Rapid detection and identification of human adenovirus species by adenoplex, a multiplex PCR-enzyme hybridization assay

Human adenoviruses (AdV) have been implicated in a wide variety of diseases and are ubiquitous in populations worldwide. These agents are of concern particularly in immunocompromised patients, children, and military recruits, resulting in severe disease or death. Clinical diagnosis of AdV is usually achieved through routine viral cell culture, which can take weeks for results. Immunofluorescence and enzyme-linked immunosorbent assay-based techniques are more timely but lack sensitivity. The ability to distinguish between the six different AdV species (A to F) is diagnostically relevant, as infections with specific AdV species are often associated with unique clinical outcomes and epidemiological features. Therefore, we developed a multiplex PCR-enzyme hybridization assay, the Adenoplex, using primers to the fiber gene that can simultaneously detect all six AdV species A through F in a single test. The limit of detection (LOD) based on the viral 50% tissue culture infective dose/ml for AdV A, B, C, D, E, and F was 10(-2), 10(-1), 10(-1), 10(-2), 10(-1), and 10(-2), respectively. Similarly, the LOD for the six DNA controls ranged from 10(2) to 10(3) copies/ml. Twelve common respiratory pathogens were tested with the Adenoplex, and no cross-reactivity was observed. We also validated our assay using clinical specimens spiked with different concentrations of AdV strains of each species type and tested by multiplex PCR and culture. The results demonstrated an overall sensitivity and specificity of Adenoplex of 100%. This assay can be completed in as few as 5 h and provides a rapid, specific, and sensitive method to detect and subtype AdV species A through F.     

Estrogen effects on object memory and cholinergic receptors in young and old female mice.

We investigated whether object recognition memory is modulated by estrogen in young (5 month) and aged (24 month) female C57Bl/6J mice, and if cholinergic muscarinic receptors might contribute to this response. Mice that were ovariectomized, or ovariectomized plus estradiol-treated three weeks before behavioral testing or quantitative autoradiography were compared to intact mice. Memory for a previously encountered object deteriorated significantly between 3 and 6h after initial exposure, regardless of animal age. In both young and aged mice, estradiol-treated mice showed significantly greater recall than did ovariectomized mice. In both age groups, the apparent number of [(3)H]pirenzepine/M(1)-like and [(3)H]AFDX384/M(2)-like muscarinic receptor binding sites was reduced in the basal forebrain as well as its projection areas following ovariectomy, but this decrease was not alleviated by estrogen. Aging poorly affected object memory, but reduced muscarinic binding in some cortical subregions and in the caudate nucleus. These findings suggest that estrogen effects on memory in C57Bl/6J mice are not due to changes in the number of muscarinic receptors.     

Trisomy 4 as the sole cytogenetic abnormality in a Waldenström macroglobulinemia

We report a case of Waldenstr?m macroglobulinemia with trisomy 4 as the sole cytogenetic abnormality. Trisomy 4 has been reported previously in Waldenstr?m macroglobulinemia, but only in conjunction with multiple chromosomal aberrations. Trisomy 4 has been reported in other hematologic malignancies including acute myeloid and lymphoid leukemias.     

Laterality of Body Focus and Digital Skin Temperature in Patients with Raynaud''s Disease

The purpose of the present study was to assess the relationship between lateralization of body image and right versus left vasomotor activity. Fifteen right-handed female patients suffering from idiopathic Raynaud's disease demonstrated a relationship between the extent of right lateralization of body image and bilateral digital skin temperature during a controlled temperature stress test. In addition, subjects showing a reliable right side awareness demonstrated more unilateral vasospastic attacks in their right hand than their left hand white subjects showing no clear right lateralization by body image reported more left hand attacks than right hand attacks. These results were taken as consistent with previous work on the relationship between skin conductance and lateralization of body image.     

Neonatal phenotype in Kabuki syndrome

The Kabuki syndrome is a well-established pattern of human malformation with readily recognizable features, however the diagnosis is rarely made in the newborn period. The purpose of this study was to determine if there exists a neonatal phenotype for this disorder. We ascertained 16 infants evaluated in the first 28 days of life by a dysmorphologist who subsequently received the diagnosis of Kabuki syndrome. The average age of initial evaluation was 8 days and the average age of diagnosis was 2 years 6 months. Based on these findings, it is suggested that the distinctive clinical phenotype seen in older patients is also evident in the newborn period.     

