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1.
OBJECTIVE: To identify subclinical Babesia gibsoni infection in American Pit Bull Terriers from the southeastern United States and to determine the genetic sequence of parasite DNA isolated from these dogs. DESIGN: Case series. ANIMALS: 33 American Pit Bull Terriers and 87 dogs of various other breeds. PROCEDURE: Blood smears were examined for microscopic evidence of the parasite, and DNA was extracted from blood samples and used in a polymerase chain reaction (PCR) assay designed to amplify the small subunit ribosomal RNA gene sequence of B. gibsoni. Amplification products of the expected size were sequenced, and sequences were compared with published sequences for B. gibsoni isolates. Hematocrit, platelet count, mean platelet volume, WBC count, and eosinophil count were compared between dogs with positive PCR assay results and dogs with negative results. RESULTS: Results of the PCR assay were positive for 18 of the 33 (55%) American Pit Bull Terriers, including all 10 dogs with microscopic evidence of parasitemia. Only 1 of these dogs was clinically ill at the time blood samples were collected. Results of microscopic evaluation of blood smears and of the PCR assay were negative for the 87 other dogs. Hematocrit and platelet count were significantly lower in dogs with positive PCR assay results than in dogs with negative results. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that American Pit Bull Terriers in the southeastern United States may be subclinically infected with B. gibsoni. However, subclinical infection was not identified in dogs of other breeds from the same geographic area.  相似文献   

2.
Polymerase chain reaction (PCR) assays for the immunoglobulin and T-cell receptor genes were utilized to determine phenotype and clonality from lymph node cytologic smears and peripheral blood lymphocytes from 10 dogs with lymphoma, before chemotherapy and during remission. Results were compared with those from 13 dogs with a cytologic diagnosis of lymph node hyperplasia. Clonality was identified in 7 of the lymphomas on the basis of either lymph node cytology or peripheral blood lymphocytes before treatment. No lymph node hyperplasia samples were clonal. In 6 of the dogs with lymphoma, clonality was demonstrated during clinical remission. Detection of PCR clonality during clinical remission is an effective means of identifying minimal residual disease in canine lymphoma and thus additional work is warranted to determine if molecular remission is prognostic or predictive for outcome in well-controlled and well-defined lymphoma subtypes.  相似文献   

3.
Abstract: A 1‐year‐old intact female miniature Dachshund was presented with hematochezia, vomiting, and diarrhea of more than 1‐week duration. An abdominal mass was palpated, which at exploratory surgery was found to be a 7‐cm‐long thickened section of ileum. The thickened ileum was resected. Impression smears revealed numerous small‐ to medium‐sized lymphocytes, with a smaller number of cells resembling Mott cells. The Mott‐like cells contained multiple pale vacuoles that were positive for periodic acid‐Schiff (PAS) in wet‐fixed smears, consistent with Russell bodies. Histologic evaluation of the surgically excised ileum revealed 2 populations of neoplastic lymphoid cells. The majority were uniform medium‐sized lymphocytes with hyperchromatic oval or round nuclei and inconspicuous nucleoli. The remaining cells resembled Mott cells, which contained several PAS‐positive eosinophilic globules in the cytoplasm, occasionally compressing the nucleus. The majority of neoplastic cells stained positively for vimentin, CD20, CD79a, and Pax‐5, but were negative for CD3 and lysozyme; 43.5% of cells stained positively for Ki‐67. The Mott cells were strongly positive for immunoglobulin but were negative for Pax‐5. Using electron microscopy, a homogenous substance of intermediate electron density was observed frequently in the cisternae of rough endoplasmic reticulum in the cytoplasm of the Mott cells, and rarely in the perinuclear cisternae of the lymphoid cells, corresponding to the site of immunoglobulin staining. Monoclonal rearrangement of immunoglobulin heavy‐chain (IgH) gene was observed by PCR testing for lymphocyte–antigen receptor rearrangement. The morphologic features, immunophenotype, and IgH gene rearrangement verified the lymphoid cells were neoplastic (mature cell type) and had a B‐cell phenotype, with evidence of immunoglobulin production and differentiation into Mott cells. This case was unusual because of the age of the dog and because most intestinal lymphomas are T‐cell phenotype. The Mott cell morphology also differed from typical mature B‐cell lymphoma types and may be a unique B‐cell lymphoma variant.  相似文献   

