Evaluation of an automated magnetic bead-based DNA extraction and real-time PCR in fecal samples as a pre-screening test for detection of Echinococcus multilocularis and Echinococcus canadensis in coyotes

Efficient and sensitive diagnostic tools are essential for the study of the eco-epidemiology of Echinococcus species. We evaluated an automated magnetic bead-based DNA extraction commercial kit followed by qPCR (MB-qPCR), for the detection of Echinococcus multilocularis and Echinococcus canadensis in coyote (Canis latrans) fecal samples. The diagnostic sensitivity was determined by validating the method against the scraping, filtration, and counting technique (SFCT) for samples collected in Canada. From the 60 samples tested, 27 out of 31 SFCT positives samples for Echinococcus cestodes were positive in the MB-qPCR for E. multilocularis, with a sensitivity of 87.1% (95% CI 70.2 to 96.4%). Two samples were also positive for E. canadensis in the MB-qPCR and confirmed by morphological identification of adult worms. The agreement of the MB-qPCR and the SFCT was statistically significant with a kappa value of 0.67 (95% CI 0.48–0.85; p value < 0.001). The magnetic bead-based DNA extraction followed by qPCR proved to have a sensitivity comparable to the SFCT to detect E. multilocularis. Although the diagnostic sensitivity for E. canadensis was not estimated, MB-qPCR identified E. canadensis cases previously overlooked when using SFCT. We propose a combination of molecular and morphological identification using the MB-qPCR and the SFCT to detect both parasites, allowing for a more efficient large-scale surveillance, and detecting co-infections of Echinococcus species that can be difficult to identify when only based on morphology.

     

Brushing with a potassium nitrate dentifrice to reduce bleaching sensitivity

OBJECTIVE: This research systematically evaluated the use of a clinically proven desensitizing dentifrice prior to a bleaching regimen in a randomized, multi-center, parallel group, open label clinical study following Good Clinical Practice guidelines. METHODOLOGY: Fourteen dental offices in West Palm Beach, Florida participated in the study during April/May 2004. Fourteen days prior to bleaching, impressions and oral soft tissue assessments were performed, and patients were randomized to either a KNO3 plus fluoride dentifrice (Sensodyne Fresh Mint), or a standard fluoride dentifrice (Crest Regular), brushing 2x per day. On Day 14, patients returned to the dental office for their custom tray and the dispensation of a bleaching kit (Day White Excel 3; 9.5% hydrogen peroxide and KNO3). This was used daily according to the manufacturer's instructions for 30 minutes, and normal oral hygiene continued to be performed using the assigned toothbrush and dentifrice, brushing 2x per day. At the end of each bleaching day, patients answered diary questions about the occurrence and intensity of sensitivity. At the conclusion of the 14-day bleaching period (Day 28), patients returned to their dental office for re-examination, returning all products and diaries. Within seven days of completing the study, patients answered a telephone patient satisfaction survey. RESULTS: A total of 202 patients in fourteen (14) dental offices completed all aspects of the study and were used for the analysis. The professionally dispensed bleaching product provided an improvement of approximately 4.4 Vita shades, regardless of whether it was used with the KNO3 plus fluoride (Sensodyne) or a standard fluoride (Crest) dentifrice. The patient perception of increased sensitivity caused by the bleaching treatment was low but measurable. In the first week of the bleaching, significantly more patients using the KNO3 plus fluoride dentifrice were free from sensitivity (58%) than the standard fluoride dentifrice group (42%). During the 14-day bleaching treatment period, the KNO3 dentifrice patients experienced significantly more "sensitivity free days" (average = 10.1) compared to the standard fluoride dentifrice group (average = 8.6). CONCLUSION: The use of the KNO3 plus fluoride dentifrice (Sensodyne), two weeks prior to and throughout bleaching, may be a useful adjunct for the management of sensitivity caused by professionally dispensed bleaching products. With the bleaching-induced tooth sensitivity, those patients in the KNO3 plus fluoride toothpaste group were significantly more satisfied with their whitening experience and willing to repeat the bleaching treatment.     

The selective poly(ADP-ribose) polymerase-1(2) inhibitor, CEP-8983, increases the sensitivity of chemoresistant tumor cells to temozolomide and irinotecan but does not potentiate myelotoxicity