Changes in estrogen and progestin receptor binding resulting from retrochiasmatic knife cuts

Retrochiasmatic frontolateral knife cuts (FLC) or sham operations (Sham) were performed with a Halasz-type knife. All animals were primed with estrogen plus 0.5 mg progesterone (P) and tested for lordosis both before and after surgery. Two weeks after the last test they received estradiol (E₂) in Silastic capsules and were sacrificed 2 days later for determination of either nuclear estrogen receptors or cytosol progestin receptor binding in brain and pituitary (PIT). Rats which had received FLC showed significantly lower lordosis quotients relative to Shams, and relative to their own pre-surgery scores. Nuclear E₂-receptor binding was significantly reduced in the hypothalamus (HYPO) following FLC, but not in preoptic area (POA) or PIT. No changes in cytosol P-receptor binding were observed in HYPO, POA or PIT following FLC. Our results suggest a positive correlation between the number of hypothalamic E₂-receptors and the capacity to display lordosis, and emphasize the importance of anterolateral connections to the HYPO for the progesterone-induced facilitation of lordosis.     

Telomerase activity and proliferation index in aggressive mature B-cell lymphoma: comparison to germinal center phenotypic markers

Cell proliferation may be evaluated by various methods, including Ki-67 immunohistochemistry and measures of telomerase activity. Both methods would theoretically show comparable increases in a given case. To evaluate the relationship between these 2 markers of proliferation in aggressive mature B-cell lymphomas, 48 cases were studied. The study group included 5 cases of mantle cell lymphoma (MCL); 6 cases of Burkitt's/Burkitt's-like lymphoma (BL); 9 cases of follicular lymphoma, grade 3 (FLC); and 28 cases of diffuse large B-cell lymphoma (DLC). Telomerase activity was measured as total product generated (TPG) units, and TPG results for the aforementioned cases were compared to the TPG results for 10 cases of reactive follicular hyperplasia. An overlap in TPG scores between reactive cases and lymphoma cases was found. Significant differences in both log TPG (P = 0.0443) and Ki-67 (P = 0.0006) were seen in the different lymphoma types. A positive correlation between Ki-67 percentage and TPG score was identified in FLC (r = 0.9281; P = 0.0003), but a poor correlation between these 2 indicators was seen in the other lymphoma types. Cluster analysis identified distinct patterns for MCL, FLC, and BL, but heterogeneous patterns for DLC. Because increases in both Ki-67 proliferation and telomerase activity are reported in normal germinal centers (GCs), these tests were also evaluated for usefulness as markers of a GC cell phenotype. Among the FLC and DLC cases, features of a GC phenotype significantly correlated with increased Ki-67 percentage (P = 0.0152), but not with increased log TPG. An elevated log TPG correlated with CD10 expression, and elevated Ki-67 percentage correlated with both CD10 and BCL-6 expression. TPG level and Ki-67 percentage did not correlate with the presence of t(14;18) or BCL-2 protein expression. Although the proliferation patterns were fairly distinctive for MCL, FLC, and BL, these studies show that markers of cell proliferation do not by themselves,identify distinct subtypes of large cell lymphomas. With the exception of FLC, the tumors exhibited poor correlation between telomerase activity and Ki-67 proliferation index. These tests did show some correlation with expression of GC cell phenotypic markers, however.     

Ultrastructural Morphometric Study of Follicular Center Lymphocytes: II. Analyses of Cleaved-Cell Populations Do Not Support the Lukes-Collins' Concept

Ultrastructural morphometric analysis was carried out on six cases of lymph node biopsies with reactive hyperplasia to establish the frequency and depth of invaginations in nuclear profiles situated in the mantle zones and follicular centers. The frequency distribution of the depth of invaginations was similar in nuclear profiles whether in the small lymphocytes of mantle zones or the small, partially transformed (centrocytes) and fully transformed (centroblasts) lymphocytes of follicular centers. Invaginated and cleaved lymphocytes were not confined to the partially transformed (centrocytic) lymphocytes of follicular centers, and nuclear profiles with invaginations bore no resemblance to those depicted in the Lukes-Collins model. A considerable proportion of mantle zone lymphocyte nuclear profiles had invaginations (ranging from 7.5% to 53.6%) and there was no difference between the frequency of deep indentations or clefts in mantle zone lymphocytes (8.1 ± 5.4%) and the small unstimulated (9.3 ± 5.3%) and partially transformed (8.4 ± 1.4%) lymphocytes in follicular centers. Computer modeling of stylized nuclei with conical indentations indicated that all lymphocytic nuclei likely have multiple invaginations or groove-like creases.