4.
We report on a study that evaluated the usefulness of PCR for the routine detection of Babesia equi in horses. The blood from a total of 105 horses comprising both sick and apparently healthy animals were examined for the presence of B. equi using both Wright-Giemsa-stained blood smears and PCR. Microscopic analysis of Giemsa-stained blood smears revealed 10/105 animals positive for Babesia, compared to 16/105 for the primary PCR and 36/105 for the nested PCR. Three of the 10 samples positive by Wright-Giemsa-stain were negative by PCR for B. equi. However, evidence is presented that these samples contained B. caballi and not B. equi. The Wright-Giemsa-stain was shown to identify Babesia in mostly clinically ill animals while the nested PCR detected the organism in a large number of apparently healthy animals. The results of this study suggest that the nested PCR is superior to both Wright-Giemsa-stained and primary PCR methods, and should be considered for the routine detection of B. equi in both healthy and clinically ill horses.  相似文献   

5.
In lymphoid neoplasia, molecular assays to confirm clonality rely on the fact that lymphoid cells normally contain DNA regions with unique sequences, resulting from recombination of the V, D, and J genes. The purpose of this study was to determine the utility of polymerase chain reaction (PCR) for antigen receptor rearrangements (PARR) for molecular staging and predicting prognosis in canine lymphoma. We hypothesized that the PARR assay would offer a sensitive method for detecting neoplastic cells in blood, and that the presence of such cells would be a negative prognostic finding compared with dogs with no detectable circulating tumor cells. Eighty-six patients with histologically or cytologically confirmed lymphoma were studied from initial diagnosis until death or euthanasia. All patients had PARR assays of a representative tumor-infiltrated lymph node and peripheral whole blood. Sixty-two patients had clonal rearrangements detected in the lymph node and were able to be staged by PARR. Seventeen patients (27%) had no detectable tumor in their blood and 45 (73%) were blood positive. Our findings showed that (1) PARR correlated with clinical stage in that the PARR assay was more likely to detect tumor cells in blood in stage 5 lymphomas, (2) PARR was more sensitive for detecting circulating tumor cells than visual assessment of blood or bone marrow because 80% of stage 3 lymphomas were blood-PARR-positive, and (3) PCR stage was not prognostic for disease-free interval (DFI) or survival.  相似文献   

6.
The aim of this study was to determine diameters of blood cells, differential counts of peripheral blood leukocytes, alpha-naphthyl acetate esterase (ANAE), acid phosphatase (ACP-ase) activity of some leukocyte types, and enzymatic positivity percentages of peripheral blood lymphocytes in two hedgehogs species, Hemiechinus auritus, the long-eared hedgehog, and Erinaceus concolor, the southern white-breasted hedgehog. Air-dried peripheral blood smears were stained with May-Grünwald-Giemsa stain. ANAE and ACP-ase were stained in glutaraldehyde-acetone-fixed smears. ANAE-positive lymphocytes displayed a dot-like positivity pattern characterized with 1-5 reddish brown cytoplasmic granules, whereas ACP-ase positive lymphocytes displayed a dot-like positivity pattern characterized with 1-3 pinkish cytoplasmic granules. Monocytes gave a diffuse and strong reaction while neutrophils displayed a weak positive reaction for ANAE and ACP-ase. No difference was observed in mean diameters of peripheral blood cells of these species. It was found that lymphocytes made up the majority (64.3% and 65.5%) of leukocytes, followed by neutrophils (23.9% and 23.3%), eosinophils (9.0% and 7.6%), monocytes (1.8% and 2.3%), and basophils (1.0% and 1.3%) in H. auritus and E. concolor, respectively. Mean ANAE positivity oflymphocytes was 36.6% and 51.3% and ACP-ase positivity was 32.1% and 37.5% for H. auritus and E. concolor, respectively. The ANAE positivity of lymphocytes in E. concolor was significantly (P < 0.05) higher than that of H. auritus.  相似文献   

7.
The purpose of this study was to determine the percentage of peripheral blood T lymphocytes and the localization of ANAE enzyme at the electron microscopic level in Van cats by using an alpha-naphtyl acetate esterase procedure. Peripheral blood samples taken from 20 Van cats were used. The percentage of ANAE positive lymphocytes was 83.0%. Neutrophilic gra-nulocytes gave a negative reaction, whereas monocytes, eosinophilic granulocytes showed a diffuse granular positivity. In the electron microscopic examination, ANAE positive reactions were seen in lysosomal granules found in lymphocytes.  相似文献   