The effect of the potent and selective poly(ADP-ribose) (PAR) polymerase-1 [and PAR polymerase-2] inhibitor CEP-8983 on the ability to sensitize chemoresistant glioblastoma (RG2), rhabdomyosarcoma (RH18), neuroblastoma (NB1691), and colon carcinoma (HT29) tumor cells to temozolomide- and camptothecin-induced cytotoxicity, DNA damage, and G(2)-M arrest and on the potentiation of chemotherapy-induced myelotoxicity was evaluated using in vitro assays. In addition, the effect of the prodrug CEP-9722 in combination with temozolomide and/or irinotecan on PAR accumulation and tumor growth was also determined using glioblastoma and/or colon carcinoma xenografts relative to chemotherapy alone. CEP-8983 sensitized carcinoma cells to the growth-inhibitory effects of temozolomide and/or SN38 increased the fraction of and/or lengthened duration of time tumor cells accumulated in chemotherapy-induced G(2)-M arrest and sensitized tumor cells to chemotherapy-induced DNA damage and apoptosis. A granulocyte-macrophage colony-forming unit colony formation assay showed that coincubation of CEP-8983 with temozolomide or topotecan did not potentiate chemotherapy-associated myelotoxicity. CEP-9722 (136 mg/kg) administered with temozolomide (68 mg/kg for 5 days) or irinotecan (10 mg/kg for 5 days) inhibited significantly the growth of RG2 tumors (60%) or HT29 tumors (80%) compared with temozolomide or irinotecan monotherapy, respectively. In addition, CEP-9722 showed "stand alone" antitumor efficacy in these preclinical xenografts. In vivo biochemical efficacy studies showed that CEP-9722 attenuated PAR accumulation in glioma xenografts in a dose- and time-related manner. These data indicate that CEP-8983 and its prodrug are effective chemosensitizing agents when administered in combination with select chemotherapeutic agents against chemoresistant tumors.     

Translocation t(2;7)(p12;q21-22) with dysregulation of the CDK6 gene mapping to 7q21-22 in a non-Hodgkin's lymphoma with leukemia

BACKGROUND AND OBJECTIVES: A female patient presented with splenomegaly and lymphocytosis with atypical lymphoid cell morphology. We identified t(2;7)(p12;q21) prompting studies of the translocation breakpoint and its consequences on protein expression to confirm or otherwise the recently reported involvement of CDK6 and IG k genes in the t(2;7) leading to over-expression of CDK6 protein. DESIGN AND METHODS: A variety of clinical and laboratory techniques including cell marker, cytogenetic and histologic studies were applied in order to establish the diagnosis. Fluorescence in situ hybridization (FISH) and Southern blotting were used for mapping the translocation breakpoint and Western blotting for assessing protein expression. RESULTS: Immunophenotyping showed the presence of a B-cell population with strong expression of FMC7, CD22, CD79b, CD5 and k restricted surface immunoglobulins. Based on morphology and immunophenotypic markers the diagnosis of B-cell non-Hodgkin's lymphoma was made. Karyotyping revealed a clone with t(2;7)(p12;q21-22). Evidence for clonal evolution with additional abnormalities including a deletion of the TP53 was present. We established by FISH and Southern blotting that the breakpoint on 7q21-22 fell in a region 66kb telomeric to the previously reported breakpoint for the t(2;7) and was the same as that observed in a t(7;21). CDK6 protein was over-expressed. The patient received alkylating agents and splenectomy and is alive but the lymphocytosis persists with evidence of disease progression. INTERPRETATIONS AND CONCLUSIONS: We have demonstrated that CDK6 expression is dysregulated even when the breakpoint on 7q21-22 is located 66kb upstream from the coding region. Interestingly, the precise assignment of the lymphoma type in our case was not possible even when the splenic histology was analyzed.     

The relationship of body mass index and abdominal fat on the radiation dose received during routine computed tomographic imaging of the abdomen and pelvis

Purpose

To determine the relationship of increasing body mass index (BMI) and abdominal fat on the effective dose acquired from computed tomography (CT) abdomen and pelvis scans.

Methods

Over 6 months, dose-length product and total milliamp-seconds (mAs) from routine CT abdomen and pelvis scans of 100 patients were recorded. The scans were performed on a 64-slice CT scanner by using an automatic exposure control system. Effective dose (mSv) based on dose-length product, BMI, periumbilical fat thickness, and intra-abdominal fat were documented for each patient. BMI, periumbilical fat thickness, and intra-abdominal fat were compared with effective dose.

Results

Thirty-nine men and 61 women were included in the study (mean age, 56.3 years). The mean BMI was 26.2 kg/m². The mean effective dose was 10.3 mSv. The mean periumbilical fat thickness was 2.4 cm. Sixty-five patients had a small amount of intra-abdominal fat, and 35 had a large amount of intra-abdominal fat. The effective dose increased with increasing BMI (P < .001) and increasing amounts of intra-abdominal fat (P < .001). For every kilogram of weight, there is a 0.13 mSv increase in effective dose, which is equal to 6.5 chest radiographs per CT examination. For an increase in BMI by 5 kg/m², there is a 1.95 mSv increase in effective dose, which is equal to 97.5 chest radiographs per CT examination.

Conclusion

Increasing BMI and abdominal fat significantly increases the effective dose received from CT abdomen and pelvis scans.