Systemic distribution of woodchuck hepatitis virus in the tissues of experimentally infected woodchucks

To better assess the extent of the tissue tropism of mammalian hepadnaviruses, 10 tissues from each of six woodchucks were examined for the presence and state of woodchuck hepatitis virus (WHV) nucleic acids 15 months after experimental WHV infection. The tissues examined were peripheral blood lymphocytes, lymph node, spleen, bone marrow, thymus, pancreas, kidney, ovary, testis, and liver. Tissue samples from three chronically infected animals and three animals with serologic patterns of recovery (serum: WHsAg-, anti-WHs+, anti-WHs+, WHV DNA-) from acute WHV infection were analyzed in parallel by in situ hybridization and Southern and Northern blot techniques. WHV nucleic acids were detected in several individual tissues from each animal examined, although not all tissues in every animal contained WHV. Substantial differences were observed among the various tissues and animals with respect to the frequency, level, and intratissue distribution of WHV nucleic acids, as well as the presence of different viral genomic forms. Active WHV DNA replication was present only in the liver and spleen of the chronically infected animals. No evidence of ongoing WHV DNA replication was found in any of the tissues from the recovered animals. WHV DNA was homogeneously distributed among all hepatocytes in the livers of the chronic carriers. By contrast, WHV DNA in all the extrahepatic tissues, and in the livers of the recovered animals, was detected only in scattered foci of cells.     

Detailed investigation of factors influencing amplification efficiency and allele drop-out in single cell PCR: implications for preimplantation genetic diagnosis

Preimplantation genetic diagnosis (PGD) of single gene disorders relies on PCR-based tests performed on single cells (polar bodies or blastomeres). Despite the use of increasingly robust protocols, allele drop-out (ADO; the failure to amplify one of the two alleles in a heterozygous cell) remains a significant problem for diagnosis using single cell PCR. In extreme cases ADO can affect >40% of amplifications and has already caused several PGD misdiagnoses. We suggest that an improved understanding of the origins of ADO will allow development of more reliable PCR assays. In this study we carefully varied reaction conditions in >3000 single cell amplifications, allowing factors influencing ADO rates to be identified. ADO was found to be affected by amplicon size, amount of DNA degradation, freezing and thawing, the PCR programme, and the number of cells simultaneously amplified. Factors found to have little or no affect on ADO were local DNA sequence, denaturing temperature (94 or 96 degrees C) and cell type. Consideration of the causal factors identified during this study should permit the design of PGD protocols that experience little ADO, thus improving the accuracy of PGD for single gene disorders.     

Individual T cells hold unexpected clues to the nature of anergy and memory

The primary interest of our laboratory is understanding how the signals that a T cell receives influence its behavior during an immune response. Recently, we have focused our attention on the examination of T cell responses at an individual cell level. This article outlines our approach, presents our initial findings, and discusses the implications of these findings relative to our current models of T cell activation.     

Validation of a brief screening instrument for substance abuse and mental illness in HIV-positive patients

BACKGROUND: Substance abuse (SA) and mental illness (MI) commonly co-occur with HIV infection in the United States and have important implications for clinical management of HIV/AIDS. Yet SA/MI often go untreated due in part to a lack of practical, validated screening tools. SETTING: HIV clinic in academic medical center. METHODS: The 16-item SA/MI Symptoms Screener (SAMISS) targets SA/MI in HIV-positive patients. Consecutive consenting HIV-positive patients completed the SAMISS and then a reference standard diagnostic tool, SCID, the Structured Clinical Interview for DSM-IV (Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition). RESULTS: Twenty percent of participants (29/148) had an SA diagnosis and 41% (59/143) had an MI diagnosis in the past year on the SCID; 48% (68/143) had 1 or both. Thirty-seven percent (55/148) screened positive for SA and 69% (99/143) screened positive for MI on the SAMISS. The SAMISS had 86% (95% CI: 68%-96%) sensitivity and 75% (66%-82%) specificity for SA and 95% (86%-99%) sensitivity and 49% (38%-60%) specificity for MI. Patients with SA were likely to show up as false positives for MI and vice versa. CONCLUSION: The SAMISS functioned well as a first-line screening tool for SA/MI in this HIV clinic population. It missed few cases and was easily incorporated into a busy clinical setting. Persons screening positive require a more rigorous confirmatory psychiatric evaluation.     

The Site of Vascular Response to the α-Toxin of Clostridium Perfringens Type A in Skeletal Muscle

The vascular response to the α-toxin of Clostridium perfringens type A, was observed topographically in the cremaster muscle of the rat, in terms of exudation, labelling of damaged vessels by circulating carbon, and in addition histologically for patency of the vascular plexus.It was confirmed that the permeability response is biphasic. The short-lived immediate phase corresponds to that of venular labelling, and the delayed phase reaches its peak rather later than the corresponding phase of capillary labelling. The intensity and extent of these responses are determined by the degree of injury, but their shape and timing, especially in the immediate phase, vary almost as consistently with the duration of exposure to circulating dye or carbon.After the standard dose of toxin, vascular patency is largely unaffected until 24 hours. Apparently irreversible vascular occlusion occurs rather earlier with larger doses. A three-fold reduction of the standard dose proportionately reduces both exudation and capillary labelling but leaves immediate venular labelling unaffected, suggesting that the latter is not dose dependent and therefore non-specific. Prolongation of moderate venular labelling into the middle of the delayed phase may occur at this dosage. Its absence after the standard dose suggests that delayed inhibition of venular reactivity may be occurring.Irregular labelling of venules and small veins persists throughout the delayed phase with doses 2·4 or more times the standard dose. A brief ultramicroscopic survey revealed appearances in both venules and capillaries at 1-2 hours after injury closely comparable to those which have been described for Cl. oedematiens toxin at 6-24 hours.In rats given carbon during the delayed phase, the effective vascular patency 10 minutes later includes half of the labelled capillaries up to 10 μm in diameter. This proportion is little affected by toxin dose, but intensely so when the carbon clearance time is increased, suggesting that such injured microvessels may be a major source of plasma protein exudation.     

Is fecundability associated with month of birth? An analysis of 19th and early 20th century family reconstitution data from The Netherlands

The relationship between fecundability and month of birth was investigated in a cohort of 1526 women who married between 1802 and 1929, using only women whose first marriage occurred before the age of 35 years. On the basis of their time to pregnancy (TTP, calculated as time between wedding and first birth minus gestational length), women were categorized into two groups: fecunds (TTP up to 12 months or prenuptial conceptions, n = 1348) and subfecunds (TTP >18 months, n = 118). By use of logistic regression, cosinor functions with a period of 1 year or 6 months and variable shift and amplitude were fitted through the monthly odds of subfecunds versus fecunds. The best fitting curve was unimodal, with a zenith in September (P = 0.13 for H0: no differences). Exclusion of childless women (n = 36, minimum follow-up 5 years) from the subfecunds led to a similar curve (P < 0.01), while childless women, as compared with fecunds, showed a birth distribution that was best represented with a bimodal curve with zeniths in January and July (P = 0.06). This study provides evidence for the existence of differences in fecundability by month of birth. The cause of this relationship is unclear, but may lie in a melatonin-dependent circannual variability of the quality of the oocyte.