Gas chromatographic determination of N-nitrosamines in beverages following automatic solid-phase extraction

A semiautomatic method for the determination of seven N-nitrosamines in beverages by gas chromatography with nitrogen-phosphorus detection is proposed. Beverage samples are aspirated into a solid-phase extraction module for preconcentration and cleanup. The influence of the experimental conditions was examined by using various sorbents among which LiChrolut EN was found to provide quantitative elution and the highest preconcentration factors of all. The proposed method is sensitive, with limits of detection between 7 and 33 ng/kg, and precise, with relative standard deviations from 4.3% to 6.0%. The recoveries of N-nitrosamines from beverage samples spiked with 0.5 or 1 microg/kg concentrations of these compounds ranged from 95% to 102%. The method was successfully applied to the determination of residues of the studied N-nitrosamines in beverages including beer, wine, liquor, whisky, cognac, rum, vodka, grape juice, cider, tonic water, and soft drinks. The analytes were only detected in beer samples, positives being confirmed by gas chromatography coupled with impact ionization mass spectrometry.     

Detection of Leishmania infantum DNA by real-time PCR in canine oral and conjunctival swabs and comparison with other diagnostic techniques

The use of non invasive sampling, such as collection of conjunctival swabs, as a diagnostic tool for the detection of Leishmania DNA is of interest. The purpose of this study was to evaluate the diagnostic utility of detecting Leishmania infection with the use of conjunctival swab samples in dogs living in a highly endemic area for leishmaniosis and to investigate, for the first time, the presence of Leishmania DNA in oral swabs in the same population. One hundred sixty-three dogs living outdoor and recruited in various provinces of Sicily were studied. Leishmania infantum indirect fluorescent antibody test (IFAT), delayed-type hypersensitivity reaction to leishmanin (DTH) and real-time PCR of blood (BL), lymph node (LN), conjunctival (CS) and oral swab (OS) samples were performed. The positive PCR percentages in LN, CS, OS and BL samples were: 24.5%, 22.1%, 8.7% and 5.5%, respectively. Serological and DTH positive percentages were 27.0% and 73.8%, respectively. Seropositive and LN-PCR positive dogs had a high likelihood to be positive by CS-PCR. The similar positive PCR percentages found in CS and LN samples suggest the use of CS-PCR as non-invasive alternative technique to LN-PCR for the detection of Leishmania infection in dogs. In addition, this study demonstrated, for the first time, the presence of Leishmania DNA in oral swabs in dogs.     

IMAGING DIAGNOSIS—MAGNETIC RESONANCE IMAGING FINDINGS OF AN INTRACRANIAL EPIDURAL TUBERCULOMA IN A DOG

Magnetic resonance (MR) imaging is highly sensitive for detecting tuberculomas in human patients but the specificity of the MR imaging features is low. Misdiagnosis with intracranial neoplasia is common, especially with dural‐based lesions or lesions located in the epidural space. We describe the MR imaging characteristics of an intracranial epidural tuberculoma caused by Mycobacterium tuberculosis infection in a dog. The intracranial mass and skull flat bone lysis and erosion are similar to those described in human caseating tuberculomas and can mimic intracranial neoplastic disease.     

Hematologic abnormalities and flow cytometric immunophenotyping results in dogs with hematopoietic neoplasia: 210 cases (2002–2006)

Background: Growing interest in veterinary oncohematology has facilitated the recent development and advancement of new techniques, such as flow cytometry, for immunophenotyping hematopoietic neoplasia in animals. Objective: The aim of this retrospective study was to characterize hematologic abnormalities and flow cytometric immunophenotyping (FCI) results in cases of hematopoietic neoplasia in dogs. Methods: Signalment, CBC data, and FCI results were obtained for 210 dogs with blood samples submitted to our laboratory. Immunophenotyping was carried out using an Epics XL‐MCL flow cytometer and a panel of 10 antibodies (CD45, CD3, CD4, CD8, CD79, CD21, CD14, CD34, CD41/61, CD61). The prevalence and severity of hematologic abnormalities was determined for the different types of hematopoietic neoplasms. Results: Based on cell morphology and phenotype, cases were classified as: acute lymphoblastic leukemia (ALL, n=51), acute myeloid leukemia (AML, n=33), chronic lymphocytic leukemia (CLL, n=61), and leukemic high‐grade lymphoma (L‐HGL, n=65). Most cases of ALL (47/51) and L‐HGL (41/65) had a B‐cell phenotype, while most cases of CLL (54/61) had a T‐cell phenotype, with a high prevalence of the large granular lymphocyte subtype (49/61). Anemia was found in 85% of all cases and was significantly more severe in ALL and AML compared with CLL and L‐HGL. Neutropenia was seen in 64–78% of acute leukemias (AML and ALL) in contrast to no cases of CLL and 11% of L‐HGL. Thrombocytopenia was seen in 88–90% of acute leukemias in contrast to 15% of CLL and 40% of L‐HGL. Thrombocytopenia was more prevalent (71% vs 22%) and significantly more severe in T‐cell vs B‐cell L‐HGL. Conclusion: A standard CBC is useful in suggesting the type of hemoproliferative disorder and may also help to predict the phenotype, especially in cases of L‐HGL.     

Influence of the Th2 immune response established by Nippostrongylus brasiliensis infection on the protection offered by different vaccines against Chlamydophila abortus infection

Chlamydophila abortus is the aetiological agent of enzootic abortion in small ruminants in which it infects the placenta to cause abortion during the last trimester of gestation. In a mouse model, a Th1 immune response involving IFN-γ production and CD8⁺ T cells is necessary for the infection to be resolved. The authors previously demonstrated that infection with Nippostrongylus brasiliensis, a rodent gastrointestinal nematode extensively used in experimental models to induce Th2 responses, alters the specific immune response against C. abortus infection, increasing bacterial multiplication in liver and reducing specific IFN-γ production. The aim of the present work was to clarify whether a Th2 immune response has any influence on the success of vaccination using both inactivated and attenuated vaccines. The results showed that the Th2 response established prior to vaccination did not influence the induction of protection offered by the vaccines. However, the effectiveness of this protective response can be altered, depending on the adjuvant employed in the inactivated vaccines, when the Th2 response is established after vaccination, just before challenge with C. abortus.     

Papular dermatitis due to Leishmania spp. infection in dogs with parasite-specific cellular immune responses

Papular dermatitis due to Leishmania spp. infection was diagnosed in three boxers and two Rottweilers with Leishmania-specific cellular immunity. Diagnosis was based on histological and immunohistochemical examination of papules in four dogs and on cytological examination in one dog. Serum protein electrophoresis was within reference ranges and low antibody levels to Leishmania infantum were detected. Delayed-type hypersensitivity (DTH) reaction to leishmanin was evaluated before treatment in three dogs with positive results. After meglumine antimoniate therapy for 3 to 4 weeks and allopurinol treatment for 6 to 10 months, all dogs were clinically normal, had positive DTH reactions to leishmanin and reduced antibody titres. In conclusion, we suggest that this previously unreported cutaneous presentation of canine leishmaniosis appears to be associated with specific immunocompetence and, consequently, with a favourable prognosis.