8.
OBJECTIVE: To determine whether a polymerase chain reaction (PCR) assay could be used to detect Eperythrozoon wenyoni in the blood of cattle. DESIGN: Prospective study. ANIMALS: 95 cattle from various herds in Alabama and Georgia and 96 bulls enrolled in Auburn University's Alabama Beef Cattle Improvement Association Bull Test program. PROCEDURE: Blood samples were collected by means of venipuncture of the median caudal vein and submitted for a CBC and PCR assay. Blood smears were made immediately after blood collection and examined by means of light microscopy. RESULTS: Three of 95 cattle from herds in Alabama and Georgia and 5 of 96 bulls enrolled in the Bull Test program had positive PCR assay results. Organisms were seen in blood smears from only 5 of these 8 animals. Organisms were not seen in blood smears from any animals for which results of the PCR assay were negative. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that a PCR assay may be an effective method for detecting E wenyoni infection in cattle and that the PCR assay may be a more sensitive test than evaluation of blood smears.  相似文献   

9.
Eighteen cats surviving natural infection with Cytauxzoon felis were identified. All cats came from a limited geographic area in northwestern Arkansas and northeastern Oklahoma. Clinical signs in most cats were similar to those described for cytauxzoonosis; however, 4 cats were asymptomatic. All cases were initially diagnosed by microscopic identification of signet ring-shaped piroplasms in erythrocytes of peripheral blood smears. Four of 4 cats tested had detectable serum antibodies to C felis. Four different cats were positive by polymerase chain reaction (PCR). Partial sequencing of the PCR product from 1 cat revealed >99% homology with the reported sequence of C felis. Repeated examination of blood smears from 12 cats revealed that the erythroparasitemia was generally persistent for the duration of follow-up (3-154 days). Survival did not seem dependent on treatment, as only 1 cat was treated with a drug with potential antiprotozoal activity (imidocarb dipropionate), and 4 cats received no treatment. The findings of this study may indicate the existence of a less virulent strain of C felis.  相似文献   

10.
The purpose of this study was the comparison of the diagnostic sensitivity between buffy coat (BC), peripheral blood (PB), lymph node (LN), bone marrow (BM) and short-term culture (P-D) cytology that has been based on the detection of Ehrlichia canis morulae, in the acute phase of canine monocytic ehrlichiosis (CME). Their cellular localization, total numbers and microscopic differentials were also investigated. The highest sensitivities were achieved after evaluating 1000 oil immersion fields (OIFs) in BC (66%) and an equal number in LN (60.9%) smears, separately or together (74%). The morulae were more often detected into lymphocytes than monocytes. The highest total number of morulae (n=143) were found in P-D smears. Finally, to avoid false positive diagnoses, platelets, lymphocytic azurophilic granules, lymphoglandular bodies and phagocytosed nuclear material should not be confused with the morulae.  相似文献   

11.
Abstract: An 11‐year‐old spayed‐female German Shepherd dog was presented to the Veterinary Medical Teaching Hospital at Kansas State University with a history of weight loss, anorexia, depression, and lethargy for 2–3 weeks. Radiographic examination revealed a mass in the spleen and several round radiodense foci in the liver. CBC results included normocytic normochromic anemia, marked thrombocytopenia, and low numbers of neoplastic cells that frequently had cytoplasmic projections or blebs. A bone marrow aspirate contained about 80% neoplastic megakaryoblasts with the same microscopic features as those observed in peripheral blood. Using flow cytometry, cells of large size were identified in peripheral blood that expressed CD41/61, CD45, CD61, and CD62P (P‐selectin) and were negative for markers of T cells, B cells, monocyte/macrophages, and dendritic cells. Because of the poor prognosis, euthanasia and subsequently necropsy were performed. On histopathologic examination, neoplastic megakaryoblasts were identified in spleen, liver, mesenteric lymph node, and the pulmonary vasculature. Using immunohistochemistry, the neoplastic megakaryoblasts weakly expressed von Willebrand factor. Based on microscopic and immunophenotypic findings, a diagnosis of acute megakaryoblastic leukemia (AMegL) was made. To our knowledge, this is the first report of AMegL in a domestic animal in which immunophenotyping by flow cytometry and a panel of antibodies against CD41/61, CD61, and CD62P were used to support the diagnosis.  相似文献   

