in all classes of cattle tested was 4.73%. The period seroprevalence in dairy cattle was 9.58%, in beef cattle it was 3.95%, and in cattle of unknown breed it was 4.72%. It was concluded that the seroprevalence of Johne's disease in cull beef and dairy cattle in Georgia is economically significant.
Simple Capillary Electrophoretic Determination of Soluble Oxalate and Nitrate in Forage Grasses
Authors: Miyazaki Shigeru , Yamanaka Noriko , Guruge Keerthi Siri
Published: May 2011

A simple capillary electrophoretic method is described for the simultaneous determination of soluble oxalate and nitrate in forage grasses. Grass samples were ground and extracted with water. The extracts were filtered and submitted to capillary electrophoresis. Electrophoresis was performed in a 75 μm ×50 cm fused silica capillary with 30 mM sodium sulfate containing an electroosmotic flow modifier under constant voltage at —8 kV Separated oxalate and nitrate were detected with direct UV absorption at 214 nm. The present method can be used for routine monitoring of the concentration of soluble oxalate and nitrate in grasses.
Nested Multiplex RT-PCR for Detection and Differentiation of West Nile Virus and Eastern Equine Encephalomyelitis Virus in Brain Tissues
Authors: Johnson Donna J. , Ostlund Eileen N. , Schmitt Beverly J.
Published: May 2011

A traditional nested reverse transcription-polymerase chain reaction (RT-PCR) assay specific for eastern equine encephalomyelitis (EEE) virus was designed to multiplex with a previously described West Nile (WN) virus nested RT-PCR assay. Differentiation of EEE and WN was based on base pair size of the amplified product. One hundred fifty-seven mammalian and avian brain tissues were tested by EEE/WN nested multiplex RT-PCR, EEE nested RT-PCR, and WN nested RT-PCR, and results were compared with other diagnostic test results from the same animals. Serological and virus isolation testing confirmed the results of the multiplex PCR assay. When compared with cell culture virus isolation, the multiplex assay was shown to be more sensitive in detecting the presence of EEE or WN virus in brain tissues. The multiplex assay was shown to be sensitive and specific for North American EEE and WN and provided a rapid means of identifying both viruses in brain tissues. No apparent sacrifice in sensitivity was observed in the multiplex procedure compared with the individual EEE and WN nested RT-PCR assays. Data collected from an additional 485 multiplex RT-PCR tests conducted during the summer and fall of 2002 further support the validity of the procedure.