12.
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13.
Babesia equi and Babesia caballi are tick-borne haemoparasites that may cause babesiosis of Equidae. In southern Europe B. equi is enzootic and infections may occur asymptomatically and more frequently than those due to B. caballi. Complement fixation test (CFT) is the official serological test for the diagnosis of equine babesiosis, but it has low sensitivity during early and latent stages of the disease. With the aim of developing more sensitive and rapid direct diagnostic alternatives, PCR systems that amplified DNA targets of 664 or 659 bp regions of the 16S rRNA genes were designed and demonstrated to specifically detect the genomes of B. equi and B. caballi, respectively. An approximated parasitaemia of 0.000083% was detected by the PCR system for B. equi compared with reported limits of 0.001% for microscopic examination of stained blood smears, and up to 0.00025% for DNA probes. Although the sensitivity of the PCR system for B. caballi could not be estimated, samples with microscopically undetectable parasitaemia as well as those with 0.017% parasitised red blood cells were detected. DNA extracts of blood collected with EDTA as an anticoagulant from 23 horses from Portugal were tested with both PCR systems. Of these samples, 22 were positive for B. equi and 8 were positive for B. caballi with PCR tests and intraerythrocytic parasites were seen in all samples. Antibodies against both parasites were not detected by CFT in several cases, but in these cases the presence of either or both parasites was apparent by PCR tests. The PCR systems may be useful in the diagnosis of equine babesiosis covering a wider range of clinical disease, as useful adjuncts to serological, microscopic, and cultural methods, especially for the import and export testing of horses.  相似文献   

14.
A nested polymerase chain reaction (PCR) assay was developed using primers selected from the genomic DNA of Leishmania infantum and applied to the diagnosis of leishmaniosis in peripheral blood in dogs. Blood of 39 dogs of different breeds, all sampled in Catalonia (Spain), were tested for leishmaniosis by enzyme linked immunosorbent assay (ELISA), western blotting (WB) and peripheral blood mononuclear cell (PBMC) culture and nested PCR. Twenty negative controls (healthy dogs less than 1-year-old that had not been exposed to a sandfly season) were also studied. Nineteen of the 39 dogs studied were positive by ELISA and/or WB, and 18 of these had a positive PBMC nested PCR. PBMC nested PCR was negative in all the remaining animals that were negative by serological examination, including the 20 negative controls. Parasitological examination and nested PCR of bone marrow and lymph node aspirate from the 19 dogs positive by serological examination, were also positive. These results indicate that PBMC nested PCR is a sensitive and specific tool to diagnose leishmaniosis in dogs. The use of PBMC has the advantage over bone marrow and lymph node aspirates in that it is a less invasive sample.  相似文献   

15.
Tissues from 4 aborted polar foxes (3 samples of brain and 4 samples of liver) were selected for Toxoplasma gondii PCR assay. Positive results of serological tests of mothers and immunofluorescence test (IFT) of fetal organ smears were the criteria of sample selection. Five sets of primers designed from B1 gene and ITS1 sequences of T. gondii were used for detection of the parasite in fetal fox tissues. All used primer sets successfully amplified T. gondii DNA in PCR from organs which were positive by IFT. Single tube nested PCR also showed positive result from a sample negative by IFT, but this product was not confirmed. The studies showed usefullness of PCR for routine diagnosis of toxoplasmosis in carnivores.  相似文献   

16.
A 12‐year‐old castrated male mixed breed dog was presented with anorexia, lethargy, intermittent vomiting, diarrhea, and weight loss. Clinicopathologic and imaging abnormalities included pancytopenia, icterus, and splenomegaly with multiple minute hypoechogenic nodules. Bone marrow (BM) smears revealed 2.5% hemophagocytic macrophages. In addition, an increased number of small to intermediate lymphocytes (16.3%) and plasma cells (3.2%) were recognized in the BM smears. More than 80% of the lymphocytes contained multiple small intracytoplasmic magenta granules. Histopathologic findings of the spleen revealed hemophagocytosis. Large granular lymphocytes (LGLs) were not found on the liver cytology or splenic histopathology at this time. PCR for antigen receptor rearrangement (PARR) analysis showed a clonal reaction in the T‐cell receptor ? (TCR?) gene in the BM sample. The dog was diagnosed with hemophagocytic syndrome (HPS). The dog was maintained in good condition with immunosuppressive therapy. However, the dog developed hepatic LGL lymphoma 7 months later. At this time, PARR analysis showed a clonal TCR? gene rearrangement in the hepatic LGL lymphoma samples. The BM and liver sample clonal rearrangements showed 100% homology, indicating that the small to intermediate granular lymphocytes in the BM at the HPS stage had progressed to hepatic LGL lymphoma. To our knowledge, this is the first report of canine secondary HPS caused by the occurrence of a BM LGL lymphoma clone that progressed to hepatic LGL lymphoma.  相似文献   

17.
Sheep-associated malignant catarrhal fever (SA-MCF), a frequently fatal disease primarily of certain ruminants, is caused by ovine herpesvirus 2 (OvHV-2). Molecular diagnosis of SA-MCF in affected animals has relied on detection of OvHV-2 DNA using a nested PCR, which has significant potential for amplicon contamination as a routine method in diagnostic laboratories. In this report, a nonnested and a previously developed real-time PCR were validated for detection of OvHV-2 DNA in samples from clinically affected animals. Three sets of blood or tissue samples were collected: 1) 97 samples from 97 naturally affected animals with evidence of clinical SA-MCF; 2) 200 samples from 8 animals with experimentally induced SA-MCF; and 3) 100 samples from 100 animals without any evidence of clinical SA-MCF. Among 97 positive samples defined by nested PCR from clinically affected animals, 95 (98%) were positive by nonnested PCR and 93 (96%) were positive by real-time PCR, respectively. One hundred percent of the samples from the animals with experimentally induced MCF were positive by real-time PCR, while 99% were positive by nonnested PCR. Neither nonnested PCR nor real-time PCR yielded a positive result on any of the 100 nested PCR-negative samples from animals without evidence of clinical MCF. The data confirmed that both nonnested and real-time PCR maintained high specificity and sensitivity for the detection of OvHV-2 DNA in clinical samples.  相似文献   

18.
Porcine infectious anemia is a well-known disease that occurs worldwide and is caused by the unculturable hemotrophic bacterium Mycoplasma suis. The actual prevalence and impact of M. suis infections, however, remain fairly unknown. This study examined the prevalence of M. suis in post-weaning pigs by employing a quantitative real-time LightCycler PCR. M. suis infections were detected in 164 out of 1176 feeder pigs (20-30 kg; 13.9%) as well as on 79 out of 196 pig farms (40.3%). The comparison of PCR results with microscopic investigation of acridine-orange-stained blood smears revealed a considerably lower sensitivity of the microscopic method: only 35 out of 1176 blood smears were microscopically positive. The microscopic detection of M. suis was shown to be closely linked to the bacterial load in the blood. M. suis infection is associated with significantly decreased hematocrit, erythrocyte counts and hemoglobin concentrations as well as significantly higher bilirubin concentrations. Furthermore, M. suis blood loads were significantly associated with erythrocyte count, hematocrit, hemoglobin, glucose and iron concentrations indicating that high M. suis loads are connected with clinical anemia. In conclusion, this study has shown, that M. suis infections are often under-diagnosed in pig husbandry and can therefore lead to considerable economic profit losses in pig husbandry. Furthermore, our study has shown that the LightCycler PCR could be an appropriate tool for a sufficiently coherent identification of M. suis in latent carrier animals in view of introducing effective treatment and disease control measures.  相似文献   

19.
An 8-y-old spayed female Beagle dog was presented with peripheral lymphadenomegaly. Lymph node cytology and flow cytometry led to the diagnosis of large B-cell lymphoma (LBCL). We detected minimal percentages of LBCL cells in peripheral blood and bone marrow samples. However, a monomorphic population of neoplastic cells different from those found in the lymph node was found in the bone marrow. T-cell acute lymphoblastic leukemia was suspected based on flow cytometric immunophenotyping. PCR for antigen receptor rearrangement (PARR) revealed clonal rearrangement of both B-cell and T-cell receptors, and the presence of both neoplastic clones in the lymph node, peripheral blood, and bone marrow. The dog was treated with multi-agent chemotherapy but died 46 d following diagnosis. Tumor staging and patient classification are needed to accurately establish a prognosis and select the most appropriate therapeutic protocol.  相似文献   

20.
Lymphoblastic leukaemia, preceded by a significantly increasing percentage of prolymphocytes in peripheral blood smears for from 12 to 68 weeks before death was a feature of sheep which developed lymphosarcoma following inoculation with the Australian strain of bovine leucosis virus (BLV). Lymphocytosis and/or the appearance of immature cells were a reliable predictor of tumour formation in sheep, but not in cattle. There was a terminal lymphoblastic leukaemia in only 43 of 84 cattle with lymphosarcoma. Differences in the morphological appearance and glycogen content of the leukaemic lymphoblasts of sheep and cattle were observed. In spite of these differences the high frequency of lymphocytosis and lymphosarcoma in experimentally infected sheep suggests that they could be a useful model for studying the pathological and immunological responses to BLV infection.  相似文献   

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